TW202500169A - Compositions and methods for inhibiting expression of complement factor b (cfb) - Google Patents

Abstract

Compositions and methods useful for inhibiting the expression of complement factor B (CFB) gene and for treating CFB-associated diseases and conditions are provided. Provided are CFB dsRNA agents, CFB antisense polynucleotide agents, compositions comprising CFB dsRNA agents, and compositions comprising CFB antisense polynucleotide agents useful for inhibiting CFB expression in cells and subjects.

Description

Compositions and methods for inhibiting complement factor B (CFB) expression

The present invention relates in part to compositions and methods useful for inhibiting expression of the complement factor B (CFB) gene.

Complements were first discovered in the 1890s when they were found to aid or "complement" the antibacterial activity of heat-stable antibodies present in normal serum. The complement system or pathway is part of the host's innate immune system that defends against invading pathogens. It is primarily composed of more than 30 proteins that are either present in the blood as soluble proteins or as membrane-associated proteins.

Three major pathways of complement activation have been recognized, termed the classical pathway, the alternative pathway, and the lectin pathway. Activation of complements results in a sequential cascade of enzymatic reactions, termed the complement activation pathway, leading to the formation of the potent allergic toxins C3a and C5a, which trigger a variety of physiological responses ranging from chemoattraction to apoptosis. Initially, complements were thought to play an important role in innate immunity, generating a powerful and rapid response to invading pathogens. However, it has recently become increasingly apparent that complements also play an important role in adaptive immunity involving T cells and B cells, aiding in the elimination of pathogens, maintaining immunological memory to prevent reinvasion by pathogens, and participating in a variety of human pathological conditions.

Functionally, complement activation occurs natively at low levels (spontaneous cleavage of C3 to produce C3a and C3b) and is enhanced in the presence of microorganisms by an enzyme cascade that converts the inactive form of the enzyme (the zymogen) into its active counterpart. One type of C3 convertase is a complex of C3b and complement factor B (CFB, Factor B). Once formed, the C3 convertase can convert large amounts of C3 into its cleavage products, C3a and C3b, in a short period of time. The specific C3 convertase is a complex of C3b and Factor B that was originally described in the context of the alternative pathway but may also be formed in the context of the other two pathways. In the alternative pathway, Factor B is also a component of the C5 convertase, a complex that converts C5 (a downstream component of the pathway) into its active form.

Complement factor B (also called CFB or "factor B") is involved in activating the alternative pathway. Binding of CFB to C3b (e.g., on the cell surface) renders CFB susceptible to cleavage by factor D, forming the serine protease C3Bb, itself a C3 convertase, leading to an amplification loop of C3 activation. CFB is synthesized primarily in the liver, with lower levels also occurring in several extrahepatic sites.

Several diseases are associated with aberrant acquisition or genetic activation of complement pathways and abnormal or overexpression of CFB. For example, C3 glomerulopathy, systemic lupus erythematosus (SLE), lupus nephritis, IgA nephropathy, diabetic nephropathy, polycystic nephropathy, membranous nephropathy, age-related macular degeneration, atypical uremic hemolytic syndrome, thrombotic microangiopathy, myasthenia gravis, ischemia and reperfusion injury, paroxysmal nocturnal hemoglobinuria, rheumatoid arthritis, immune complex-mediated glomerulonephritis (IC-mediated GN), post-infectious glomerulonephritis (PIGN), ischemia/reperfusion injury, antineutrophil cytoplasmic autoantibody-associated vasculitis (ANCA-AV), dysbacteriosis periodontitis, malaria anemia, and hyperlipidemia.

Currently, there are few treatments for diseases, conditions, and syndromes mediated by the complement system. One of these is the monoclonal humanized antibody eculizumab. Although eculizumab has been shown to be effective in treating paroxysmal nocturnal hemoglobinuria (PNH), atypical uremic hemolytic syndrome (aHUS), and myasthenia gravis, and is currently being evaluated in clinical trials for the treatment of other complement-related diseases, eculizumab therapy requires high-dose infusions every week followed by maintenance infusions every two weeks, which is costly. Therefore, there is a high need for medical treatments for complement-mediated or related diseases. C3 is a key factor in the activation of the complement pathway. Therefore, inhibiting the expression of factors involved in C3 activation, such as CFB, provides a promising therapeutic strategy for many complement-mediated diseases. Targeting CFB expression or activity by means of antisense oligonucleotides, double-stranded siRNA, or small molecule inhibitors against CFB has been proposed as a potential therapeutic strategy for the treatment of various complement-mediated diseases.

Thus, the CFB RNAi agents disclosed herein are useful for treating diseases, disorders and conditions associated with complement activation, for example, through activation of complement factor B activity.

In general, the present disclosure features novel CFB gene-specific RNAi agents, compositions comprising the CFB RNAi agents, and methods of inhibiting CFB gene expression in vitro and/or in vivo using the CFB RNAi agents and compositions comprising the CFB RNAi agents described herein. The CFB RNAi agents described herein can selectively and effectively reduce, inhibit, or silence the expression of a CFB gene in a subject (e.g., a human or animal subject).

According to one aspect of the present invention, a double-stranded ribonucleic acid (dsRNA) agent for inhibiting CFB expression is provided, wherein the dsRNA agent comprises a sense chain and an antisense chain, wherein the sense chain comprises at least 15 consecutive nucleotides that differ from the nucleotide sequence of SEQ ID NO: 1, 3 or 5 by no more than 1, 2 or 3 nucleotides, and the antisense chain comprises at least 15 consecutive nucleotides that differ from the nucleotide sequence of SEQ ID NO: 2, 4 or 6 by no more than 1, 2 or 3 nucleotides, wherein the sense chain and the antisense chain can partially complement each other, substantially complement each other or completely complement each other.

In some embodiments, the dsRNA agent comprises a sense strand and an antisense strand that form a duplex region, wherein the antisense strand comprises a region complementary to a CFB RNA transcript comprising at least 15 consecutive nucleotides that differ by no more than 1, 2, or 3 nucleotides from any one of the antisense sequences listed in Tables 1-3. In some embodiments, the dsRNA agent comprises a sense strand and an antisense strand that form a duplex region, wherein the antisense strand comprises a region complementary to a CFB RNA transcript comprising at least 15 consecutive nucleotides from any one of the antisense sequences listed in any one of Tables 1-3.

In some embodiments, the dsRNA agent comprises a sense strand and an antisense strand, wherein nucleotide positions 2 to 18 of the antisense strand comprise a region complementary to a CFB RNA transcript, wherein the complementary region comprises at least 15, 16, 17, 18, 19, 20, or 21 consecutive nucleotides that differ by 0, 1, 2, or 3 nucleotides from one of the antisense sequences listed in one of Tables 1-3, and optionally comprises a targeting ligand.

In some embodiments, a double-stranded ribonucleic acid (dsRNA) agent for inhibiting CFB expression is provided, wherein the dsRNA agent comprises a sense strand and an antisense strand, wherein the sense strand comprises at least 15, 16, 17, 18, 19, 20, or 21 consecutive nucleotides that differ by 0, 1, 2, or 3 nucleotides from any one of nucleotides 483-513, 486-516, 491-521, 483-521, 513-543, 987-1017, 989-1019, 1317-1347, 2237-2267, 2439-2469 in SEQ ID NO: 1, and the antisense ... The corresponding nucleotide sequences in NO:2 differ by 0, 1, 2 or 3 nucleotides in at least 15, 16, 17, 18, 19, 20 or 21 consecutive nucleotides, wherein the sense strand and the antisense strand may partially, substantially or completely complement each other.

In some embodiments, a double-stranded ribonucleic acid (dsRNA) agent for inhibiting CFB expression is provided, wherein the sense strand comprises at least 15, 16, 17, 18, 19, 20 or 21 consecutive nucleotides that differ from any one of nucleotides 488-508, 491-511, 496-516, 488-516, 518-538, 992-1012, 994-1014, 1322-1342, 2242-2262, 2444-2464 in SEQ ID NO: 1 by 0, 1, 2 or 3 nucleotides, and the antisense strand comprises at least 15, 16, 17, 18, 19, 20 or 21 consecutive nucleotides that differ from the corresponding nucleotide sequence in SEQ ID NO: 2 by 0, 1, 2 or 3 nucleotides.

In some embodiments, a double-stranded ribonucleic acid (dsRNA) agent for inhibiting CFB expression is provided, wherein the sense strand comprises at least 15, 16, 17, 18 or 19 consecutive nucleotides that differ by 0, 1, 2 or 3 nucleotides from any one of nucleotides 490-508, 493-511, 498-516, 490-516, 520-538, 994-1012, 996-1014, 1324-1342, 2244-2262, 2446-2464 in SEQ ID NO: 1, and the antisense strand comprises at least 15, 16, 17, 18 or 19 consecutive nucleotides that differ by 0, 1, 2, 3 nucleotides from the corresponding nucleotide sequence from SEQ ID NO: 2.

In some embodiments, a double-stranded ribonucleic acid (dsRNA) agent for inhibiting CFB expression is provided, wherein the sense strand comprises at least 15, 16, 17, 18 or 19 consecutive nucleotides that differ by 0, 1, 2 or 3 nucleotides from any one of nucleotides 489-507, 492-510, 497-515, 489-515, 519-537, 993-1011, 995-1013, 1323-1341, 2243-2261, 2445-2463 in SEQ ID NO: 1, and the antisense strand comprises at least 15, 16, 17, 18 or 19 consecutive nucleotides that differ by 0, 1, 2, 3 nucleotides from the corresponding nucleotide sequence from SEQ ID NO: 2.

In some embodiments, the CFB RNA transcript is SEQ ID NO:1.

In some embodiments, the antisense strand of the dsRNA agent is at least substantially complementary to one of the target regions of SEQ ID NO: 1 and is provided in any of Tables 1-3. In some embodiments, the antisense strand of the dsRNA agent is fully complementary to any of the target regions of SEQ ID NO: 1 and is provided in any of Tables 1-3. In some embodiments, the dsRNA agent comprises a sense strand sequence listed in any of Tables 1-3, wherein the sense strand sequence is at least substantially complementary to the antisense strand sequence in the dsRNA agent. In certain embodiments, the dsRNA agent comprises a sense strand sequence listed in any of Tables 1-3, wherein the sense strand sequence is fully complementary to the antisense strand sequence in the dsRNA agent. In some embodiments, the dsRNA agent comprises an antisense strand sequence listed in any of Tables 1-3. In some embodiments, the dsRNA agent comprises a sequence listed as a duplex sequence in any one of Tables 1-3.

In some embodiments, the sense and antisense strands of a dsRNA agent can be partially, substantially, or completely complementary to each other.

In some embodiments, the dsRNA reagent includes at least one modified nucleotide. In certain embodiments, all or substantially all nucleotides of the antisense strand are modified nucleotides. In some embodiments, at least one modified nucleotide includes: 2'-O-methyl nucleotides, 2'-fluoro nucleotides, 2'-deoxy nucleotides, 2'3'-seco nucleotide mimetics, locked nucleotides, unblocked nucleic acid nucleotides (UNA), glycol nucleic acid nucleotides (GNA), 2'-F-arabinose nucleotides, 2'-methoxyethyl nucleotides, debasic nucleotides, ribitol, reverse nucleotides, reverse debasic nucleotides, reverse 2'-OMe nucleotides, reverse 2'-deoxy nucleotides, isomannitol nucleotides, 2'-amino modified nucleotides, 2'-alkyl modified nucleotides, morpholino nucleotides and 3'-OMe nucleotides, nucleotides comprising a 5'-phosphorothioate group, or a terminal nucleotide linked to a cholesterol derivative or a dodecanoic acid bisdecylamide group, a 2'-amino modified nucleotide, a phosphoramidite, or a nucleotide comprising an unnatural base.

In some embodiments, the dsRNA agent comprises an E-vinylphosphonate nucleotide at the 5' end of the guide strand.

In some embodiments, the dsRNA agent comprises at least one phosphorothioate nucleoside interlink. In some embodiments, the sense chain comprises at least one phosphorothioate nucleoside interlink. In some embodiments, the antisense chain comprises at least one phosphorothioate nucleoside interlink. In some embodiments, the sense chain comprises 1, 2, 3, 4, 5 or 6 phosphorothioate nucleoside interlinks. In some embodiments, the antisense chain comprises 1, 2, 3, 4, 5 or 6 phosphorothioate nucleoside interlinks. In some embodiments, the 5' end of the antisense chain comprises 2 phosphorothioate nucleoside interlinks. In some embodiments, the 3' end of the antisense chain comprises 2 phosphorothioate nucleoside interlinks. In some embodiments, the 5' end of the antisense strand and the 3' end of the antisense strand independently comprise two phosphorothioate internucleoside bonds.

In some embodiments, all or substantially all nucleotides of the sense strand and the antisense strand are modified nucleotides. In some embodiments, the antisense strand comprises 15 or more modified nucleotides independently selected from 2'-O-methyl nucleotides, 2'-fluoro nucleotides and UNA modified nucleotides, wherein less than 6 modified nucleotides are 2'-fluoro nucleotides. In some embodiments, the antisense strand comprises 3 or 5 2'-fluoro nucleotides, preferably, the antisense strand comprises 5 2'-fluoro nucleotides. In some embodiments, the sense strand comprises 15 or more modified nucleotides independently selected from 2'-O-methyl nucleotides and 2'-fluoro nucleotides, wherein less than 4 modified nucleotides are 2'-fluoro nucleotides. In certain embodiments, the sense strand comprises 3 2'-fluoro nucleotides. In some embodiments, the antisense strand comprises 15 or more modified nucleotides independently selected from 2'-O-methyl nucleotides and 2'-fluoro nucleotides, wherein at least 14 modified nucleotides are 2'-O-methyl nucleotides and are located at positions 2, 5, 7, 11, 12, 14, 16 and/or 18, counting from the first matching position at the 5' end of the antisense strand, are independently 2'-fluoro nucleotides. In some embodiments, the antisense strand comprises at least one UNA modified nucleotide and 5 2'-fluoro nucleotides. In some embodiments, the antisense strand comprises one UNA modified nucleotide at position 7 and 5 2'-fluoro nucleotides at positions 2, 5, 12, 14 and 16, counting from the first matching position at the 5' end, and the rest are 2'-O-methyl nucleotides. In some embodiments, the antisense strand comprises one UNA modified nucleotide at position 7 and five 2'-fluoro nucleotides at positions 2, 5, 12, 14 and 18, counting from the first matching position at the 5' end, and the rest are 2'-O-methyl nucleotides. In some embodiments, the antisense strand comprises one UNA modified nucleotide at position 7 and five 2'-fluoro nucleotides at positions 2, 5, 11, 14 and 16, counting from the first matching position at the 5' end, and the rest are 2'-O-methyl nucleotides. In some embodiments, the antisense strand comprises five 2'-fluoro nucleotides at positions 2, 7, 12, 14 and 16, counting from the first matching position at the 5' end, and the rest are 2'-O-methyl nucleotides. In some embodiments, the antisense strand comprises 5 2'-fluoro nucleotides at positions 2, 7, 11, 14, and 16, counting from the first matching position at the 5' end, and the remaining 2'-O-methyl nucleotides. In some embodiments, the antisense strand comprises 5 2'-fluoro nucleotides at positions 2, 5, 12, 14, and 16, counting from the first matching position at the 5' end, and the remaining 2'-O-methyl nucleotides. In some embodiments, the antisense strand comprises 5 2'-fluoro nucleotides at positions 2, 5, 12, 14, and 18, counting from the first matching position at the 5' end, and the remaining 2'-O-methyl nucleotides. In some embodiments, the sense strand comprises 15 or more modified nucleotides independently selected from 2'-O-methyl nucleotides and 2'-fluoro nucleotides, preferably, wherein at least 18 of the modified nucleotides are 2'-O-methyl nucleotides and the nucleotides at positions 9, 11 and/or 13, counting from the first matching position at the 3' end of the sense strand, are 2'-fluoro nucleotides. In some embodiments, the sense strand comprises at least 18 modified nucleotides that are 2'-O-methyl nucleotides, and the nucleotides at positions 8, 11 and/or 13, counting from the first matching position at the 3' end of the sense strand, are 2'-fluoro nucleotides. In some embodiments, the modified sense strand is a modified sense strand sequence listed in one of Tables 2-3. In some embodiments, the modified antisense strand is a modified antisense strand sequence listed in one of Tables 2-3.

In some embodiments, the dsRNA reagent includes at least one modified nucleotide and further includes one or more targeting groups or linking groups. In some embodiments, the one or more targeting groups or linking groups are conjugated to a sense chain. In some embodiments, the targeting group or linking group includes N-acetylgalactosamine (GalNAc).

In some embodiments, the targeting group has the following structure: Formula 1; n'' is independently selected from 1 or 2.

In some embodiments, the targeting group has the following structure: GLO-1 GLS-1* GLO-2 GLS-2* GLO-3 GLS-3* GLO-4 GLS-4* GLO-5 GLS-5* GLO-6 GLS-6* GLO-7 GLS-7* GLO-8 GLS-8* GLO-9 GLS-9* GLO-10 GLS-10* GLO-11 GLS-11* GLO-12 GLS-12* GLO-13 GLS-13* GLO-14 GLS-14* GLO-15 GLS-15* GLO-16 GLS-16*.

In certain embodiments, the dsRNA agent comprises a targeting group conjugated to the 5'-end of the sense strand. In certain embodiments, the dsRNA agent comprises a targeting group conjugated to the 3'-end of the sense strand.

In some embodiments, the antisense strand comprises an inverted abatic residue at the 3'-terminus.

In some embodiments, the sense chain comprises one or two reverse abatic residues and/or one or two imann residues at the 3' or/and 5' termini. In some embodiments, each end of the sense chain comprises a reverse abatic residue. In some embodiments, each end of the sense chain comprises an imann residue. In some embodiments, the 5' terminus of the sense chain comprises a reverse abatic residue or an imann residue, wherein the reverse abatic residue or the imann residue is linked to the adjacent nucleotide at the 5' terminus of the nucleotide sequence of the sense chain via a phosphorothioate bond. In some embodiments, the sense chain further comprises a targeting group connected to a reverse abaci group or imann residue at the 5' end of the sense chain, wherein the targeting group is connected to an adjacent reverse abaci group or imann residue via a phosphorothioate bond, and optionally the targeting group is N-acetylgalactosamine (GalNAc). In some embodiments, a reverse abaci group is included at the 5' end of the sense chain, wherein the reverse abaci group is connected to an adjacent nucleotide at the 5' end of the nucleotide sequence of the sense chain via a phosphorothioate bond. In certain embodiments, the sense chain further comprises a targeting group linked to the reverse abasic residue at the 5' end of the sense chain, wherein the targeting group is linked to the adjacent reverse abasic residue via a phosphorothioate bond, and optionally the targeting group is N-acetylgalactosamine (GalNAc), and the length of each chain is independently 21 nucleotides.

In some embodiments, the dsRNA agent has two blunt ends. In some embodiments, at least one strand comprises a 3' overhang of at least 1 nucleotide. In some embodiments, at least one strand comprises a 3' overhang of at least 2 nucleotides.

In some embodiments, the dsRNA comprises a duplex selected from AV02373, AV02375, AV02379, AV02388, AV02411, AV02464, AV02554, AV02584, AV06327, AV06328, AV06329, and wherein the duplex optionally includes a targeting ligand. In some embodiments, the dsRNA comprises a duplex selected from AD01093, AD01093-1, AD01093-2, AD01094, AD01096, AD01393, AD01393-1, AD01396, AD01399, AD01412, AD01420, AD01420-1.

According to another aspect of the present invention, a double-stranded ribonucleic acid (dsRNA) agent for inhibiting CFB expression is provided, wherein the dsRNA agent comprises a sense strand and an antisense strand, wherein the sense strand is complementary to the antisense strand, wherein the antisense strand comprises a region complementary to a portion of a CFB RNA transcript, wherein each strand is about 15 to about 30 nucleotides in length, wherein the sense strand comprises a sequence that can be represented by formula (I): 5′-(N′ _L ) _n′ N′ _L N′ _L N′ _L N′ _L N′ _F N′ L _N ′ _F N′ _L N′ _N1 N′ _N2 N′ _L N′ _L N′ L N′ _L N _{′ L (} N′ _L ) _m′ -3′ (I); wherein: each N′ _F represents a 2′-fluorine-modified nucleotide; each N′ _N1 and _N′N2 independently represent modified or unmodified nucleotides; each _N′L independently represents a modified or unmodified nucleotide, but does not represent a 2'-fluoro-modified nucleotide, and m′ and n′ are each independently an integer from 0 to 7.

In some embodiments, _N'N1 and _N'N2 contain only one 2'-fluorine modified nucleotide.

In some embodiments, N′ and _N1 independently represent 2'-fluoro modified nucleotides.

In some embodiments, N′ and _N2 independently represent 2′-fluoro modified nucleotides.

In some embodiments, m' is 2 and n' is 4, or m' is 2 and n' is 2. In some embodiments, m' is 1 and n' is 4, or m' is 1 and n' is 2. In some embodiments, m' is 0 and n' is 4, or m' is 0 and n' is 2.

In some embodiments, the dsRNA agent includes a targeting group conjugated to the 5'-end of the sense chain, preferably, the targeting group is selected from any one of the aforementioned GLO-1 to GLO-16 and GLS-1* to GLS-16*, more preferably, the targeting group is the aforementioned GLS-15*. In certain embodiments, the dsRNA agent includes a targeting group conjugated to the 3'-end of the sense chain. In certain embodiments, the antisense chain includes a reverse abacillary residue at the 3'-end. In certain embodiments, the sense chain includes one or two reverse abacillary residues and/or one or two imann residues at the 3' or/and 5' ends. In certain embodiments, each 3' and 5' end of the sense chain independently includes a reverse abacillary residue. In certain embodiments, each 3' and 5' end of the sense chain independently comprises an imann residue. In certain embodiments, the sense chain includes two inverted abatic residues at the 3' and 5' ends, and any residue at the 3' or 5' end is further conjugated to a targeting group, which is preferably the aforementioned GLS-15*. In certain embodiments, the sense chain includes two imann residues at the 3' and 5' ends, and any residue at the 3' or 5' end is further conjugated to a targeting group, which is preferably the aforementioned GLS-15*. In certain embodiments, each end of the sense strand comprises a reverse abaci or imann residue at the 5' end of the sense strand, wherein the reverse abaci or imann residue is linked to the adjacent nucleotide at the 5' end of the nucleotide sequence of the sense strand via a phosphorothioate bond. In certain embodiments, the sense strand further comprises a targeting group linked to the reverse abaci or imann residue at the 5' end of the sense strand, wherein the targeting group is linked to the adjacent reverse abaci or imann residue via a phosphorothioate bond, and optionally the targeting group is N-acetylgalactosamine (GalNAc). In some embodiments, the 5' end of the sense strand includes a reverse abatic residue, wherein the reverse abatic residue is linked to the adjacent nucleotide at the 5' end of the nucleotide sequence of the sense strand through a phosphorothioate bond. In some embodiments, the sense strand further includes a targeting group linked to the reverse abatic residue at the 5' end of the sense strand, wherein the targeting group is linked to the adjacent reverse abatic residue through a phosphorothioate bond, and optionally the targeting group is N-acetylgalactosamine (GalNAc), and each strand is independently 21 nucleotides. In some embodiments, the antisense strand of the dsRNA agent is at least substantially complementary to any one of the target regions of SEQ ID NO: 1, and is provided in any one of Tables 1-3.

According to another aspect of the present invention, a double-stranded ribonucleic acid (dsRNA) agent for inhibiting CFB expression is provided, wherein the dsRNA agent comprises a sense strand and an antisense strand, wherein the sense strand is complementary to _{the antisense} strand, wherein the antisense strand comprises a region complementary to the CFB RNA transcript, wherein each strand is about 18 to about 30 nucleotides in length, wherein the antisense strand comprises a sequence that can be represented by formula (II): 3′-( _NL ) _{nNM1N L NM2N L} N _{FN L} N _{M3N M4N L} N _L N _L N _{M5N L} N _{M6N L} N _L N _{FN L} -5′ (II); wherein: each _NF represents a 2′-fluorine-modified nucleotide; _NM1 , _NM2 , _NM3 , _NM4 , _NM5 and N Each _M6 independently represents a modified or unmodified nucleotide; each _NL independently represents a modified or unmodified nucleotide, but not a 2'-fluoro-modified nucleotide, and n is an integer from 0 to 7.

In some embodiments, N _M1 , N _M2 , N _M3 , N _M4 , N _M5 and N _M6 have only three 2'-fluoro modified nucleotides.

In some embodiments, N _M2 , N _M3 , and N _M5 each independently represent a 2'-fluoro modified nucleotide.

In some embodiments, N _M2 , N _M4 , and N _M5 each independently represent a 2'-fluoro modified nucleotide.

In some embodiments, N _M1 , N _M3 , and N _M6 each independently represent a 2'-fluoro modified nucleotide.

In some embodiments, N _M2 , N _M3 , and N _M6 each independently represent a 2'-fluoro modified nucleotide.

In some embodiments, N _M2 , N _{M4 ,} and N _M6 each independently represent a 2'-fluoro modified nucleotide.

In some embodiments, _NM1 , _NM3 , and _NM6 each independently represent a 2'-fluoro modified nucleotide and _NM5 represents a UNA modified nucleotide.

In some embodiments, N _M2 , N _M3 , and N _M6 each independently represent a 2'-fluoro modified nucleotide and N _M5 represents a UNA modified nucleotide.

In some embodiments, N _M2 , N _M4 , and N _M6 each independently represent a 2'-fluoro modified nucleotide and N _M5 represents a UNA modified nucleotide.

In some embodiments, n is 1, or n is 2, or n is 3, or n is 5. In some embodiments, the antisense strand of the dsRNA agent is at least substantially complementary to any of the target regions of SEQ ID NO: 1 and is provided in any of Tables 1-3.

According to another aspect of the present invention, a double-stranded ribonucleic acid (dsRNA) agent for inhibiting CFB expression is provided, wherein the dsRNA agent comprises a sense chain and an antisense chain, wherein the sense chain and the antisense chain form a dsRNA duplex, wherein the sense chain is complementary to the antisense chain, wherein the antisense chain comprises a region complementary to the CFB RNA transcript, wherein the complementary region comprises at least 15 consecutive nucleotides, wherein the dsRNA duplex comprises a duplex represented by formula (III): Sense chain: 5′-(N′ _L ) _n′ N′ _L N′ _L N′ _L N′ _L N′ _F N′ _L N′ _F N′ _L N′ _N1 N′ _N2 N′ _L N′ L N′ _L N′ _L N _{′ L (} N′ _L ) _m′ -3′ Antisense chain: 3′-( _{NL ) nNM1NLNM2NLNFNLNM3NM4NLNLNLNM5NLNM6NLNLNFNL - 5} ′ ( _III ) wherein: the length of each chain is about _{18 to} about 30 nucleotides; each _NF and _N'F independently _represents a _2' - _{fluorine -} modified nucleotide; _NM1 , _{NM2 , NM3} , _NM4 , _{NM5 , NM6} , _N'N1 , _and N'N2 each _{independently represent} a modified or unmodified nucleotide; each _NL and _N'L independently represent a modified or unmodified _nucleotide , but not a 2'-fluorine-modified nucleotide, _and m', n' and n each independently represent an integer _{from 0} to 7.

In some embodiments, N _M1 , N _M2 , N _M3 , N _M4 , N _M5 and N _M6 have only three 2'-fluorine modified nucleotides, and _N'N1 and _N'N2 include only one 2'-fluorine modified nucleotide.

In some embodiments, m' is 2 and n' is 4, m' is 2 and n' is 6, or m' is 2 and n' is 2. In some embodiments, m' is 1 and n' is 4, or m' is 1 and n' is 2. In some embodiments, m' is 0 and n' is 4, or m' is 0 and n' is 2. In some embodiments, n is 1, or n is 2, or n is 3, or n is 5.

In some embodiments, N' and _N1 independently represent 2'-fluoro modified nucleotides.

In some embodiments, N' and _N2 independently represent 2'-fluoro modified nucleotides.

In some embodiments, N _M2 , N _M3 , and N _M5 each independently represent a 2'-fluoro modified nucleotide.

In some embodiments, N _M2 , N _M4 , and N _M5 each independently represent a 2'-fluoro modified nucleotide.

In some embodiments, N _M1 , N _M3 , and N _M6 each independently represent a 2'-fluoro modified nucleotide.

In some embodiments, N _M2 , N _M3 , and N _M6 each independently represent a 2'-fluoro modified nucleotide.

In some embodiments, N _M2 , N _{M4 ,} and N _M6 each independently represent a 2'-fluoro modified nucleotide.

In some embodiments, _NM1 , _NM3 , and _NM6 each independently represent a 2'-fluoro modified nucleotide and _NM5 represents a UNA modified nucleotide.

In some embodiments, N _M2 , N _M3 , and N _M6 each independently represent a 2'-fluoro modified nucleotide, and N _M5 represents a UNA modified nucleotide.

In some embodiments, N _M2 , N _M4 , and N _M6 each independently represent a 2'-fluoro modified nucleotide, and N _M5 represents a UNA modified nucleotide.

In some embodiments, the dsRNA agent includes a targeting group conjugated to the 5'-end of the sense chain, preferably, the targeting group is selected from any one of the aforementioned GLO-1 to GLO-16 and GLS-1* to GLS-16*, more preferably, the targeting group is the aforementioned GLS-15*. In certain embodiments, the dsRNA agent includes a targeting group conjugated to the 5'-end of the sense chain. In certain embodiments, the antisense chain includes a reverse abacillary residue at the 3'-end. In certain embodiments, the sense chain includes one or two reverse abacillary residues and/or one or two imann residues at the 3' or/and 5' ends. In certain embodiments, each 3' and 5' end of the sense chain independently includes a reverse abacillary residue. In certain embodiments, each 3' and 5' end of the sense chain independently comprises an imann residue. In certain embodiments, the sense chain includes two reverse abamatic residues at the 3' and 5' ends, and any residue at the 3' or 5' end is further conjugated to a targeting group, which is preferably the aforementioned GLS-15*. In certain embodiments, the sense chain includes two imann residues at the 3' and 5' ends, and any residue at the 3' or 5' end is further conjugated to a targeting group, which is preferably the aforementioned GLS-15*. In certain embodiments, the dsRNA agent has two blunt ends. In certain embodiments, at least one chain comprises a 3' overhang of at least 1 nucleotide. In certain embodiments, at least one chain comprises a 3' overhang of at least 2 nucleotides. In certain embodiments, each end of the sense strand comprises a reverse abaci or imann residue at the 5' end of the sense strand, wherein the reverse abaci or imann residue is linked to the adjacent nucleotide at the 5' end of the nucleotide sequence of the sense strand via a phosphorothioate bond. In certain embodiments, the sense strand further comprises a targeting group linked to the reverse abaci or imann residue at the 5' end of the sense strand, wherein the targeting group is linked to the adjacent reverse abaci or imann residue via a phosphorothioate bond, and optionally the targeting group is N-acetylgalactosamine (GalNAc). In some embodiments, the 5' end of the sense chain includes a reverse abaci residue, wherein the reverse abaci residue is connected to the adjacent nucleotide at the 5' end of the nucleotide sequence of the sense chain through a thiophosphate bond. In some embodiments, the sense chain also includes a targeting group connected to the reverse abaci residue at the 5' end of the sense chain, wherein the targeting group is connected to the adjacent reverse abaci residue through a thiophosphate bond, and optionally the targeting group is N-acetylgalactosamine (GalNAc), and each chain is independently 21 nucleotides. In some embodiments, the antisense chain of the dsRNA agent is at least substantially complementary to any one of the target regions of SEQ ID NO: 1, and is provided in any one of Tables 1-3.

According to one aspect of the present invention, a composition is provided, which includes any embodiment of the aforementioned dsRNA agent of the present invention. In some embodiments, the composition also includes a pharmaceutically acceptable carrier. In some embodiments, the composition also includes one or more other therapeutic agents. In some embodiments, the composition is packaged in a kit, container, package, dispenser, prefilled syringe or vial. In some embodiments, the composition is formulated for subcutaneous administration or is formulated for intravenous (IV) administration.

According to another aspect of the present invention, a cell is provided, which includes any embodiment of the aforementioned dsRNA agent of the present invention. In some embodiments, the cell is a mammalian cell, optionally a human cell.

According to another aspect of the present invention, a method for inhibiting CFB gene expression in a cell is provided, the method comprising: (i) preparing a cell comprising an effective amount of any embodiment of the above-mentioned dsRNA agent of the present invention or any embodiment of the above-mentioned composition of the present invention. In certain embodiments, the method further comprises: (ii) maintaining the prepared cell for a sufficient time to obtain degradation of the mRNA transcript of the CFB gene, thereby inhibiting the expression of the CFB gene in the cell. In some embodiments, the cell is in a subject, and the dsRNA agent is administered to the subject subcutaneously. In some embodiments, the cell is in a subject, and the dsRNA agent is administered to the subject intravenously. In certain embodiments, the method further comprises assessing suppression of the CFB gene after administering the dsRNA agent to the subject, wherein the assessing method comprises: (i) determining one or more physiological characteristics of a CFB-related disease or condition in the subject, and (ii) comparing the determined physiological characteristics to a baseline pre-treatment physiological characteristic of the CFB-related disease or condition and/or a control physiological characteristic of the CFB-related disease or condition, wherein the comparison indicates one or more of the presence or absence of suppression of CFB gene expression in the subject. In some embodiments, the physiological characteristic is one or more of: CFB mRNA level and CFB protein level. A reduction in CFB expression can also be assessed indirectly by measuring a reduction in CFB biological activity, for example, or other pathologies associated with elevated CFB levels (preferably in the blood or kidneys), or overactivation of the complement pathway, or other therapeutic approaches that require inhibition of CFB expression. Decreased one or more of the following: CFB mRNA levels, CFB protein levels, CH50 activity (a measure of total hemolytic complement), AH50 (a measure of hemolytic activity of the alternative complement pathway), lactate dehydrogenase (LDH) (a measure of intravascular hemolysis), hemoglobin levels; levels of one or more of C3, C9, C5, C5a, C5b, and soluble C5b-9 complex.

According to another aspect of the present invention, a method for inhibiting CFB gene expression in a subject is provided, the method comprising administering to the subject an effective amount of the above-mentioned embodiment of the dsRNA agent of the present invention or the above-mentioned embodiment of the composition of the present invention. In some embodiments, the dsRNA agent is administered to the subject subcutaneously. In certain embodiments, the dsRNA agent is administered to the subject intravenously. In some embodiments, the method further comprises, after administration of the dsRNA agent, assessing suppression of the CFB gene, wherein the assessing method comprises: (i) determining one or more physiological characteristics of a CFB-related disease or condition in the subject, and (ii) comparing the determined physiological characteristics to a baseline pre-treatment physiological characteristic of the CFB-related disease or condition and/or a control physiological characteristic of the CFB-related disease or condition, wherein the comparison indicates the presence or absence of one or more of suppression of CFB gene expression in the subject. In some embodiments, expression of the CFB gene can be assessed based on the level or change in level of any variable associated with CFB gene expression, such as CFB mRNA level, CFB protein level. Reduction in CFB expression can also be assessed indirectly by measuring a reduction in CFB biological activity, for example, or other pathologies associated with elevated CFB levels, preferably in the blood or kidneys, or overactivation of the complement pathway, or other therapeutic approaches requiring inhibition of CFB expression, CFB mRNA levels, CFB protein levels, CH50 activity (a measure of total hemolytic complement), AH50 (a measure of hemolytic activity of the alternative complement pathway), lactate dehydrogenase (LDH) (a measure of intravascular hemolysis), hemoglobin levels; the levels of one or more of C3, C9, C5, C5a, C5b, and soluble C5b-9 complex.

According to another aspect of the present invention, a method for treating a disease or condition associated with the presence of CFB protein is provided, the method comprising: administering to a subject an effective amount of any of the above-mentioned dsRNA agent embodiments of the present invention, or any of the above-mentioned compositions of the present invention, for inhibiting CFB gene expression. In some embodiments, the disease, condition or condition associated with CFB is selected from: autoimmune diseases, complement system dysfunction, including abnormal upregulation of complement components, such as CFB, C3 glomerulopathy (C3G), systemic lupus erythematosus (SLE), lupus nephritis, Ig-mediated renal lesions, such as IgA nephropathy and primary membranous nephropathy, nephropathy, diabetic nephropathy, polycystic nephropathy, membranous nephropathy, age-related Macular degeneration (AMD), including dry AMD and geographic atrophy, typical or infectious hemolytic uremic syndrome (tHUS), atypical uremic hemolytic syndrome (aHUS), asthma, psoriasis, thrombotic microangiopathy, ischemia-reperfusion injury, paroxysmal nocturnal hemoglobinuria (PNH), rheumatic diseases, rheumatoid arthritis, multiple sclerosis (MS), neuromyelitis optica (NMO), immune complex Compound-mediated glomerulonephritis (IC-mediated GN), post-infectious glomerulonephritis (PIGN), anti-neutrophil cytoplasmic autoantibody-associated vasculitis (ANCA-AV), antiphospholipid antibody syndrome (APS), periodontal disease, malaria-induced anemia, dermatomyositis, sphaeroidal sphaeroides, Shiga toxin-induced Escherichia coli-associated hemolytic uremic syndrome, myasthenia gravis (MG), neuromyelitis optica (NMO), dense sediment coronary artery disease, dermatomyositis, Graves' disease, atherosclerosis, Alzheimer's disease, systemic inflammatory response sepsis, septic shock, spinal cord injury, glomerulonephritis, Hashimoto's thyroiditis, type I diabetes mellitus, psoriasis, urticaria, autoimmune hemolytic anemia (AIHA), cold agglutinin disease, humoral and vascular transplant rejection, graft dysfunction, myocardial infarction, graft allergy, hyperlipidemia, and sepsis.

In some embodiments, the method further comprises: administering an additional treatment regimen to the subject. In some embodiments, the additional treatment regimen comprises treatment of a CFB-related disease or condition. In certain embodiments, the additional treatment regimen comprises: administering to the subject one or more CFB antisense polynucleotides of the present invention, administering to the subject a non-CFB dsRNA therapeutic agent, and behavioral changes to the subject. In some embodiments, the additional therapeutic agent is selected from the group consisting of: oligonucleotides, small molecules, monoclonal antibodies, polyclonal antibodies, and peptides. Preferably, the additional therapeutic agent is a C5 inhibitor, such as an anti-complement component C5 antibody or an antigen-binding fragment thereof (e.g., eculizumab, ravulizumab-cwvz or polzelimab (REGN3918)) or a C5 peptide inhibitor (e.g., zulucoplan). Eculizumab is a humanized monoclonal IgG2/4, kappa light chain antibody that specifically binds to the complement component C5 with high affinity and inhibits the cleavage of C5 into C5a and C5b, thereby inhibiting the formation of the terminal complement complex C5b-9. Ravulizumab-cwvz is a humanized IgG2/4 monoclonal antibody that specifically binds complement component C5 with high affinity and inhibits the cleavage of C5 into C5a and C5b, thereby inhibiting the generation of the terminal complement complex C5b-9. Pozelimab (also known as H4H12166P, described in US20170355757) is a fully human IgG4 monoclonal antibody designed to block complement factor C5. Zilucoplan is a synthetic macrocyclic peptide that binds complement component 5 (C5) with subnanomolar affinity and allosterically inhibits its cleavage into C5a and C5b after activation of the classical, alternative or lectin pathways. Preferably, the additional therapeutic agent is a C3 peptide inhibitor or an analog thereof. In one embodiment, the C3 peptide inhibitor is compstatin. Compstatin is a cyclic tridecapeptide with potent and selective C3 inhibitory activity.

In some embodiments, the dsRNA agent is administered to the subject subcutaneously. In certain embodiments, the dsRNA agent is administered to the subject by intravenous injection. In some embodiments, the method further comprises determining the efficacy of the administered double-stranded RNA (dsRNA) agent on the subject.

In some embodiments, a method of determining a therapeutic efficacy of a treatment in a subject comprises: (i) determining one or more physiological characteristics of a CFB-related disease or condition in the subject, and (ii) comparing the determined physiological characteristics to a baseline, pre-treatment physiological characteristic of the CFB-related disease or condition, wherein the comparison indicates one or more of the presence, absence, and level of efficacy of a double-stranded RNA (dsRNA) agent administered to the subject.

In some embodiments, expression of a CFB gene can be assessed based on the level or change in level of any variable associated with CFB gene expression, such as a subject's CFB mRNA level, CFB protein level, or CH50 activity (a measure of total hemolytic complement), AH50 (a measure of hemolytic activity of the alternative complement pathway), lactate dehydrogenase (LDH) (a measure of intravascular hemolysis), hemoglobin level; the level of one or more of C3, C9, C5, C5a, C5b, and soluble C5b-9 complex.

According to another aspect of the invention, a method of reducing the level of CFB protein in a subject compared to the baseline pre-treatment level of CFB protein in the subject is provided, the method comprising administering to the subject an effective amount of any of the aforementioned dsRNA agent embodiments of the invention or any of the aforementioned composition embodiments of the invention to reduce the level of CFB gene expression. In some embodiments, the dsRNA agent is administered to the subject subcutaneously or by IV administration.

According to another aspect of the invention, a method of altering the physiological characteristics of a CFB-related disease or condition in a subject compared to a baseline pre-treatment physiological characteristic of the CFB-related disease or condition in the subject is provided, the method comprising administering to the subject an effective amount of any of the aforementioned dsRNA agent embodiments of the invention or any of the aforementioned composition embodiments of the invention to alter the physiological characteristics of the CFB-related disease or condition in the subject. In some embodiments, the dsRNA agent is administered subcutaneously to the subject or administered to the subject by IV administration. In certain embodiments, the physiological characteristics and symptoms are one or more of: CFB mRNA levels, CFB protein levels, or CH50 activity (a measure of total hemolytic complement), AH50 (a measure of hemolytic activity of the alternative complement pathway), lactate dehydrogenase (LDH) (a measure of intravascular hemolysis), hemoglobin levels in a subject; the levels of any one or more of C3, C9, C5, C5a, C5b, and soluble C5b-9 complex.

According to another aspect of the present invention, a method for treating a disease or condition associated with the CFB protein using the aforementioned dsRNA agent is provided. In some embodiments, the disease or condition is one or more of the following: autoimmune disease, complement system dysfunction (including abnormal upregulation of complement components, such as CFB), C3 glomerulopathy (C3G), , systemic lupus erythematosus (SLE), lupus nephritis; Ig-mediated renal diseases (e.g., IgA nephropathy and primary membranous nephropathy), nephropathy, diabetic nephropathy, polycystic nephropathy, membranous nephropathy; age-related macular degeneration (AMD), including dry AMD and geographic atrophy, typical or infectious hemolytic uremic syndrome (tHUS), atypical uremic hemolytic syndrome (aHUS), asthma, psoriasis, thrombotic microangiopathy, ischemia and reperfusion injury, paroxysmal nocturnal hemoglobinuria (PNH), rheumatism, rheumatoid arthritis, multiple sclerosis (MS), neuromyelitis optica (NMO), immune complex-mediated glomerulonephritis (IC-mediated GN), post-infectious glomerulonephritis (PIGN), antineutrophil cytoplasmic autoantibody-associated vasculitis (ANCA-AV), antiphospholipid antibody syndrome (APS), periodontal disease, malaria-induced anemia, dermatomyositis, scrofuloid herpes, Shiga toxin-induced Escherichia coli-associated hemolytic uremic syndrome, myasthenia gravis (MG), neuromyelitis optica (NMO), dense deposit disease, coronary artery disease, dermatomyositis, Graves' disease, atherosclerosis, Alzheimer's disease, systemic inflammatory response sepsis, septic shock, spinal cord injury, glomerulonephritis, Hashimoto's thyroiditis, type I diabetes mellitus, psoriasis, urticaria, autoimmune hemolytic anemia (AIHA), cold agglutinin disease, humoral and vascular transplant rejection, graft dysfunction, myocardial infarction, transplant sensitization, hyperlipidemia, and sepsis.

According to another aspect of the present invention, an antisense polynucleotide agent for inhibiting CFB protein expression is provided, the agent comprising 10 to 30 consecutive nucleotides, wherein at least one consecutive nucleotide is a modified nucleotide, and wherein the nucleotide sequence of the agent is about 80% complementary to the equivalent region of the nucleotide sequence of SEQ ID NO: 1 over its entire length. In some embodiments, the equivalent region is any one of the target regions of SEQ ID NO: 1, and the complementary sequence is a sequence provided in one of Tables 1-3. In certain embodiments, the antisense polynucleotide agent comprises one of the antisense sequences provided in one of Tables 1-3.

According to another aspect of the present invention, a composition comprising any of the embodiments of the antisense polynucleotide agents described above is provided. In some embodiments, the composition further comprises a pharmaceutically acceptable carrier. In some embodiments, the composition further comprises one or more additional therapeutic agents for treating CFB-related diseases or conditions. In certain embodiments, the composition is packaged in a kit, container, package, dispenser, prefilled syringe or vial. In certain embodiments, the composition is formulated for subcutaneous or intravenous administration.

According to another aspect of the present invention, a cell comprising any one of the embodiments of the antisense polynucleotide agent described above is provided. In some embodiments, the cell is a mammalian cell, optionally a human cell.

According to another aspect of the present invention, a method for inhibiting the expression of a CFB gene in a cell is provided, the method comprising: (i) preparing a cell comprising an effective amount of any of the embodiments of the aforementioned antisense polynucleotide reagent. In some embodiments, the method further comprises (ii) maintaining the cell prepared in (i) for a time sufficient to obtain degradation of the mRNA transcript of the CFB gene, thereby inhibiting the expression of the CFB gene in the cell.

According to another aspect of the present invention, a method for inhibiting CFB gene expression in a subject is provided, the method comprising administering to the subject an effective amount of any of the aforementioned antisense polynucleotide reagent embodiments.

According to another aspect of the present invention, a method for treating a disease or condition associated with the presence of CFB protein is provided, the method comprising administering to a subject an effective amount of any of the above-mentioned antisense polynucleotide agent embodiments or any of the above-mentioned compositions of the present invention embodiments to inhibit CFB gene expression. In certain embodiments, the disease or condition is one or more of the following: autoimmune disease, complement system dysfunction (including abnormal upregulation of complement components, such as CFB), C3 glomerulopathy (C3G), systemic lupus erythematosus (SLE), lupus nephritis, Ig-mediated renal lesions (such as IgA nephropathy and primary membranous nephropathy), nephropathy, diabetic nephropathy, polycystic nephropathy, membranous nephropathy, age-related jaundice, Advanced macular degeneration (AMD), including dry AMD and geographic atrophy, typical or infectious hemolytic uremic syndrome (tHUS), atypical uremic hemolytic syndrome (aHUS), asthma, psoriasis, thrombotic microangiopathy, ischemia and reperfusion injury, paroxysmal nocturnal hemoglobinuria (PNH), rheumatism, rheumatoid arthritis, multiple sclerosis (MS), neuromyelitis optica (NMO), immune complex-mediated glomerulonephritis (IC-mediated GN), post-infectious glomerulonephritis (PIGN), antineutrophil cytoplasmic autoantibody-associated vasculitis (ANCA-AV), antiphospholipid antibody syndrome (APS), dysbacteriosis periodontitis, malaria-induced anemia, dermatomyositis, ulcerative colitis, Shiga toxin-induced Escherichia coli-associated hemolytic uremic syndrome, myasthenia gravis (MG), neuromyelitis optica (NMO), dense deposits disease, coronary artery disease, dermatomyositis, Graves' disease, atherosclerosis, Alzheimer's disease, systemic inflammatory response sepsis, septic shock, spinal cord injury, glomerulonephritis, Hashimoto's thyroiditis, type I diabetes, psoriasis, ulcers, autoimmune hemolytic anemia (AIHA), cold agglutinin disease, humoral and vascular transplant rejection, graft dysfunction, myocardial infarction, transplant sensitization, hyperlipidemia, and sepsis.

According to another aspect of the invention, a method of reducing the level of CFB protein in a subject compared to the baseline pre-treatment level of CFB protein in the subject is provided, the method comprising administering to the subject an effective amount of any of the aforementioned antisense polynucleotide reagent embodiments or any of the aforementioned composition embodiments of the invention to reduce the level of CFB gene expression. In certain embodiments, the antisense polynucleotide reagent is administered to the subject subcutaneously or by IV administration.

According to another aspect of the present invention, an antisense polynucleotide reagent for inhibiting CFB gene expression is provided, the reagent comprising 10 to 30 consecutive nucleotides, wherein at least one of the consecutive nucleotides is a modified nucleotide, and wherein the nucleotide sequence has about 80% or about 85% complementarity with the equivalent region of the nucleotide sequence of SEQ ID NO: 1 over its entire length.

According to another aspect of the present invention, a method for altering the physiological characteristics of a CFB-related disease or condition in a subject is provided, the method comprising administering to the subject an effective amount of any of the above-mentioned antisense polynucleotide agent embodiments or any of the above-mentioned composition embodiments of the present invention to alter the physiological characteristics of the CFB disease or condition in the subject, compared to the baseline pre-treatment physiological characteristics of the CFB-related disease or condition in the subject. In some embodiments, the antisense polynucleotide agent is administered to the subject by subcutaneous or intravenous injection. In certain embodiments, the physiological characteristics and symptoms are one or more of the following: CFB mRNA levels, CFB protein levels, or CH50 activity (a measure of total hemolytic complement), AH50 (a measure of hemolytic activity of the alternative complement pathway), lactate dehydrogenase (LDH) (a measure of intravascular hemolysis), hemoglobin levels; levels of one or more of C3, C9, C5, C5a, C5b, and soluble C5b-9 complex in the subject .

SEQ ID NO: 1 and SEQ ID NO: 2 (reverse complement) are Homo sapiens CFB mRNA [NCBI Reference Sequence: NM_001710.6].

SEQ ID NO: 3 and SEQ ID NO: 4 (reverse complement) are macaque (rhesus monkey) CFB mRNA [NCBI Reference Sequence: XM_015136029.2].

SEQ ID NO: 5 and SEQ ID NO: 6 (reverse complement) are Mus musculus (subtype 1) CFB mRNA [NCBI Reference Sequence: NM_008198.3].

SEQ ID NOs: 7-468, 1489-1595 are shown in Table 1 and are sense chain sequences.

SEQ ID NOs: 469-930, 1596-1702 are shown in Table 1 and are antisense chain sequences.

SEQ ID NOs: 931-1392, 1703-1810 are chemically modified sequences as shown in Table 2.

SEQ ID NOs: 1393-1488, 1811-1879, 1882-1950 are shown in Table 3. The delivery molecule is indicated as "GLX-__" at the 3' end or 5' end of each sense strand.

The present invention includes, in part, RNAi agents, such as but not limited to double-stranded (ds) RNAi agents, which are capable of inhibiting CFB gene expression. The present invention also includes, in part, compositions comprising CFB RNAi agents and methods of using the compositions. The CFB RNAi agents disclosed herein can be attached to delivery compounds for delivery to cells, including hepatocytes. The pharmaceutical compositions of the present invention can include at least one dsRNA CFB agent and a delivery compound. In some embodiments of the compositions and methods of the present invention, the delivery compound is a delivery compound containing GalNAc. The CFB RNAi agent delivered to the cell can inhibit CFB gene expression, thereby reducing the activity of the CFB protein product of the gene in the cell. The dsRNAi reagents of the present invention can be used to treat CFB-related diseases and disorders.

In some embodiments of the invention, reducing CFB expression in a cell or subject treats a disease or condition associated with CFB expression in a cell or subject, respectively. Non-limiting examples of diseases and conditions that can be treated by reducing CFB activity include: alleviation or amelioration of one or more symptoms associated with unwanted or excessive CFB expression, CH50 activity (a measure of total hemolytic complement), AH50 (a measure of hemolytic activity of the alternative complement pathway), lactate dehydrogenase (LDH) (a measure of intravascular hemolysis), hemoglobin levels; levels of one or more of C3, C9, C5, C5a, C5b, and soluble C5b-9 complex. "Treatment" can also refer to prolonging survival compared to expected survival without treatment.

As used herein, "G", "C", "A" and "U" generally represent nucleotides containing guanine, cytosine, adenine and uracil as bases, respectively. However, it should be understood that the term "ribonucleotide" or "nucleotide" can also refer to modified nucleotides (as further described below) or alternative replacement moieties. Those skilled in the art understand that guanine, cytosine, adenine and uracil can be replaced by other moieties without significantly changing the base pairing properties of the oligonucleotide containing nucleotides with such replacement moieties. For example, but not limited to, nucleotides containing inosine as a base can be paired with nucleotide bases containing adenine, cytosine or uracil. Therefore, in the nucleotide sequences of the present invention, nucleotides containing uracil, guanine or adenine can be replaced by nucleotides containing, for example, inosine. Sequences containing such replacement moieties are embodiments of the present invention.

As used herein, "complement factor B" is used interchangeably with the term "factor B" or "CFB" and refers to a naturally occurring gene encoding a complement factor B protein from any vertebrate or mammal, including but not limited to humans, cows, chickens, rodents, mice, rats, pigs, sheep, primates, monkeys and guinea pigs, unless otherwise indicated. The term also refers to fragments and variants of native CFB that retain at least one in vivo or in vitro activity of native CFB. The amino acid and complete coding sequence of the reference sequence of the human CFB gene can be found in, for example, GenBank Ref Seq Accession No. NM_001710.6 (SEQ ID NO: 1 and SEQ ID NO: 2), Macaca mulatta (rhesus monkey) CFB mRNA GenBank Ref Seq Accession No. XM_015136029.2 (SEQ ID NO: 3 and SEQ ID NO: 4), Mus musculus (isoform 1) NM_008198.3 (SEQ ID NO: 5 and SEQ ID NO: 6). Other examples of CFB mRNA sequences are readily available using public databases such as GenBank, UniProt, Ensembl, and OMIM.

Described below are compositions comprising single stranded CFB (ssRNA) and dsRNA agents to inhibit CFB gene expression, as well as compositions and methods for treating diseases and disorders caused or regulated by CFB gene expression. The term "RNAi" is also known in the art and may be referred to as "siRNA".

As used herein, the term "RNAi" refers to a reagent that contains RNA and mediates targeted cleavage of RNA transcripts through an RNA-induced silencing complex (RISC) pathway. As known in the art, the RNAi target region refers to a continuous portion of the nucleotide sequence of an mRNA molecule formed during gene transcription, including messenger RNA (mRNA) as an RNA processing product of the primary transcription product. The targeted portion of the sequence will be at least long enough to serve as a substrate for RNAi directed cleavage at or near the portion. The target sequence can be 8-30 nucleotides long (including end values), 10-30 nucleotides long (including end values), 12-25 nucleotides long (including end values), 15-23 nucleotides long (including end values), 16-23 nucleotides long (including end values), or 18-23 nucleotides long (including end values), including all shorter lengths within each specified range. In some embodiments of the invention, the target sequence is 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, or 26 nucleotides in length. In certain embodiments, the target sequence is between 9 and 26 nucleotides in length (inclusive), including all subranges and integers therebetween. For example, although not intended to be limiting, in certain embodiments of the invention, the target sequence is 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleotides in length, and its sequence is completely or at least substantially complementary to at least a portion of the RNA transcript of the CFB gene. Some aspects of the invention include pharmaceutical compositions comprising one or more CFB dsRNA agents and a pharmaceutically acceptable carrier. In certain embodiments of the invention, the CFB RNAi described herein inhibits the expression of CFB protein.

As used herein, "dsRNA agents" refer to compositions containing RNA or RNA-like (e.g., chemically modified RNA) oligonucleotide molecules that are capable of degrading or inhibiting translation of messenger RNA (mRNA) transcripts of target mRNAs in a sequence-specific manner. Although not wishing to be limited to a particular theory, the dsRNA agents of the present invention may act through an RNA interference mechanism (i.e., by inducing RNA interference by interacting with the RNA interference pathway mechanism of mammalian cells (RNA-induced silencing complex or RISC)) or through any alternative mechanism or pathway. Methods for silencing genes in plant, invertebrate and vertebrate cells are well known in the art [see, e.g., (Sharp et al., Genes Dev. 2001, 15:485; Bernstein, et al., (2001) Nature 409:363; Nykanen, et al., (2001) Cell 107:309; and Elbashir, et al., (2001) Genes Dev. 15:188)], the disclosures of each of which are incorporated herein by reference in their entirety. Gene silencing procedures known in the art can be used in conjunction with the disclosure provided herein to inhibit the expression of CFB.

The dsRNA agents disclosed herein consist of a sense strand and an antisense strand, including but not limited to: short interfering RNA (siRNA), RNAi agents, microRNA (miRNA), short hairpin RNA (shRNA) and Dicer substrates. The antisense strand of the dsRNA agents described herein is at least partially complementary to the target mRNA. It is known in the art that dsRNA duplex structures of different lengths can be used to inhibit target gene expression. For example, dsRNAs with duplex structures of 19, 20, 21, 22 and 23 base pairs are known to effectively induce RNA interference (Elbashir et al., EMBO 2001, 20: 6877-6888). It is also known in the art that shorter or longer RNA duplex structures can also effectively induce RNA interference. In some embodiments, the length of the sense strand and the antisense strand can be the same or different. In some embodiments, the length of each strand is no more than 40 nucleotides. In some embodiments, the length of each strand is no more than 30 nucleotides. In some embodiments, the length of each strand is no more than 25 nucleotides. In some embodiments, the length of each strand is no more than 23 nucleotides. In some embodiments, the length of each strand is no more than 21 nucleotides. In some embodiments, the length of the sense strand and the antisense strand of the RNAi agent can be 15 to 49 nucleotides, respectively. In some embodiments, the length of the antisense strand is independently 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleotides. In some embodiments, the length of the sense strand is independently 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, or 49 nucleotides. In some embodiments, the length of the sense strand and the antisense strand are both 21 nucleotides. In some embodiments, the sense strand is complementary or substantially complementary to the antisense strand, and the length of the complementary region is 15 to 23 nucleotides. In some embodiments, the length of the complementary region is 19-21 nucleotides. In some embodiments, the length of the complementary region is 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30 nucleotides. In certain embodiments of the present invention, the CFB dsRNA may include at least one strand of at least 21 nt in length, or may have a shorter duplex based on one of the sequences listed in any one of Tables 1-3, but it may also be effective to reduce 1, 2, 3 or 4 nucleotides at one or both ends, respectively, compared to the dsRNA listed in Tables 1-3. In some embodiments of the present invention, the CFB dsRNA agent may have a partial sequence of at least 15, 16, 17, 18, 19, 20 or more consecutive nucleotides from one or more sequences in Tables 1-3, and its ability to inhibit CFB gene expression differs from the level of inhibition produced by the dsRNA containing the complete sequence by no more than 5%, 10%, 15%, 20%, 25% or 30%. The sense sequences, antisense sequences and duplexes disclosed in Tables 1-3 may be referred to herein as "parent" sequences, which means that the sequences disclosed in Tables 1-3 may be modified, shortened, lengthened, including substitutions, etc. as described herein, and the resulting sequences retain all or at least a portion of the efficacy of their parent sequences in the methods and compositions of the present invention. The sense strand and antisense strand contained in the dsRNA of the present invention are independently selected. As used herein, the term "independent selection" means that each of two or more similar elements can be selected independently of the selection of other elements. For example, although not intended to be limiting, when preparing the dsRNA of the present invention, "elements" of the two chains can be selected to be included in the double chain. One selected element, the sense sequence, can be SEQ ID NO: 932 (as shown in Table 2), while the other selected element, the antisense sequence, can be SEQ ID NO: 1163, or can be SEQ ID NO: 1163 that has been modified, shortened, lengthened, and/or includes 1, 2, or 3 substitutions compared to its parent sequence SEQ ID NO: 1163. It should be understood that the double chain of the present invention does not necessarily include the sense and antisense sequences shown in pairs in the double chain in Tables 1-3 at the same time. Each sense and antisense strand sequence in the table is followed by its SEQ ID NO.

Certain embodiments of the compositions and methods of the present invention include single-stranded RNA in the composition and/or administered to a subject. For example, the antisense chain listed in any of Tables 1-3 can be a composition or a composition administered to a subject to reduce the CFB polypeptide activity and/or the expression of the CFB gene in the subject. Table 1 shows certain CFB dsRNA agent antisense chains and sense chain core extension base sequences. Single-stranded antisense molecules that may be included in certain compositions of the present invention and/or administered in certain methods of the present invention are referred to herein as "single-stranded antisense agents" or "antisense polynucleotide agents". Single-stranded sense molecules that may be included in certain compositions of the present invention and/or administered in certain methods of the present invention are referred to herein as "single-stranded sense agents" or "sense polynucleotide agents". The term "base sequence" is used herein to refer to a polynucleotide sequence without chemical modification or delivery compounds. For example, the sense strand GACAAUGUGAGUGAUGAGAUA (SEQ ID NO: 22) shown in Table 1 is the base sequence of SEQ ID NO: 946 in Table 2 and SEQ ID NO: 1393 in Table 3, wherein SEQ ID NO: 946 and SEQ ID NO: 1393 show their chemical modifications and delivery compounds. Sequences disclosed herein can be assigned identifiers. For example, a single-strand sense sequence can be identified by "sense strand SS#"; a single-strand antisense sequence can be identified by "antisense strand AS#", and a duplex comprising a sense strand and an antisense strand can be identified by "duplex AD#/AV#".

Table 1 includes the sense chain and the antisense chain, and provides the identification number of the duplex formed by the sense chain and the antisense chain on the same line in Table 1. In certain embodiments of the present invention, the antisense sequence includes a nucleobase u or a nucleobase a at position 1 of the antisense sequence. In certain embodiments of the present invention, the antisense sequence includes a nucleobase u at position 1 of the antisense sequence. As used herein, the term "matching position" in the sense chain and the antisense chain is the position of "pairing" in each chain when the two chains are a duplex. For example, in a sense chain of 21 nucleobases and an antisense chain of 21 nucleobases, the nucleobase at position 1 of the sense chain and the nucleobase at position 21 of the antisense chain are in a "matching position". In yet another non-limiting example, in a 23-nucleobase sense chain and a 23-nucleobase antisense chain, nucleobase 2 of the sense chain and nucleobase 22 of the antisense chain are in a matching position. In yet another non-limiting example, in an 18-nucleobase sense chain and an 18-nucleobase antisense chain, nucleobase 1 of the sense chain and nucleobase 18 of the antisense chain are in a matching position, and nucleobase 4 of the sense chain and nucleobase 15 of the antisense chain are in a matching position. Those skilled in the art know how to identify matching positions in sense chains and antisense chains that are or will be double chains and paired chains.

The last column in Table 1 represents the duplex AV# of a duplex that includes sense and antisense sequences in the same table row. For example, Table 1 discloses a duplex designated as Duplex AV02358.um that includes sense strand SEQ ID NO: 7 and antisense strand SEQ ID NO: 469. Thus, each row in Table 1 identifies a duplex of the present invention, each duplex comprising the sense and antisense sequences shown in the same row, wherein the identifier assigned to each duplex is shown in the last column of that row.

In some embodiments of the methods of the present invention, an RNAi agent comprising a polynucleotide sequence shown in any one of Tables 1-3 is administered to a subject. In some embodiments of the present invention, the RNAi agent administered to a subject comprises a duplex comprising at least one base sequence listed in Table 1, comprising 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24 sequence modifications. In some embodiments of the methods of the present invention, an RNAi agent comprising a polynucleotide sequence shown in any one of Tables 1-3 is attached to a delivery molecule, a non-limiting example of a delivery molecule is a delivery compound comprising a GalNAc compound or a GLS-15* compound.

Table 1: Unmodified CFB RNAi Antisense and Sense Strand Sequences. All sequences are shown in 5' to 3' orientation. Duplex AV# is the duplex identification number assigned to both strands in the same row of the table. Double chain AV# Sense chain Serial Number Antisense chain Serial Number AV02358.um GAGGUCUAGGUCUGGAGUUUA 7 UAAACUCCAGACCUAGACCUC 469 AV02359.um GAGGUCUGGAGUUUCAGCUUA 8 UAAGCUGAAACUCCAGACCUC 470 AV02360.um GGGUCUGGAGUUUCAGCUUGA 9 UCAAGCUGAAACUCCAGACCC 471 AV02361.um CGUCUGGAGUUUCAGCUUGGA 10 UCCAAGCUGAAACUCCAGACG 472 AV02362.um CGAGUUUCAGCUUGGACACUA 11 UAGUGUCCAAGCUGAAACUCG 473 AV02363.um CAGUUUCAGCUUGGACACUGA 12 UCAGUGUCCAAGCUGAAACUG 474 AV02364.um GCCUUCCGACUUCUCCAAGAA 13 UUCUUGGAGAAGUCGGAAGGC 475 AV02365.um GCGUGUGUCCUUCUGGCUUCA 14 UGAAGCCAGAAGGACACACGC 476 AV02366.um CUGUGUCCUUCUGGCUUCUAA 15 UUAGAAGCCAGAAGGACACAG 477 AV02367.um CGAAGGCAGAGUGCAGAGCAA 16 UUGCUCUGCACUCUGCCUUCG 478 AV02368.um CACUUCGAGAACGGGGAAUAA 17 UUAUUCCCCGUUCUCGAAGUG 479 AV02369.um GCCUACUACAAUGUGAGUGAA 18 UUCACUCACAUUGUAGUAGGC 480 AV02370.um GUACUACAAUGUGAGUGAUGA 19 UCAUCACUCACAUUGUAGUAC 481 AV02371.um GACUACAAUGUGAGUGAUGA 20 UUCAUCACUCACAUUGUAGUC 482 AV02372.um GCUACAAUGUGAGUGAUGAGA twenty one UCUCAUCACUCACAUUGUAGC 483 AV02373.um GACAAUGUGAGUGAUGAGAUA twenty two UAUCUCAUCACUCACAUUGUC 484 AV02374.um GAAUGUGAGUGAUGAGAUCUA twenty three UAGAUCUCAUCACUCACAUUC 485 AV02375.um GAUGUGAGUGAUGAGAUCUCA twenty four UGAGAUCUCAUCACUCACAUC 486 AV02376.um GUGUGAGUGAUGAGAUCUCUA 25 UAGAGAUCUCAUCACUCACAC 487 AV02377.um GGUGAGUGAUGAGAUCUCUUA 26 UAAGAGAUCUCAUCACUCACC 488 AV02378.um GGAGUGAUGAGAUCUCUUUCA 27 UGAAAGAGAUCUCAUCACUCC 489 AV02379.um CAGUGAUGAGAUCUCUUUCCA 28 UGGAAAGAGAUCUCAUCACUG 490 AV02380.um GGUGAUGAGAUCUCUUUCCAA 29 UUGGAAAGAGAUCUCAUCACC 491 AV02381.um CUGAUGAGAUCUCUUUCCACA 30 UGUGGAAAGAGAUCUCAUCAG 492 AV02382.um GGAGAUCUCUUUUCCACUGCUA 31 UAGCAGUGGAAAGAGAUCUCC 493 AV02383.um GGAUCUCUUUCCACUGCUAUA 32 UAUAGCAGUGGAAAGAGAUCC 494 AV02384.um GCUCUUUCCACUGCUAUGACA 33 UGUCAUAGCAGUGGAAAGAGC 495 AV02385.um GUCCACUGCUAUGACGGUUAA 34 UUAACCGUCAUAGCAGUGGAC 496 AV02386.um GCACUGCUAUGACGGUUACAA 35 UUGUAACCGUCAUAGCAGUGC 497 AV02387.um GCUGCUAUGACGGUUACACUA 36 UAGUGUAACCGUCAUAGCAGC 498 AV02388.um GGCUAUGACGGUUACACUCUA 37 UAGAGUGUAACCGUCAUAGCC 499 AV02389.um CCUAUGACGGUUACACUCUCA 38 UGAGAGUGUAACCGUCAUAGG 500 AV02390.um GCAGCGAUCUGUGACAACGGA 39 UCCGUUGUCACAGAUCGCUGC 501 AV02391.um GCAGUACCGCCUUGAAGACAA 40 UUGUCUUCAAGGCGGUACUGC 502 AV02392.um GGAAGACAGCGUCACCUACCA 41 UGGUAGGUGACGCUGUCUUCC 503 AV02393.um GCUGCCAAGACUCCUUCAUGA 42 UCAUGAAGGAGUCUUGGCAGC 504 AV02394.um CUGGCCGAAGCUUUCCUGUCA 43 UGACAGGAAAGCUUCGGCCAG 505 AV02395.um CACAGAGACCAUAGAAGGAGA 44 UCUCCUUCUAUGGUCUCUGUG 506 AV02396.um GGACCAUAGAAGGAGUCGAUA 45 UAUCGACUCCUUCUAUGGUCC 507 AV02397.um GCCCUUCAGGCUCCAUGAACA 46 UGUUCAUGGAGCCUGAAGGGC 508 AV02398.um GUCAGGCUCCAUGAACAUCUA 47 UAGAUGUUCAUGGAGCCUGAC 509 AV02399.um GUCCAUGAACAUCUACCUGGA 48 UCCAGGUAGAUGUUCAUGGAC 510 AV02400.um CAACAUCUACCUGGUGCUAGA 49 UCUAGCACCAGGUAGAUGUUG 511 AV02401.um GUACCUGGUGCUAGAUGGAUA 50 UAUCCAUCUAGCACCAGGUAC 512 AV02402.um GUGGUGCUAGAUGGAUCAGAA 51 UUCUGAUCCAUCUAGCACCAC 513 AV02403.um CGCCAGCAACUUCACAGGAGA 52 UCUCCUGUGAAGUUGCUGGCG 514 AV02404.um GAACUUCACAGGAGCCAAAAA 53 UUUUUGGCUCCUGUGAAGUUC 515 AV02405.um GCUUCACAGGAGCCAAAAAGA 54 UCUUUUUGGCUCCUGUGAAGC 516 AV02406.um GUUCACAGGAGCCAAAAAGUA 55 UACUUUUUGGCUCCUGUGAAC 517 AV02407.um GAGGAGCCAAAAAGUGUCUAA 56 UUAGACACUUUUUGGCUCCUC 518 AV02408.um GGGAGCCAAAAAGUGUCUAGA 57 UCUAGACACUUUUUGGCUCCC 519 AV02409.um CAGCCAAAAAGUGUCUAGUCA 58 UGACUAGACACUUUUUGGCUG 520 AV02410.um GGCCAAAAAGUGUCUAGUCAA 59 UUGACUAGACACUUUUUGGCC 521 AV02411.um CCCAAAAAGUGUCUAGUCAAA 60 UUUGACUAGACACUUUUUGGG 522 AV02412.um GAAAAAGUGUCUAGUCAACUA 61 UAGUUGACUAGACACUUUUUC 523 AV02413.um GAAAGUGUCUAGUCAACUUAA 62 UUAAGUUGACUAGACACUUUC 524 AV02414.um GAAGUGUCUAGUCAACUUAAA 63 UUUAAGUUGACUAGACACUUC 525 AV02415.um CUGUCUAGUCAACUUAAUUGA 64 UCAAUUAAGUUGACUAGACAG 526 AV02416.um GGUCUAGUCAACUUAAUUGAA 65 UUCAAUUAAGUUGACUAGACC 527 AV02417.um CUCUAGUCAACUUAAUUGAGA 66 UCUCAAUUAAGUUGACUAGAG 528 AV02418.um GCUAGUCAACUUAAUUGAGAA 67 UUCUCAAUUAAGUUGACUAGC 529 AV02419.um GUAGUCAACUUAAUUGAGAAA 68 UUUCUCAAUUAAGUUGACUAC 530 AV02420.um CUCAACUUAAUUGAGAAGGUA 69 UACCUUCUCAAUUAAGUUGAG 531 AV02421.um GCAACUUAAUUGAGAAGGUGA 70 UCACCUUCUCAAUUAAGUUGC 532 AV02422.um GAUUGAGAAGGUGGCAAGUUA 71 UAACUUGCCACCUUCUCAAUC 533 AV02423.um GUUGAGAAGGUGGCAAGUUAA 72 UUAACUUGCCACCUUCUCAAC 534 AV02424.um GGAGAAGGUGGCAAGUUAUGA 73 UCAUAACUUGCCACCUUCUCC 535 AV02425.um GAGGUGGCAAGUUAUGGUGUA 74 UACACCAUAACUUGCCACCUC 536 AV02426.um GGGUGGCAAGUUAUGGUGUGA 75 UCACACCAUAACUUGCCACCC 537 AV02427.um CUGGCAAGUUAUGGUGUGAAA 76 UUUCACACCAUAACUUGCCAG 538 AV02428.um CCAAGUUAUGGUGUGAAGCCA 77 UGGCUUCACACCAUAACUUGG 539 AV02429.um GAGUUAUGGUGUGAAGCCAAA 78 UUUGGCUUCACACCAUAACUC 540 AV02430.um GAUGGUGUGAAGCCAAGAUAA 79 UUAUCUUGGCUUCACACCAUC 541 AV02431.um GAGAUAUGGUCUAGUGACAUA 80 UAUGUCACUAGACCAUAUCUC 542 AV02432.um GGAUAUGGUCUAGUGACAUAA 81 UUAUGUCACUAGACCAUAUCC 543 AV02433.um CAUAUGGUCUAGUGACAUAUA 82 UAUAUGUCACUAGACCAUAUG 544 AV02434.um GUAUGGUCUAGUGACAUAUGA 83 UCAUAUGUCACUAGACCAUAC 545 AV02435.um GAUGGUCUAGUGACAUAUGCA 84 UGCAUAUGUCACUAGACCAUC 546 AV02436.um CUCUAGUGACAUAUGCCACAA 85 UUGUGGCAUAUGUCACUAGAG 547 AV02437.um GUAGUGACAUAUGCCACAUAA 86 UUAUGUGGCAUAUGUCACUAC 548 AV02438.um GAGUGACAUAUGCCACAUACA 87 UGUAUGUGGCAUAUGUCACUC 549 AV02439.um GCCAAAAUUUGGGUCAAAGUA 88 UACUUUGACCCAAAUUUUGGC 550 AV02440.um GAAAAUUUGGGUCAAAGUGUA 89 UACACUUUGACCCAAAUUUUC 551 AV02441.um GAAUGAAAUCAAUUAUGAAGA 90 UCUUCAUAAUUGAUUUCAUUC 552 AV02442.um GAUGAAAUCAAUUAUGAAGAA 91 UUCUUCAUAAUUGAUUUCAUC 553 AV02443.um GUGAAAUCAAUUAUGAAGACA 92 UGUCUUCAUAAUUGAUUUCAC 554 AV02444.um CAAAUCAAUUAUGAAGACCAA 93 UUGGUCUUCAUAAUUGAUUUG 555 AV02445.um GAUCAAUUAUGAAGACCACAA 94 UUGUGGUCUUCAUAAUUGAUC 556 AV02446.um GUCAAUUAUGAAGACCACAAA 95 UUUGUGGUCUUCAUAAUUGAC 557 AV02447.um GAUUAUGAAGACCACAAGUUA 96 UAACUUGUGGUCUUCAUAAUC 558 AV02448.um GUAUGAAGACCACAAGUUGAA 97 UUCAACUUGUGGUCUUCAUAC 559 AV02449.um GAUGAAGACCACAAGUUGAAA 98 UUUCAACUUGUGGUCUUCAUC 560 AV02450.um GGAAGACCACAAGUUGAAGUA 99 UACUUCAACUUGUGGUCUUCC 561 AV02451.um GAGACCACAAGUUGAAGUCAA 100 UUGACUUCAACUUGUGGUCUC 562 AV02452.um GCAAGUUGAAGUCAGGGACUA 101 UAGUCCCUGACUUCAACUUGC 563 AV02453.um GAAGUUGAAGUCAGGGACUAA 102 UUAGUCCCUGACUUCAACUUC 564 AV02454.um GAGUUGAAGUCAGGGACUAAA 103 UUUAGUCCCUGACUUCAACUC 565 AV02455.um CUUGAAGUCAGGGACUAACAA 104 UUGUUAGUCCCUGACUUCAAG 566 AV02456.um GAGUCAGGGACUAACACCAAA 105 UUUGGUGUUAGUCCCUGACUC 567 AV02457.um CUCAGGGACUAACACCAAGAA 106 UUCUUGGUGUUAGUCCCUGAG 568 AV02458.um GGACUGAUGGAUUGCACAACA 107 UGUUGUGCAAUCCAUCAGUCC 569 AV02459.um CACUGAUGGAUUGCACAACAA 108 UUGUUGUGCAAUCCAUCAGUG 570 AV02460.um CACCCAAUUACUGUCAUUGAA 109 UUCAAUGACAGUAAUUGGGUG 571 AV02461.um GCCAAUUACUGUCAUUGAUGA 110 UCAUCAAUGACAGUAAUUGGC 572 AV02462.um GCAAUUACUGUCAUUGAUGAA 111 UUCAUCAAUGACAGUAAUUGC 573 AV02463.um GAUUACUGUCAUUGAUGAGAA 112 UUCUCAUCAAUGACAGUAAUC 574 AV02464.um GUUACUGUCAUUGAUGAGAUA 113 UAUCUCAUCAAUGACAGUAAC 575 AV02465.um GACUGUCAUUGAUGAGAUCCA 114 UGGAUCUCAUCAAUGACAGUC 576 AV02466.um GAUUGGCAAGGAUCGCAAAAA 115 UUUUUGCGAUCCUUGCCAAUC 577 AV02467.um GCAAGGGAGGAUUAUCUGGAA 116 UUCCAGAUAAUCCUCCCUUGC 578 AV02468.um GGGGAGGAUUAUCUGGAUGUA 117 UACAUCCAGAUAAUCCUCCCC 579 AV02469.um CGGAGGAUUAUCUGGAUGUCA 118 UGACAUCCAGAUAAUCCUCCG 580 AV02470.um CGAGGAUUAUCUGGAUGUCUA 119 UAGACAUCCAGAUAAUCCUCG 581 AV02471.um CAGGAUUAUCUGGAUGUCUAA 120 UUAGACAUCCAGAUAAUCCUG 582 AV02472.um GGGAUUAUCUGGAUGUCUAUA 121 UAUAGACAUCCAGAUAAUCCC 583 AV02473.um CGAUUAUCUGGAUGUCUAUGA 122 UCAUAGACAUCCAGAUAAUCG 584 AV02474.um GUAUCUGGAUGUCUAUGUGUA 123 UACACAUAGACAUCCAGAUAC 585 AV02475.um GAUCUGGAUGUCUAUGUGUUA 124 UAACACAUAGACAUCCAGAUC 586 AV02476.um GUCUGGAUGUCUAUGUGUUUA 125 UAAACACAUAGACAUCCAGAC 587 AV02477.um GUGGAUGUCUAUGUGUUUGGA 126 UCCAAACACAUAGACAUCCAC 588 AV02478.um CGUGAACCAAGUGAACAUCAA 127 UUGAUGUUCACUUGGUUCACG 589 AV02479.um CUGAACCAAGUGAACAUCAAA 128 UUUGAUGUUCACUUGGUUCAG 590 AV02480.um GGAACCAAGUGAACAUCAAUA 129 UAUUGAUGUUCACUUGGUUCC 591 AV02481.um CAACCAAGUGAACAUCAAUGA 130 UCAUUGAUGUUCACUUGGUUG 592 AV02482.um GACCAAGUGAACAUCAAUGCA 131 UGCAUUGAUGUUCACUUGGUC 593 AV02483.um GCAAGUGAACAUCAAUGCUUA 132 UAAGCAUUGAUGUUCACUUGC 594 AV02484.um GAAGUGAACAUCAAUGCUUUA 133 UAAAGCAUUGAUGUUCACUUC 595 AV02485.um GGUGAACAUCAAUGCUUUGGA 134 UCCAAAGCAUUGAUGUUCACC 596 AV02486.um CUGAACAUCAAUGCUUUGGCA 135 UGCCAAAGCAUUGAUGUUCAG 597 AV02487.um CAACAUCAAUGCUUUGGCUUA 136 UAAGCCAAAGCAUUGAUGUUG 598 AV02488.um GACAUCAAUGCUUUGGCUUCA 137 UGAAGCCAAAGCAUUGAUGUC 599 AV02489.um GAAUGCUUUGGCUUCCAAGAA 138 UUCUUGGAAGCCAAAGCAUUC 600 AV02490.um GUGCUUUGGCUUCCAAGAAAA 139 UUUUCUUGGAAGCCAAAGCAC 601 AV02491.um GUUGGCUUCCAAGAAAGACAA 140 UUGUCUUUCUUGGAAGCCAAC 602 AV02492.um CGCUUCCAAGAAAGACAAUGA 141 UCAUUGUCUUUCUUGGAAGCG 603 AV02493.um CCUUCCAAGAAAGACAAUGA 142 UUCAUUGUCUUUCUUGGAAGG 604 AV02494.um GUUCCAAGAAAGACAAUGAGA 143 UCUCAUUGUCUUUCUUGGAAC 605 AV02495.um GCCAAGAAAGACAAUGAGCAA 144 UUGCUCAUUGUCUUUCUUGGC 606 AV02496.um GCAAGAAAGACAAUGAGCAAA 145 UUUGCUCAUUGUCUUUCUUGC 607 AV02497.um GAGAAAGACAAUGAGCAACAA 146 UUGUUGCUCAUUGUCUUUCUC 608 AV02498.um CAAAGACAAUGAGCAACAUGA 147 UCAUGUUGCUCAUUGUCUUUG 609 AV02499.um GAAGACAAUGAGCAACAUGUA 148 UACAUGUUGCUCAUUGUCUUC 610 AV02500.um GAGACAAUGAGCAACAUGUGA 149 UCACAUGUUGCUCAUUGUCUC 611 AV02501.um CACAAUGAGCAACAUGUGUUA 150 UAACACAUGUUGCUCAUUGUG 612 AV02502.um GAAUGAGCAACAUGUGUUCAA 151 UUGAACACAUGUUGCUCAUUC 613 AV02503.um GAUGAGCAACAUGUGUUCAAA 152 UUUGAACACAUGUUGCUCAUC 614 AV02504.um GUGAGCAACAUGUGUUCAAA 153 UUUUGAACACAUGUUGCUCAC 615 AV02505.um GGAGCAACAUGUGUUCAAAGA 154 UCUUUGAACACAUGUUGCUCC 616 AV02506.um CAGCAACAUGUGUUCAAAGUA 155 UACUUUGAACACAUGUUGCUG 617 AV02507.um GAGUCAAGGAUAUGGAAAACA 156 UGUUUUCCAUAUCCUUGACUC 618 AV02508.um GGUCAAGGAUAUGGAAAACCA 157 UGGUUUUCCAUAUCCUUGACC 619 AV02509.um GAGGAUAUGGAAAACCUGGAA 158 UUCCAGGUUUUCCAUAUCCUC 620 AV02510.um CAUAUGGAAAACCUGGAAGAA 159 UUCUUCCAGGUUUUCCAUAUG 621 AV02511.um CAGCUGCUCCCUGCACAGGAA 160 UUCCUGUGCAGGGAGCAGCUG 622 AV02512.um CCACAGGAUAUCAAAGCUCUA 161 UAGAGCUUUGAUAUCCUGUGG 623 AV02513.um GACAGGAUAUCAAAGCUCUGA 162 UCAGAGCUUUGAUAUCCUGUC 624 AV02514.um GCAGGAUAUCAAAGCUCUGUA 163 UACAGAGCUUUGAUAUCCUGC 625 AV02515.um GAGGAUAUCAAAGCUCUGUUA 164 UAACAGAGCUUUGAUAUCCUC 626 AV02516.um GGGAUAUCAAAGCUCUGUUUA 165 UAAACAGAGCUUUGAUAUCCC 627 AV02517.um CGAUAUCAAAGCUCUGUUUGA 166 UCAAACAGAGCUUUGAUAUCG 628 AV02518.um CAUAUCAAAGCUCUGUUUGUA 167 UACAAACAGAGCUUUGAUAUG 629 AV02519.um GUAUCAAAGCUCUGUUUGUGA 168 UCACAAACAGAGCUUUGAUAC 630 AV02520.um GAUCAAAGCUCUGUUUGUGUA 169 UACACAAACAGAGCUUUGAUC 631 AV02521.um GCAAAGCUCUGUUUGUGUCUA 170 UAGACACAAACAGAGCUUUGC 632 AV02522.um GAAAGCUCUGUUUGUGUCUGA 171 UCAGACACAAACAGAGCUUUC 633 AV02523.um GAAGCUCUGUUUGUGUCUGAA 172 UUCAGACACAAACAGAGCUUC 634 AV02524.um GGCUCUGUUUGUGUCUGAGGA 173 UCCUCAGACACAAACAGAGCC 635 AV02525.um GUCUGUUUGUGUCUGAGGAGA 174 UCUCCUCAGACACAAACAGAC 636 AV02526.um GGGAAGGAGGUCUACAUCAAA 175 UUUGAUGUAGACCUCCUUCCC 637 AV02527.um CGAAGGAGGUCUACAUCAAGA 176 UCUUGAUGUAGACCUCCUUCG 638 AV02528.um CAAGGAGGUCUACAUCAAGAA 177 UUCUUGAUGUAGACCUCCUUG 639 AV02529.um CGAGGUCUACAUCAAGAAUGA 178 UCAUUCUUGAUGUAGACCUCG 640 AV02530.um CAGGUCUACAUCAAGAAUGGA 179 UCCAUUCUUGAUGUAGACCUG 641 AV02531.um GUGUGAGAGAGAUGCUCAAUA 180 UAUUGAGCAUCUCUCUCUCACAC 642 AV02532.um GGUGAGAGAGAUGCUCAAUAA 181 UUAUUGAGCAUCUCUCUCACC 643 AV02533.um CUGAGAGAGAUGCUCAAUAUA 182 UAUAUUGAGCAUCUCUCUCUCAG 644 AV02534.um GGAGAGAGAUGCUCAAUAUGA 183 UCAUAUUGAGCAUCUCUCUCUCC 645 AV02535.um CAGAGAGAUGCUCAAUAUGCA 184 UGCAUAUUGAGCAUCUCUCUG 646 AV02536.um GAGGCUAUGACAAAGUCAAGA 185 UCUUGACUUUGUCAUAGCCUC 647 AV02537.um GUAUGACAAAGUCAAGGACAA 186 UUGUCCUUGACUUUGUCAUAC 648 AV02538.um GAUGACAAAGUCAAGGACAUA 187 UAUGUCCUUGACUUUGUCAUC 649 AV02539.um GGACAAAGUCAAGGACAUCUA 188 UAGAUGUCCUUGACUUUGUCC 650 AV02540.um CUGGUCACCCCUCGGUUCCUA 189 UAGGAACCGAGGGGUGACCAG 651 AV02541.um GCCUCGGUUCCUUUGUACUGA 190 UCAGUACAAAGGAACCGAGGC 652 AV02542.um GCUCGGUUCCUUUGUACUGGA 191 UCCAGUACAAAGGAACCGAGC 653 AV02543.um GUCCUUUGUACUGGAGGAGUA 192 UACUCCUCCAGUACAAAGGAC 654 AV02544.um CAGUGAGUCCCUAUGCUGACA 193 UGUCAGCAUAGGGACUCACUG 655 AV02545.um GCAAUACUUGCAGAGGUGAUA 194 UAUCACCUCUGCAAGUAUUGC 656 AV02546.um GUACUUGCAGAGGUGAUUCUA 195 UAGAAUCACCUCUGCAAGUAC 657 AV02547.um GCCCUUGAUAGUUCACAAGAA 196 UUCUUGUGAACUAUCAAGGGC 658 AV02548.um GCCUUGAUAGUUCACAAGAGA 197 UCUCUUGUGAACUAUCAAGGC 659 AV02549.um GCUUGAUAGUUCACAAGAGAA 198 UUCUCUUGUGAACUAUCAAGC 660 AV02550.um GGAUAGUUCACAAGAGAAGUA 199 UACUUCUCUUGUGAACUAUCC 661 AV02551.um GGUUCACAAGAGAAGUCGUUA 200 UAACGACUUCUCUUGUGAACC 662 AV02552.um CUUCACAAGAGAAGUCGUUUA 201 UAAACGACUUCUCUUGUGAAG 663 AV02553.um GUCACAAGAGAAGUCGUUUCA 202 UGAAACGACUUCUCUUGUGAC 664 AV02554.um GCACAAGAGAAGUCGUUUCAA 203 UUGAAACGACUUCUCUUGUGC 665 AV02555.um GACAAGAGAAGUCGUUUCAUA 204 UAUGAAACGACUUCUCUUGUC 666 AV02556.um GCAAGAGAAGUCGUUUCAUUA 205 UAAUGAAACGACUUCUCUUGC 667 AV02557.um GAAGAGAAGUCGUUUCAUUCA 206 UGAAUGAAACGACUUCUCUUC 668 AV02558.um GAGAGAAGUCGUUUCAUUCAA 207 UUGAAUGAAACGACUUCUCUC 669 AV02559.um GGAGAAGUCGUUUCAUUCAAA 208 UUUGAAUGAAACGACUUCUCC 670 AV02560.um CAGAAGUCGUUUCAUUCAAGA 209 UCUUGAAUGAAACGACUUCUG 671 AV02561.um GGAAGUCGUUUCAUUCAAGUA 210 UACUUGAAUGAAACGACUUCC 672 AV02562.um CAAGUCGUUUCAUUCAAGUUA 211 UAACUUGAAUGAAACGACUUG 673 AV02563.um GAGUCGUUUCAUUCAAGUUGA 212 UCAACUUGAAUGAAACGACUC 674 AV02564.um GGUCGUUUCAUUCAAGUUGGA 213 UCCAACUUGAAUGAAACGACC 675 AV02565.um CUCGUUUCAUUCAAGUUGGUA 214 UACCAACUUGAAUGAAACGAG 676 AV02566.um GCGUUUCAUUCAAGUUGGUGA 215 UCACCAACUUGAAUGAAACGC 677 AV02567.um GGUUUCAUUCAAGUUGGUGUA 216 UACACCAACUUGAAUGAAACC 678 AV02568.um CUGUAAUCAGCUGGGGAGUAA 217 UUACUCCCCAGCUGAUUACAG 679 AV02569.um GUGGGGAGUAGUGGAUGUCUA 218 UAGACAUCCACUACUCCCCAC 680 AV02570.um GGUAGUGGAUGUCUGCAAAAA 219 UUUUUGCAGACAUCCACUACC 681 AV02571.um GAGUGGAUGUCUGCAAAAACA 220 UGUUUUUGCAGACAUCCACUC 682 AV02572.um CGAUGUCUGCAAAAACCAGAA 221 UUCUGGUUUUUGCAGACAUCG 683 AV02573.um CAUGUCUGCAAAAACCAGAAA 222 UUUCUGGUUUUUGCAGACAUG 684 AV02574.um GUGUCUGCAAAAACCAGAAGA 223 UCUUCUGGUUUUUGCAGACAC 685 AV02575.um GAAAACCAGAAGCGGCAAAAA 224 UUUUUGCCGCUUCUGGUUUUC 686 AV02576.um GUGAAGGAGAAACUCCAAGAA 225 UUCUUGGAGUUUCUCCUUCAC 687 AV02577.um GGAAGGAGAAACUCCAAGAUA 226 UAUCUUGGAGUUUCUCCUUCC 688 AV02578.um GGGAGAAACUCCAAGAUGAGA 227 UCUCAUCUUGGAGUUUCUCCC 689 AV02579.um CAAACUCCAAGAUGAGGAUUA 228 UAAUCCUCAUCUUGGAGUUUG 690 AV02580.um GACUCCAAGAUGAGGAUUUGA 229 UCAAAUCCUCAUCUUGGAGUC 691 AV02581.um GAAGAUGAGGAUUUGGGUUUA 230 UAAACCCAAAUCCUCAUCUUC 692 AV02582.um GGAUGAGGAUUUGGGUUUUCA 231 UGAAAACCCAAAUCCUCAUCC 693 AV02583.um CAUGAGGAUUUGGGUUUUCUA 232 UAGAAAACCCAAAUCCUCAUG 694 AV02584.um CCGUGGGAUUGAAUUAAAACA 233 UGUUUUAAUUCAAUCCCACGG 695 AV02585.um CUGGGAUUGAAUUAAAACAGA 234 UCUGUUUUAAUUCAAUCCCAG 696 AV02586.um GGGGAUUGAAUUAAAACAGCA 235 UGCUGUUUUAAUUCAAUCCCC 697 AV02587.um CGGAUUGAAUUAAAACAGCUA 236 UAGCUGUUUUAAUUCAAUCCG 698 AV02588.um GAUUAAAACAGCUGCGACAAA 237 UUUGUCGCAGCUGUUUUAAUC 699 AV02358-19-1.um GGUCUAGGUCUGGAGUUUA 238 UAAACUCCAGACCUAGACC 700 AV02359-19-1.um GGUCUGGAGUUUCAGCUUA 239 UAAGCUGAAACUCCAGACC 701 AV02360-19-1.um GUCUGGAGUUUCAGCUUGA 240 UCAAGCUGAAACUCCAGAC 702 AV02361-19-1.um UCUGGAGUUUCAGCUUGGA 241 UCCAAGCUGAAACUCCAGA 703 AV02362-19-1.um AGUUUCAGCUUGGACACUA 242 UAGUGUCCAAGCUGAAACU 704 AV02363-19-1.um GUUUCAGCUUGGACACUGA 243 UCAGUGUCCAAGCUGAAAC 705 AV02364-19-1.um CUUCCGACUUCUCCAAGAA 244 UUCUUGGAGAAGUCGGAAG 706 AV02365-19-1.um GUGUGUCCUUCUGGCUUCA 245 UGAAGCCAGAAGGACACAC 707 AV02366-19-1.um GUGUCCUUCUGGCUUCUAA 246 UUAGAAGCCAGAAGGACAC 708 AV02367-19-1.um AAGGCAGAGUGCAGAGCAA 247 UUGCUCUGCACUCUGCCUU 709 AV02368-19-1.um CUUCGAGAACGGGGAAUAA 248 UUAUUCCCCGUUCUCGAAG 710 AV02369-19-1.um CUACUACAAUGUGAGUGAA 249 UUCACUCACAUUGUAGUAG 711 AV02370-19-1.um ACUACAAUGUGAGUGAUGA 250 UCAUCACUCACAUUGUAGU 712 AV02371-19-1.um CUACAAUGUGAGUGAUGA 251 UUCAUCACUCACAUUGUAG 713 AV02372-19-1.um UACAAUGUGAGUGAUGAGA 252 UCUCAUCACUCACAUUGUA 714 AV02373-19-1.um CAAUGUGAGUGAUGAGAUA 253 UAUCUCAUCACUCACAUUG 715 AV02374-19-1.um AUGUGAGUGAUGAGAUCUA 254 UAGAUCUCAUCACUCACAU 716 AV02375-19-1.um UGUGAGUGAUGAGAUCUCA 255 UGAGAUCUCAUCACUCA 717 AV02376-19-1.um GUGAGUGAUGAGAUCUCUA 256 UAGAGAUCUCAUCACUCAC 718 AV02377-19-1.um UGAGUGAUGAGAUCUCUUA 257 UAAGAGAUCUCAUCACUCA 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428 UCUCUUGUGAACUAUCAAG 890 AV02549-19-1.um UUGAUAGUUCACAAGAGAA 429 UUCUCUUGUGAACUAUCAA 891 AV02550-19-1.um AUAGUUCACAAGAGAAGUA 430 UACUUCUCUUGUGAACUAU 892 AV02551-19-1.um UUCACAAGAGAAGUCGUUA 431 UAACGACUUCUCUUGUGAA 893 AV02552-19-1.um UCACAAGAGAAGUCGUUUA 432 UAAACGACUUCUCUUGUGA 894 AV02553-19-1.um CACAAGAGAAGUCGUUUCA 433 UGAAACGACUUCUCUUGUG 895 AV02554-19-1.um ACAAGAGAAGUCGUUUCAA 434 UUGAAACGACUUCUCUUGU 896 AV02555-19-1.um CAAGAGAAGUCGUUUCAUA 435 UAUGAAACGACUUCUCUUG 897 AV02556-19-1.um AAGAGAAGUCGUUUCAUUA 436 UAAUGAAACGACUUCUCUU 898 AV02557-19-1.um AGAGAAGUCGUUUCAUUCA 437 UGAAUGAAACGACUUCUCU 899 AV02558-19-1.um GAGAAGUCGUUUCAUUCAA 438 UUGAAUGAAACGACUUCUC 900 AV02559-19-1.um AGAAGUCGUUUCAUUCAAA 439 UUUGAAUGAAACGACUUCU 901 AV02560-19-1.um GAAGUCGUUUCAUUCAAGA 440 UCUUGAAUGAAACGACUUC 902 AV02561-19-1.um AAGUCGUUUCAUUCAAGUA 441 UACUUGAAUGAAACGACUU 903 AV02562-19-1.um AGUCGUUUCAUUCAAGUUA 442 UAACUUGAAUGAAACGACU 904 AV02563-19-1.um GUCGUUUCAUUCAAGUUGA 443 UCAACUUGAAUGAAACGAC 905 AV02564-19-1.um UCGUUUCAUUCAAGUUGGA 444 UCCAACUUGAAUGAAACGA 906 AV02565-19-1.um CGUUUCAUUCAAGUUGGUA 445 UACCAACUUGAAUGAAACG 907 AV02566-19-1.um GUUUCAUUCAAGUUGGUGA 446 UCACCAACUUGAAUGAAAC 908 AV02567-19-1.um UUUCAUUCAAGUUGGUGUA 447 UACACCAACUUGAAUGAAA 909 AV02568-19-1.um GUAAUCAGCUGGGGAGUAA 448 UUACUCCCCAGCUGAUUAC 910 AV02569-19-1.um GGGGAGUAGUGGAUGUCUA 449 UAGACAUCCACUACUCCCC 911 AV02570-19-1.um UAGUGGAUGUCUGCAAAAA 450 UUUUUGCAGACAUCCACUA 912 AV02571-19-1.um GUGGAUGUCUGCAAAAACA 451 UGUUUUUGCAGACAUCCAC 913 AV02572-19-1.um AUGUCUGCAAAAACCAGAA 452 UUCUGGUUUUUGCAGACAU 914 AV02573-19-1.um UGUCUGCAAAAACCAGAAA 453 UUUCUGGUUUUUGCAGACA 915 AV02574-19-1.um GUCUGCAAAAACCAGAAGA 454 UCUUCUGGUUUUUGCAGAC 916 AV02575-19-1.um AAACCAGAAGCGGCAAAAA 455 UUUUUGCCGCUUCUGGUUU 917 AV02576-19-1.um GAAGGAGAAACUCCAAGAA 456 UUCUUGGAGUUUCUCCUUC 918 AV02577-19-1.um AAGGAGAAACUCCAAGAUA 457 UAUCUUGGAGUUUCUCCUU 919 AV02578-19-1.um GAGAAACUCCAAGAUGAGA 458 UCUCAUCUUGGAGUUUCUC 920 AV02579-19-1.um AACUCCAAGAUGAGGAUUA 459 UAAUCCUCAUCUUGGAGUU 921 AV02580-19-1.um CUCCAAGAUGAGGAUUUGA 460 UCAAAUCCUCAUCUUGGAG 922 AV02581-19-1.um AGAUGAGGAUUUGGGUUUA 461 UAAACCCAAAUCCUCAUCU 923 AV02582-19-1.um AUGAGGAUUUGGGUUUUCA 462 UGAAAACCCAAAUCCUCAU 924 AV02583-19-1.um UGAGGAUUUGGGUUUUCUA 463 UAGAAAACCCAAAUCCUCA 925 AV02584-19-1.um GUGGGAUUGAAUUAAAACA 464 UGUUUUAAUUCAAUCCCAC 926 AV02585-19-1.um GGGAUUGAAUUAAAACAGA 465 UCUGUUUUAAUUCAAUCCC 927 AV02586-19-1.um GGAUUGAAUUAAAACAGCA 466 UGCUGUUUUAAUUCAAUCC 928 AV02587-19-1.um GAUUGAAUUAAAACAGCUA 467 UAGCUGUUUUAAUUCAAUC 929 AV02588-19-1.um UUAAAACAGCUGCGACAAA 468 UUUGUCGCAGCUGUUUUAA 930 AV02022.um GACUUUCACAUCAACCUCUUU 1489 AAAGAGGUUGAUGUGAAAGUC 1596 AV02023.um CUGAAGCAGACAGCAGUAAUU 1490 AAUUACUGCUGUCUGCUUCAG 1597 AV02024.um UCCUUCCGACUUCUCCAAGAU 1491 AUCUUGGAGAAGUCGGAAGGA 1598 AV02025.um UCGGUUCCUUUGUACUGGAGU 1492 ACUCCAGAGUACAAAGGAACCGA 1599 AV02026.um ACUUGCUAUACAUUGGCAAGU 1493 ACUUGCCAAUGUAUAGCAAGU 1600 AV02027.um AUGAGCUGGCCAGAUGACGUU 1494 AACGUCAUCUGGCCAGCUCAU 1601 AV05135.um GACGUGUGUCCUUCUGGCUUA 1495 UAAGCCAGAAGGACACACGUC 1602 AV05136.um GCCUACUACAAUGUGAGUGA 1496 UCACUCACAUUGUAGUAGGGC 1603 AV05137.um GUACAAUGUGAGUGAUGAGAA 1497 UUCUCAUCACUCACAUUGUAC 1604 AV05138.um GCAAUUGUGAGUGAUGAGAUCA 1498 UGAUCUCAUCACUCACAUUGC 1605 AV05139.um GUCUUUCCACUGCUAUGACGA 1499 UCGUCAUAGCAGUGGAAAGAC 1606 AV05140.um GACUGCUAUGACGGUUACACA 1500 UGUGUAACCGUCAUAGCAGUC 1607 AV05141.um GUCUCCGGGGCUCUGCCAAUA 1501 UAUUGGCAGAGCCCCGGAGAC 1608 AV05142.um GCCGGGGCUCUGCCAAUCGCA 1502 UGCGAUUGGCAGAGCCCCGGC 1609 AV05143.um GGACAGCGAUCUGUGACAACA 1503 UGUUGUCACAGAUCGCUGUCC 1610 AV05144.um CCCAAGACUCCUUCAUGUACA 1504 UGUACAUGAAGGAGUCUUGGG 1611 AV05145.um GCAAGACUCCUUCAUGUACGA 1505 UCGUACAUGAAGGAGUCUUGC 1612 AV05146.um GGACUCCUUCAUGUACGACAA 1506 UUGUCGUACAUGAAGGAGUCC 1613 AV05147.um GAUCUACCUGGUGCUAGAUGA 1507 UCAUCUAGCACCAGGUAGAUC 1614 AV05148.um GGAUGGAUCAGACAGCAUUGA 1508 UCAAUGCUGUCUGAUCCAUCC 1615 AV05149.um GCACAGGAGCCAAAAAGUGUA 1509 UACACUUUUUGGCUCCUGUGC 1616 AV05150.um GACAGGAGCCAAAAAGUGUCA 1510 UGACACUUUUUGGCUCCUGUC 1617 AV05151.um GCAGGAGCCAAAAAGUGUCUA 1511 UAGACACUUUUUGGCUCCUGC 1618 AV05152.um GCAAAAAGUGUCUAGUCAACA 1512 UGUUGACUAGACACUUUUUGC 1619 AV05153.um GAGUGUCUAGUCAACUUAAUA 1513 UAUUAAGUUGACUAGACACUC 1620 AV05154.um GGUGUCUAGUCAACUUAAUUA 1514 UAAUUAAGUUGACUAGACACC 1621 AV05155.um GAGUCAACUUAAUUGAGAAGA 1515 UCUUCUCAAUUAAGUUGACUC 1622 AV05156.um GGUCAACUUAAUUGAGAAGGA 1516 UCCUUCUCAAUUAAGUUGACC 1623 AV05157.um GUAUGGUGUGAAGCCAAGAUA 1517 UAUCUUGGCUUCACACCAUAC 1624 AV05158.um GCCAGGCAGUGUACAGCAUGA 1518 UCAUGCUGUACACUGCCUGGC 1625 AV05159.um CCAGUGUACAGCAUGAUGAGA 1519 UCUCAUCAUGCUGUACACUGG 1626 AV05160.um GCUGAUGGAUUGCACAACAUA 1520 UAUGUUGUGCAAUCCAUCAGC 1627 AV05161.um GUGAUGGAUUGCACAACAUGA 1521 UCAUGUUGUGCAAUCCAUCAC 1628 AV05162.um GCCCAAUUACUGUCAUUGAUA 1522 UAUCAAUGACAGUAAUUGGGC 1629 AV05163.um GAAGGGAGGAUUAUCUGGAUA 1523 UAUCCAGAUAAUCCUCCCUUC 1630 AV05164.um GCCAAGUGAACAUCAAUGCUA 1524 UAGCAUUGAUGUUCACUUGGC 1631 AV05165.um GGAAAGACAAUGAGCAACAUA 1525 UAUGUUGCUCAUUGUCUUUCC 1632 AV05166.um GACAUCAAGAAUGGGGAUAAA 1526 UUUAUCCCCAUUCUUGAUGUC 1633 AV05167.um GCAUCAAGAAUGGGGAUAAGA 1527 UCUUAUCCCCAUUCUUGAUGC 1634 AV05168.um GAUCAAGAAUGGGGAUAAGAA 1528 UUCUUAUCCCCAUUCUUGAUC 1635 AV05169.um GUCAAGAAUGGGGAUAAGAAA 1529 UUUCUUAUCCCCAUUCUUGAC 1636 AV05170.um GCAAGAAUGGGGAUAAGAAAA 1530 UUUUCUUAUCCCCAUUCUUGC 1637 AV05171.um GAAGAAUGGGGAUAAGAAAGA 1531 UCUUUCUUAUCCCCAUUCUUC 1638 AV05172.um GAGAAUGGGGAUAAGAAAGGA 1532 UCCUUUCUUAUCCCCAUUCUC 1639 AV05173.um CGCUAUGACAAAGUCAAGGAA 1533 UUCCUUGACUUUGUCAUAGCG 1640 AV05174.um GAUGCUGACCCCAAUACUUGA 1534 UCAAGUAUUGGGGUCAGCAUC 1641 AV05175.um CGAGUAGUGGAUGUCUGCAAA 1535 UUUGCAGACAUCCACUACUCG 1642 AV05176.um CUGGAUGUCUGCAAAAACCAA 1536 UUGGUUUUUGCAGACAUCCAG 1643 AV05177.um GCAAGAUGAGGAUUUGGGUUA 1537 UAACCCAAAUCCUCAUCUUGC 1644 AV05178.um GAGGGGUUUCCUGCUGGACAA 1538 UUGUCCAGCAGGAAACCCCUC 1645 AV06327.um CAAUGUGAGUGAUGAGAUU 1539 AAUCUCAUCACUCACAUUG 1646 AV06328.um GACAAUGUGAGUGAUGAGAUU 1540 AAUCUCAUCACUCACAUUGUC 1647 AV06329.um GAAAAAGUGUCUAGUCAACUA 1541 UAGUUGACUAGACACUUUUUC 1648 AV06330.um CCGCGGGAUUGAAUUAAAACA 1542 UGUUUUAAUUCAAUCCCGCGG 1649 AV02022-19-1.um CUUUCACAUCAACCUCUUU 1543 AAAGAGGUUGAUGUGAAAG 1650 AV02023-19-1.um GAAGCAGACAGCAGUAAUU 1544 AAUUACUGCUGUCUGCUUC 1651 AV02024-19-1.um CUUCCGACUUCUCCAAGAU 1545 AUCUUGGAGAAGUCGGAAG 1652 AV02025-19-1.um GGUUCCUUUGUACUGGAGU 1546 ACUCCAGAGUACAAAGGAACC 1653 AV02026-19-1.um UUGCUAUACAUUGGCAAGU 1547 ACUUGCCAAUGUAUAGCAA 1654 AV02027-19-1.um GAGCUGGCCAGAUGACGUU 1548 AACGUCAUCUGGCCAGCUC 1655 AV05135-19-1.um CGUGUGUCCUUCUGGCUUA 1549 UAAGCCAGAAGGACACACG 1656 AV05136-19-1.um CCUACUACAAUGUGAGUGA 1550 UCACUCACAUUGUAGUAGG 1657 AV05137-19-1.um ACAAUGUGAGUGAUGAGA 1551 UUCUCAUCACUCACAUUGU 1658 AV05138-19-1.um AAUGUGAGUGAUGAGAUCA 1552 UGAUCUCAUCACUCACAUU 1659 AV05139-19-1.um CUUUCCACUGCUAUGACGA 1553 UCGUCAUAGCAGUGGAAAG 1660 AV05140-19-1.um CUGCUAUGACGGUUACACA 1554 UGUGUAACCGUCAUAGCAG 1661 AV05141-19-1.um CUCCGGGGCUCUGCCAAUA 1555 UAUUGGCAGAGCCCCGGAG 1662 AV05142-19-1.um CGGGGCUCUGCCAAUCGCA 1556 UGCGAUUGGCAGAGCCCCG 1663 AV05143-19-1.um ACAGCGAUCUGUGACAACA 1557 UGUUGUCACAGAUUCGCUGU 1664 AV05144-19-1.um CAAGACUCCUUCAUGUACA 1558 UGUACAUGAAGGAGUCUUG 1665 AV05145-19-1.um AAGACUCCUUCAUGUACGA 1559 UCGUACAUGAAGGAGUCUU 1666 AV05146-19-1.um ACUCCUUCAUGUACGACAA 1560 UUGUCGUACAUGAAGGAGU 1667 AV05147-19-1.um UCUACCUGGUGCUAGAUGA 1561 UCAUCUAGCACCAGGUAGA 1668 AV05148-19-1.um AUGGAUCAGACAGCAUUGA 1562 UCAAUGCUGUCUGAUCCAU 1669 AV05149-19-1.um ACAGGAGCCAAAAAGUGUA 1563 UACACUUUUUGGCUCCUGU 1670 AV05150-19-1.um CAGGAGCCAAAAAGUGUCA 1564 UGACACUUUUUGGCUCCUG 1671 AV05151-19-1.um AGGAGCCAAAAAGUGUCUA 1565 UAGACACUUUUUGGCUCCU 1672 AV05152-19-1.um AAAAAGUGUCUAGUCAACA 1566 UGUUGACUAGACACUUUUU 1673 AV05153-19-1.um GUGUCUAGUCAACUUAAUA 1567 UAUUAAGUUGACUAGACAC 1674 AV05154-19-1.um UGUCUAGUCAACUUAAUUA 1568 UAAUUAAGUUGACUAGACA 1675 AV05155-19-1.um GUCAACUUAAUUGAGAAGA 1569 UCUUCUCAAUUAAGUUGAC 1676 AV05156-19-1.um UCAACUUAAUUGAGAAGGA 1570 UCCUUCUCAAUUAAGUUGA 1677 AV05157-19-1.um AUGGUGUGAAGCCAAGAUA 1571 UAUCUUGGCUUCACACCAU 1678 AV05158-19-1.um CAGGCAGUGUACAGCAUGA 1572 UCAUGCUGUACACUGCCUG 1679 AV05159-19-1.um AGUGUACAGCAUGAUGAGA 1573 UCUCAUCAUGCUGUACACU 1680 AV05160-19-1.um UGAUGGAUUGCACAACAUA 1574 UAUGUUGUGCAAUCCAUCA 1681 AV05161-19-1.um GAUGGAUUGCACAACAUGA 1575 UCAUGUUGUGCAAUCCAUC 1682 AV05162-19-1.um CCAAUUACUGUCAUUGAUA 1576 UAUCAAUGACAGUAAUUGG 1683 AV05163-19-1.um AGGGAGGAUUAUCUGGAUA 1577 UAUCCAGAUAAUCCUCCCU 1684 AV05164-19-1.um CAAGUGAACAUCAAUGCUA 1578 UAGCAUUGAUGUUCACUUG 1685 AV05165-19-1.um AAAGACAAUGAGCAACAUA 1579 UAUGUUGCUCAUUGUCUUU 1686 AV05166-19-1.um CAUCAAGAAUGGGGAUAAA 1580 UUUAUCCCCAUUCUUGAUG 1687 AV05167-19-1.um AUCAAGAAUGGGGAUAAGA 1581 UCUUAUCCCCAUUCUUGAU 1688 AV05168-19-1.um UCAAGAAUGGGGAUAAGAA 1582 UUCUUAUCCCCAUUCUUGA 1689 AV05169-19-1.um CAAGAAUGGGGAUAAGAAA 1583 UUUCUUAUCCCCAUUCUUG 1690 AV05170-19-1.um AAGAAUGGGGAUAAGAAAA 1584 UUUUCUUAUCCCCAUUCUU 1691 AV05171-19-1.um AGAAUGGGGAUAAGAAAGA 1585 UCUUUCUUAUCCCCAUUCU 1692 AV05172-19-1.um GAAUGGGGAUAAGAAAGGA 1586 UCCUUUCUUAUCCCCAUUC 1693 AV05173-19-1.um CUAUGACAAAGUCAAGGAA 1587 UUCCUUGACUUUGUCAUAG 1694 AV05174-19-1.um UGCUGACCCCAAUACUUGA 1588 UCAAGUAUUGGGGUCAGCA 1695 AV05175-19-1.um AGUAGUGGAUGUCUGCAAA 1589 UUUGCAGACAUCCACUACU 1696 AV05176-19-1.um GGAUGUCUGCAAAAACCAA 1590 UUGGUUUUUGCAGACAUCC 1697 AV05177-19-1.um AAGAUGAGGAUUUGGGUUA 1591 UAACCCAAAUCCUCAUCUU 1698 AV05178-19-1.um GGGGUUUCCUGCUGGACAA 1592 UUGUCCAGCAGGAAACCCC 1699 AV06328-19-1.um CAAUGUGAGUGAUGAGAUU 1593 AAUCUCAUCACUCACAUUG 1700 AV06329-19-1.um AAAAGUGUCUAGUCAACUA 1594 UAGUUGACUAGACACUUUU 1701 AV06330-19-1.um GCGGGAUUGAAUUAAAACA 1595 UGUUUUAAUUCAAUCCCGC 1702

The sense chain sequences of AV02373.um, AV02375.um, AV02379.um, AV02388.um, AV02554.um, and AV02584.um are shown in Table 1, corresponding to the nucleotide sequences at positions 488-508, 491-511, 496-516, 518-538, 2242-2262, and 2444-2464 of SEQ ID NO: 1, respectively.

Table 2 lists certain chemically modified CFB RNAi agent antisense and sense strand sequences of the present invention. In some embodiments of the methods of the present invention, an RNAi agent having a polynucleotide sequence shown in Table 2 is administered to a cell and/or a subject. In some embodiments of the methods of the present invention, an RNAi agent having a polynucleotide sequence shown in Table 2 is administered to a subject. In some embodiments of the present invention, the RNAi agent administered to a subject comprises the duplex identified in the first row of Table 2, and comprises sequence modifications in the sense strand and antisense strand sequences shown in the third and sixth columns of the same row of Table 2, respectively. In some embodiments of the methods of the present invention, the sequences shown in Table 2 may be attached to (also referred to herein as "linked to") a compound capable of delivering the RNAi agent to cells and/or tissues of a subject. Non-limiting examples of delivery compounds that can be used in certain embodiments of the present invention are GalNAc-containing compounds or GLS-15*-containing compounds. In Table 2, the first column represents the duplex AV# of the base sequence shown in Table 1. Table 2 discloses the duplex AV# and also shows the chemical modifications contained in the sense and antisense sequences of the duplex. For example, Table 1 shows the base single strand sequences SEQ ID NO: 7 (sense) and SEQ ID NO: 469 (antisense), which together form a duplex duplex identified as: Duplex AV# AV02358.um, while Table 2 lists duplex AV# AV02358, indicating that duplexes of SEQ ID NO: 931 and SEQ ID NO: 1162 comprise the base sequences of SEQ ID NO: 7 and SEQ ID NO: 469, respectively, but have the chemical modifications shown in the sense and antisense sequences shown in the third and sixth columns, respectively. "Sense Strand SS#" in the second column of Table 2 is the assigned identifier for the sense sequence (including modifications) shown in the third column of the same row. "Antisense AS#" in the fifth column of Table 2 is the assigned identifier for the antisense sequence (including modifications) shown in the sixth column.

Table 2 provides the antisense and sense strand sequences of CFB RNAi agents with chemical modifications. All sequences are represented from 5' to 3'. These sequences were used in certain in vitro assay studies described herein. Double chain AV# Meaningful chain SS# Sense chain sequence Serial Number Antisense chain AS# Antisense chain sequence Serial Number AV02358 AV02358-SS g*a*ggucuaGfgUfcUfggaguu*u*a 931 AV02358-AS u*Af*aacuCfcagaCfcUfaGfacc*u*c 1162 AV02359 AV02359-SS g*a*ggucugGfaGfuUfucagcu*u*a 932 AV02359-AS u*Af*agcuGfaaacUfcCfaGfacc*u*c 1163 AV02360 AV02360-SS g*g*gucuggAfgUfuUfcagcuu*g*a 933 AV02360-AS u*Cf*aaGfc(uUNA)gaaaCfuCfcAfgac*c*c 1164 AV02361 AV02361-SS c*g*ucuggaGfuUfuCfagcuug*g*a 934 AV02361-AS u*Cf*caAfg(cUNA)ugaaAfcUfcCfaga*c*g 1165 AV02362 AV02362-SS c*g*aguuucAfgCfuUfggacac*u*a 935 AV02362-AS u*Af*guGfu(cUNA)caagCfuGfaAfacu*c*g 1166 AV02363 AV02363-SS c*a*guuucaGfcUfuGfgacacu*g*a 936 AV02363-AS u*Cf*agUfg(uUNA)ccaaGfcUfgAfaac*u*g 1167 AV02364 AV02364-SS g*c*cuuccgAfcUfuCfuccaag*a*a 937 AV02364-AS u*Uf*cuugGfagaaGfuCfgGfaag*g*c 1168 AV02365 AV02365-SS g*c*guguguCfcUfuCfuggcuu*c*a 938 AV02365-AS u*Gf*aaGfc(cUNA)agaaGfgAfcAfcac*g*c 1169 AV02366 AV02366-SS c*u*guguccUfuCfuGfgcuucu*a*a 939 AV02366-AS u*Uf*agaaGfccagAfaGfgAfcac*a*g 1170 AV02367 AV02367-SS c*g*aaggcaGfaGfuGfcagagc*a*a 940 AV02367-AS u*Uf*gcUfc(uUNA)gcacUfcUfgCfcuu*c*g 1171 AV02368 AV02368-SS c*a*cuucgaGfaAfcGfgggaau*a*a 941 AV02368-AS u*Uf*auucCfccguUfcUfcGfaag*u*g 1172 AV02369 AV02369-SS g*c*cuacuaCfaAfuGfugagug*a*a 942 AV02369-AS u*Uf*cacuCfacauUfgUfaGfuag*g*c 1173 AV02370 AV02370-SS g*u*acuacaAfuGfuGfagugau*g*a 943 AV02370-AS u*Cf*aucaCfucacAfuUfgUfagu*a*c 1174 AV02371 AV02371-SS g*a*cuacaaUfgUfgAfgugaug*a*a 944 AV02371-AS u*Uf*caucAfcucaCfaUfuGfuag*u*c 1175 AV02372 AV02372-SS g*c*uacaauGfuGfaGfugauga*g*a 945 AV02372-AS u*Cf*ucauCfacucAfcAfuUfgua*g*c 1176 AV02373 AV02373-SS g*a*caauguGfaGfuGfaugaga*u*a 946 AV02373-AS u*Af*ucucAfucacUfcAfcAfuug*u*c 1177 AV02374 AV02374-SS g*a*augugaGfuGfaUfgagauc*u*a 947 AV02374-AS u*Af*gaucUfcaucAfcUfcAfcau*u*c 1178 AV02375 AV02375-SS g*a*ugugagUfgAfuGfagaucu*c*a 948 AV02375-AS u*Gf*agauCfucauCfaCfuCfaca*u*c 1179 AV02376 AV02376-SS g*u*gugaguGfaUfgAfgaucuc*u*a 949 AV02376-AS u*Af*gagaUfcucaUfcAfcUfcac*a*c 1180 AV02377 AV02377-SS g*g*ugagugAfuGfaGfaucucu*u*a 950 AV02377-AS u*Af*agagAfucucAfuCfaCfuca*c*c 1181 AV02378 AV02378-SS g*g*agugauGfaGfaUfcucuuu*c*a 951 AV02378-AS u*Gf*aaagAfgaucUfcAfuCfacu*c*c 1182 AV02379 AV02379-SS c*a*gugaugAfgAfuCfucuuuc*c*a 952 AV02379-AS u*Gf*gaaaGfagauCfuCfaUfcac*u*g 1183 AV02380 AV02380-SS g*g*ugaugaGfaUfcUfcuuucc*a*a 953 AV02380-AS u*Uf*ggaaAfgagaUfcUfcAfuca*c*c 1184 AV02381 AV02381-SS c*u*gaugagAfuCfuCfuuucca*c*a 954 AV02381-AS u*Gf*uggaAfagagAfuCfuCfauc*a*g 1185 AV02382 AV02382-SS g*g*agaucuCfuUfuCfcacugc*u*a 955 AV02382-AS u*Af*gcAfg(uUNA)ggaaAfgAfgAfucu*c*c 1186 AV02383 AV02383-SS g*g*aucucuUfuCfcAfcugcua*u*a 956 AV02383-AS u*Af*uagcAfguggAfaAfgAfgau*c*c 1187 AV02384 AV02384-SS g*c*ucuuucCfaCfuGfcuauga*c*a 957 AV02384-AS u*Gf*ucauAfgcagUfgGfaAfaga*g*c 1188 AV02385 AV02385-SS g*u*ccacugCfuAfuGfacgguu*a*a 958 AV02385-AS u*Uf*aaccGfucauAfgCfaGfugg*a*c 1189 AV02386 AV02386-SS g*c*acugcuAfuGfaCfgguuac*a*a 959 AV02386-AS u*Uf*guaaCfcgucAfuAfgCfagu*g*c 1190 AV02387 AV02387-SS g*c*ugcuauGfaCfgGfuuacac*u*a 960 AV02387-AS u*Af*guguAfaccgUfcAfuAfgca*g*c 1191 AV02388 AV02388-SS g*g*cuaugaCfgGfuUfacacuc*u*a 961 AV02388-AS u*Af*gaguGfuaacCfgUfcAfuag*c*c 1192 AV02389 AV02389-SS c*c*uaugacGfgUfuAfcacucu*c*a 962 AV02389-AS u*Gf*agAfg(uUNA)guaaCfcGfuCfaua*g*g 1193 AV02390 AV02390-SS g*c*agcgauCfuGfuGfacaacg*g*a 963 AV02390-AS u*Cf*cgUfu(gUNA)ucacAfgAfuCfgcu*g*c 1194 AV02391 AV02391-SS g*c*aguaccGfcCfuUfgaagac*a*a 964 AV02391-AS u*Uf*gucuUfcaagGfcGfgUfacu*g*c 1195 AV02392 AV02392-SS g*g*aagacaGfcGfuCfaccuac*c*a 965 AV02392-AS u*Gf*guAfg(gUNA)ugacGfcUfgUfcuu*c*c 1196 AV02393 AV02393-SS g*c*ugccaaGfaCfuCfcuucau*g*a 966 AV02393-AS u*Cf*augaAfggagUfcUfuGfgca*g*c 1197 AV02394 AV02394-SS c*u*ggccgaAfgCfuUfuccugu*c*a 967 AV02394-AS u*Gf*acAfg(gUNA)aaagCfuUfcGfgcc*a*g 1198 AV02395 AV02395-SS c*a*cagagaCfcAfuAfgaagga*g*a 968 AV02395-AS u*Cf*ucCfu(uUNA)cuauGfgUfcUfcug*u*g 1199 AV02396 AV02396-SS 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AV05163-SS g*a*agggagGfaUfuAfucugga*u*a 1737 AV05163-AS u*Af*uccaGfauaaUfcCfuCfccu*u*c 1791 AV05164 AV05164-SS g*c*caagugAfaCfaUfcaaugc*u*a 1738 AV05164-AS u*Af*gcauUfgaugUfuCfaCfuug*g*c 1792 AV05165 AV05165-SS g*g*aaagacAfaUfgAfgcaaca*u*a 1739 AV05165-AS u*Af*uguuGfcucaUfuGfuCfuuu*c*c 1793 AV05166 AV05166-SS g*a*caucaaGfaAfuGfgggaua*a*a 1740 AV05166-AS u*Uf*uaucCfccauUfcUfuGfaug*u*c 1794 AV05167 AV05167-SS g*c*aucaagAfaUfgGfggauaa*g*a 1741 AV05167-AS u*Cf*uuauCfcccaUfuCfuUfgau*g*c 1795 AV05168 AV05168-SS g*a*ucaagaAfuGfgGfgauaag*a*a 1742 AV05168-AS u*Uf*cuuaUfccccAfuUfcUfuga*u*c 1796 AV05169 AV05169-SS g*u*caagaaUfgGfgGfauaaga*a*a 1743 AV05169-AS u*Uf*ucuuAfucccCfaUfuCfuug*a*c 1797 AV05170 AV05170-SS g*c*aagaauGfgGfgAfuaagaa*a*a 1744 AV05170-AS u*Uf*uucuUfauccCfcAfuUfcuu*g*c 1798 AV05171 AV05171-SS g*a*agaaugGfgGfaUfaagaaa*g*a 1745 AV05171-AS u*Cf*uuucUfuaucCfcCfaUfucu*u*c 1799 AV05172 AV05172-SS g*a*gaauggGfgAfuAfagaaag*g*a 1746 AV05172-AS u*Cf*cuuuCfuuauCfcCfcAfuuc*u*c 1800 AV05173 AV05173-SS c*g*cuaugaCfaAfaGfucaagg*a*a 1747 AV05173-AS u*Uf*ccuuGfacuuUfgUfcAfuag*c*g 1801 AV05174 AV05174-SS g*a*ugcugaCfcCfcAfauacuu*g*a 1748 AV05174-AS u*Cf*aaguAfuuggGfgUfcAfgca*u*c 1802 AV05175 AV05175-SS c*g*aguaguGfgAfuGfucugca*a*a 1749 AV05175-AS u*Uf*ugcaGfacauCfcAfcUfacu*c*g 1803 AV05176 AV05176-SS c*u*ggauguCfuGfcAfaaaacc*a*a 1750 AV05176-AS u*Uf*gguuUfuugcAfgAfcAfucc*a*g 1804 AV05177 AV05177-SS g*c*aagaugAfgGfaUfuugggu*u*a 1751 AV05177-AS u*Af*acccAfaaucCfuCfaUfcuu*g*c 1805 AV05178 AV05178-SS g*a*gggguuUfcCfuGfcuggac*a*a 1752 AV05178-AS u*Uf*guCfc(aUNA)gcagGfaAfaCfccc*u*c 1806 AV06327 AV06327-SS c*a*auguGfaGfuGfaugagau*u 1753 AV06327-AS a*Af*ucucAfucacUfcAfcAfu*u*g 1807 AV06328 AV06328-SS g*a*caauguGfaGfuGfaugaga*u*u 1754 AV06328-AS a*Af*ucucAfucacUfcAfcAfuug*u*c 1808 AV06329 AV06329-SS g*a*aaaaguGfuCfuAfgucaac*u*a 1755 AV06329-AS u*Af*guugAfcuagAfcAfcUfuuu*u*c 1809 AV06330 AV06330-SS c*c*gcgggaUfuGfaAfuuaaaa*c*a 1756 AV06330-AS u*Gf*uuuuAfauuCfaaUfcCfcgc*g*g 1810

Table 3 shows certain chemically modified CFBRNAi agent antisense and sense strand sequences of the present invention. In some embodiments of the methods of the present invention, the RNAi agents shown in Table 3 are administered to cells and/or subjects. In some embodiments of the methods of the present invention, RNAi agents having polynucleotide sequences shown in Table 3 are administered to subjects. In some embodiments of the present invention, the RNAi agent administered to the subject includes the duplex identified in a row of the first column of Table 3, and includes sequence modifications and/or delivery compounds shown in the sense strand of the third column and the antisense strand of the sixth column of the same row in Table 3, respectively. These sequences are used in certain in vivo test studies described elsewhere herein. In some embodiments of the methods of the present invention, the sequences shown in Table 3 can be attached (also referred to herein as "bound to") a compound for delivery, a non-limiting example of which is a GalNAc-containing compound, wherein the delivery compound is identified as "GLX-n" on the sense strand in the third column of Table 3. As used herein, "GLX-n" is used to represent a "GLS-n*" or "GLO-n" delivery compound ("X" can be "S" or "O"), and GLX-0 can be any "GLS-n*" and "GLO-n" delivery compound that can be attached to the 3' end of an oligonucleotide during synthesis. As used herein and shown in Table 3, "GLX-n" is used to indicate that the attached GalNAc-containing compound is any one of the following compounds: GLS-1*, GLS-2*, GLS-3*, GLS-4*, GLS-5*, GLS-6*, GLS-7*, GLS-8*, GLS-9*, GLS-10*, GLS-11*, GLS-12*, GLS-13*, GLS-14*, GLS-15*, GLS-16*, GLO-1, GLO-2, GLO-3, GLO-4, GLO-5, GLO-6, GLO-7, GLO-8, GLO-9, GLO-10, GLO-11, GLO-12, GLO-13, GLO-14, GLO-15 and GLO-16, the structures of each of which are provided elsewhere herein. Those skilled in the art will be able to prepare and use the dsRNA compounds of the invention wherein the attached delivery compound is any one of GLS-1*, GLS-2*, GLS-3*, GLS-4*, GLS-5*, GLS-6*, GLS-7*, GLS-8*, GLS-9*, GLS-10*, GLS-11*, GLS-12*, GLS-13*, GLS-14*, GLS-15*, GLS-16*, GLO-1, GLO-2, GLO-3, GLO-4, GLO-5, GLO-6, GLO-7, GLO-8, GLO-9, GLO-10, GLO-11, GLO-12, GLO-13, GLO-14, GLO-15 and GLO-16. The first column of Table 3 provides the duplex AD# assigned to the duplex of the sense and antisense sequences in the row. For example, duplex AD#AD01093 is a duplex of sense chain SEQIDNO:1393 and antisense chain SEQIDNO:1441. Each row in Table 3 provides a sense chain and an antisense chain, and discloses the duplex of the sense chain and antisense chain shown. The "sense chain SS#" in the second column of Table 3 is the assigned identifier of the sense sequence (including modifications) shown in the third column of the same row. The "antisense chain AS#" in the fifth column of Table 3 is the assigned identifier of the antisense sequence (including modifications) shown in the sixth column. The identifiers for certain linked GalNAc-containing "GLO-n" or "GLS-n*" compounds are shown as GLS-5*, GLS-15*, or GLX-0, and it should be understood that another "GLO-n" or "GLS-n*" compound can be substituted for the compound shown as GLO-0, and the resulting compounds are included in the embodiments of the methods and/or compositions of the present invention.

Table 3 provides chemically modified CFB RNAi agent antisense and sense strand sequences. All sequences are shown 5' to 3'. The sequences were used in certain in vivo testing studies described elsewhere herein. The delivery molecules used in the in vivo studies were indicated as "GLO-n" or "GLS-n*" at the 3' or 5' end of each sense strand. Dual Chain AD# Meaningful chain SS# Sense chain sequence Serial Number Antisense chain AS# Antisense chain sequence Serial Number AD01093 AD01093-SS (GLS-15*)(imann*)gacaauguGfaGfuGfaugagau*a*(imann) 1393 AD01093-AS u*Af*ucucAfucacUfcAfcAfuug*u*c 1441 AD01094 AD01094-SS (GLS-15*)(imann*)cccaaaaaGfuGfuCfuagucaa*a*(imann) 1394 AD01094-AS u*Uf*ugacUfagacAfcUfuUfuug*g*g 1442 AD01095 AD01095-SS (GLS-15*)(imann*)gccaauuaCfuGfuCfauugaug*a*(imann) 1395 AD01095-AS u*Cf*aucaAfugacAfgUfaAfuug*g*c 1443 AD01096 AD01096-SS (GLS-15*)(imann*)guuacuguCfaUfuGfaugagau*a*(imann) 1396 AD01096-AS u*Af*ucucAfucaaUfgAfcAfgua*a*c 1444 AD01097 AD01097-SS (GLS-15*)(imann*)gucuggauGfuCfuAfuguguuu*a*(imann) 1397 AD01097-AS u*Af*aacaCfauagAfcAfuCfcag*a*c 1445 AD01098 AD01098-SS (GLS-15*)(imann*)gagaaagaCfaAfuGfagcaaca*a*(imann) 1398 AD01098-AS u*Uf*guugCfucauUfgUfcUfuuc*u*c 1446 AD01099 AD01099-SS (GLS-15*)(imann*)gaugagcaAfcAfuGfuguucaa*a*(imann) 1399 AD01099-AS u*Uf*ugaaCfacauGfuUfgCfuca*u*c 1447 AD01100 AD01100-SS (GLS-15*)(imann*)cauaucaaAfgCfuCfuguuugu*a*(imann) 1400 AD01100-AS u*Af*caaaCfagagCfuUfuGfaua*u*g 1448 AD01101 AD01101-SS (GLS-15*)(imann*)ggagaaguCfgUfuUfcauucaa*a*(imann) 1401 AD01101-AS u*Uf*ugaaUfgaaaCfgAfcUfucu*c*c 1449 AD01102 AD01102-SS (GLS-15*)(imann*)gagucguuUfcAfuUfcaaguug*a*(imann) 1402 AD01102-AS u*Cf*aacuUfgaauGfaAfaCfgac*u*c 1450 AD01103 AD01103-SS (GLS-15*)(imann*)ggucguuuCfaUfuCfaaguugg*a*(imann) 1403 AD01103-AS u*Cf*caacUfugaaUfgAfaAfcga*c*c 1451 AD01104 AD01104-SS (GLS-15*)(imann*)gaagaugaGfgAfuUfuggguuu*a*(imann) 1404 AD01104-AS u*Af*aaccCfaaauCfcUfcAfucu*u*c 1452 AD01389 AD01389-SS (GLS-15*)(imann*)gaggucuaGfgUfcUfggaguuu*a*(imann) 1405 AD01389-AS u*Af*aacuCfcagAfccUfaGfacc*u*c 1453 AD01390 AD01390-SS (GLS-15*)(imann*)gaggucugGfaGfuUfucagcuu*a*(imann) 1406 AD01390-AS u*Af*agcuGfaaaCfucCfaGfacc*u*c 1454 AD01391 AD01391-SS (GLS-15*)(imann*)gacuacaUfgUfgAfgugauga*a*(imann) 1407 AD01391-AS u*Uf*caucAfcucAfcaUfuGfuag*u*c 1455 AD01392 AD01392-SS (GLS-15*)(imann*)gaaugugaGfuGfaUfgagaucu*a*(imann) 1408 AD01392-AS u*Af*gaucUfcauCfacUfcAfcau*u*c 1456 AD01393 AD01393-SS (GLS-15*)(imann*)gaugugagUfgAfuGfagaucuc*a*(imann) 1409 AD01393-AS u*Gf*agauCfucaUfcaCfuCfaca*u*c 1457 AD01394 AD01394-SS (GLS-15*)(imann*)gugugaguGfaUfgAfgaucucu*a*(imann) 1410 AD01394-AS u*Af*gagaUfcucAfucAfcUfcac*a*c 1458 AD01395 AD01395-SS (GLS-15*)(imann*)ggagugauGfaGfaUfcucuuuc*a*(imann) 1411 AD01395-AS u*Gf*aaagAfgauCfucAfuCfacu*c*c 1459 AD01396 AD01396-SS (GLS-15*)(imann*)cagugaugAfgAfuCfucuuucc*a*(imann) 1412 AD01396-AS u*Gf*gaaaGfagaUfcuCfaUfcac*u*g 1460 AD01397 AD01397-SS (GLS-15*)(imann*)ggugaugaGfaUfcUfcuuucca*a*(imann) 1413 AD01397-AS u*Uf*ggaaAfgagAfucUfcAfuca*c*c 1461 AD01398 AD01398-SS (GLS-15*)(imann*)cugaugagAfuCfuCfuuuccac*a*(imann) 1414 AD01398-AS u*Gf*uggaAfagaGfauCfuCfauc*a*g 1462 AD01399 AD01399-SS (GLS-15*)(imann*)ggcuaugaCfgGfuUfacacucu*a*(imann) 1415 AD01399-AS u*Af*gaguGfuaaCfcgUfcAfuag*c*c 1463 AD01400 AD01400-SS (GLS-15*)(imann*)ggccaaaaAfgUfgUfcuaguca*a*(imann) 1416 AD01400-AS u*Uf*gacuAfgacAfcuUfuUfugg*c*c 1464 AD01401 AD01401-SS (GLS-15*)(imann*)gaagugucUfaGfuCfaacuuaa*a*(imann) 1417 AD01401-AS u*Uf*uaagUfugaCfuaGfaCfacu*u*c 1465 AD01402 AD01402-SS (GLS-15*)(imann*)ggggaggaUfuAfuCfuggaugu*a*(imann) 1418 AD01402-AS u*Af*caucCfagaUfaaUfcCfucc*c*c 1466 AD01403 AD01403-SS (GLS-15*)(imann*)gagacaauGfaGfcAfacaugug*a*(imann) 1419 AD01403-AS u*Cf*acauGfuugCfucAfuUfguc*u*c 1467 AD01404 AD01404-SS (GLS-15*)(imann*)gugagcaaCfaUfgUfguucaaa*a*(imann) 1420 AD01404-AS u*Uf*uugaAfcacAfugUfuGfcuc*a*c 1468 AD01405 AD01405-SS (GLS-15*)(imann*)gaggauauGfgAfaAfaccugga*a*(imann) 1421 AD01405-AS u*Uf*ccAfg(gUNA)uuuUfccAfuAfucc*u*c 1469 AD01406 AD01406-SS (GLS-15*)(imann*)gaggauauCfaAfaGfcucuguu*a*(imann) 1422 AD01406-AS u*Af*acagAfgcuUfugAfuAfucc*u*c 1470 AD01407 AD01407-SS (GLS-15*)(imann*)gcaaagcuCfuGfuUfugugucu*a*(imann) 1423 AD01407-AS u*Af*gacaCfaaaCfagAfgCfuuu*g*c 1471 AD01408 AD01408-SS (GLS-15*)(imann*)gaagcucuGfuUfuGfugucuga*a*(imann) 1424 AD01408-AS u*Uf*cagaCfacaAfacAfgAfgcu*u*c 1472 AD01409 AD01409-SS (GLS-15*)(imann*)gguucacaAfgAfgAfagucguu*a*(imann) 1425 AD01409-AS u*Af*acgaCfuucUfcuUfgUfgaa*c*c 1473 AD01410 AD01410-SS (GLS-15*)(imann*)cuucacaaGfaGfaAfgucguuu*a*(imann) 1426 AD01410-AS u*Af*aacgAfcuuCfucUfuGfuga*a*g 1474 AD01411 AD01411-SS (GLS-15*)(imann*)gucacaagAfgAfaGfucguuuc*a*(imann) 1427 AD01411-AS u*Gf*aaacGfacuUfcuCfuUfgug*a*c 1475 AD01412 AD01412-SS (GLS-15*)(imann*)gcacaagaGfaAfgUfcguuuca*a*(imann) 1428 AD01412-AS u*Uf*gaaaCfgacUfucUfcUfugu*g*c 1476 AD01413 AD01413-SS (GLS-15*)(imann*)gacaagagAfaGfuCfguuucau*a*(imann) 1429 AD01413-AS u*Af*ugaaAfcgaCfuuCfuCfuug*u*c 1477 AD01414 AD01414-SS (GLS-15*)(imann*)gcaagagaAfgUfcGfuuucauu*a*(imann) 1430 AD01414-AS u*Af*augaAfacgAfcuUfcUfcuu*g*c 1478 AD01415 AD01415-SS (GLS-15*)(imann*)gagagaagUfcGfuUfucauuca*a*(imann) 1431 AD01415-AS u*Uf*gaauGfaaaCfgaCfuUfcuc*u*c 1479 AD01416 AD01416-SS (GLS-15*)(imann*)ggaagucgUfuUfcAfuucaagu*a*(imann) 1432 AD01416-AS u*Af*cuugAfaugAfaaCfgAfcuu*c*c 1480 AD01417 AD01417-SS (GLS-15*)(imann*)cgaugucuGfcAfaAfaaccaga*a*(imann) 1433 AD01417-AS u*Uf*cuggUfuuuUfgcAfgAfcau*c*g 1481 AD01418 AD01418-SS (GLS-15*)(imann*)ggaaggagAfaAfcUfccaagau*a*(imann) 1434 AD01418-AS u*Af*ucuuGfgagUfuuCfuCfcuu*c*c 1482 AD01419 AD01419-SS (GLS-15*)(imann*)caugaggaUfuUfgGfguuuucu*a*(imann) 1435 AD01419-AS u*Af*gaaaAfcccAfaaUfcCfuca*u*g 1483 AD01420 AD01420-SS (GLS-15*)(imann*)ccgugggaUfuGfaAfuuaaaac*a*(imann) 1436 AD01420-AS u*Gf*uuuuAfauuCfaaUfcCfcac*g*g 1484 AD01421 AD01421-SS (GLS-15*)(imann*)cugggauuGfaAfuUfaaaacag*a*(imann) 1437 AD01421-AS u*Cf*uguuUfuaaUfucAfaUfccc*a*g 1485 AD01422 AD01422-SS (GLS-15*)(imann*)ggggauugAfaUfuAfaaacagc*a*(imann) 1438 AD01422-AS u*Gf*cuguUfuuaAfuuCfaAfucc*c*c 1486 AD01423 AD01423-SS (GLS-15*)(imann*)cggauugaAfuUfaAfaacagcu*a*(imann) 1439 AD01423-AS u*Af*gcugUfuuuAfauUfcAfauc*c*g 1487 AD01420-1 AD01420-1-SS (GLS-15*)(imann*)ccgcgggaUfuGfaAfuuaaaac*a*(imann) 1440 AD01420-1-AS u*Gf*uuuuAfauuCfaaUfcCfcgc*g*g 1488 AD01080 AD01080-SS a*a*gagaAfgUfCfGfuuucaucau(L96) 1811 AD01080-AS a*Uf*gaaUfgAfAfacgaCfuUfcucuu*g*u 1882 AD01696 AD01696-SS (GLS-15*)(imann*)gccuacuaCfaAfuGfugaguga*a*(imann) 1812 AD01696-AS u*Uf*cacuCfacaUfugUfaGfuag*g*c 1883 AD01697 AD01697-SS (GLS-15*)(imann*)guacuacaAfuGfuGfagugaug*a*(imann) 1813 AD01697-AS u*Cf*aucaCfucaCfauUfgUfagu*a*c 1884 AD01698 AD01698-SS (GLS-15*)(imann*)gcuacaauGfuGfaGfugaugag*a*(imann) 1814 AD01698-AS u*Cf*ucauCfacuCfacAfuUfgua*g*c 1885 AD01699 AD01699-SS (GLS-15*)(imann*)ggugagugAfuGfaGfaucucuu*a*(imann) 1815 AD01699-AS u*Af*agagAfucuCfauCfaCfuca*c*c 1886 AD01700 AD01700-SS (GLS-15*)(imann*)ggaucucuUfuCfcAfcugcuau*a*(imann) 1816 AD01700-AS u*Af*uagcAfgugGfaaAfgAfgau*c*c 1887 AD01701 AD01701-SS (GLS-15*)(imann*)gcacugcuAfuGfaCfgguuaca*a*(imann) 1817 AD01701-AS u*Uf*guaaCfcguCfauAfgCfagu*g*c 1888 AD01702 AD01702-SS (GLS-15*)(imann*)gcugcuauGfaCfgGfuuacacu*a*(imann) 1818 AD01702-AS u*Af*guguAfaccGfucAfuAfgca*g*c 1889 AD01703 AD01703-SS (GLS-15*)(imann*)cagccaaaAfaGfuGfucuaguc*a*(imann) 1819 AD01703-AS u*Gf*acuaGfacaCfuuUfuUfggc*u*g 1890 AD01704 AD01704-SS (GLS-15*)(imann*)gaaaaaguGfuCfuAfgucaacu*a*(imann) 1820 AD01704-AS u*Af*guugAfcuaGfacAfcUfuuu*u*c 1891 AD01705 AD01705-SS (GLS-15*)(imann*)gaaaguguCfuAfgUfcaacuua*a*(imann) 1821 AD01705-AS u*Uf*aaguUfgacUfagAfcAfcuu*u*c 1892 AD01706 AD01706-SS (GLS-15*)(imann*)cucaacuuAfaUfuGfagaaggu*a*(imann) 1822 AD01706-AS u*Af*ccuuCfucaAfuuAfaGfuug*a*g 1893 AD01707 AD01707-SS (GLS-15*)(imann*)gauugagaAfgGfuGfgcaaguu*a*(imann) 1823 AD01707-AS u*Af*acuuGfccaCfcuUfcUfcaa*u*c 1894 AD01708 AD01708-SS (GLS-15*)(imann*)gagauaugGfuCfuAfgugacau*a*(imann) 1824 AD01708-AS u*Af*ugucAfcuaGfacCfaUfauc*u*c 1895 AD01709 AD01709-SS (GLS-15*)(imann*)guagugacAfuAfuGfccacaua*a*(imann) 1825 AD01709-AS u*Uf*auguGfgcaUfauGfuCfacu*a*c 1896 AD01710 AD01710-SS (GLS-15*)(imann*)gaugaaauCfaAfuUfaugaaga*a*(imann) 1826 AD01710-AS u*Uf*cuucAfuaaUfugAfuUfuca*u*c 1897 AD01711 AD01711-SS (GLS-15*)(imann*)gucaauuaUfgAfaGfaccacaa*a*(imann) 1827 AD01711-AS u*Uf*ugugGfucuUfcaUfaAfuug*a*c 1898 AD01712 AD01712-SS (GLS-15*)(imann*)gaugaagaCfcAfcAfaguugaa*a*(imann) 1828 AD01712-AS u*Uf*ucaaCfuugUfggUfcUfuca*u*c 1899 AD01713 AD01713-SS (GLS-15*)(imann*)gagaccacAfaGfuUfgaaguca*a*(imann) 1829 AD01713-AS u*Uf*gacuUfcaaCfuuGfuGfguc*u*c 1900 AD01714 AD01714-SS (GLS-15*)(imann*)cuugaaguCfaGfgGfacuaaca*a*(imann) 1830 AD01714-AS u*Uf*guuaGfuccCfugAfcUfuca*a*g 1901 AD01715 AD01715-SS (GLS-15*)(imann*)gauuacugUfcAfuUfgaugaga*a*(imann) 1831 AD01715-AS u*Uf*cucaUfcaaUfgaCfaGfuaa*u*c 1902 AD01716 AD01716-SS (GLS-15*)(imann*)gauuggcaAfgGfaUfcgcaaaa*a*(imann) 1832 AD01716-AS u*Uf*uuugCfgauCfcuUfgCfcaa*u*c 1903 AD01717 AD01717-SS (GLS-15*)(imann*)cgaggauuAfuCfuGfgaugucu*a*(imann) 1833 AD01717-AS u*Af*gacaUfccaGfauAfaUfccu*c*g 1904 AD01718 AD01718-SS (GLS-15*)(imann*)guggauguCfuAfuGfuguuugg*a*(imann) 1834 AD01718-AS u*Cf*caaaCfacaUfagAfcAfucc*a*c 1905 AD01719 AD01719-SS (GLS-15*)(imann*)gcaagugaAfcAfuCfaaugcuu*a*(imann) 1835 AD01719-AS u*Af*agcaUfugaUfguUfcAfcuu*g*c 1906 AD01720 AD01720-SS (GLS-15*)(imann*)caacaucaAfuGfcUfuuggcuu*a*(imann) 1836 AD01720-AS u*Af*agccAfaagCfauUfgAfugu*u*g 1907 AD01721 AD01721-SS (GLS-15*)(imann*)gugcuuugGfcUfuCfcaagaaa*a*(imann) 1837 AD01721-AS u*Uf*uucuUfggaAfgcCfaAfagc*a*c 1908 AD01722 AD01722-SS (GLS-15*)(imann*)gaagacaaUfgAfgCfaacaugu*a*(imann) 1838 AD01722-AS u*Af*caugUfugcUfcaUfuGfucu*u*c 1909 AD01723 AD01723-SS (GLS-15*)(imann*)cacaaugaGfcAfaCfauguguu*a*(imann) 1839 AD01723-AS u*Af*acacAfuguUfgcUfcAfuug*u*g 1910 AD01724 AD01724-SS (GLS-15*)(imann*)gaaugagcAfaCfaUfguguuca*a*(imann) 1840 AD01724-AS u*Uf*gaacAfcauGfuuGfcUfcau*u*c 1911 AD01725 AD01725-SS (GLS-15*)(imann*)ggagcaacAfuGfuGfuucaaag*a*(imann) 1841 AD01725-AS u*Cf*uuugAfacaCfauGfuUfgcu*c*c 1912 AD01726 AD01726-SS (GLS-15*)(imann*)cagcaacaUfgUfgUfucaaagu*a*(imann) 1842 AD01726-AS u*Af*cuuuGfaacAfcaUfgUfugc*u*g 1913 AD01727 AD01727-SS (GLS-15*)(imann*)ggucaaggAfuAfuGfgaaaacc*a*(imann) 1843 AD01727-AS u*Gf*guuuUfccaUfauCfcUfuga*c*c 1914 AD01728 AD01728-SS (GLS-15*)(imann*)gggauaucAfaAfgCfucuguuu*a*(imann) 1844 AD01728-AS u*Af*aacaGfagcUfuuGfaUfauc*c*c 1915 AD01729 AD01729-SS (GLS-15*)(imann*)guaugacaAfaGfuCfaaggaca*a*(imann) 1845 AD01729-AS u*Uf*guccUfugaCfuuUfgUfcau*a*c 1916 AD01730 AD01730-SS (GLS-15*)(imann*)gccuugauAfgUfuCfacaagag*a*(imann) 1846 AD01730-AS u*Cf*ucuuGfugaAfcuAfuCfaag*g*c 1917 AD01731 AD01731-SS (GLS-15*)(imann*)gaguggauGfuCfuGfcaaaaac*a*(imann) 1847 AD01731-AS u*Gf*uuuuUfgcaGfacAfuCfcac*u*c 1918 AD01732 AD01732-SS (GLS-15*)(imann*)ggaugaggAfuUfuGfgguuuuc*a*(imann) 1848 AD01732-AS u*Gf*aaaaCfccaAfauCfcUfcau*c*c 1919 AD01093-1 AD01093-1-SS (GLS-15*)(invab*)caauguGfaGfuGfaugagau*u*(invab) 1849 AD01093-1-AS a*Af*ucucAfucacUfcAfcAfu*u*g 1920 AD01393-1 AD01393-1-SS (GLS-15*)(invab*)gaugugagUfgAfuGfagaucuc*a*(invab) 1850 AD01393-1-AS u*Gf*agauCfucauCfaCfuCfaca*u*c 1921 AD01399-1 AD01399-1-SS (GLS-15*)(invab*)ggcuaugaCfgGfuUfacacucu*a*(invab) 1851 AD01399-1-AS u*Af*gaguGfuaacCfgUfcAfuag*c*c 1922 AD02225 AD02225-SS (GLS-15*)g*gaaggaUfuAfUfCfUfggaugucu*a*(invab) 1852 AD02225-AS u*Af*gAfcAfuCfcAfgAfuAfaUfcCfuUf*c*c 1923 AD01704-1 AD01704-1-SS (GLS-15*)(invab*)gaaaaaguGfuCfuAfgucaacu*a*(invab) 1853 AD01704-1-AS u*Af*guugAfcuagAfcAfcUfuuu*u*c 1924 AD02226 AD02226-SS (GLS-15*)(invab*)cuguguccUfuCfuGfgcuucua*a*(invab) 1854 AD02226-AS u*Uf*agaaGfccagAfaGfgAfcac*a*g 1925 AD02227 AD02227-SS (GLS-15*)(invab*)cgaaggcaGfaGfuGfcagagca*a*(invab) 1855 AD02227-AS u*Uf*gcUfc(uUNA)gcacUfcUfgCfcuu*c*g 1926 AD02228 AD02228-SS (GLS-15*)(invab*)ggagaucuCfuUfuCfcacugcu*a*(invab) 1856 AD02228-AS u*Af*gcAfg(uUNA)ggaaAfgAfgAfucu*c*c 1927 AD02229 AD02229-SS (GLS-15*)(invab*)cucuagucAfaCfuUfaauugag*a*(invab) 1857 AD02229-AS u*Cf*ucaaUfuaagUfuGfaCfuag*a*g 1928 AD02230 AD02230-SS (GLS-15*)(invab*)guauggucUfaGfuGfacauaug*a*(invab) 1858 AD02230-AS u*Cf*auauGfucacUfaGfaCfcau*a*c 1929 AD02231 AD02231-SS (GLS-15*)(invab*)cacugaugGfaUfuGfcacaaca*a*(invab) 1859 AD02231-AS u*Uf*guugUfgcaaUfcCfaUfcag*u*g 1930 AD02232 AD02232-SS (GLS-15*)(invab*)cacccaauUfaCfuGfucauuga*a*(invab) 1860 AD02232-AS u*Uf*caauGfacagUfaAfuUfggg*u*g 1931 AD02233 AD02233-SS (GLS-15*)(invab*)gacugucaUfuGfaUfgagaucc*a*(invab) 1861 AD02233-AS u*Gf*gaUfc(uUNA)caucAfaUfgAfcag*u*c 1932 AD02234 AD02234-SS (GLS-15*)(invab*)caggauuaUfcUfgGfaugucua*a*(invab) 1862 AD02234-AS u*Uf*agacAfuccaGfaUfaAfucc*u*g 1933 AD02235 AD02235-SS (GLS-15*)(invab*)gggauuauCfuGfgAfugucuau*a*(invab) 1863 AD02235-AS u*Af*uagaCfauccAfgAfuAfauc*c*c 1934 AD02236 AD02236-SS (GLS-15*)(invab*)cgauuaucUfgGfaUfgucuaug*a*(invab) 1864 AD02236-AS u*Cf*auagAfcaucCfaGfaUfaau*c*g 1935 AD02237 AD02237-SS (GLS-15*)(invab*)cgugaaccAfaGfuGfaacauca*a*(invab) 1865 AD02237-AS u*Uf*gaugUfucacUfuGfgUfuca*c*g 1936 AD02238 AD02238-SS (GLS-15*)(invab*)cgauaucaAfaGfcUfcuguuug*a*(invab) 1866 AD02238-AS u*Cf*aaacAfgagcUfuUfgAfuau*c*g 1937 AD02239 AD02239-SS (GLS-15*)(invab*)cagagagaUfgCfuCfaauaugc*a*(invab) 1867 AD02239-AS u*Gf*cauaUfugagCfaUfcUfcuc*u*g 1938 AD02240 AD02240-SS (GLS-15*)(invab*)gcccuugaUfaGfuUfcacaaga*a*(invab) 1868 AD02240-AS u*Uf*cuugUfgaacUfaUfcAfagg*g*c 1939 AD02241 AD02241-SS (GLS-15*)(invab*)gguuucauUfcAfaGfuuggugu*a*(invab) 1869 AD02241-AS u*Af*caccAfacuuGfaAfuGfaaa*c*c 1940 AD02339 AD02339-SS (GLS-15*)(invab*)guacaaugUfgAfgUfgaugaga*a*(invab) 1870 AD02339-AS u*Uf*cucaUfcacuCfaCfaUfugu*a*c 1941 AD02340 AD02340-SS (GLS-15*)(invab*)gacugcuaUfgAfcGfguuacac*a*(invab) 1871 AD02340-AS u*Gf*uguaAfccguCfaUfaGfcag*u*c 1942 AD02341 AD02341-SS (GLS-15*)(invab*)gcacaggaGfcCfaAfaaagugu*a*(invab) 1872 AD02341-AS u*Af*cacuUfuuugGfcUfcCfugu*g*c 1943 AD02342 AD02342-SS (GLS-15*)(invab*)gcaaaaagUfgUfcUfagucaac*a*(invab) 1873 AD02342-AS u*Gf*uugaCfuagaCfaCfuUfuuu*g*c 1944 AD02343 AD02343-SS (GLS-15*)(invab*)ggugucuaGfuCfaAfcuuaauu*a*(invab) 1874 AD02343-AS u*Af*auuaAfguugAfcUfaGfaca*c*c 1945 AD02344 AD02344-SS (GLS-15*)(invab*)gacaucaaGfaAfuGfgggauaa*a*(invab) 1875 AD02344-AS u*Uf*uaucCfccauUfcUfuGfaug*u*c 1946 AD02345 AD02345-SS (GLS-15*)(invab*)cgaguaguGfgAfuGfucugcaa*a*(invab) 1876 AD02345-AS u*Uf*ugcaGfacauCfcAfcUfacu*c*g 1947 AD02346 AD02346-SS (GLS-15*)(invab*)gcaagaugAfgGfaUfuuggguu*a*(invab) 1877 AD02346-AS u*Af*acccAfaaucCfuCfaUfcuu*g*c 1948 AD02347 AD02347-SS (GLS-15*)(invab*)gagggguuUfcCfuGfcuggaca*a*(invab) 1878 AD02347-AS u*Uf*guCfc(aUNA)gcagGfaAfaCfccc*u*c 1949 AD01093-2 AD01093-2-SS (GLS-15*)g*acaauguGfaGfuGfaugagau*u*(invab) 1879 AD01093-2-AS a*Af*ucucAfucacUfcAfcAfuug*u*c 1950

AD01786 comprises the following nucleotide sequence: u*Uf*gAfaUfgAfaAfcGfa*Cf*u*Uf*c*Uf*cGfuUfuCfaUfuCf*a*Af*(L-96) (SEQ ID NO 1880); AD01787 comprises the following nucleotide sequence: (GLO-15) (MOE-A*)(MOE-T*)(MOE-m5C*)(MOE-m5C*)(MOE-m5C*)(dA*) (m5dC*)(dG*)(m5dC*) (m5dC*)(m5dC*)(m5dC*)(dT*)(dG*)(dT*) (MOE-m5C*)(MOE-m5C*)(MOE-A*)(MOE-G*)(MOE-m5C) (SEQ ID NO 1881); L-96 refers to Jayaprakash, et al. al., (2014) J. Am. Chem. Soc., 136, 16958−16961. Table 4: Abbreviations for nucleotide monomers used in nucleotide sequences. It will be understood that these monomers, when present in an oligonucleotide, are linked to one another via 5'-3' phosphodiester bonds. It will also be understood that when a nucleotide contains a 2'-fluorine modification, the fluorine replaces the hydroxyl group at that position of the parent nucleotide (i.e., it is a 2'-deoxy-2'-fluoro nucleotide). It will also be understood that the abbreviations correspond to omitting the phosphate at the 3'-end when it is a 3'-terminal nucleotide (i.e., they are 3'-OH). The skilled person will understand that these abbreviations are merely identification symbols and will be able to determine the connection pattern of the nucleotide sequences containing these abbreviations. Abbreviation Nucleotides A Adenosine-3'-phosphate C Cytidine-3'-phosphate G Guanosine-3'-phosphate T 5-Methyluridine-3'-phosphate U Uridine-3'-phosphate a 2'-O-methyladenosine-3'-phosphate c 2'-O-methylcytidine-3'-phosphate g 2'-O-methylguanosine-3'-phosphate t 2'-O-methyl-5-methyluridine-3'-phosphate u 2'-O-methyluridine-3'-phosphate A 2'-Fluoroadenosine-3'-phosphate Cf 2'-Fluorocytidine-3'-phosphate Gf 2'-Fluoroguanosine-3'-phosphate Tf 2'-Fluoro-5-methyluridine-3'-phosphate Uf 2'-Fluorouridine-3'-phosphate Af* 2'-Fluoroadenosine-3'-phosphorothioate Cf* 2'-Fluorocytidine-3'-phosphorothioate Gf* 2'-Fluoroguanosine-3'-phosphorothioate Tf* 2'-Fluoro-5-methyluridine-3'-phosphorothioate Uf* 2'-Fluorouridine-3'-phosphorothioate a* 2'-O-methyladenosine-3'-phosphorothioate c* 2'-O-methylcytidine-3'-phosphorothioate g* 2'-O-methylguanosine-3'-phosphorothioate t* 2'-O-methyl-5-methyluridine-3'-phosphorothioate u* 2'-O-methyluridine-3'-phosphorothioate ai 2', 3'-seco-adenosine-3'-phosphate cUA 2', 3'-seco-cytidine-3'-phosphate g 2', 3'-seco-guanosine-3'-phosphate tUNA 2', 3'-seco-5-methyluridine-3'-phosphate uUNA 2', 3'- seco-uridine-3'-phosphate Vp Vinyl-phosphate invab Inverted abasic-5'-phosphate. When located at the 3' end of the chain, it is linked to the adjacent nucleotide via the substituent at its 3' position (3'-3'linkage); when located at the 5' end of the chain, it is linked to the adjacent nucleotide via the substituent at its 5' position (5'-5' internucleotide linkage). It should also be understood that when in the 3'-terminal position, the corresponding invabs of the abbreviation omit the 5'-phosphate (i.e., they are 5'-OH). invab* Inverted abasic-5'-phosphorothioate. When located at the 3' end of the chain, it is linked to the adjacent nucleotide via the substituent at its 3' position (3'-3'linkage); when located at the 5' end of the chain, it is linked to the adjacent nucleotide via the substituent at its 5' position (5'-5' internucleotide linkage). It should also be understood that the abbreviation corresponds to invab*, which omits the 5'-phosphorothioate (i.e., they are 5'-OH) when found at the 3'-terminal position. imann imannWhen at the end of each chain: When imann is in the middle of each chain: imann* Isomannitol sulphothioate imann*When at the end of each chain imann* When in the middle of each chain MOE-A* 2'-Methoxyethyl-adenosine-3'-phosphorothioate MOE-T* 2'-Methoxyethyl-5-methyluridine-3'-phosphorothioate MOE-m5C* 2'-Methoxyethyl-(5-methyl-cytidine)-3'-phosphorothioate m5dC* 2'-Deoxy-(5-methyl-cytidine)-3'-phosphorothioate dA* 2'-deoxyadenosine-3'-phosphorothioate dG* 2'-deoxyguanosine-3'-phosphorothioate dT* 2'-Deoxythymidine-3'-phosphorothioate

In certain embodiments of the invention, the dsRNA (also referred to herein as a "duplex") is a dsRNA disclosed in one of Tables 1-3. Each row in Tables 1-3 discloses a duplex comprising the sense and antisense sequences in that row of the table. In addition to the duplexes disclosed in Tables 1-3, it should be understood that in some embodiments, the duplexes of the invention may include the sense and antisense sequences shown in Tables 1-3 that differ from the sequences shown in Tables 1-3 by zero, one, two, or three nucleotides. Thus, as non-limiting examples, in some embodiments, the antisense strand in the duplex of the present invention can be SEQ ID NO: 484, 490, 497, 665 or 667, which differs from the nucleotides in SEQ ID 484, 490, 497, 665 or 667 by zero, one, two or three nucleotides, respectively.

It should be understood that the sense chain sequence and the antisense chain sequence in the duplex of the present invention can be selected independently. Therefore, the dsRNA of the present invention can include the sense chain and the antisense chain of the duplex disclosed in a row in Table 1-3. Or, in the dsRNA of the present invention, one or both of the sense chain and the antisense chain selected in the dsRNA can include the sequence shown in Table 1-3, but one or both of the sense chain and the antisense chain include 1,2,3 or more nucleobase substitutions from the parent sequence. In some embodiments, the selected sequence can be longer or shorter than its parent sequence. Therefore, the dsRNA agent included in the present invention can but need not include the exact sequence of the sense chain and the antisense chain pair disclosed as a duplex in Table 1-3.

In some embodiments, the dsRNA agent comprises a sense strand and an antisense strand, nucleotide positions 2 to 18 in the antisense strand comprising a region complementary to a CFB RNA transcript, wherein the complementary region comprises at least 15 consecutive nucleotides that differ by 0, 1, 2, or 3 nucleotides from one of the antisense sequences listed in one of Tables 1-3, and optionally comprises a targeting ligand. In some cases, the region complementary to the CFB RNA transcript comprises at least 15, 16, 17, 18, or 19 consecutive nucleotides that differ by no more than 3 nucleotides from one of the antisense sequences listed in one of Tables 1-3. In some embodiments of the dsRNA agents of the invention, the antisense strand of the dsRNA is at least substantially complementary to any one of the target regions of SEQ ID NO: 1 and is provided in any one of Tables 1-3. In some embodiments, the antisense strand of the dsRNA agent of the present invention is fully complementary to any of the target regions of SEQ ID NO: 1 and is provided in any of Tables 1-3. In some embodiments, the dsRNA agent includes a sense strand sequence listed in any of Tables 1-3, and the sense strand sequence is at least substantially complementary to the antisense strand sequence in the dsRNA agent. In other embodiments, the dsRNA agent of the present invention comprises a sense strand sequence listed in any of Tables 1-3, and the sense strand sequence is fully complementary to the antisense strand sequence in the dsRNA agent. In some cases, the dsRNA agent of the present invention comprises an antisense strand sequence listed in any of Tables 1-3. Some embodiments of the dsRNA agents of the present invention comprise a sense sequence and an antisense sequence disclosed as a duplex in any one of Tables 1-3 . As described herein, it should be understood that the sense strand and the antisense strand in the duplex of the present invention can be independently selected.

It is known to those skilled in the art that mismatches in dsRNA are tolerated for efficacy, especially mismatches in the terminal regions of the dsRNA. Some mismatches are more tolerated, such as mismatches of the dsRNA G:U and A:C pairs, which are tolerated for efficacy (Du et el., A systematic analysis of the silencing effects of an active siRNA at all single-nucleotide mismatched target sites. Nucleic Acids Res. 2005 Mar 21;33(5):1671-7. Doi: 10.1093/nar/gki312. Nucleic Acids Res. 2005;33(11):3698). In some embodiments of the methods and compounds of the present invention, the CFB dsRNA agent may contain one or more mismatches with the CFB target sequence. In some embodiments, the CFB dsRNA agents of the present invention do not include mismatches. In some embodiments, the CFB dsRNA agents of the present invention contain no more than 1 mismatch. In some embodiments, the CFB dsRNA agents of the present invention contain no more than 2 mismatches. In some embodiments, the CFB dsRNA agents of the present invention contain no more than 3 mismatches. In some embodiments of the present invention, the antisense strand of the CFB dsRNA agent contains mismatches with the CFB target sequence, and these mismatches are not located in the center of the complementary region. In some embodiments, the antisense strand of a CFB dsRNA agent comprises 1, 2, 3, 4 or more mismatches that are located within the last 5, 4, 3, 2 or 1 nucleotides of one or both of the 5' or 3' ends of the complementary region. The methods described herein and/or methods known in the art can be used to determine whether a CFB dsRNA agent comprising mismatches to a CFB target sequence is effective in inhibiting expression of a CFB gene. Complementarity

As used herein, unless otherwise indicated, the term "complementary" when used to describe a first nucleotide sequence (e.g., a CFB dsRNA agent sense strand or a targeted CFB mRNA) relative to a second nucleotide sequence (e.g., a CFB dsRNA agent antisense strand or a single-stranded antisense polynucleotide) refers to the ability of an oligonucleotide or polynucleotide comprising the first nucleotide sequence to hybridize [form base-pair hydrogen bonds under mammalian physiological conditions (or similar conditions in vitro )] with an oligonucleotide or polynucleotide comprising the second nucleotide sequence and form a double chain or double helical structure under certain conditions. Other conditions, such as physiologically relevant conditions that may be encountered in an organism, may also be applicable. A skilled person will be able to determine the most appropriate set of conditions for testing the complementarity of two sequences based on the ultimate application of the hybridized nucleotides. Complementary sequences include Watson-Crick base pairs or non-Watson-Crick base pairs, including natural or modified nucleotides or nucleotide mimetics, at least to the extent that the above hybridization requirements are met. Sequence identity or complementarity is independent of modification.

Complementary sequences, for example, within the CFB dsRNA described herein, include base pairing of an oligonucleotide or polynucleotide comprising a first nucleotide sequence with an oligonucleotide or polynucleotide comprising a second nucleotide sequence over the entire length of one or both nucleotide sequences. Such sequences may be referred to herein as "completely complementary" to one another. It should be understood that in embodiments, when two oligonucleotides are designed to form one or more single-stranded overhangs upon hybridization, such overhangs are not considered mismatches in the determination of complementarity herein. For example, a CFB dsRNA agent comprises an oligonucleotide of 19 nucleotides in length and another oligonucleotide of 20 nucleotides in length, wherein the longer oligonucleotide comprises a sequence of 19 nucleotides that is completely complementary to the shorter oligonucleotide, but for the purposes described herein, may still be referred to as "completely complementary". Therefore, "complete complementation" as used herein means that all bases (100%) in the contiguous sequence of the first polynucleotide will hybridize with the same number of bases in the contiguous sequence of the second polynucleotide. The contiguous sequence may include all or part of the first or second nucleotide sequence.

As used herein, the term "substantially complementary" means that in a hybridized nucleobase sequence pair, at least about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99%, but not all, of the bases in the contiguous sequence of a first polynucleotide will hybridize with the same number of bases in the contiguous sequence of a second polynucleotide. The term "substantially complementary" may be used to refer to a first sequence relative to a second sequence, such as up to 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30 base pairs (bp) of the two sequences of the duplex comprising one or more, such as at least 1, 2, 3, 4 or 5 mismatched base pairs after hybridization, while retaining the ability to hybridize under conditions most relevant to its ultimate application, such as inhibition of CFB gene expression via the RISC pathway.

The term "partial complement" can be used herein to refer to a hybridized nucleobase sequence pair, wherein at least 75% (but not all) of the bases in the contiguous sequence of the first polynucleotide will hybridize with the same number of bases in the contiguous sequence of the second polynucleotide. In some embodiments, "partial complement" refers to at least 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% of the bases in the contiguous sequence of the first polynucleotide will hybridize with the same number of bases in the contiguous sequence of the second polynucleotide.

The terms "complementary", "completely complementary", "substantially complementary" and "partially complementary" as used herein refer to base matching between the sense strand and the antisense strand of a CFB dsRNA agent, between the antisense strand of a CFB dsRNA agent and a target CFB mRNA sequence, or between a single-stranded antisense oligonucleotide and a target CFB mRNA sequence. It should be understood that the term "antisense strand of a CFB dsRNA agent" can refer to the same sequence of a "CFB antisense polynucleotide agent".

As used herein, the term "substantially identical" or "substantial identity" used in reference to nucleic acid sequences refers to sequences having at least about 85% sequence identity or greater, preferably at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% compared to a reference sequence. The percentage of sequence identity is determined by comparing the two optimally aligned sequences within a comparison window. The percentage is calculated by determining the number of positions at which the same nucleic acid base occurs in the two sequences to generate the number of matching positions, dividing the number of matching positions by the total number of positions in the comparison window, and then multiplying the result by 100 to obtain the percentage of sequence identity. The invention disclosed herein encompasses nucleotide sequences substantially identical to those disclosed herein, such as in Tables 1-3. In some embodiments, the sequences disclosed herein are exactly the same, or at least about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to those disclosed herein, e.g., in Tables 1-3.

As used herein, the term "strand comprising a sequence" means an oligonucleotide comprising a nucleotide strand described by a sequence referred to using standard nucleotide nomenclature. As used herein, the term "double-stranded RNA" or "dsRNA" refers to an RNAi comprising an RNA molecule or a complex of molecules having a hybrid duplex region comprising two antiparallel and substantially or completely complementary nucleic acid strands, which are referred to as having "sense" and "antisense" orientations relative to the target CFB RNA. The duplex region can be any length that allows for specific degradation of the desired target CFB RNA via the RISC pathway, but typically ranges from 9 to 30 base pairs in length, such as 15-30 base pairs in length. Considering duplexes between 9 and 30 base pairs, the duplexes can be any length within that range, for example 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30, and any subranges therein, including but not limited to 15-30 base pairs, 15-26 base pairs, 15-23 base pairs, 15-22 base pairs, 15-21 base pairs, 15-20 base pairs, 15-19 base pairs, 15-18 base pairs, 15-17 base pairs, 18-30 base pairs, 18-26 base pairs, 18- 23 base pairs, 18-22 base pairs, 18-21 base pairs, 18-20 base pairs, 19-30 base pairs, 19-26 base pairs, 19-23 base pairs, 19-22 base pairs, 19-21 base pairs, 19-20 base pairs, 20-30 base pairs, 20-26 Base pairs, 20-25 base pairs, 20-24 base pairs, 20-23 base pairs, 20-22 base pairs, 20-21 base pairs, 21-30 base pairs, 21-26 base pairs, 21-25 base pairs, 21-24 base pairs, 21-23 base pairs or 21-22 base pairs. The length of CFB dsRNA reagents produced in cells by treatment with Dicer and similar enzymes is generally in the range of 19-22 base pairs. One strand of the duplex region of the CFB dsDNA agent contains a sequence that is substantially complementary to a region of the target CFB RNA. The two chains forming the duplex structure may be from a single RNA molecule having at least one self-complementary region, or may be formed by two or more separate RNA molecules. When the duplex region is formed by two chains of a single molecule, the molecule may have a duplex region separated by a single-stranded nucleotide chain (referred to herein as a "hairpin loop") between the 3' end of one chain forming the duplex structure and the 5' end of the corresponding other chain. In some embodiments of the invention, the hairpin loop comprises at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more unpaired nucleotides. When the two substantially complementary strands of a CFB dsRNA agent consist of separate RNA molecules, these molecules need not but can be covalently linked. When the two strands are covalently linked by means other than a hairpin loop, the linking structure is referred to as a "linker." The term "siRNA" is also used herein to refer to the dsRNA agents described herein.

In some embodiments of the invention, a CFB dsRNA agent may include sense and antisense sequences with unpaired nucleotides or nucleotide analogs at one or both ends of the dsRNA agent. Ends without unpaired nucleotides are referred to as "blunt ends" and have no nucleotide overhangs. If both ends of a dsRNA agent are blunt ends, the dsRNA is referred to as "blunt ends". In some embodiments of the invention, the first end of the dsRNA agent is blunt, in some embodiments, the second end of the dsRNA agent is blunt, and in certain embodiments of the invention, both ends of the CFB dsRNA agent are blunt.

In some embodiments of the dsRNA reagent of the present invention, the dsRNA does not have one or two blunt ends. In this case, the end of the dsRNA reagent chain has at least one unpaired nucleotide. For example, when the 3' end of one dsRNA chain extends beyond the 5' end of the other chain, or vice versa, there is a nucleotide overhang. The dsRNA can include an overhang of at least 1, 2, 3, 4, 5, 6 or more nucleotides. The nucleotide overhang can include or be composed of nucleotide/nucleoside analogs (including deoxynucleotides/nucleosides). It should be understood that in some embodiments, the nucleotide overhang is located on the sense strand of the dsRNA reagent, on the antisense strand of the dsRNA reagent, or at both ends of the dsRNA reagent, and the nucleotides of the overhang may be present at the 5' end, 3' end, or both ends of the dsRNA antisense strand or sense strand. In certain embodiments of the present invention, one or more nucleotides in the overhang are substituted with nucleoside phosphorothioates.

As used herein, the term "antisense strand" or "guide strand" refers to the strand of a CFB dsRNA agent that includes a region that is substantially complementary to the CFB target sequence. As used herein, the term "sense strand" or "passenger strand" refers to the strand of a CFB dsRNA agent that includes a region that is substantially complementary to the antisense strand region of the CFB dsRNA agent. Modifications

In some embodiments of the present invention, the RNA of the CFB RNAi agent is chemically modified to enhance stability and/or one or more other beneficial properties. The nucleic acids in certain embodiments of the present invention can be synthesized and/or modified by methods well established in the art, for example, "Current protocols in Nucleic Acid Chemistry," Beaucage, S. L. et al. (Eds.), John Wiley & Sons, Inc., New York, N.Y., USA, which is incorporated herein by reference. Modifications that may be present in certain embodiments of the CFB dsRNA agents of the invention include, for example, (a) terminal modifications, such as 5'-terminal modifications (phosphorylation, conjugation, reverse ligation, etc.), 3'-terminal modifications (conjugation, DNA nucleotides, reverse ligation, etc.); (b) base modifications, replacement of bases with stable bases, destabilizing bases, or base pairs with an expanded repertoire of partners, removal of bases (abasic nucleotides) or conjugated bases, (c) sugar modifications (e.g., at the 2' position or the 4' position) or replacement of sugars, and (d) backbone modifications, including modification or replacement of phosphodiester bonds. Specific examples of RNA compounds that can be used in certain embodiments of the CFB dsRNA agents, CFB antisense polynucleotides, and CFB sense polynucleotides of the present invention include, but are not limited to, RNAs that contain modified backbones or do not contain natural internucleoside bonds. As a non-limiting example, RNAs with modified backbones may not have phosphorus atoms in the backbone. RNAs that do not have phosphorus atoms in their internucleoside backbones may be referred to as oligonucleosides. In certain embodiments of the present invention, modified RNAs have phosphorus atoms in their internucleoside backbones.

It should be understood that the term "RNA molecule" or "RNA" or "ribonucleic acid molecule" encompasses not only RNA molecules expressed or found in nature, but also RNA analogs and derivatives comprising one or more ribonucleotide/ribonucleoside analogs or derivatives described herein or known in the art. The terms "ribonucleoside" and "ribonucleotide" are used interchangeably herein. RNA molecules can be modified in the nucleobase structure or the ribose-phosphate backbone structure, for example as described below, and the molecule comprising the ribonucleoside analog or derivative must retain the ability to form a double chain. As non-limiting examples, the RNA molecule may further include at least one modified ribonucleoside, including but not limited to a 2'-O-methyl modified nucleoside, a nucleoside containing a 5' phosphorothioate group, a terminal nucleoside linked to a cholesterol derivative or a dodecanoic acid bisdecylamide group, a locked nucleoside, a debasic nucleoside, a 2'-deoxy-2'-fluoro modified nucleoside, a 2'-amino modified nucleoside, a 2'-alkyl modified nucleoside, a morpholino nucleoside, a phosphoramidate, or a nucleoside containing an unnatural base, or any combination thereof. In some embodiments of the invention, the RNA molecule comprises at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or up to the full-length CFB dsRNA agent molecule ribonucleosides that are modified ribonucleosides. The modification of each of these multiple modified ribonucleosides in the RNA molecule does not have to be the same.

In some embodiments, the dsRNA agents, CFB antisense polynucleotides and/or CFB sense polynucleotides of the present invention may comprise one or more independently selected modified nucleotides and/or one or more independently selected non-phosphodiester bonds. Herein, the term "independently selected" used to refer to selected elements (e.g., modified nucleotides, non-phosphodiester bonds, etc.) means that two or more selected elements may be, but need not be, identical to each other.

As used herein, "nucleotide bases", "nucleotides" or "nucleobases" are heterocyclic pyrimidine or purine compounds that are standard components of all nucleic acids and include the forming nucleotides adenine, guanine, cytosine, thymine and uracil. Nucleobases can be further modified to include, but are not intended to be limited to: universal bases, hydrophobic bases, mixed bases, size-expanded bases and fluorinated bases. The term "ribonucleotide" or "nucleotide" can be used herein to refer to unmodified nucleotides, modified nucleotides or substitute replacement moieties. Those skilled in the art will recognize that guanine, cytosine, adenine and uracil can be substituted with other moieties without substantially changing the base pairing properties of the oligonucleotides containing nucleotides with such replacement moieties.

In one embodiment, the modified RNA contemplated for use in the methods and compositions described herein is a peptide nucleic acid (PNA) that has the ability to form a desired double-stranded structure and allows or mediates specific degradation of the target RNA through the RISC pathway. In certain embodiments of the present invention, the CFB RNA interferor includes a single-stranded RNA that interacts with a target CFB RNA sequence to direct the cleavage of the target CFB RNA.

The modified RNA backbone can include, for example, phosphorothioates, chiral phosphorothioates, phosphorodithioates, phosphotriesters, aminoalkylphosphotriesters, methyl and other alkylphosphonates, including 3'-alkylenephosphonates and chiral phosphonates, phosphinates, phosphoramidates, including 3'-aminophosphoramidates and aminoalkylphosphoramidates, thiophosphoramidates, thioalkylphosphotriesters and borophosphates with normal 3'-5' connection, 2'-5' connection analogs of these, and those with reversed polarity, wherein adjacent nucleoside unit pairs are connected at 3'-5' to 5'-3' connection or 2'-5' to 5'-2' connection. Various salts, mixed salts and free acid forms are also included. Methods for preparing phosphorus-containing linkages are routinely practiced in the art, and such methods can be used to prepare certain modified CFB dsRNA reagents, certain modified CFB antisense polynucleotides and/or certain modified CFB sense polynucleotides of the present invention.

The modified RNA backbone that does not contain a phosphorus atom has a backbone formed by short-chain alkyl or cycloalkyl nucleoside bonds, mixed heteroatom and alkyl or cycloalkyl nucleoside bonds, or one or more short-chain heteroatom or heterocyclic nucleoside bonds. These include backbones with morpholino bonds (formed in part by the sugar portion of the nucleoside); siloxane backbones; sulfide, sulfonyl and sulfonyl backbones; formyl and thioformyl backbones; methyleneformyl and thioformyl backbones; olefin-containing backbones; sulfamate backbones; methyleneimino and methylenehydrazino backbones; sulfonate and sulfonamide backbones; amide backbones; and other backbones with mixed N, O, S and _CH2 components. Methods for preparing modified RNA backbones that do not include phosphorus atoms are routine in the art, and such methods can be used to prepare certain modified CFB dsRNA agents, certain modified CFB antisense polynucleotides, and/or certain modified CFB sense polynucleotides of the present invention.

In certain embodiments of the present invention, RNA mimics are included in CFB dsRNA, CFB antisense polynucleotides and/or CFB sense polynucleotides, such as but not limited to: replacing the sugar and nucleoside bonds of the nucleotide units, i.e., the main chain, with new groups. In such embodiments, the base units are retained to hybridize with appropriate CFB nucleic acid target compounds. One such oligomeric compound, an RNA mimetic that has been shown to have excellent hybridization properties, is called a peptide nucleic acid (PNA). In PNA compounds, the sugar main chain of RNA is replaced with an amide-containing main chain, particularly an aminoethylglycine main chain. The nucleobase is retained and is directly or indirectly bound to the nitrogen-nitrogen atom of the amide portion of the main chain. Methods for preparing RNA mimetics are routinely practiced in the art, and such methods can be used to prepare certain modified CFB dsRNA agents of the present invention.

Some embodiments of the invention include RNAs having phosphorothioate backbones and oligonucleosides having heteroatom backbones, and particularly _-CH2- NH- _CH2- , _-CH2 -N( _CH3 )-O- _CH2- [referred to as a methylene(methylimino) or MMI backbone], -CH2 _- ON( _CH3 ) _-CH2- , _-CH2- N( _CH3 )-N( _CH3 ) _-CH2- , and -N( _CH3 ) _-CH2- [where the natural phosphodiester backbone is represented as -OPO- _CH2- ]. Methods for preparing RNAs having phosphorothioate backbones and oligonucleosides having heteroatom backbones are routinely practiced in the art, and such methods can be used to prepare certain modified CFB dsRNA reagents, certain CFB antisense polynucleotides and/or certain CFB sense polynucleotides of the present invention.

The modified RNA may also contain one or more substituted sugar moieties. The CFB dsRNA, CFB antisense polynucleotides and/or CFB sense polynucleotides of the present invention may comprise one of the following at the 2' position: OH; F; O-, S- or N-alkyl; O-, S- or N-alkenyl; O-, S- or N-alkynyl; or O-alkyl-O-alkyl, wherein the alkyl, alkenyl and alkynyl groups may be substituted or unsubstituted C ₁ to C ₁₀ alkyl or C ₂ to C ₁₀ alkenyl and alkynyl groups. Exemplary suitable modifications include O[( _CH2 ) _nO ] _mCH3 , O( _CH2 ) _{nOCH3 ,} O( _CH2 ) _nNH2 , O(CH2) _{nCH3 , O(CH2)nONH2, and O(CH2)nON[(CH2)nCH3 ) ] 2 , wherein n and m are from 1 to} about 10. In other embodiments, the dsRNA includes one of the following at the 2' position: _C1 to _C10 lower alkyl, substituted lower alkyl, alkaryl, aralkyl, O-alkaryl or O-aralkyl, SH, _SCH3 , OCN, Cl, Br, CN, _CF3 , _OCF3 , _SOCH3 , _{SO2CH3, ONO2 , NO2} , _N3 , _NH2 , heterocycloalkyl, heterocycloalkaryl, aminoalkylamino, _{polyalkylamino} , substituted silyl, RNA cleavage group, reporter group, intercalator, group for improving the pharmacokinetic properties of CFB dsRNA agent, or group for improving the pharmacodynamic properties of CFB dsRNA agent, CFB antisense polynucleotide and/or CFB sense polynucleotide, as well as other substituents with similar properties. In some embodiments, the modification includes 2'-methoxyethoxy (2'-O-CH ₂ CH ₂ OCH ₃ , also known as 2'-O-(2-methoxyethyl) or 2'-MOE) (Martin et al., Helv. Chim. Acta, 1995, 78:486-504), i.e., alkoxy-alkoxy. Another exemplary modification is 2'-dimethylaminoethoxyethoxy, i.e., O(CH ₂ ) ₂ ON(CH ₃ ) ₂ group, also known as 2'-DMAOE, as described in the Examples below, and 2'-dimethylaminoethoxyethoxy (also known as 2'-dimethylaminoethoxyethoxy) (also known as 2'-DMAOE). Known in the art as 2'-O-dimethylaminoethoxyethyl or 2'-DMAEOE), i.e., 2'-O-CH ₂ -O-CH ₂ -N(CH ₂ ) ₂ . Methods for preparing modified RNAs such as those described are routinely practiced in the art, and such methods can be used to prepare certain modified CFB dsRNA agents of the invention.

Other modifications include 2'-methoxy (2'-OCH ₃ ), 2'-aminopropoxy (2'-OCH ₂ CH ₂ CH ₂ NH ₂ ) and 2'-fluoro (2'-F). Similar modifications can also be made at other positions on the RNA of the CFB dsRNA agent, CFB antisense polynucleotide and/or CFB sense polynucleotide of the present invention, particularly the 3' position of the sugar on the 3' terminal nucleotide or 2'-5' linked CFB dsRNA, CFB antisense polynucleotide or CFB sense polynucleotide, and the 5' position of the 5' terminal nucleotide. The CFB dsRNA agent, CFB antisense polynucleotide and/or CFB sense polynucleotide can also have a sugar mimetic, such as a cyclobutyl moiety in place of the pentofuranosyl sugar. Methods for preparing modified RNA, such as those described, are routinely practiced in the art, and such methods can be used to prepare certain modified CFB dsRNA reagents, CFB antisense polynucleotides and/or CFB sense polynucleotides of the invention.

In some embodiments, CFB dsRNA agents, CFB antisense polynucleotides and/or CFB sense polynucleotides may include nucleobase (commonly referred to in the art as "base") modifications or substitutions. As used herein, "unmodified" or "natural" nucleobases include the purine bases adenine and guanine, and the pyrimidine bases thymine, cytosine and uracil. Modified nucleobases include other synthetic and natural nucleobases such as 5-methylcytosine (5-Me-C), 5-hydroxymethylcytosine, xanthine, hypoxanthine, 2-aminoadenine, 6-methyl and other alkyl derivatives of adenine and guanine, 2-propyl and other alkyl derivatives of adenine and guanine, 2-thiouracil, 2-thiothymine and 2-thiocytosine, 5-halogenated uracil and cytosine, 5-propynyl uracil and cytosine, 6-azouracil, cytosine and thymine, 5-uracil (pseudouracil), 4-thiouracil, 8-halogenated, 8-amino, 8-thiol, 8-thioalkyl, 8-hydroxyaldehyde, other 8-substituted adenines and guanines, 5-halogenated, especially 5-bromo, 5-trifluoromethyl and other 5-substituted uracils, cytosine, 7-methylguanine and 7-methyladenine, 8-azaguanine and 8-azaadenine, 7-deazaguanine and 7-azaadenine and 3-deazaguanine and 3-deazaadenine. Additional nucleobases that can be included in certain embodiments of the CFB dsRNA agents of the present invention are known in the art, see, for example: Modified Nucleosides in Biochemistry, Biotechnology and Medicine, Herdewijn, P. Ed. Wiley-VCH, 2008; The Concise Encyclopedia Of Polymer Science And Engineering, pages 858-859, Kroschwitz, J. L, Ed. John Wiley & Sons, 1990, English et al., Angewandte Chemie, International Edition, 1991, 30, 613, Sanghvi, Y S., Chapter 15, dsRNA Research and Applications, pages 289-302, Crooke, S. T. and Lebleu, B., Ed., CRC Press, 1993. Methods for preparing dsRNA, CFB antisense polynucleotides and/or CFB sense polynucleotides comprising nucleobase modifications and/or substitutions, such as those described herein, are routinely practiced in the art, and such methods can be used to prepare certain modified CFB dsRNA reagents, CFB sense polynucleotides and/or CFB antisense polynucleotides of the invention.

Certain embodiments of the CFB dsRNA agents, CFB antisense polynucleotides and/or CFB sense polynucleotides of the present invention include RNA modified to include one or more locking nucleic acids (LNAs). Locking nucleic acids are nucleotides with a modified ribose moiety that includes an additional bridge connecting the 2' and 4' carbons. This structure effectively "locks" the ribose in a 3'-endo conformation. The addition of a locking nucleic acid to the CFB dsRNA agent, CFB antisense polynucleotide and/or CFB sense polynucleotide of the present invention can increase stability in serum and reduce off-target effects (Elmen, J. et al., (2005) Nucleic Acids Research 33(1):439-447; Mook, O R. et al., (2007) Mol Canc Ther 6(3):833-843; Grunweller, A. et al., (2003) Nucleic Acids Research 31(12):3185-3193). Methods for preparing dsRNA agents comprising locked nucleic acids, CFB antisense polynucleotides and/or CFB sense polynucleotides are routinely practiced in the art, and such methods can be used to prepare certain modified CFB dsRNA agents of the present invention.

Certain embodiments of the CFB dsRNA compounds, sense polynucleotides and/or antisense polynucleotides of the present invention comprise at least one modified nucleotide, wherein the at least one modified nucleotide comprises: 2'-O-methyl nucleotides, 2'-fluoro nucleotides, 2'-deoxy nucleotides, 2'3'-seco nucleotide mimetics, locked nucleotides, 2'-F-arabino nucleotides, 2'-methoxyethyl nucleotides, 2'-amino modified nucleotides, 2'-alkyl modified nucleotides, morpholino nucleotides and 3'-OMe nucleotides, nucleotides comprising a 5'-phosphorothioate group, nucleotides comprising vinylphosphonate, nucleotides comprising adenosine-glycol nucleic acid (GNA), nucleotides comprising thymidine-glycol nucleic acid (GNA) S-isomers, nucleotides comprising 2'-deoxythymidine-3' phosphate, nucleotides comprising 2'-deoxyguanosine-3'-phosphate, nucleotides comprising 2'-deoxyadenosine-3'-phosphate nucleotides, 2'-deoxycytidine-3'-phosphate nucleotides, 2'-deoxyuridine-3'-phosphate nucleotides, or terminal nucleotides linked to a cholesterol derivative or dodecanoic acid didecylamide group, 2'-amino modified nucleotides, phosphoramidates, or nucleotides containing unnatural bases. In some embodiments, the CFB dsRNA compound comprises an E-vinylphosphonate nucleotide at the 5' end of the antisense strand (also referred to herein as the guide strand).

Certain embodiments of the CFB dsRNA compounds, the 3' and 5' ends of the sense polynucleotides and/or the 3' end of the antisense polynucleotides of the present invention include at least one modified nucleotide, wherein the at least one modified nucleotide includes: a debasic nucleotide, a ribitol, an inverted nucleotide, an inverted debasic nucleotide, an inverted 2'-OMe nucleotide, an inverted 2'-deoxynucleotide. It is known to those skilled in the art that including a debasic or inverted debasic nucleotide at the end of an oligonucleotide can enhance stability (Czauderna et al. Structural variations and stabilizing modifications of synthetic siRNAs in mammalian cells. Nucleic Acids Res. 2003; 31(11): 2705-2716. doi: 10.1093/nar/gkg393). In some embodiments, the CFB dsRNA compound comprises one or more inverted debasing residues (invab) at the 3' end or the 5' end, or at both the 3' end and the 5' end. Exemplary inverted debasing residues (invab) include, but are not limited to, the following:

Certain embodiments of the CFB dsRNA compounds, 3' and 5' sense polynucleotides and/or 3' antisense polynucleotides of the present invention include at least one modified nucleotide, wherein the at least one modified nucleotide includes: an isomannitol residue or a stereoisomer of the isomannitol residue. Specific examples of the isomannitol residue or the stereoisomer of the isomannitol residue include, but are not limited to: , , , , , , , , , , and , wherein the phrase "Olig" each independently refers to a polynucleotide portion. Exemplary isomannose residues (imann) include but are not limited to the following: or .

In certain embodiments, the isomannoside nucleotides may also be conjugated to one or more targeting groups or delivery molecules, such as a GalNAc moiety.

Certain embodiments of CFB dsRNA compounds and antisense polynucleotides of the present invention include at least one modified nucleotide, wherein the at least one modified nucleotide comprises an unlocked nucleic acid nucleotide (UNA) or/and a glycol nucleic acid nucleotide (GNA). It is known to those skilled in the art that UNA and GNA are thermally unstable chemical modifications that can significantly improve the off-target profile of siRNA compounds (Janas et al., Selection of GalNAc-conjugate siRNAs with limit off-target-driven rat. Nat Commun. 2018;9(1):723. doi:10.1038/s41467-018-02989-4; Laursen et al., Enhancement of siRNA in vitro and in vivo performance using unlocked nucleic acid (UNA). Mol BioSyst. 2010;6:862–70).

Another modification that may be included in the RNA of certain embodiments of the CFB dsRNA agents, CFB antisense polynucleotides and/or CFB sense polynucleotides of the present invention includes chemically linking one or more ligands, moieties or conjugates to the RNA, which ligands, moieties or conjugates enhance one or more characteristics of the CFB dsRNA agent, CFB antisense polynucleotide and/or CFB sense polynucleotide, respectively. Non-limiting examples of characteristics that may be enhanced are: CFB dsRNA agent, CFB antisense polynucleotide and/or CFB sense polynucleotide activity, cellular distribution, delivery of the CFB dsRNA agent, pharmacokinetic properties of the CFB dsRNA agent, and cellular uptake of the CFB dsRNA agent. In some embodiments of the present invention, the CFB dsRNA agent comprises one or more targeting groups or linking groups, which are conjugated to the sense chain in certain embodiments of the CFB dsRNA agent of the present invention. A non-limiting example of a targeting group is a compound comprising N-acetyl-galactosamine (GalNAc). The terms "targeting group", "targeting agent", "linking agent", "targeting compound", "delivery molecule", "delivery compound" and "targeting ligand" can be used interchangeably herein. In certain embodiments of the present invention, the CFB dsRNA agent comprises a targeting compound conjugated to the 5'-end of the sense chain. In certain embodiments of the present invention, the CFB dsRNA agent comprises a targeting compound conjugated to the 3'-end of the sense chain. In some embodiments of the present invention, the CFB dsRNA agent comprises a targeting group containing GalNAc. In some embodiments of the present invention, the CFB dsRNA agent does not include a targeting compound conjugated to one or both of the 3'-end and the 5'-end of the sense chain. In some embodiments of the present invention, the CFB dsRNA agent does not include a targeting compound conjugated to one or both of the 5'-end and the 3'-end of the sense chain.

Additional targeting agents and linking agents are well known in the art. For example, targeting agents and linking agents that can be used in certain embodiments of the present invention include, but are not limited to, lipid moieties, such as cholesterol moieties (Letsinger et al., Proc. Natl. Acid. USA, 1989, 86: 6553-6556), bile acid (Manoharan et al., Biorg. Med. Chem. Let., 1994, 4: 1053-1060), thioethers, such as beryl-S-tritylthiol (Manoharan et al., Ann. N.Y. Acad. Sci., 1992, 660: 306-309; Manoharan et al., Biorg. Med. Chem. Let., 1993, 3:2765-2770), a thiocholesterol (Oberhauser et al., Nucl. Acids Res., 1992, 20:533-538), an aliphatic chain, such as dodecanediol or an undecyl residue (Saison-Behmoaras et al., EMBO J, 1991, 10:1111-1118; Kabanov et al., FEBS Lett., 1990, 259:327-330; Svinarchuk et al., Biochimie, 1993, 75:49-54), a phospholipid, such as di-hexadecyl-rac-glycerol or triethylammonium 1,2-di-O-hexadecyl-rac-glycero-3-phosphonate (Manoharan et al., Tetrahedron Lett., 1995, 36:3651-3654; Shea et al., Nucl. Acids Res., 1990, 18:3777-3783), polyamine or polyethylene glycol chain (Manoharan et al., Nucleosides & Nucleotides, 1995, 14:969-973) or adamantane acetic acid (Manoharan et al., Tetrahedron Lett., 1995, 36:3651-3654), palmityl moiety (Mishra et al., Biochim. Biophys. Acta, 1995, 1264:229-237), or octadecylamine or hexylamino-carbonyloxycholesterol moiety (Crooke et al., J. Pharmacol. Exp. Ther., 1996, 277:923-937).

Certain embodiments of compositions comprising CFB dsRNA agents, CFB antisense polynucleotides, and/or CFB sense polynucleotides may comprise ligands that alter the distribution, targeting, etc., of CFB dsRNA agents. In some embodiments of compositions comprising CFB dsRNA agents of the invention, the ligand increases affinity for a selected target (e.g., a molecule, a cell or cell type, a compartment, such as a cell or organ compartment, a tissue, an organ, or a body region) compared to species in which such ligand is absent. Ligands useful in the compositions and/or methods of the invention may be naturally occurring substances, such as proteins (e.g., human serum albumin (HSA), low-density lipoprotein (LDL), or globulins); carbohydrates (e.g., dextran, pullulan, chitin, chitosan, inulin, cyclodextrin, or hyaluronic acid); or lipids. The ligand may also be a recombinant or synthetic molecule, such as a synthetic polymer, such as a synthetic polyamino acid or polyamine. Examples of polyamino acids are polylysine (PLL), poly-L-aspartic acid, poly-L-glutamic acid, styrene-maleic anhydride copolymer, poly(L-lactide-co-glycolic acid) copolymer, divinyl ether-maleic anhydride copolymer, N-(2-hydroxypropyl)methacrylamide copolymer (HMPA), polyethylene glycol (PEG), polyvinyl alcohol (PVA), polyurethane, poly(2-ethylacrylic acid), N-isopropylacrylamide polymer or polyphosphazene. Examples of polyamines include: polyethyleneimine, polylysine (PLL), spermine, spermidine, polyamines, pseudopeptide polyamines, peptide mimetic polyamines, dendritic polyamines, arginine, amidine, protamine, cationic lipids, cationic porphyrins, quaternary salts of polyamines or alpha helical peptides.

The ligands included in the compositions and/or methods of the invention may comprise a targeting group, non-limiting examples of which are cell or tissue targeting agents, such as lectins, glycoproteins, lipids, or proteins, such as antibodies that bind to specific cell types (e.g., kidney cells or liver cells). The targeting group can be thyroid stimulating hormone, melanocyte stimulating hormone, lectin, glycoprotein, surfactant protein A, mucin carbohydrate, polyvalent lactose, polyvalent galactose, N-acetylgalactosamine, N-acetylglucosamine polyvalent mannose, polyvalent fucose, glycosylated polyamino acid, polyvalent galactose, transferrin, bisphosphonate, polyglutamic acid, polyaspartic acid, lipid, cholesterol, steroid, bile acid, folic acid, vitamin B12, vitamin A, biotin or RGD peptide or RGD peptide mimetic.

Other examples of ligands include dyes, intercalating agents (e.g., acridine), crosslinkers (e.g., psoralen, mitomycin C), porphyrins (TPPC4, texaphylline, saphthrin), polycyclic aromatic hydrocarbons (e.g., phenazine, dihydrophenazine), artificial endonucleases (e.g., EDTA), lipophilic molecules (e.g., cholesterol, bile acid, adamantaneacetic acid, 1-pyrenebutyric acid, dihydrotestosterone, 1,3-bis-O(hexadecyl)glycerol, Geranyloxyhexyl, hexadecylglycerol, borneol, menthol, 1,3-propylene glycol, heptadecyl, palmitic acid, myristic acid, O3-(oleyl)cholestyric acid, O3-(oleyl)cholestyric acid, dimethoxytrityl or phenoxazine) and peptide conjugates (e.g., tripeptide, Tat peptide), alkylating agents, phosphates, amino, hydroxyl, PEG (e.g., PEG-40K), MPEG, [MPEG] _2. Polyamino, alkyl, substituted alkyl, radiolabel, enzyme, hapten (e.g., biotin), transport/absorption enhancer (e.g., aspirin, vitamin E, folic acid), synthetic ribonuclease (e.g., imidazole, bisimidazole, histamine, imidazole cluster, acridine-imidazole conjugate, tetrazamacrocyclic Eu3+ complex), dinitrophenyl, HRP or AP.

The ligand included in the compositions and/or methods of the present invention can be a protein, such as a glycoprotein or a peptide, such as a molecule with a specific affinity for a co-ligand, or an antibody, such as an antibody that binds to a specific cell type (e.g., cancer cells, endothelial cells, heart cells, or bone cells). The ligand useful in embodiments of the compositions and/or methods of the present invention can be a hormone or a hormone receptor. The ligand useful in embodiments of the compositions and/or methods of the present invention can be a lipid, a lectin, a carbohydrate, a vitamin, a cofactor, a multivalent lactose, a multivalent galactose, N-acetylgalactosamine, N-acetylglucosamine, a multivalent mannose, or a multivalent fucose. Ligands useful in embodiments of the compositions and/or methods of the invention can be substances that increase the entry of CFB dsRNA agents into cells, for example, by disrupting the cell's cytoskeleton, for example, by disrupting the cell's microtubules, microfilaments and/or intermediate filaments. Non-limiting examples of agents of this type are: taxon, vincristine, vinblastine, cytorelaxin, nocodazole, japlakinolide, latrunculin A, phalloidin, swinholide A, indanocine, and myoservin.

In some embodiments, the ligand linked to the CFB dsRNA agent of the present invention is used as a pharmacokinetic (PK) modulator. Examples of PK modulators that can be used in the compositions and methods of the present invention include, but are not limited to, lipophilic substances, bile acid, steroids, phospholipid analogs, peptides, protein binders, PEG, vitamins, cholesterol, fatty acids, bile acid, cholic acid, dialkyl glycerides, diacyl glycerides, phospholipids, sphingolipids, naproxen, ibuprofen, vitamin E, biotin, aptamers that bind to serum proteins, and the like. It is also known that oligonucleotides containing a large number of phosphorothioate bonds can bind serum proteins, so short oligonucleotides containing multiple phosphorothioate bonds in the backbone (e.g., oligonucleotides of about 5 bases, 10 bases, 15 bases, or 20 bases) can also be used as ligands in the compositions and/or methods of the present invention. CFB dsRNA Agent Compositions

In some embodiments of the present invention, the CFB dsRNA agent is present in a composition. The composition of the present invention may include one or more CFB dsRNA agents, and optionally one or more pharmaceutically acceptable carriers, delivery agents, targeting agents, detectable markers, etc. According to some embodiments of the present method, non-limiting examples of potentially useful targeting agents are agents that direct the CFB dsRNA agent of the present invention to cells to be treated and/or to cells to be treated. The selection of the targeting agent will depend on the following factors: the nature of the CFB-related disease or condition, and the type of cell being targeted. In a non-limiting example, in some embodiments of the present invention, it may be desirable to direct the CFB dsRNA agent to hepatocytes and/or to hepatocytes. It should be understood that in some embodiments of the methods of the invention, the therapeutic agent includes a CFB dsRNA agent with only a delivery agent, such as a delivery agent comprising N-acetylgalactosamine (GalNAc), without any additional elements. For example, in some aspects of the invention, a CFB dsRNA agent can be linked to a delivery compound comprising GalNAc and contained in a composition comprising a pharmaceutically acceptable carrier and administered to a cell or subject without any detectable label or targeting agent, etc., being linked to the CFB dsRNA agent.

When the CFB dsRNA agents of the present invention are administered together with and/or attached to one or more delivery agents, targeting agents, labeling agents, etc., those skilled in the art will understand and be able to select and use agents suitable for the methods of the present invention. Labeling agents can be used in certain methods of the present invention to determine the location of CFB dsRNA agents in cells and tissues, and can be used to determine the cell, tissue or organ location of therapeutic compositions containing CFB dsRNA agents that have been administered in the methods of the present invention. Procedures for attaching and using labeling agents (e.g., enzyme labels, dyes, radioactive labels, etc.) are well known in the art. It should be understood that in some embodiments of the compositions and methods of the present invention, the labeling agent is attached to one or both of the sense polynucleotide and the antisense polynucleotide contained in the CFB dsRNA agent. Delivery of CFB dsRNA Agents and CFB Antisense Polynucleotide Agents

Certain embodiments of the methods of the present invention include delivering the CFB dsRNA agent to cells. As used herein, the term "delivery" refers to promoting or affecting the uptake or absorption of cells. The absorption or uptake of the CFB dsRNA agent can occur through independent diffusion or active cell processes, or through the use of delivery agents, targeting agents, etc. that can be associated with the CFB dsRNA agent of the present invention. Delivery methods suitable for the methods of the present invention include, but are not limited to: in vivo delivery, in which the CFB dsRNA agent is injected into a tissue site or administered systemically. In some embodiments of the present invention, the CFB dsRNA agent is linked to a delivery agent.

Non-limiting examples of methods that can be used to deliver CFB dsRNA agents to cells, tissues and/or subjects include: CFB dsRNA-GalNAc complexes, SAMiRNA technology, LNP-based delivery methods, and naked RNA delivery. These and other delivery methods have been successfully used in the art to deliver therapeutic RNAi agents to treat various diseases and conditions, such as, but not limited to: liver disease, acute intermittent porphyria (AIP), hemophilia, pulmonary fibrosis, etc. Details of various delivery methods can be found in the following publications: Nikam, R.R. & K. R. Gore (2018) Nucleic Acid Ther, 28 (4), 209-224 Aug 2018; Springer A.D. & S.F. Dowdy (2018) Nucleic Acid Ther. Jun 1; 28(3): 109–118; Lee, K. et al., (2018) Arch Pharm Res, 41(9), 867-874; and Nair, J.K. et al., (2014) J. Am. Chem. Soc. 136:16958-16961, all of which are incorporated herein by reference.

Some embodiments of the present invention include using lipid nanoparticles (LNPs) to deliver CFB dsRNA agents of the present invention to cells, tissues and/or subjects. LNPs are generally used to deliver CFB ds RNA agents in vivo , including therapeutic CFB dsRNA agents. One advantage of using LNPs or other delivery agents is that the stability of the CFB RNA agent is increased when the CFB RNA agent is delivered to a subject using LNPs or other delivery agents. In some embodiments of the present invention, the LNPs include cationic LNPs carrying one or more CFB RNAi molecules of the present invention. LNPs containing CFB RNAi molecules are administered to a subject, and the LNPs and their attached CFB RNAi molecules are taken up by cells via endocytosis, and their presence leads to the release of RNAi trigger molecules, thereby mediating RNAi.

Another non-limiting example of a delivery agent that can be used in embodiments of the present invention to deliver the CFB dsRNA agent of the present invention to cells, tissues and/or subjects is a reagent comprising GalNAc, which is attached to the following CFB dsRNA agent: The present invention and delivers the CFB dsRNA agent to cells, tissues and/or subjects. Examples of certain additional delivery agents comprising GalNAc that can be used in certain embodiments of the methods and compositions of the present invention are disclosed in PCT application: WO2020191183A1 (the entire contents of which are incorporated herein). A non-limiting example of a GalNAc targeting ligand that can be used in the compositions and methods of the present invention to deliver the CFB dsRNA agent to cells is a targeting ligand cluster. Examples of targeting ligand clusters presented herein are referred to as: GalNAc ligands with phosphodiester linkages (GLO) and GalNAc ligands with phosphorothioate linkages (GLS). The term "GLX-n" may be used herein to indicate that the GalNAc-containing compound that is linked is compound GLS-1*, GLS-2*, GLS-3*, GLS-4*, GLS-5*, GLS-6*, GLS-7*, GLS-8*, GLS-9*, GLS-10*, GLS-11*, GLS-12*, GLS-13*, GLS-14*, GLS-15*, GLS-16*, Any one of GLO-1, GLO-2, GLO-3, GLO-4, GLO-5, GLO-6, GLO-7, GLO-8, GLO-9, GLO-10, GLO-11, GLO-12, GLO-13, GLO-14, GLO-15 and GLO-16, each of which has a structure as shown below, wherein the connection position of the GalNAc targeting ligand to the RNAi agent of the present invention is the rightmost side of each (shown by “ ⸾ ”). It should be understood that any RNAi and dsRNA molecules of the present invention can be attached to GLS-1*, GLS-2*, GLS-3*, GLS-4*, GLS-5*, GLS-6*, GLS-7*, GLS-8*, GLS-9*, GLS-10*, GLS-11*, GLS-12*, GLS-13*, GLS-14*, GLS-15*, GLS-16*, GLO-1, GLO-2, GLO-3, GLO-4, GLO-5, GLO-6, GLO-7, GLO-8, GLO-9, GLO-10, GLO-11, GLO-12, GLO-13, GLO-14, GLO-15 and GLO-16. The structures of GLO-1 to GLO-16 and GLS-1* to GLS-16* are shown below. GLO-1 GLS-1* GLO-2 GLS-2* GLO-3 GLS-3* GLO-4 GLS-4* GLO-5 GLS-5* GLO-6 GLS-6* GLO-7 GLS-7* GLO-8 GLS-8* GLO-9 GLS-9* GLO-10 GLS-10* GLO-11 GLS-11* GLO-12 GLS-12* GLO-13 GLS-13* GLO-14 GLS-14* GLO-15 GLS-15* GLO-16 . GLS-16*

In certain embodiments, the aforementioned isomannose nucleotides may also be conjugated to one or more GalNAc targeting ligands. Specific examples of isomannose nucleotides conjugated to GalNAc targeting ligands include, but are not limited to: , wherein the phrase "olig" each independently refers to a polynucleotide portion.

In some embodiments of the invention, in vivo delivery can also be performed by a β-glucan delivery system, such as those described in U.S. Patent No. 5,100,000. U.S. Patent Nos. 5,032,401 and 5,607,677 and U.S. Publication No. 2005/0281781, the entire contents of which are incorporated herein by reference. CFB RNAi agents can also be introduced into cells in vitro using methods known in the art such as electroporation and lipofection. In certain embodiments of the methods of the invention, CFB dsRNA is delivered without a targeting agent. These RNAs can be delivered as "naked" RNA molecules. As a non-limiting example, the CFB dsRNA of the present invention can be administered to a subject in a pharmaceutical composition comprising an RNAi agent but not comprising a targeting agent, such as a GalNAc targeting compound, to treat a CFB-related disease or disorder in the subject, such as a cardiovascular disease.

In addition to certain delivery modes described herein, it will be appreciated that RNAi delivery modes, such as but not limited to those described herein and those used in the art, can be used in conjunction with the embodiments of the CFB RNAi agents and treatment methods described herein.

The CFB dsRNA reagents of the present invention can be administered to a subject in an amount and manner effective to reduce the level and activity of CFB polypeptides in cells and/or subjects. In some embodiments of the methods of the present invention, one or more CFB dsRNA reagents are administered to cells and/or subjects to treat a disease or condition associated with CFB expression and activity. In some embodiments, the methods of the present invention include administering one or more CFB dsRNA reagents to a subject in need of such treatment to reduce a disease or condition associated with CFB expression in the subject. The CFB dsRNA reagents or CFB antisense polynucleotide reagents of the present invention can be administered to reduce CFB expression and/or activity in one or more of in vitro , ex vivo , and in vivo cells.

In some embodiments of the present invention, the level of CFB polypeptide in the cell is reduced and thus its activity is reduced by delivering (e.g., introducing) a CFB dsRNA agent or a CFB antisense polynucleotide agent into the cell. Targeting agents and methods can be used to help deliver CFB dsRNA agents or CFB antisense polynucleotide agents to specific cell types, cell subtypes, organs, spatial regions, and/or subcellular regions within cells within a subject. CFB dsRNA agents can be administered alone or in combination with one or more additional CFB dsRNA agents in certain methods of the present invention. In some embodiments, 2, 3, 4 or more independently selected CFB dsRNA agents are administered to a subject.

In certain embodiments of the invention, a CFB dsRNA agent is administered to a subject to treat a CFB-related disease or condition in combination with one or more additional treatment regimens for treating a CFB-related disease or condition. Non-limiting examples of additional treatment regimens are: administration of one or more CFB antisense polynucleotides of the invention, administration of non-CFB dsRNA therapeutics, and behavioral changes. Additional treatment regimens may be administered at one or more times before, simultaneously with, and after administration of a CFB dsRNA agent of the invention. It should be understood that, as used herein, within five minutes of time zero, within 10 minutes of time zero, within 30 minutes of time zero, within 45 minutes of time zero, and within 60 minutes of time zero, "time zero" is the time at which the CFB dsRNA agent of the present invention is administered to a subject. Non-limiting examples of non-CFB dsRNA therapeutic agents are: C5 inhibitors, such as anti-complement component C5 antibodies or antigen-binding fragments thereof (e.g., eculizumab, ravulizumab-cwvz or bozelimumab (REGN3918)) or C5 inhibitor peptide inhibitors (e.g., zilucoplan). Eculizumab is a humanized monoclonal IgG2/4, kappa light chain antibody that specifically binds to complement component C5 with high affinity and inhibits the cleavage of C5 into C5a and C5b, thereby inhibiting the formation of the terminal complement complex C5b-9. Ravulizumab-cwvz is a humanized IgG2/4 monoclonal antibody that specifically binds to complement component C5 with high affinity and inhibits the cleavage of C5 into C5a and C5b, thereby inhibiting the formation of the terminal complement complex C5b-9. Pozelimab (also known as H4H12166P, described in US20170355757) is a fully human IgG4 monoclonal antibody designed to block complement factor C5. Zilucoplan is a synthetic macrocyclic peptide that binds complement component 5 (C5) with subnanomolar affinity and allosterically inhibits its cleavage into C5a and C5b following classical, alternative or lectin pathway activation. Preferably, the additional therapeutic agent is a C3 peptide inhibitor or an analog thereof. In one embodiment, the C3 peptide inhibitor is compstatin. Compstatin is a cyclic tridecapeptide with potent and selective C3 inhibitory activity. These and other therapeutic agents and behavioral changes are known in the art and are used to treat CFB-related diseases or conditions in subjects and can be administered to subjects in combination with administration of one or more CFB dsRNA agents of the present invention to treat CFB-related diseases or conditions. The CFB dsRNA agents of the present invention administered to cells or subjects to treat CFB-related diseases or disorders may act in a synergistic manner with one or more other therapeutic agents or activities and increase the effectiveness of the one or more therapeutic agents or activities and/or increase the effectiveness of the CFB dsRNA agent in treating CFB-related diseases or disorders.

The treatment methods of the present invention include administering CFB dsRNA agents, which can be used before the onset of a CFB-related disease or condition and/or while a CFB-related disease or condition exists, including the early, middle and late stages of the disease or condition and all times before and after these stages. The methods of the present invention can also be used to treat subjects who have previously used one or more other therapeutic agents and/or therapeutic activities to treat a CFB-related disease or condition, and these therapeutic agents and/or therapeutic activities were unsuccessful, had a low success rate and/or were no longer successful in treating the subject's CFB-related disease or condition. Vector-Encoded dsRNA

In certain embodiments of the present invention, a vector can be used to deliver a CFB dsRNA agent to a cell. The CFB dsRNA agent transcription unit can be contained in a DNA or RNA vector. The preparation and use of such vectors encoding transgenes for delivery of sequences to cells and/or subjects are well known in the art. The vector can be used in the method of the present invention, which results in transient expression of CFB dsRNA for example, at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more hours, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 weeks or longer. The length of transient expression can be determined using conventional methods based on factors such as, but not limited to, the specific vector construct selected and the target cell and/or tissue. Such transgenes can be introduced as linear constructs, circular plasmids or viral vectors, which can be integrating or non-integrating vectors. Transgenes can also be constructed to allow them to be inherited as extrachromosomal plasmids (Gassmann, et al., Proc. Natl. Acad. Sci. USA (1995) 92: 1292).

Single or multiple chains of CFB dsRNA agents can be transcribed from a promoter on an expression vector. When two separate chains are to be expressed to produce, for example, dsRNA, two separate expression vectors can be introduced into cells together using methods such as transfection or infection. In certain embodiments, each separate chain of the CFB dsRNA agent of the present invention can be transcribed from a promoter contained on the same expression vector. In certain embodiments of the present invention, the CFB dsRNA agent is expressed as an inverted repeat polynucleotide connected by a linker polynucleotide sequence, so that the CFB dsRNA agent has a stem and loop structure.

Non-limiting examples of RNA expression vectors are DNA plasmids or viral vectors. Expression vectors useful in embodiments of the present invention may be compatible with eukaryotic cells. Eukaryotic cell expression vectors are routinely used in the art and are available from many commercial sources. Delivery of CFB dsRNA expression vectors may be systemic, such as by intravenous or intramuscular administration, by administration to target cells removed from a subject and then reintroduced into the subject, or by any other means that allows introduction of desired target cells.

Viral vector systems that may be included in embodiments of the method include, but are not limited to: (a) adenoviral vectors; (b) retroviral vectors, including, but not limited to, lentiviral vectors, Moloney murine leukemia virus, etc.; (c) adeno-associated viral vectors; (d) herpes simplex virus vectors; (e) SV 40 vectors; (f) polyoma virus vectors; (g) papilloma virus vectors; (h) picornavirus vectors; (i) poxvirus vectors, such as orthopoxvirus vectors or avipoxvirus vectors, such as canarypoxvirus vectors or chickenpoxvirus vectors; and (j) helper-dependent or intestinal adenovirus. Constructs for recombinant expression of CFB dsRNA agents may include regulatory elements, such as promoters, enhancers, etc., which may be selected to provide constitutive or regulated/induced expression. The use of viral vector systems, promoters and enhancers, etc. is routine in the art and can be used in conjunction with the methods and compositions described herein.

Certain embodiments of the present invention include the use of viral vectors to deliver CFB dsRNA reagents into cells. Many adenovirus-based delivery systems are routinely used in the art for delivery to, for example, the lungs, liver, central nervous system, endothelial cells, and muscle. Non-limiting examples of viral vectors that can be used in the methods of the present invention are: AAV vectors, poxviruses such as vaccinia virus, Modified Virus Ankara (MVA), NYVAC, avian pox such as chicken pox or canary pox.

Certain embodiments of the present invention include methods of delivering CFB dsRNA agents to cells using vectors, and such vectors may be in a pharmaceutically acceptable carrier that may, but need not, include a slow-release matrix in which the gene delivery vector is embedded. In some embodiments, the vector used to deliver CFB dsRNA may be produced by recombinant cells, and the pharmaceutical compositions of the present invention may include one or more cells that produce the CFB dsRNA delivery system. Pharmaceutical compositions containing CFB dsRNA or ssRNA agents

Certain embodiments of the present invention include the use of a pharmaceutical composition containing a CFB dsRNA agent or a CFB antisense polynucleotide agent and a pharmaceutically acceptable carrier. The pharmaceutical composition containing a CFB dsRNA agent or a CFB antisense polynucleotide agent can be used in the methods of the present invention to reduce CFB gene expression and CFB activity in cells, and can be used to treat CFB-related diseases or conditions. Such pharmaceutical compositions can be formulated according to the mode of administration. Non-limiting examples of formulations for delivery modes are: compositions formulated for subcutaneous delivery, compositions formulated for systemic administration via parenteral delivery, compositions formulated for intravenous (IV) delivery, compositions formulated for intrathecal delivery, compositions formulated for direct delivery to the brain, and the like. Administration of the pharmaceutical compositions of the present invention to deliver CFB dsRNA agents or CFB antisense polynucleotide agents to cells can be performed using one or more methods, such as: topical (e.g., via a transdermal patch), pulmonary, for example, by inhalation or insufflation of powders or aerosols, including via a nebulizer; intratracheal, intranasal, epidermal and transdermal, oral or parenteral. Parenteral administration includes intravenous, intraarterial, subcutaneous, intraperitoneal or intramuscular injection or infusion; subcutaneous, for example, via an implant device; or intracranial, for example, via intraparenchymal, intrathecal or intraventricular administration. CFB dsRNA agents or CFB antisense polynucleotide agents can also be delivered directly to target tissues, such as directly to the liver, directly to the kidneys, etc. It should be understood that "delivering a CFB dsRNA agent" or "delivering a CFB antisense polynucleotide agent" into a cell includes directly delivering a CFB dsRNA agent or a CFB antisense polynucleotide agent and expressing the CFB dsRNA agent in the cell from an encoding vector delivered to the cell, or causing the CFB dsRNA or CFB antisense polynucleotide agent to be present in the cell by any suitable means. Methods for preparing and using agents and delivering inhibitory RNA are well known in the art and are routinely used.

As used herein, a "pharmaceutical composition" comprises a pharmacologically effective amount of the CFB dsRNA agent or CFB antisense polynucleotide agent of the present invention and a pharmaceutically acceptable carrier. The term "pharmaceutically acceptable carrier" refers to a carrier for administering a therapeutic agent. Such carriers include, but are not limited to, saline, buffered saline, glucose, water, glycerol, ethanol, and combinations thereof. The term explicitly excludes cell culture media. For drugs administered orally, pharmaceutically acceptable carriers include, but are not limited to, pharmaceutically acceptable excipients, such as inert diluents, disintegrants, binders, lubricants, sweeteners, flavorings, coloring agents, and preservatives. Suitable inert diluents include sodium and calcium carbonates, sodium and calcium phosphates, and lactose, while corn starch and alginic acid are suitable disintegrants. Binders may include starch and gelatin, while lubricants, if present, are typically magnesium stearate, stearic acid or talc. If desired, tablets may be coated with materials such as glyceryl monostearate or glyceryl distearate to delay absorption in the gastrointestinal tract. The agents included in the pharmaceutical formulations are further described below.

As used herein, terms such as "pharmacologically effective amount,""therapeutically effective amount," and "effective amount" refer to an amount of a CFB dsRNA agent or CFB antisense polynucleotide agent of the present invention that produces a desired pharmacological, therapeutic, or preventive result. For example, if a given clinical treatment is considered effective when a measurable parameter associated with a disease or condition is reduced by at least 10%, then the therapeutically effective amount of a drug for treating that disease or condition is the amount necessary to reduce that parameter by at least 10%. For example, a therapeutically effective amount of a CFB dsRNA agent or CFB antisense polynucleotide agent can reduce CFB polypeptide levels by at least 10%. Effective amount

The methods of the present invention, in certain aspects, include contacting a cell with an effective amount of a CFB dsRNA agent or a CFB antisense polynucleotide agent to reduce CFB gene expression in the contacted cell. Certain embodiments of the methods of the present invention include administering an effective amount of a CFB dsRNA agent or a CFB antisense polynucleotide agent to a subject to reduce CFB gene expression and treat a CFB-related disease or condition in the subject. An "effective amount" for reducing CFB expression and/or treating a CFB-related disease or condition is an amount necessary or sufficient to achieve a desired biological effect. For example, an effective amount of a CFB dsRNA agent or a CFB antisense polynucleotide agent for treating a CFB-related disease or condition may be an amount necessary to (i) slow or stop the progression of the disease or condition; or (ii) reverse, reduce, or eliminate one or more symptoms of the disease or condition. In some aspects of the invention, an effective amount is an amount that produces a therapeutic response that prevents and/or treats a CFB-related disease or condition when the CFB dsRNA agent or CFB antisense polynucleotide agent is administered to a subject in need of treatment for the disease or condition. According to some aspects of the invention, an effective amount is an amount that produces a therapeutic response that prevents and/or treats the disease or condition when the CFB dsRNA agent or CFB antisense polynucleotide agent of the invention is combined with or co-administered to a subject as a treatment method for another CFB-related disease or condition. In some embodiments of the invention, the biological effect of treating a subject with the CFB dsRNA agent or CFB antisense polynucleotide agent of the invention can be to improve and/or completely eliminate symptoms caused by a CFB-related disease or condition. In some embodiments of the invention, the biological effect is complete elimination of the CFB-related disease or condition, for example, as indicated by a diagnostic test that the subject is free of the CFB-related disease or condition. Non-limiting examples of physiological symptoms that can be detected include a decrease in CFB levels in the liver of a subject after administration of an agent of the invention. Other methods known in the art for assessing the status of a CFB-related disease or condition can be used to determine the effect of the agents and/or methods of the invention on a CFB-related disease or condition.

Typically, the effective amount of a CFB dsRNA agent or CFB antisense polynucleotide agent that reduces CFB polypeptide activity to a level that treats a CFB-related disease or condition will be determined in a clinical trial, establishing the effective dose in a test population versus a control population in a blinded study. In some embodiments, the effective amount will be the amount that results in the desired response, such as a reduction in the amount of CFB-related disease or condition in cells, tissues, and/or subjects suffering from the disease or condition. Thus, the effective amount of a CFB dsRNA agent or CFB antisense polynucleotide agent for treating a CFB-related disease or condition that can be treated by reducing CFB polypeptide activity can be an amount that, when administered, reduces the amount of CFB polypeptide activity in a subject to less than the amount present in the cell, tissue, and/or subject when the CFB dsRNA agent or CFB antisense polynucleotide agent is not administered. In certain aspects of the invention, the level of CFB polypeptide activity and/or CFB gene expression present in cells, tissues, and/or subjects that have not been exposed to or administered the CFB dsRNA agent or CFB antisense polynucleotide agent of the invention is referred to as a "control" amount. In some embodiments of the methods of the invention, the control amount of the subject is the amount of the subject before treatment, in other words, the level of the subject before administration of the CFB agent can be the control level of the subject and compared with the level of CFB polypeptide activity and/or CFB gene expression in the subject after administration of the siRNA to the subject. In the case of treating a CFB-related disease or condition, the desired response can be a reduction or elimination of one or more symptoms of the disease or condition in the cell, tissue, and/or subject. The reduction or elimination may be temporary or permanent. It should be understood that the status of a CFB-related disease or disorder can be monitored using methods of determining CFB polypeptide activity, CFB gene expression, symptom assessment, clinical testing, etc. In some aspects of the invention, the desired response to treatment is "delay" of a CFB-related disease or disorder or even prevention of the onset of the disease or disorder.

The effective amount of a compound that reduces CFB polypeptide activity can also be determined by evaluating the physiological effects of administering a CFB dsRNA agent or a CFB antisense polynucleotide agent to cells or subjects (e.g., reduction in CFB-related diseases or conditions after administration). The efficacy of the CFB dsRNA agent or CFB antisense polynucleotide agent of the present invention (which can be administered in the form of a pharmaceutical compound of the present invention) can be determined using assays and/or symptom monitoring of subjects, and whether there is a response to treatment. Non-limiting examples are one or more tests known in the art for CH50 activity (a measure of total hemolytic complement), AH50 (a measure of complement alternative pathway hemolytic activity), lactate dehydrogenase (LDH) (a measure of intravascular hemolysis), hemoglobin levels; levels of one or more of C3, C9, C5, C5a, C5b, and soluble C5b-9 complex. Another non-limiting example is that one or more liver function tests known in the art can be used to determine the status of CFB-related lipid imbalance in a subject before and after treatment of the subject with the CFB dsRNA agent of the present invention.

Some embodiments of the invention include methods of determining the efficacy of a dsRNA agent or CFB antisense polynucleotide agent of the invention administered to a subject by treating a CFB-related disease or condition by assessing and/or monitoring one or more "physiological characteristics" of the CFB-related disease or condition in the subject. Non-limiting examples of physiological characteristics of a CFB-related disease or condition are CFB mRNA levels, CFB protein levels, or CH50 activity (a measure of total hemolytic complement), AH50 (a measure of hemolytic activity of the alternative complement pathway), lactate dehydrogenase (LDH) (a measure of intravascular hemolysis), hemoglobin levels; levels of one or more of C3, C9, C5, C5a, C5b, and soluble C5b-9 complex. Standard methods for determining such physiological characteristics are known in the art and include, but are not limited to, blood tests, imaging studies, physical examinations, etc.

It should be understood that the amount of CFB dsRNA agent or CFB antisense polynucleotide agent administered to a subject can be modified based at least in part on the determination of the disease and/or condition state and/or physiological characteristics of the subject. The therapeutic amount can be changed, for example, by increasing or decreasing the amount of CFB-dsRNA agent or CFB antisense polynucleotide agent, by changing the composition of the CFB dsRNA agent or CFB antisense polynucleotide agent, respectively, by changing the route of administration, by changing the time of administration, etc. The effective amount of a CFB dsRNA agent or CFB antisense polynucleotide agent will vary with the specific condition being treated, the age and physical condition of the subject being treated, the severity of the condition, the duration of treatment, the nature of concurrent treatments (if any), the specific route of administration, and other factors within the knowledge and expertise of a health practitioner. For example, the effective amount may depend on the desired CFB polypeptide activity and/or CFB gene expression level for effective treatment of a CFB-related disease or condition. A skilled person can empirically determine the effective amount of a specific CFB dsRNA agent or CFB antisense polynucleotide agent used in the methods of the present invention without undue experimentation. In conjunction with the teachings provided herein, by selecting from the various CFB dsRNA agents or CFB antisense polynucleotide agents of the present invention, and weighing factors such as potency, relative bioavailability, patient weight, severity of adverse side effects, and preferred modes of administration, an effective preventive or therapeutic regimen can be planned to effectively treat a particular subject. As used in the embodiments of the present invention, an effective amount of a CFB dsRNA agent or CFB antisense polynucleotide agent of the present invention can be an amount that produces a desired biological effect in a cell when in contact with the cell.

It should be recognized that CFB gene silencing can be determined in any cell expressing CFB, either constitutively or by genome engineering, and by any appropriate assay. In some embodiments of the invention, CFB gene expression is reduced by at least 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% by administering a CFB dsRNA agent of the invention. In some embodiments of the invention, CFB gene expression is reduced by between 5% and 10%, 5% and 25%, 10% and 50%, 10% and 75%, 25% and 75%, 25% and 100%, or 50% and 100% by administering a CFB dsRNA agent of the invention. Dosage

CFB dsRNA agents and CFB antisense polynucleotide agents are delivered in a pharmaceutical composition in an amount sufficient to inhibit CFB gene expression. In certain embodiments of the present invention, the dosage of CFB dsRNA agents or CFB antisense polynucleotide agents is in the range of 0.01 to 200.0 mg per kg of body weight per day of the recipient, usually in the range of 1 to 50 mg, 5 to 40 mg/kg of body weight, 10 to 30 mg/kg of body weight, 1 to 20 mg/kg of body weight, 1 to 10 mg/kg of body weight, 4 to 15 mg/kg of body weight per day (including the end values). For example, the CFB The dsRNA agent or CFB antisense polynucleotide agent can be administered in the following single dose/body weight amount: about 0.01 mg/kg, 0.05 mg/kg, 0.1 mg/kg, 0.2 mg/kg, 0.3 mg/kg, 0.4 mg/kg, 0.5 mg/kg, 1 mg/kg, 1.1 mg/kg, 1.2 mg/kg, 1.3 mg/kg, 1.4 mg/kg, 1.5 mg/kg, 1.6 mg/kg, 1.7 mg/kg, 1.8 mg/kg, 1.9 mg/kg, 2 mg/kg, 2.1 mg/kg, 2.2 mg/kg, 2.3 mg/kg, 2.4 mg/kg, 2.5 mg/kg, 2.6 mg/kg, 2.7 mg/kg, 2.8 mg/kg, 2.9 mg/kg, 3.0 mg/kg, 3.1 mg/kg, 3.2 mg/kg, 3.3 mg/kg. g/kg, 3.4mg/kg, 3.5mg/kg, 3.6mg/kg, 3.7mg/kg, 3.8mg/kg, 3.9mg/kg, 4mg/kg, 4.1mg/kg, 4 .2mg/kg, 4.3mg/kg, 4.4mg/kg, 4.5mg/kg, 4.6mg/kg, 4.7mg/kg, 4.8mg/kg, 4.9mg/kg, 5mg/k g, 5.1mg/kg, 5.2mg/kg, 5.3mg/kg, 5.4mg/kg, 5.5mg/kg, 5.6mg/kg, 5.7mg/kg, 5.8mg/kg, 5. 9mg/kg, 6mg/kg, 6.1mg/kg, 6.2mg/kg, 6.3mg/kg, 6.4mg/kg, 6.5mg/kg, 6.6mg/kg, 6.7mg/kg, 6.8mg/kg, 6.9mg/kg, 7mg/kg, 7.1mg/kg, 7.2mg/kg, 7.3mg/kg, 7.4mg/kg, 7.5mg/kg, 7.6mg/ kg, 7.7mg/kg, 7.8mg/kg, 7.9mg/kg, 8mg/kg, 8.1mg/kg, 8.2mg/kg, 8.3mg/kg, 8.4mg/kg, 8.5 mg/kg, 8.6mg/kg, 8.7mg/kg, 8.8mg/kg, 8.9mg/kg, 9mg/kg, 9.1mg/kg, 9.2mg/kg, 9.3mg/kg, 9.4mg/kg, 9.5mg/kg, 9.6mg/kg, 9.7mg/kg, 9.8mg/kg, 9.9mg/kg, 10mg/kg, 11mg/kg, 12mg/kg , 13mg/kg, 14mg/kg, 15mg/kg, 16mg/kg, 17mg/kg, 18mg/kg, 19mg/kg, 20mg/kg, 21mg/kg, 22mg/kg, 23mg/kg, 24mg/kg, 25mg/kg, 26mg/kg, 27mg/kg, 28mg/kg, 29mg/kg, 30mg/kg, 31mg/kg , 32mg/kg, 33mg/kg, 34mg/kg, 35mg/kg, 36mg/kg, 37mg/kg, 38mg/kg, 39mg/kg, 40mg/kg, 41mg/kg, 42mg/kg, 43mg/kg, 44mg/kg, 45mg/kg, 46mg/kg, 47mg/kg, 48mg/kg, 49mg/kg to 50mg/kg.

Various factors may be considered when determining the dosage and administration schedule of the CFB dsRNA agents of the present invention. The absolute amount of the CFB dsRNA agent or CFB antisense polynucleotide agent administered will depend on various factors, including concurrent treatments, number of doses, and individual subject parameters, including age, physical condition, body shape, and weight. These factors are well known to those of ordinary skill in the art and can be resolved with routine experimentation. In some embodiments, a maximum dose may be used, i.e., the highest safe dose based on reasonable medical judgment.

In some embodiments, the methods of the present invention may include administering 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more doses of CFB dsRNA or CFB antisense polynucleotide to a subject. In some cases, a pharmaceutical compound (e.g., a CFB dsRNA or CFB antisense polynucleotide) may be administered to a subject at least daily, every other day, weekly, every other week, monthly, etc. The dose may be administered once or more per day, such as 2, 3, 4, 5 or more times in a 24-hour period. The pharmaceutical composition of the present invention may be administered once per day, or the CFB dsRNA or CFB antisense polynucleotide may be administered as two, three or more divided doses at appropriate intervals throughout the day, or even administered by continuous infusion or by controlled release formulation delivery. In some embodiments of the methods of the invention, the pharmaceutical compositions of the invention are administered to a subject once or more per day, once or more per week, once or more per month, or once or more per year.

In certain aspects, the methods of the present invention include administering a drug compound alone, in combination with one or more other CFB dsRNA agents or CFB antisense polynucleotide agents, and/or in combination with other drug therapies or therapeutic activities or regimens administered to a subject suffering from a CFB-related disease or condition. The drug compound can be administered in the form of a pharmaceutical composition. The pharmaceutical composition used in the methods of the present invention can be sterile and contain an amount of a CFB dsRNA agent or a CFB antisense polynucleotide agent that reduces the activity of the CFB polypeptide to a level sufficient to produce a desired response in a weight or volume unit suitable for administration to a subject. The dosage of the pharmaceutical composition comprising a CFB dsRNA agent or a CFB antisense polynucleotide agent to reduce the activity of a CFB protein administered to a subject can be selected according to different parameters, particularly according to the mode of administration used and the state of the subject. Other factors include the desired duration of treatment . If a subject does not respond adequately to the initial dose, a higher dose may be administered as tolerated by the patient (or the dose may be effectively increased by a different, more localized route of delivery).

As used herein, "CFB-related diseases," "CFB-related diseases and conditions," and "diseases and conditions caused and/or regulated by CFB" are intended to include any disease associated with the CFB gene or protein. These diseases may be caused by overproduction of CFB protein, mutations in the CFB gene, abnormal cleavage of CFB protein, abnormal interactions between CFB and other proteins or other endogenous or exogenous substances. Exemplary CFB-related diseases include, but are not limited to, autoimmune diseases, complement system dysfunction (including abnormal upregulation of complement components, such as CFB), C3 glomerulopathy (C3G), systemic lupus erythematosus (SLE), lupus nephritis, Ig-mediated renal diseases (such as IgA nephropathy and primary membranous nephropathy), nephropathy, diabetic nephropathy, polycystic nephropathy, membranous nephropathy, age-related macular degeneration (AMD), These include dry AMD and geographic atrophy, typical or infectious hemolytic uremic syndrome (tHUS), atypical uremic hemolytic syndrome (aHUS), asthma, psoriasis, thrombotic microangiopathy, ischemia and reperfusion injury, paroxysmal nocturnal hemoglobinuria (PNH), rheumatism, rheumatoid arthritis, multiple sclerosis (MS), neuromyelitis optica (NMO), immune complex-mediated glomerulonephritis (ICG-mediated glomerulonephritis), and inflammatory bowel disease (EMD). mediated GN), postinfectious glomerulonephritis (PIGN), antineutrophil cytoplasmic autoantibody-associated vasculitis (ANCA-AV), antiphospholipid antibody syndrome (APS), dysbacteriosis periodontitis, malaria-induced anemia, dermatomyositis, ulcerative colitis, Shigella toxin-induced Escherichia coli-associated hemolytic uremic syndrome, myasthenia gravis (MG), neuromyelitis optica (NMO), dense deposit disease, coronary artery disease , dermatomyositis, Graves' disease, atherosclerosis, Alzheimer's disease, systemic inflammatory response sepsis, septic shock, spinal cord injury, glomerulonephritis, Hashimoto's thyroiditis, type I diabetes mellitus, psoriasis, ulcers, autoimmune hemolytic anemia (AIHA), cold agglutinin disease, humoral and vascular transplant rejection, graft dysfunction, myocardial infarction, transplant sensitization, hyperlipidemia, sepsis, or lipid imbalances associated with CFB.

In certain aspects of the invention, a CFB dsRNA agent or CFB antisense polynucleotide agent of the invention may be administered to a subject at one or more times before or after diagnosis of a CFB-related disease or condition. In some aspects of the invention, a subject is at risk of having or developing a CFB-related disease or condition. A subject at risk of developing a CFB-related disease or condition is a subject with an increased likelihood of developing a CFB-related disease or condition compared to a control risk of developing a CFB-related disease or condition. In some embodiments of the invention, the risk level may be statistically significant compared to a control level of risk. At-risk subjects may include, for example, subjects who are or will be: subjects with pre-existing diseases and/or genetic abnormalities that make the subject more susceptible to a CFB-related disease or condition than a control subject without a pre-existing disease or genetic abnormality; subjects with a family and/or personal history of a CFB-related disease or condition; and subjects who have previously been treated for a CFB-related disease or condition. It should be understood that a pre-existing disease and/or genetic abnormality that makes a subject more susceptible to a CFB-related disease or condition may be a disease or genetic abnormality that, when present, has been pre-identified as being associated with a higher likelihood of developing a CFB-related disease or condition.

It should be understood that a CFB dsRNA agent or a CFB antisense polynucleotide agent can be administered to a subject according to the medical condition of the individual subject. For example, the medical care provided to the subject can evaluate the CFB level measured in a sample obtained from the subject and determine that it is desirable to reduce the CFB level of the subject by administering the CFB dsRNA agent or CFB antisense polynucleotide agent of the present invention. In this example, even if the subject is not diagnosed with a CFB-related disease (such as a disease disclosed herein), the CFB level can be regarded as a physiological characteristic of a CFB-related disease. A medical care provider can monitor changes in the CFB level of the subject as a measure of the efficacy of the CFB dsRNA agent or CFB antisense polynucleotide agent of the present invention administered. In a non-limiting example, a biological sample (e.g., a blood or serum sample) can be obtained from a subject and the subject's CFB level can be determined in the sample. A CFB dsRNA agent or a CFB antisense polynucleotide agent is administered to the subject, and a blood sample is obtained from the subject after administration, and the CFB level is determined using the sample, and the result is compared with the result determined in the subject's pre-administration (pre-) sample. A decrease in the CFB level in the subject's post-administration sample compared to the pre-administration level indicates that the administered CFB dsRNA agent or CFB antisense polynucleotide agent is effective in reducing the subject's lipid level.

Certain embodiments of the methods of the invention include modulating treatment, which includes administering to a subject a dsRNA agent or CFB antisense polynucleotide agent of the invention based at least in part on an assessment of changes in one or more physiological characteristics of a CFB-related disease or condition in the subject resulting from the treatment. For example, in some embodiments of the invention, the effect of an administered dsRNA agent or CFB antisense polynucleotide agent of the invention can be determined and used to assist in modulating the amount of the dsRNA agent or CFB antisense polynucleotide agent subsequently administered to the subject. In a non-limiting example, a dsRNA agent or CFB antisense polynucleotide agent of the present invention is administered to a subject, the subject's CFB level is determined after administration, and based at least in part on the determined level, a higher amount of the dsRNA agent or CFB antisense polynucleotide agent is determined to be desirable in order to increase the physiological effect of the administered agent, such as to reduce or further reduce the subject's CFB level. In another non-limiting example, a dsRNA agent or CFB antisense polynucleotide agent of the present invention is administered to a subject, the subject's CFB level is determined after administration, and based at least in part on the determined level, a lower amount of the dsRNA agent or CFB antisense polynucleotide agent is desired to be administered to the subject.

Thus, some embodiments of the invention include assessing changes in one or more physiological characteristics resulting from a subject's prior treatment to adjust the amount of the dsRNA agent or CFB antisense polynucleotide agent of the invention subsequently administered to the subject. Some embodiments of the methods of the invention include performing 1, 2, 3, 4, 5, 6 or more assays for physiological characteristics of a CFB-related disease or condition to assess and/or monitor the efficacy of the administered CFB dsRNA agent or CFB antisense polynucleotide agent of the invention, and optionally using the assays to adjust one or more of the dose, administration regimen, and/or administration frequency of the dsRNA agent or CFB antisense polynucleotide agent of the invention to treat a CFB-related disease or condition in a subject. In some embodiments of the methods of the invention, the desired result of administering an effective amount of a dsRNA agent or CFB antisense polynucleotide agent of the invention to a subject is a decrease in the subject's CFB mRNA level, CFB protein level in the subject, or CH50 activity (a measure of total hemolytic complement), AH50 (a measure of hemolytic activity of the alternative complement pathway), lactate dehydrogenase (LDH) (a measure of intravascular hemolysis), hemoglobin level; any one or more of C3, C9, C5, C5a, C5b, and soluble C5b-9 complex.

As used herein, the terms "treat," "therapeutic," or "treating" when used with respect to a CFB-related disease or disorder may refer to prophylactic treatment to reduce the likelihood of a subject developing a CFB-related disease or disorder, and may also refer to treatment after a subject has a CFB-related disease or disorder to eliminate the CFB-related disease or disorder or to reduce the level of the CFB-related disease or disorder, to prevent the CFB-related disease or disorder from becoming more advanced (e.g., more severe), and/or to slow the progression of the CFB-related disease or disorder in the subject, compared to a subject in the absence of a treatment that reduces the activity of a CFB polypeptide in the subject.

Certain embodiments of the agents, compositions and methods of the present invention can be used to inhibit CFB gene expression. The terms "inhibit", "silencing", "reducing", "down-regulating" and "knockdown" herein with respect to CFB gene expression refer to when a cell, cell group, tissue, organ or subject is contacted with (e.g., treated with) a CFB dsRNA agent or a CFB antisense polynucleotide agent of the present invention, the expression of the CFB gene is reduced as measured by one or more of the following in the cell, cell group, tissue, organ or subject that transcribes the CFB gene: the level of RNA transcribed from the gene, the level of CFB activity expressed and the level of CFB polypeptide, protein or protein subunit translated from mRNA, respectively, compared to a control level of RNA transcribed from the CFB gene, the level of CFB activity expressed and the level of CFB polypeptide, protein or protein subunit translated from mRNA. In some embodiments, the control level is the level in a cell, tissue, organ, or subject that has not been contacted with (e.g., treated with) a CFB dsRNA agent or a CFB antisense polynucleotide agent.

A variety of administration routes of CFB dsRNA agents or CFB antisense polynucleotide agents can be used in the methods of the present invention. The specific delivery mode selected will depend at least in part on the specific condition being treated and the dose required for therapeutic efficacy. In general, the methods of the present invention can be implemented using any medically acceptable mode of administration, which means any mode that produces an effective therapeutic level for a CFB-related disease or condition without causing clinically unacceptable side effects. In some embodiments of the present invention, CFB dsRNA agents or CFB antisense polynucleotide agents can be administered via oral, enteral, mucosal, subcutaneous and/or parenteral routes. The term "parenteral" includes subcutaneous, intravenous, intrathecal, intramuscular, intraperitoneal and intrasternal injection or infusion techniques. Other routes include, but are not limited to, nasal (e.g., through a nasogastric tube), transdermal, vaginal, rectal, sublingual, and inhalation. The delivery route of the present invention may include intrathecal, intraventricular, or intracranial. In some embodiments of the present invention, a CFB dsRNA agent or a CFB antisense polynucleotide agent may be placed in a sustained-release matrix and administered by placing the matrix in a subject. In some aspects of the present invention, nanoparticles coated with a delivery agent targeting specific cells or organelles may be used to deliver a CFB dsRNA agent or a CFB antisense polynucleotide agent to a subject's cells. Various delivery modes, methods, and reagents are known in the art. Non-limiting examples of delivery methods and delivery agents are also provided elsewhere herein. In some aspects of the present invention, the term "delivery" related to CFB dsRNA agents or CFB antisense polynucleotide agents can refer to the administration of one or more "naked" CFB dsRNA agents or CFB antisense polynucleotide agent sequences to cells or subjects, and in certain aspects of the present invention, "delivery" refers to administration to cells or subjects by transfection, delivering cells containing CFB dsRNA agents or CFB antisense polynucleotide agents to subjects, and delivering vectors encoding CFB dsRNA agents or CFB antisense polynucleotide agents to subjects. Delivery of CFB dsRNA agents or CFB antisense polynucleotide agents using transfection means can include administering vectors to cells and/or subjects.

In some methods of the present invention, one or more CFB dsRNA agents or CFB antisense polynucleotide agents can be administered in a formulation that can be administered in a pharmaceutically acceptable solution that can typically contain pharmaceutically acceptable concentrations of salt, buffers, preservatives, compatible carriers, adjuvants, and optional other therapeutic ingredients. In some embodiments of the present invention, CFB dsRNA agents or CFB antisense polynucleotide agents can be formulated with another therapeutic agent for simultaneous administration. According to the methods of the present invention, CFB dsRNA agents or CFB antisense polynucleotide agents can be administered in a pharmaceutical composition. In general, a pharmaceutical composition comprises a CFB dsRNA agent or a CFB antisense polynucleotide agent and an optional pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers are well known to those of ordinary skill in the art. As used herein, a pharmaceutically acceptable carrier refers to a non-toxic material that does not interfere with the effectiveness of the biological activity of the active ingredient, such as the ability of a CFB dsRNA agent or a CFB antisense polynucleotide agent to inhibit the expression of a CFB gene in a cell or subject. A variety of methods for administering and delivering a CFB dsRNA agent or a CFB antisense polynucleotide agent for treatment are known in the art and can be used in the methods of the present invention.

Pharmaceutically acceptable carriers include diluents, fillers, salts, buffers, stabilizers, solubilizers and other materials well known in the art. Exemplary pharmaceutically acceptable carriers are described in U.S. Patent No. 5,211,657, and other carriers are known to those skilled in the art. Such formulations may generally contain salts, buffers, preservatives, compatible carriers and optional other therapeutic agents. When used in medicines, salts should be pharmaceutically acceptable, but non-pharmaceutically acceptable salts can be conveniently used to prepare pharmaceutically acceptable salts thereof and are not excluded from the scope of the present invention. Such pharmacologically and pharmaceutically acceptable salts include, but are not limited to, salts prepared from the following acids: hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, maleic acid, acetic acid, salicylic acid, citric acid, formic acid, malonic acid, succinic acid, etc. In addition, pharmaceutically acceptable salts can be prepared as alkaline metal or alkaline earth salts, such as sodium salts, potassium salts or calcium salts.

Some embodiments of the methods of the present invention include administering one or more CFB dsRNA reagents or CFB antisense polynucleotide reagents directly to a tissue. In some embodiments, the tissue to which the compound is administered is a tissue in which a CFB-related disease or condition exists or may occur, a non-limiting example of which is the heart. Direct tissue administration can be achieved by direct injection or other means. Many orally delivered compounds naturally travel and pass through the liver and kidneys, and some embodiments of the treatment methods of the present invention include orally administering one or more CFB dsRNA agents to a subject. CFB dsRNA agents or CFB antisense polynucleotide agents, alone or in combination with other therapeutic agents, can be administered once, or alternatively, they can be administered multiple times. If multiple administrations are given, the CFB dsRNA agent or CFB antisense polynucleotide agent may be administered by different routes. For example, although not intended to be limiting, the first (or first few) administrations may be subcutaneous, and one or more additional administrations may be oral and/or systemic.

For embodiments of the present invention where systemic administration of CFB dsRNA agents or CFB antisense polynucleotide agents is desired, CFB dsRNA agents or CFB antisense polynucleotide agents may be formulated for parenteral administration by injection, such as by bolus injection or continuous infusion. Injectable formulations may be in unit dose form, such as ampoules or multi-dose containers, with or without added preservatives. CFB dsRNA agent formulations (also referred to as pharmaceutical compositions) may be in the form of suspensions, solutions or emulsions in oily or aqueous carriers, and may contain formulation agents, such as suspending agents, stabilizers and/or dispersants.

Preparations for parenteral administration include sterile aqueous or non-aqueous solutions, suspensions and emulsions. Examples of non-aqueous solvents include propylene glycol, polyethylene glycol, vegetable oils (such as olive oil) and injectable organic esters (such as ethyl oleate). Aqueous carriers include water, alcohol/aqueous solutions, emulsions or suspensions, including saline and buffered media. Parenteral carriers include sodium chloride solution, Ringer's dextrose, dextrose and sodium chloride, lactated Ringer's solution or non-volatile oils. Intravenous carriers include liquids and nutritional supplements, electrolyte supplements (such as supplements based on Ringer's dextrose), etc. Preservatives and other additives may also be present, such as antimicrobial agents, antioxidants, chelating agents, and inert gases. Other forms of administration (such as intravenous administration) will have lower doses. If the subject does not respond adequately to the initial dose, a higher dose may be used within the patient's tolerance range (or the administered dose may actually be increased by a different, more localized delivery route). Multiple doses may be used daily as needed to achieve appropriate systemic or local levels of one or more CFB dsRNA agents or CFB antisense polynucleotide agents and to achieve an appropriate reduction in CFB protein activity.

In other embodiments, the methods of the invention include the use of a delivery vehicle, such as a biocompatible microparticle, nanoparticle, or implant suitable for implantation into a recipient (e.g., a subject). Exemplary bioerosion implants that can be used according to the methods are described in PCT Publication No. WO 95/24929 (incorporated herein by reference), which describes a biocompatible, biodegradable polymer matrix for containing biomacromolecules.

Both non-biodegradable and biodegradable polymer matrices can be used in the methods of the present invention to deliver one or more CFB dsRNA reagents or CFB antisense polynucleotide reagents to a subject. In some embodiments, the matrix can be biodegradable. The matrix polymer can be a natural or synthetic polymer. The polymer can be selected based on the time period over which release is desired, typically on the order of a few hours to a year or more. Typically, release over a period of a few hours to three to twelve months can be used. The polymer is optionally in the form of a hydrogel that can absorb up to about 90% of its weight in water and is further optionally crosslinked with multivalent ions or other polymers.

Generally speaking, in some embodiments of the present invention, CFB dsRNA agents or CFB antisense polynucleotide agents can be delivered by diffusion or by degradation of a polymer matrix using a bioerodible implant. Exemplary synthetic polymers for such uses are well known in the art. Biodegradable polymers and non-biodegradable polymers can be used to deliver CFB dsRNA agents or CFB antisense polynucleotide agents by methods known in the art. Bioadhesive polymers such as bioerodible hydrogels (see H. S. Sawhney, C. P. Pathak and J. A. Hubell in Macromolecules, 1993, 26, 581-587, the teachings of which are incorporated herein by reference) can also be used to deliver CFB dsRNA agents or CFB antisense polynucleotide agents to treat CFB-related diseases or disorders. Other suitable delivery systems may include timed release, delayed release, or sustained release delivery systems. Such systems may avoid repeated administration of CFB dsRNA agents or CFB antisense polynucleotide agents, thereby increasing convenience for subjects and healthcare professionals. Many types of release delivery systems are available and are known to those of ordinary skill in the art. (See, for example, U.S. Patent Nos. 5,075,109; 4,452,775; 4,675,189; 5,736,152; 3,854,480; 5,133,974; and 5,407,686 (the teachings of each of which are incorporated herein by reference). In addition, pump-based hardware delivery systems may be used, some of which are suitable for implantation.

The use of long-term sustained release implants may be suitable for preventive treatment of subjects and for subjects at risk for recurrent CFB-related diseases or conditions. As used herein, long-term release refers to the construction and arrangement of the implant to deliver therapeutic levels of CFB dsRNA agents or CFB antisense polynucleotide agents for at least 10 days, 20 days, 30 days, 60 days, 90 days, six months, one year or longer. Long-term sustained release implants are well known to those of ordinary skill in the art and include some of the above-mentioned release systems.

Therapeutic preparations of CFB dsRNA agents or CFB antisense polynucleotide agents can be prepared for storage in the form of lyophilized preparations or aqueous solutions by mixing the molecule or compound having the desired purity with an optional pharmaceutically acceptable carrier, excipient or stabilizer [Remington's Pharmaceutical Sciences ^21st edition, (2006)]. Acceptable carriers, excipients or stabilizers are nontoxic to recipients at the dosages and concentrations employed, and include buffers such as phosphates, citrates and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (e.g., octadecyldimethylbenzylammonium chloride; hexamethylammonium chloride; benzoxamethylammonium chloride, benzethonium chloride; phenol, butyl alcohol or benzyl alcohol; alkyl parabens, such as methyl or propyl parabens; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) ) polypeptides; proteins such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates, including glucose, mannose, or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions such as sodium; metal complexes (e.g., Zn-protein complexes); and/or nonionic surfactants such as ^TWEEN® , ^PLURONICS® , or polyethylene glycol (PEG). Cells, subjects, and controls

The methods of the present invention can be used in conjunction with cells, tissues, organs and/or subjects. In some aspects of the present invention, the subject is a human or vertebrate mammal, including but not limited to dogs, cats, horses, cows, goats, mice, rats and primates, such as monkeys. Therefore, the present invention can be used to treat CFB-related diseases or conditions in human and non-human subjects. In some aspects of the present invention, the subject can be a farm animal, a zoo animal, a domesticated animal or a non-domesticated animal, and the methods of the present invention can be used in veterinary prevention and treatment programs. In some embodiments of the present invention, the subject is a human and the methods of the present invention can be used in human prevention and treatment programs.

Non-limiting examples of subjects to which the present invention may be applied are subjects diagnosed with, suspected of having, or at risk of having a disease or condition associated with higher than desired CFB expression and/or activity, also referred to as "elevated CFB expression levels." Non-limiting examples of diseases and conditions associated with higher than desired CFB expression and/or activity are described elsewhere herein. The methods of the present invention may be applied to subjects diagnosed with, or believed to be at risk of having or developing, a disease or condition associated with higher than desired CFB expression and/or activity at the time of treatment. In certain aspects of the invention, the disease or condition associated with higher than desired CFB expression and/or activity is an acute disease or condition, and in certain aspects of the invention, the disease or condition associated with higher than desired CFB expression and/or activity is a chronic disease or condition.

In a non-limiting example, the CFB dsRNA agents of the invention are administered to subjects diagnosed with, suspected of having, or at risk for statin-resistant hypercholesterolemia, a disease in which CFB expression needs to be reduced. The methods of the invention can be applied to subjects who have been diagnosed with the disease or condition at the time of treatment, or are believed to be at risk for having or developing the disease or condition.

In another non-limiting example, the CFB dsRNA agent of the present invention is administered to a subject diagnosed with, suspected of having, or at risk of hyperlipidemia, a disease in which CFB expression needs to be reduced. The methods of the present invention can be applied to subjects diagnosed with, or believed to be at risk of having or developing, such a disease or condition during treatment.

Cells to which the methods of the present invention can be applied include in vitro , in vivo , and ex vivo cells. Cells can be in a subject, in culture, and/or in suspension, or in any other suitable state or condition. Cells to which the methods of the present invention can be applied can be hepatocytes, liver cells, myocardial cells, pancreatic cells, cardiovascular cells, kidney cells, or other types of vertebrate cells, including human and non-human mammalian cells. In certain aspects of the present invention, cells to which the methods of the present invention can be applied are healthy, normal cells, which are not known to be disease cells. In certain embodiments of the present invention, the cells to which the methods and compositions of the present invention are applied are hepatocytes, liver cells, myocardial cells, pancreatic cells, cardiovascular cells and/or kidney cells. In certain aspects of the present invention, the control cells are normal cells, but it should be understood that cells with a disease or condition can also serve as control cells under certain circumstances, such as comparing the results of treating cells with a disease or condition with the results of cells with the disease or condition not being treated.

According to the methods of the present invention, the level of CFB polypeptide activity can be determined and compared to a control level of CFB polypeptide activity. The control can be a predetermined value, which can take a variety of forms. It can be a single cutoff value, for example, a median or mean value. It can be established based on comparison groups, such as a group with normal levels of CFB polypeptide and/or CFB polypeptide activity and a group with increased levels of CFB polypeptide and/or CFB polypeptide activity. Another non-limiting example of a comparison group can be a group with one or more symptoms of a CFB-related disease or condition or diagnosed with a CFB-related disease or condition; a group without one or more symptoms of the disease or condition or not diagnosed with the disease or condition; a group of subjects treated with the siRNA of the present invention; a group of subjects not treated with the siRNA of the present invention. Typically, the control may be based on apparently healthy normal individuals or apparently healthy cells of the appropriate age group. It should be understood that the control according to the present invention may be a sample of a material tested in parallel with the experimental material in addition to a predetermined value. Examples include samples from a control population or control samples produced by manufacturing to be tested in parallel with the experimental samples. In some embodiments of the present invention, the control may include cells or subjects that have not been contacted or treated with the CFB dsRNA agent of the present invention, and in such cases, the control level of CFB polypeptide and/or CFB polypeptide activity may be compared with the level of CFB polypeptide and/or CFB polypeptide activity in cells or subjects contacted with the CFB dsRNA agent or CFB antisense polynucleotide agent of the present invention.

In some embodiments of the present invention, the level of CFB polypeptide determined for a subject can be a control level for the level of CFB polypeptide determined for the same subject at a different time to be compared. In one non-limiting example, the level of CFB is determined in a biological sample obtained from a subject who has not been administered the CFB treatment of the present invention. In some embodiments, the biological sample is a serum sample. The level of CFB polypeptide determined in the sample obtained from the subject can be used as a baseline or control value for the subject. In the treatment methods of the present invention, after administering one or more CFB dsRNA agents to the subject, one or more additional serum samples can be obtained from the subject, and the level of CFB polypeptide in the subsequent one or more samples can be compared to the control/baseline level of the subject. Such a comparison can be used to assess the onset, progression or regression of a CFB-related disease or condition in a subject. For example, a level of CFB polypeptide in a baseline sample obtained from a subject that is higher than the level obtained from the same subject after administration of a CFB dsRNA agent or CFB antisense polynucleotide agent of the present invention to the subject indicates regression of a CFB-related disease or condition, and indicates the efficacy of the administered CFB dsRNA agent of the present invention for treating a CFB-related disease or condition.

In some aspects of the invention, one or more of the levels of CFB polypeptide and/or CFB polypeptide activity determined for a subject can serve as a control value for later comparison of the levels of CFB polypeptide and/or CFB activity in the subject under the same circumstances, thereby allowing assessment of changes in the subject relative to a "baseline" CFB polypeptide activity. Thus, an initial CFB polypeptide level and/or initial CFB polypeptide activity level can be present and/or determined in a subject, and the methods and compounds of the invention can be used to reduce the CFB polypeptide level and/or CFB polypeptide activity in a subject, wherein the initial level serves as a control level for the subject.

Using the methods of the present invention, the CFB dsRNA agents and/or CFB antisense polynucleotide agents of the present invention can be administered to a subject. The efficacy of the administration and treatment of the present invention can be evaluated when the level of CFB polypeptide in a serum sample obtained from a subject is reduced by at least 0.5%, 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95% or more compared to the pre-administration level of CFB polypeptide in a serum sample obtained from the subject at a previous time point, or compared to a non-contact control level (e.g., the level of CFB polypeptide in a control serum sample). It should be understood that the level of CFB polypeptide and the level of CFB polypeptide activity are both related to the level of CFB gene expression. Certain embodiments of the methods of the invention include administering to a subject a CFB dsRNA and/or CFB antisense of the invention in an amount effective to inhibit CFB gene expression and thereby reduce CFB polypeptide levels and reduce CFB polypeptide activity levels in the subject.

Some embodiments of the invention include determining the presence, absence, and/or amount (also referred to herein as level) of a CFB polypeptide in one or more biological samples obtained from one or more subjects. The determination can be used to assess the effectiveness of the treatment methods of the invention. For example, the methods and compositions of the invention can be used to determine the level of a CFB polypeptide in a biological sample obtained from a subject previously treated with a CFB dsRNA agent and/or CFB antisense agent of the invention. A level of CFB polypeptide measured in a serum sample obtained from a treated subject that is at least 0.5%, 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95% or more lower than the level of CFB polypeptide measured in the subject before treatment or compared to the level in a non-contact control biological sample indicates a level of efficacy of the treatment administered to the subject.

In some embodiments of the present invention, the physiological characteristics of a CFB-related disease or condition measured for a subject can be a control measurement for a physiological characteristic measured for the same subject at a different time to be compared. In a non-limiting example, a physiological characteristic such as CFB mRNA level, CFB protein level or CH50 activity, AH50, lactate dehydrogenase (LDH), hemoglobin level in a subject; any one or more levels of C3, C9, C5, C5a, C5b and soluble C5b-9 complex in a plasma or tissue sample are measured in a biological sample (e.g., a serum sample) obtained from a subject (who has not yet been treated with a CFB agent of the present invention). The CFB mRNA level (and/or other physiological characteristics of a CFB disease or condition) measured in a sample obtained from a subject can serve as a baseline or control value for the subject. After one or more administrations of a CFB dsRNA agent to a subject using the treatment methods of the present invention, one or more additional serum samples can be obtained from the subject, and the CFB mRNA level and/or CFB protein level in the subsequent sample or samples can be measured and compared to the control/baseline level and/or ratio of the subject, respectively. Such comparisons can be used to assess the onset, progression, or regression of a CFB-related disease or condition in a subject. For example, a CFB mRNA level in a baseline sample obtained from a subject that is higher than the CFB mRNA level measured in a sample obtained from the same subject after administration of a CFB dsRNA agent or CFB antisense polynucleotide agent of the invention to the subject indicates that the CFB-related disease or disorder is regressing and indicates the efficacy of the administered CFB dsRNA agent of the invention for treating the CFB-related disease or disorder.

In certain aspects of the invention, one or more physiological characteristic values determined for a CFB-related disease or condition in a subject can be used as a control value to later compare the physiological characteristic of the same subject, thereby allowing the subject's changes relative to the "baseline" physiological characteristic to be evaluated. Thus, a subject may have an initial physiological characteristic present and/or determined, and the methods and compounds of the invention can be used to reduce the level of CFB polypeptide and/or CFB polypeptide activity in the subject, wherein the initial physiological characteristic measurement value is used as a control for the subject.

Using the methods of the present invention, the CFB dsRNA agents and/or CFB antisense polynucleotide agents of the present invention can be administered to a subject in an effective amount to treat a CFB-related disease or condition. The effectiveness of the administration and treatment of the present invention can be assessed by determining changes in one or more physiological characteristics of a CFB-related disease or condition. In a non-limiting example, the CFB mRNA level in a serum sample obtained from a subject is reduced by at least 0.5%, 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95% or more compared to pre-administration lipids in a serum sample obtained from a subject at a previous time point, or compared to a non-contact control level (e.g., CFB mRNA level in a control serum sample). It is understood that the CFB mRNA level, CFB protein level, or lipid level, triglyceride, cholesterol level, free fatty acid level in a subject's plasma or tissue sample are all related to the CFB gene expression level. Certain embodiments of the methods of the present invention include administering to a subject an amount of the CFB dsRNA agent and/or CFB antisense agent of the present invention that is effective in inhibiting CFB gene expression, thereby reducing the subject's CFB mRNA level, CFB protein level, or otherwise having a positive effect on the subject's physiological characteristics of a CFB-related disease or condition.

Some embodiments of the invention include determining the presence, absence and/or changes in physiological characteristics of a CFB-related disease or condition using methods such as, but not limited to, the following: (1) evaluating physiological characteristics of one or more biological samples obtained from one or more subjects; (2) imaging the subject (e.g., but not limited to, obtaining liver images); (3) physical examination of the subject. This assay can be used to evaluate the efficacy of the treatment methods of the invention. Kits

Kits of CFB dsRNA reagents and/or CFB antisense polynucleotide reagents and instructions for use in the methods of the present invention are also within the scope of the present invention. The kits of the present invention may include one or more of CFB dsRNA reagents, CFB sense polynucleotides, and CFB antisense polynucleotide reagents that can be used to treat CFB-related diseases or conditions. Kits containing one or more CFB dsRNA reagents, CFB sense polynucleotides, and CFB antisense polynucleotide reagents can be prepared for use in the treatment methods of the present invention. The components of the kits of the present invention can be packaged in an aqueous medium or in a lyophilized form. The kit of the present invention may include a carrier that is compartmentalized to receive in close confinement therein one or more container means or a series of container means, such as test tubes, vials, flasks, bottles, syringes, etc. The first container means or a series of container means may contain one or more compounds, such as CFB dsRNA reagents and/or CFB sense or antisense polynucleotide reagents. The second container means or a series of container means may contain targeting agents, labeling agents, delivery agents, etc., which may be included as part of the CFB dsRNA agent and/or CFB antisense polynucleotide to be administered in one embodiment of the treatment method of the present invention.

The kit of the present invention may also include instructions. The instructions are usually in written form and are used to guide how to implement the treatment contained in the kit and make decisions based on the treatment.

The following examples are used to illustrate specific examples of the practice of the present invention and are not intended to limit the scope of the present invention. A person of ordinary skill in the art will appreciate that the present invention will be applied to various compositions and methods.

Example 1. Phosphamide Compound 2

A solution of DMTrCl (232 g, 684 mmol, 1.0 eq) in pyridine (400 mL) was added to a solution of isomannitol compound A (100 g, 684 mmol, 1.0 eq) in pyridine (600 mL), and the mixture was stirred at 25 °C for 12 hours. LC-MS showed that compound A was completely consumed and a main peak with the expected mass was detected. The resulting reaction mixture was diluted with water (500 mL), extracted with DCM (500 mL *2), and the combined organic phases were washed with brine (500 mL), dried over Na ₂ SO ₄ and concentrated in vacuo to obtain a residue. The residue was purified by column chromatography (DCM/MeOH=100/1 to 50/1, 0.1% Et ₃ N) to obtain compound B (150 g, 48.9% yield) as a yellow solid.

¹ H NMR: EC4783-404-P1B1_C (400 MHz, DMSO- d ₆ ) δ ppm 7.46 (br d, J =7.63 Hz, 2 H) 7.28 - 7.37 (m, 6 H) 7.19 - 7.25 (m, 1 H) 6.90 (br d, J =7.88 Hz, 4 H) 4.70 (d, J =6.50 Hz, 1 H) 3.99 - 4.09 (m, 6 H) 3.88 - 3.96 (m, 2 H) 3.83 (br dd, J =7.82, 6.94 Hz, 1 H) 3.74 (s, 6 H) 3.41 (br t, J =8.13 Hz, 1H) 3.05 (t, J =8.44 Hz, 1 H) 2.85 (br t, J =7.50 Hz, 1 H).

To a solution of compound B (80.0 g, 178 mmol, 1.0 eq) in DCM (800 mL) was added 2H-tetrazole (0.45 M, 436 mL, 1.1 eq ) dropwise under _N2 atmosphere at 25°C, and then a solution of compound C (80.6 g, 267 mmol, 85.0 mL, 1.5 eq ) in DCM (200 mL) was added dropwise to the mixture. The reaction mixture was stirred at 25°C for 1.0 h. LC-MS showed that compound B was completely consumed and a major peak with the desired mass was detected. The resulting reaction mixture was cooled to -20°C and poured into ice-cold saturated NaHCO ₃ (500 mL), extracted with DCM (500 mL*3), the combined organic layers were washed with saturated NaHCO ₃ / brine = 1:1 (300 mL/300 mL), dried over Na ₂ SO ₄ and concentrated in vacuo (35°C) to obtain a residue (100 mL). The residue was purified by column chromatography (Al ₂ O ₃ , DCM/MeOH = 100/1 to 50/1, 0.1% Et ₃ N) to obtain compound 2 (77 g, 119 mmol, yield 66.5%) as a white solid.

¹ H NMR: EC4783-423-P1B1_C (400 MHz, DMSO- d ₆ ) δ ppm 7.22 (br d, J=7.50 Hz, 2 H) 7.05 - 7.14 (m, 6 H) 6.96 - 7.02 (m, 1 H) 6.67 (br dd, J=8.82, 1.81 Hz, 4 H) 3.95 - 4.07 (m, 2 H) 3.73 - 3.83 (m, 1 H) 3.62 - 3.72 (m, 2 H) 3.48 - 3.53 (m, 6 H) 3.27 - 3.37 (m, 3 H) 3.11 (s, 6 H) 2.82 (td, J=8.54, 2.31 Hz, 1 H) 2.47 - 2.63 (m, 3 H) 2.28 (br d, J=1.63 Hz, 3 H) 0.82 - 1.00 (m, 13 H).

Phosphamide compound 1

To a solution of compound B (500 mg, 1.11 mmol, 1.0 eq) in DCM (5.0 mL) were added compound D (607 mg _, 3.34 mmol, 3.0 eq) and DIEA (432 mg, 3.34 mmol, 582 μL, 3.0 eq) under N2 at 0-5°C, and the mixture was stirred at 25°C for 1.0 hour. LC-MS showed that compound B was completely consumed, several new peaks were shown on LC-MS, and about 70.9% of the desired compound was detected. The resulting reaction mixture was cooled to -20°C and poured into a cold (0-5°C) saturated NaHCO ₃ (5.0 mL) solution, extracted with DCM (5.0 mL*2), the combined organic layer was washed with cold (0-5°C) saturated NaHCO ₃ / brine = 1:1 (5.0 mL/5.0 mL), dried over Na ₂ SO ₄ and concentrated in vacuo to obtain a residue (~5 mL). The residue was purified by column chromatography (basic Al ₂ O ₃ , petroleum ether/ethyl acetate = 10/1 to 5/1, 0.1% Et ₃ N) to give compound 1 (280 mg, 471 μmol, yield 42.3%) as a white solid.

¹ H NMR: EC10615-49-P1N (400 MHz, DMSO- d ₆ ) δ ppm 7.44 (br d, J =7.63 Hz, 2 H), 7.31 (br t, J =7.94 Hz, 6 H), 7.18 - 7.26 (m, 1 H), 6.89 (brd, J =8.00 Hz, 4 H), 4.08 - 4.13 (m, 1 H), 3.95 - 4.03 (m, 1 H), 3.84 - 3.93 (m, 1 H), 3.77 - 3.83 (m, 1 H), 3.74 (s, 6 H), 3.43 - 3.53 (m, 3 H), 3.38 (br d, J =6.75 Hz, 1 H), 2.94 - 3.04 (m, 1 H), 2.70 - 2.85 (m, 1 H), 1.09 - 1.15 (m, 12 H), 1.07 (br s, 3 H).

Other phosphoramidites can be prepared according to the procedures described herein and/or prior art such as, but not limited to, US 426,220 and WO 02/36743.

Example 2. Preparation of a solid support containing the phosphoramidite monomer of the present invention Represents the macroporous aminomethyl polyethylene resin carrier part

Under nitrogen protection, add dichloromethane (19.50 kg) to a 50L glass reactor, start stirring, control the temperature at 20-30°C, add DMTrimann (1.47 kg), triethylamine (1.50 kg), 4-dimethylaminopyridine (0.164 kg) and succinic anhydride (1.34 kg) to the glass reactor, keep warm at 20-30°C for 18 hours, take samples, and terminate the reaction. Add saturated sodium bicarbonate solution (22.50 kg) to the reaction system, stir for 10-20 minutes, and separate the layers. Separate the organic phase, extract the aqueous phase twice with dichloromethane, combine the organic phases, dry with anhydrous sodium sulfate, filter, and concentrate in vacuum to obtain a residue of 1.83 kg of gray to off-white solid.

Add N, N-dimethylformamide (23.50 kg) to a 100 L glass kettle and stir. The temperature is controlled at 20-30 ° C. Under nitrogen protection, add the product of the previous step, O-benzotriazole tetramethyluronium hexafluorophosphate (0.33 kg) and N, N-diisopropylethylamine (0.13 kg) to the above 100 L glass kettle through a solid addition funnel, stir for 10-30 minutes, and then discharge into a 50 L zinc barrel for standby use. The macroporous amine methyl resin (3.25 kg) (purchased from Tianjin Nankai Hecheng Technology Co., Ltd., batch number HA2X1209, loading 0.48 mmol/g) was added to the above 100L solid phase synthesis reactor through a solid addition funnel to control the temperature at 20-30°C, and N, N-dimethylformamide (21.00 kg + 21.00 kg) and the reaction solution in the zinc barrel in the previous step were added to the solid phase synthesis reactor. The system was subjected to an adiabatic reaction, and the solid load was tracked to be ≥ 250 umol/g. The load detection method was UV. Nitrogen pressure filtration was performed, and the filter cake was washed 3 times with N, N-dimethylformamide (26.00 kg + 26.10 kg + 26.00 kg), and the filter cake was retained in the kettle. Add CAP.A (50% acetonitrile and 50% acetic anhydride, 4.40kg+4.42kg+4.30kg) and CAP.B (20% pyridine and 30% N-methylimidazole and 50% acetonitrile, 4.40kg+4.40kg+4.47kg) to a 80L glass reactor and stir for 3-8min before use. Repeat this operation 3 times, cap, and add acetonitrile (18.00kg+18.00kg+18.00kg+17.50kg+17.50kg) to the solid phase synthesis reactor. Bubble nitrogen for 10-30 minutes and filter. Repeat this operation 4 times, purge the filter cake with nitrogen in the solid phase synthesis reactor for 2-4 hours, and then transfer it to a 50L pressure filter tank. The temperature was controlled at 15-30°C and the drying was continued. After drying, a yellow to white solid product was obtained with a weight of 3.516 kg.

The isomannitol residue is added to the 5'-end or 3'-end of the oligonucleotide chain by methods well known to those skilled in the art, such as the invab method, and further added to the targeting group.

Example 3. Synthesis of CFB RNAi agent.

The CFB RNAi agent duplexes shown in Tables 2-3 above were synthesized according to the following general procedure:

The sense and antisense sequences of siRNA are synthesized on an oligonucleotide synthesizer using a well-established solid phase synthesis method based on phosphoramidite chemistry. Oligonucleotide chain growth is achieved through a 4-step cycle: deprotection, condensation, capping, and an oxidation or sulfurization step for the addition of each nucleotide. The synthesis is carried out on a solid support made of controlled pore glass (CPG, 1000 Å). Monomer phosphoramidites can be purchased from commercial sources or can be the phosphoramidite compounds in Example 1. The phosphoramidite compounds herein can be linked to the 3'-end as monomeric phosphoramidites and further linked to a CPG solid support. In the case of 5' end attachment, the phosphoramidite compound can be used for the final coupling reaction and can be further conjugated to a targeting ligand if desired.

Phosphoramidites with GalNAc ligand clusters (GLS-5* or GLS-15* phosphoramidites as non-limiting examples) are disclosed in WO2023/045995A1 (incorporated herein in its entirety). Synthesis was performed at a 2 μmol scale for siRNAs used for in vitro screening (Table 2) and at a 5 μmol or greater scale for siRNAs used for in vivo testing (Table 3). A GalNAc ligand-attached CPG solid support was used with a GalNAc ligand (GLO-n phosphoramidites as non-limiting examples disclosed in WO2023/045995A1 (incorporated herein in its entirety)) conjugated to the 3'-end of the sense strand. In the case where a GalNAc ligand (GLS-5* or GLS-15* as a non-limiting example) is attached to the 5' end of the sense chain, a GLS-5* or GLS-15* phosphoamidite with a GalNAc ligand cluster is attached to the 5' end of the sense chain, and the GalNAc phosphoamidite is used for the final coupling reaction. A 3% solution of trichloroacetic acid (TCA) in dichloromethane is used for the deprotection of the 4,4'-dimethoxytrityl protecting group (DMT). 5-Ethylthio-1H-tetrazole is used as an activating agent. I ₂ in THF/Py/H ₂ O and phenylacetyl disulfide (PADS) in pyridine/MeCN are used for oxidation and sulfidation reactions, respectively. After the last solid phase synthesis step, the solid support-bound oligomers were cleaved and the protecting groups removed by treatment with a 1:1 volume of 40 wt % aqueous methylamine and 28 % ammonium hydroxide solution. To synthesize siRNA for in vitro screening, the crude mixture was concentrated. The remaining solid was dissolved in 1.0 M NaOAc and ice-cold EtOH was added to precipitate the single-chain product as the sodium salt, which was used for annealing without further purification. To synthesize siRNA for in vivo testing, the crude single-chain product was further purified by ion pair reversed phase HPLC (IP-RP-HPLC). The purified single-stranded oligonucleotide product from IP-RP-HPLC was converted to the sodium salt by dissolving in 1.0 M NaOAc and precipitating by adding ice-cold EtOH. Sense and antisense oligonucleotides were annealed by equimolar complementation in water to form a double-stranded siRNA product, which was lyophilized to provide a fluffy white solid.

In certain studies, methods for attaching a targeting group comprising GalNAc (also referred to herein as a GalNAc delivery compound) to the 5' end of the sense strand include using a GalNAc phosphoramidite (GLS-5* or GLS-15* phosphoramidite) in the final coupling step of a solid phase synthesis, using a synthetic process such as that used in oligonucleotide chain extension to add nucleotides to the 5' end of the sense strand.

In some studies, the method of attaching a targeting group comprising GalNAc to the 3' end of the sense chain includes using a solid support (CPG) comprising GLO-n. In some studies, the method of attaching a targeting group comprising GalNAc to the 3' end of the sense chain includes attaching the GalNAc targeting group to the CPG solid support via an ester bond, and using the resulting CPG to which the GalNAc targeting group is attached when synthesizing the sense chain, thereby obtaining the GalNAc targeting group attached to the 3' end of the sense chain.

The imann residue can be added to the 5' or 3' end of the oligonucleotide chain by methods well known to those skilled in the art, such as the inversion debasing residue (invab) method, and/or further added to the targeting group targeting GalNAc.

Example 4. In vitro screening of CFB siRNA duplexes

Huh7 cells were trypsinized and adjusted to the appropriate density, mixed with a complex of psiCHECK TM -2 vector plasmid and Lipofectamine 2000 (Invitrogen-11668-019), and plated into 96-well plates. At the same time as the plate, cells were transfected with test siRNA or control siRNA using Lipofectamine RNAiMax (Invitrogen-13778-150) according to the manufacturer's recommendations. siRNAs were tested in triplicate at two concentrations (1 nM and 10 nM).

Day 1, psiCHECK(TM)-2 vector transfection (one plate)

(1) Transfer 2.5 μg of psiCHECK(TM)-2 vector plasmid to an RNASE-free Eppendorf tube (Solution Mix #1)

(2) Add trypsin to a flask to dissociate Huh7 cells, count the cells using a Vi-Cell counter, and adjust the cell density to 1*10^5/ml.

(3) Transfer 7.5 µL of Lipofectamine 2000 (Invitrogen-11668-019) to solution mix #1 tube and mix well.

(4) Add the solution from step 3 to the cell suspension, mix well, and dispense the suspension into a 96-well plate (100 μl/well).

Day 2, siRNA transfection

(1) Dilute Lipofectamine® RNAiMAX reagent in Opti-MEM® medium.

(2) Dilute siRNA with RNA-free water to prepare a 12× stock solution.

(3) Mix equal volumes of diluted RNAiMax and siRNA and incubate at room temperature for 15 minutes to allow complex formation.

(4) Add 45 μl/well of the compound Lipofectamine® RNAiMAX (Opti-MEM) to 225 μl/well of fresh DMEM medium. Discard the supernatant in the test plate and add 120 μl/well of the compound mixture to a 96-well plate.

(5) No compound control wells were defined as cells transfected with psiCHECK™-2 vector and not treated with siRNA; blank controls were wells containing cells alone.

Day 3, Dual-Glo® Luciferase Assay

(1) Add reagents to the assay plate and wait 10 minutes to allow cell lysis to occur.

(2) Transfer 100 μl of cell lysate to a plate and measure firefly luminescence.

(3) Add 50 μl of Dual-Glo® Stop & Glo® Reagent to the assay plate and mix, wait 10 minutes, and then measure luminescence.

(4) Calculate relative expressions

Data Analysis

Sample well ratio = (nephron luminescence sample - background blank) / (firefly luminescence sample - background blank)

No compound control well ratio = (control nephron luminescence - background blank) / (control sample firefly luminescence - background blank)

Inhibition rate = 100-(sample well ratio/average ratio of no compound control) × 100%

The duplex sequences used correspond to those shown in Tables 5-7.

Table 5 provides the results of in vitro studies using various CFB RNAi agents to inhibit CFB expression. The duplex sequences used correspond to those shown in Table 2. Double chain AV# Average inhibition rate % (mean ± SD) 10nM 1nM AV02358 78.85±1.28 73.22±1.51 AV02359 51.86±6.47 33.86±6.33 AV02360 11.90±8.86 29.27±2.25 AV02361 20.43±8.24 18.07±2.59 AV02362 -3.31±13.26 -8.04±12.73 AV02363 12.50±5.21 7.41±7.05 AV02364 19.68±6.72 21.48±2.24 AV02365 16.14±1.38 21.47±13.95 AV02366 -5.31±2.10 -6.22±14.50 AV02367 38.35±7.42 32.70±9.53 AV02368 -10.06±14.23 6.88±3.55 AV02369 29.11±3.89 15.01±10.92 AV02370 44.96±1.52 25.15±14.65 AV02371 50.81±2.65 30.69±11.27 AV02373 62.58±1.82 49.37±1.89 AV02374 64.46±5.00 52.27±4.09 AV02375 65.09±3.06 51.65±3.69 AV02376 53.34±4.13 42.77±6.35 AV02378 56.94±7.39 32.39±7.96 AV02380 50.70±2.96 48.58±2.06 AV02381 46.90±2.81 43.81±11.94 AV02382 31.79±7.71 40.16±4.66 AV02383 20.41±1.37 20.90±5.56 AV02384 4.54±3.18 0.78±7.34 AV02385 21.05±7.43 11.25±6.36 AV02388 47.96±8.18 43.77±6.61 AV02390 2.47±2.81 -0.01±7.14 AV02391 15.85±14.28 6.04±10.57 AV02392 34.20±5.92 6.28±6.65 AV02393 22.20±8.49 5.88±3.11 AV02394 16.69±6.94 1.85±7.89 AV02396 23.26±4.61 30.83±12.27 AV02397 -16.14±2.13 -5.65±13.35 AV02398 10.04±2.64 16.44±15.84 AV02399 -7.96±14.32 -11.29±13.60 AV02400 -20.34±5.33 -2.13±7.40 AV02401 21.96±14.53 15.17±2.95 AV02402 26.15±5.27 11.66±15.49 AV02404 -6.14±0.74 3.00±6.73 AV02405 0.39±15.98 -0.40±6.66 AV02406 3.56±15.28 -4.26±9.93 AV02407 3.44±6.90 2.47±10.09 AV02408 12.58±7.37 20.65±12.20 AV02410 46.35±2.68 37.79±5.28 AV02379 50.65±4.18 41.39±4.60 AV02387 5.52±4.86 0.84±3.74 AV02395 -4.59±9.65 -0.53±9.08 AV02403 15.89±2.92 5.47±8.25 AV02411 35.83±4.25 32.71±2.81 AV02022 34.15±8.13 39.43±9.78

Table 6 provides the results of in vitro studies using various CFB RNAi agents to inhibit CFB expression. The duplex sequences used correspond to those shown in Table 2. Double chain AV# Average inhibition rate % (mean ± SD) 10nM 1nM AV02414 58.44±6.34 14.01±2.05 AV02415 24.48±14.25 5.58±1.63 AV02416 31.97±13.33 7.50±4.28 AV02417 25.95±9.30 16.15±9.01 AV02418 42.97±5.34 9.02±5.13 AV02420 35.00±8.58 8.87±4.28 AV02421 15.80±4.08 16.38±9.91 AV02422 22.05±1.77 22.55±10.49 AV02423 18.98±2.72 7.46±6.19 AV02424 10.79±3.59 7.34±14.42 AV02425 9.79±3.77 3.03±6.94 AV02426 26.73±1.61 9.47±12.32 AV02428 15.76±15.82 4.42±5.52 AV02429 25.81±7.54 2.14±6.60 AV02430 7.80±16.50 -0.10±6.59 AV02431 30.97±12.02 3.23±7.65 AV02432 18.85±12.49 1.60±4.93 AV02433 -0.87±18.80 4.83±7.06 AV02434 6.22±9.60 1.87±3.28 AV02436 47.14±6.01 28.66±6.40 AV02437 -3.38±12.51 2.88±8.50 AV02438 0.23±3.45 8.98±6.91 AV02439 -5.33±9.10 2.83±5.65 AV02440 25.25±5.37 12.04±12.79 AV02441 32.99±2.90 10.13±0.89 AV02442 44.07±4.87 21.94±5.53 AV02444 26.93±3.63 4.89±7.76 AV02445 7.21±7.63 3.21±2.44 AV02446 25.60±17.35 8.63±6.73 AV02447 32.67±13.70 13.49±17.88 AV02448 30.82±1.72 13.33±3.20 AV02449 26.82±15.54 2.02±2.10 AV02450 27.15±3.99 -2.70±9.88 AV02452 29.20±3.47 9.78±0.82 AV02453 14.98±11.54 15.17±9.57 AV02454 -4.59±6.95 1.56±8.37 AV02455 -8.24±2.81 -1.08±12.61 AV02456 -12.98±2.00 9.01±2.01 AV02457 1.59±10.72 0.21±5.39 AV02458 16.99±6.20 9.26±5.64 AV02460 18.68±13.35 -8.62±5.70 AV02461 33.01±12.98 6.27±1.79 AV02462 29.50±8.75 7.84±7.42 AV02463 46.17±10.32 12.26±1.92 AV02464 43.54±9.24 8.36±3.80 AV02465 47.17±3.43 4.34±6.00 AV02466 34.51±2.99 3.45±1.12 AV02468 48.39±1.62 25.37±9.88 AV02469 29.41±12.44 2.35±4.72 AV02470 29.53±5.19 25.58±7.58 AV02471 19.66±7.52 11.06±4.43 AV02472 8.02±20.67 13.45±2.92 AV02473 28.00±4.28 -5.08±14.15 AV02474 40.96±13.32 24.15±6.94 AV02476 -11.56±9.62 -3.91±11.40 AV02477 -3.22±10.05 -14.83±5.24 AV02478 11.53±4.95 -2.01±1.80 AV02479 -15.10±6.54 -0.21±7.74 AV02480 20.71±6.13 -1.76±5.20 AV02481 8.20±23.97 -1.31±0.29 AV02482 2.55±11.71 -1.45±2.20 AV02484 6.34±8.71 -5.34±2.44 AV02485 4.41±5.56 -12.45±7.66 AV02486 5.83±6.63 -6.34±3.51 AV02487 6.67±0.27 8.30±4.18 AV02488 2.25±4.72 -1.40±10.49 AV02489 26.38±5.66 1.24±2.50 AV02490 14.37±6.62 4.02±6.19 AV02492 24.87±2.37 11.22±7.54 AV02493 20.77±6.89 9.40±9.19 AV02494 5.73±15.89 9.71±6.95 AV02495 39.54±12.34 13.61±4.16 AV02496 18.72±12.96 14.91±8.47 AV02497 48.94±9.50 14.97±6.14 AV02498 20.15±8.36 5.40±7.22 AV02500 54.19±3.46 44.17±4.37 AV02501 37.54±4.02 26.37±3.64 AV02502 17.66±3.26 15.28±9.48 AV02503 51.59±4.08 33.57±5.52 AV02504 43.22±7.64 34.42±4.85 AV02505 35.67±4.26 21.44±6.64 AV02506 46.52±6.10 24.00±6.32 AV02419 41.18±5.85 21.14±3.65 AV02427 14.12±11.78 11.87±11.61 AV02435 16.82±3.98 7.62±12.34 AV02443 38.13±6.10 12.04±3.81 AV02451 32.60±5.25 11.45±3.55 AV02459 15.67±3.76 0.46±9.10 AV02467 43.70±7.28 11.65±8.20 AV02475 45.37±3.89 22.95±2.67 AV02483 43.19±2.67 26.00±2.35 AV02491 23.95±7.58 17.88±2.71 AV02499 32.18±8.17 19.84±9.53 AV02507 19.18±4.11 10.87±14.36 AV02358 73.38±2.07 72.16±4.50 AV02022 49.01±3.42 37.56±4.02

Table 7 provides the results of in vitro studies using various CFB RNAi agents to inhibit CFB expression. The duplex sequences used correspond to those shown in Table 2. Double chain AV# Average inhibition rate % (mean ± SD) 10nM 1nM AV02508 46.98±8.60 8.98±6.03 AV02509 57.30±10.29 27.29±1.78 AV02510 38.50±7.87 -0.29±2.61 AV02511 16.24±9.63 2.13±11.26 AV02512 38.93±3.64 3.56±7.82 AV02513 16.25±15.01 0.98±13.36 AV02514 46.23±4.52 6.42±6.00 AV02516 22.75±2.01 6.74±17.97 AV02517 -2.74±6.25 -7.02±8.09 AV02518 17.99±5.90 5.81±6.00 AV02519 22.75±12.89 4.97±11.96 AV02520 23.14±9.43 9.77±2.96 AV02521 53.81±4.58 29.86±5.76 AV02522 38.35±3.64 23.49±7.20 AV02524 2.17±10.61 -18.13±12.84 AV02525 -19.35±6.92 -25.56±10.14 AV02526 26.75±6.81 -0.94±4.85 AV02527 8.51±4.55 3.92±14.26 AV02528 28.39±11.57 13.18±1.05 AV02529 33.57±8.04 6.27±2.69 AV02530 2.70±7.71 -14.42±0.47 AV02532 23.49±8.92 -0.92±7.87 AV02533 30.79±0.98 5.18±1.68 AV02534 1.31±8.22 -16.75±2.25 AV02535 18.49±1.44 -11.67±1.48 AV02536 -14.95±14.38 -7.92±9.61 AV02537 28.50±3.60 3.72±3.00 AV02538 49.86±2.64 18.95±1.78 AV02540 -4.42±5.00 -7.62±1.91 AV02541 23.28±2.66 8.74±14.79 AV02542 27.10±2.18 9.35±6.56 AV02543 18.41±3.50 0.81±8.73 AV02544 3.25±6.19 -3.33±16.57 AV02545 32.22±6.44 3.30±10.09 AV02546 11.77±20.59 -4.39±2.08 AV02548 47.07±7.74 12.07±7.68 AV02549 49.44±3.19 18.76±1.09 AV02550 34.08±9.58 1.41±8.47 AV02551 60.99±4.13 47.11±3.68 AV02552 60.91±1.58 39.11±1.69 AV02553 50.40±2.41 24.89±9.46 AV02554 69.01±2.65 52.14±2.99 AV02556 59.17±3.92 27.20±11.71 AV02557 40.99±2.16 4.45±12.04 AV02558 58.47±4.89 18.92±7.16 AV02559 53.78±6.64 30.36±6.98 AV02560 37.82±9.83 1.54±9.12 AV02561 53.81±11.29 19.03±5.71 AV02562 31.89±8.65 8.72±4.33 AV02564 44.98±2.91 12.81±7.17 AV02565 -14.90±13.54 -36.07±2.75 AV02566 33.28±4.24 2.73±16.55 AV02567 49.58±2.60 23.31±8.41 AV02568 10.54±5.02 -11.41±8.68 AV02569 19.85±8.51 6.18±8.60 AV02570 26.19±8.99 11.37±3.77 AV02572 58.10±7.29 18.12±3.45 AV02573 16.83±8.09 0.37±1.87 AV02574 10.18±13.07 -5.82±5.91 AV02575 29.11±15.17 11.90±10.29 AV02576 34.98±3.90 1.31±6.38 AV02577 54.14±3.03 27.99±9.92 AV02578 9.53±10.81 5.25±3.20 AV02580 34.32±7.66 -2.41±7.80 AV02581 55.03±4.37 47.35±4.41 AV02582 47.06±2.46 18.61±6.79 AV02583 46.58±10.17 31.60±8.65 AV02584 55.46±8.50 35.69±3.75 AV02585 67.49±0.42 44.66±4.41 AV02586 62.75±1.95 24.02±5.23 AV02515 55.61±8.28 19.03±8.63 AV02523 51.73±3.70 24.01±7.70 AV02531 26.78±5.03 11.44±4.75 AV02539 40.98±7.01 5.43±13.15 AV02547 44.10±9.36 6.12±8.55 AV02555 58.38±2.32 38.30±5.54 AV02563 61.45±1.58 33.60±4.01 AV02571 39.49±5.19 17.90±0.82 AV02579 23.50±4.98 8.69±4.47 AV02587 61.53±3.23 37.46±4.95 AV02588 38.50±3.72 29.29±3.15 AV02358 80.02±0.46 65.46±4.78 AV02022 49.26±4.33 29.20±7.17

Example 5. In vivo testing of CFB siRNA duplex

On day 1, female C57BL/6J mice (4 per group) were infected by intravenous injection of adeno-associated virus 8 (AAV8) vector solution encoding human CFB and luciferase genes. On day 8, mice were subcutaneously administered a single dose of 2 or 3 mg/kg of CFB siRNA or PBS. Blood samples were collected on day 8, before siRNA administration, day 15, day 22, and day 29. Plasma samples were separated and the luciferase activity of plasma samples was measured according to the manufacturer's recommended protocol. Since the expression of human CFB levels correlates with the expression level of luciferase, the percentage of remaining CFB was calculated by comparing the luciferase activity before and after treatment in the siRNA-treated samples, and normalized by the change in luciferase activity in the control-treated samples during the same time period. The results are summarized in Tables 8 and 9.

Table 8. Screening of CFB siRNA in AAV-CFB transduced mice with a single 3 mpk subcutaneous dose. The percent reduction of human CFB in mouse serum was normalized to CFB expression before siRNA administration and to the PBS control group. Dual Chain AD# The remaining percentage of D15 relative to D8 (mean ± SD) The remaining percentage of D22 relative to D8 (mean ± SD) Dosage (mg/kg) AD01093 0.18±0.06 0.03±0.01 3 AD01094 0.45±0.25 0.08±0.02 3 AD01095 0.9±0.22 0.24±0.09 3 AD01096 0.36±0.04 0.1±0.01 3 AD01097 0.59±0.07 0.2±0.06 3 AD01098 0.36±0.05 0.11±0.02 3 AD01099 0.6±0.09 0.23±0.04 3 AD01100 0.95±0.11 0.35±0.02 3 AD01101 0.32±0.06 0.12±0.02 3 AD01102 0.6±0.1 0.14±0.04 3 AD01103 1.14±0.13 0.45±0.15 3 AD01104 0.36±0.17 0.26±0.05 3

Table 9. Screening of CFB siRNA with a single 2 mpk subcutaneous dose in AAV-CFB transduced mice. The percent reduction of human CFB in mouse serum was normalized to CFB expression before siRNA administration and to the PBS control group. Dual Chain AD# Percentage of D15 remaining relative to D8 (mean ± SD) The remaining percentage of D22 relative to D8 (mean ± SD) The remaining percentage of D29 relative to D8 (mean ± SD) Dosage (mg/kg) AD01080 0.59±0.47 0.23±0.03 0.23±0.04 2 AD01093 0.28±0.08 0.23±0.1 0.19±0.05 2 AD01094 0.32±0.06 0.32±0.1 0.3±0.1 2 AD01096 0.43±0.03 0.33±0.02 0.37±0.05 2 AD01389 1.07±0.09 1.05±0.03 1.02±0.03 2 AD01390 1.1±0.16 1.04±0.08 0.3±0.1 2 AD01392 0.47±0.12 0.36±0.13 0.38±0.1 2 AD01393 0.37±0.08 0.23±0.05 0.28±0.06 2 AD01394 0.58±0.13 0.56±0.06 0.51±0.08 2 AD01395 0.49±0.08 0.35±0.06 0.33±0.02 2 AD01396 0.33±0.18 0.24±0.14 0.22±0.14 2 AD01397 0.41±0.08 0.29±0.14 0.25±0.06 2 AD01398 0.47±0.03 0.28±0.1 0.31±0.07 2 AD01399 0.3±0.05 0.16±0.03 0.19±0.02 2 AD01401 0.47±0.14 0.43±0.05 0.38±0.07 2 AD01402 0.99±0.15 1.01±0.09 0.78±0.06 2 AD01403 0.59±0.11 0.65±0.12 0.53±0.12 2 AD01404 0.98±0.28 1.11±0.32 1.01±0.26 2 AD01405 1.13±0.07 1.27±0.03 1±0.07 2 AD01406 0.57±0.22 0.63±0.15 0.55±0.05 2 AD01407 0.55±0.08 0.44±0.07 0.42±0.05 2 AD01408 0.61±0.17 0.72±0.22 0.48±0.11 2 AD01409 0.58±0.23 0.53±0.13 0.47±0.12 2 AD01410 0.48±0.31 0.77±0.24 0.6±0.17 2 AD01411 0.8±0.1 0.77±0.06 0.63±0.09 2 AD01412 0.33±0.07 0.32±0.13 0.23±0.06 2 AD01413 1.43±0.48 1.44±0.28 1.2±0.32 2 AD01414 0.79±0.12 0.79±0.02 0.74±0.08 2 AD01415 0.43±0.07 0.3±0.01 0.34±0.02 2 AD01416 0.65±0.09 0.54±0.17 0.45±0.14 2 AD01417 0.68±0.15 0.5±0.05 0.45±0.02 2 AD01418 0.95±0.09 0.8±0.08 0.82±0.09 2 AD01419 1.06±0.13 0.92±0.03 0.9±0.08 2 AD01420 0.17±0.08 0.12±0.04 0.1±0.02 2 AD01421 0.27±0.06 0.23±0.05 0.2±0.03 2 AD01422 0.96±0.19 0.64±0.08 0.58±0.1 2 AD01423 0.64±0.09 0.42±0.06 0.35±0.07 2

Example 6. In vivo testing of CFB siRNA duplexes

On day 1, female C57BL/6J mice (4 per group) were infected by intravenous injection of adeno-associated virus 8 (AAV8) vector solution encoding human CFB and luciferase genes. On day 8, mice were injected subcutaneously with a single 2 mg/kg CFB siRNA dose or PBS. Blood samples were collected on day 8, before siRNA administration, day 18, day 25, and day 32. Plasma samples were separated and serum samples were collected to measure quantitative protein levels by ELISA according to the manufacturer's recommended protocol. Since the expression of human CFB levels correlates with the expression level of luciferase, the percentage of remaining CFB was calculated by comparing the samples before and after treatment in the siRNA treatment group and normalized by the ELISA changes in the samples of the control treatment group during the same time period. The results are summarized in Table 10.

Table 10. Screening of CFB siRNA in AAV-CFB transduced mice with a single 2 mpk subcutaneous dose. The percent reduction of human CFB in mouse serum was normalized to CFB expression before siRNA administration and to the PBS control group. Dual chain AD# Remaining relative to D8 (mean ± SD) Dosage mg/kg D18 D25 D32 AD01080 0.14±0.04 0.15±0.07 0.21±0.12 2 AD01093 0.12±0 0.11±0 0.11±0.01 2 AD01094 0.15±0.01 0.14±0.02 0.2±0.02 2 AD01096 0.18±0.09 0.16±0.02 0.17±0.03 2 AD01389 0.12±0.03 0.12±0.02 0.12±0.03 2 AD01390 0.2±0.05 0.21±0.07 0.24±0.09 2 AD01391 0.1±0.02 0.1±0.04 0.09±0.03 2 AD01392 0.29±0.04 0.27±0.05 0.29±0.05 2 AD01393 0.42±0.1 0.37±0.02 0.39±0.04 2 AD01394 0.82±0.1 0.8±0.16 0.94±0.12 2 AD01395 0.64±0.06 0.74±0.17 0.8±0.15 2 AD01396 0.24±0.06 0.39±0.1 0.31±0.07 2 AD01397 0.43±0.08 0.88±0.16 0.69±0.07 2 AD01398 0.48±0.07 0.69±0.11 0.56±0.1 2 AD01399 0.24±0.04 0.28±0.05 0.23±0.06 2 AD01400 0.1±0.03 0.12±0.03 0.13±0.02 2 AD01401 0.55±0.07 0.81±0.08 0.73±0.05 2 AD01402 0.48±0.02 0.76±0.04 0.62±0.1 2 AD01403 0.18±0.03 0.29±0.06 0.29±0.06 2 AD01404 0.16±0.03 0.26±0.03 0.24±0.12 2 AD01405 0.46±0.04 0.87±0.06 0.69±0.08 2 AD01406 0.31±0.02 0.47±0.05 0.36±0.02 2 AD01407 0.41±0.2 0.47±0.15 0.42±0.18 2 AD01408 0.27±0.06 0.41±0.09 0.32±0.08 2 AD01409 0.4±0.06 0.52±0.02 0.43±0.03 2 AD01410 0.51±0.1 0.64±0.17 0.59±0.11 2 AD01411 0.34±0.05 0.44±0.08 0.39±0.1 2 AD01412 0.32±0.17 0.63±0.08 0.72±0.17 2 AD01413 0.19±0.04 0.19±0.04 0.23±0.04 2 AD01414 0.65±0.09 0.67±0.08 0.59±0.13 2 AD01415 0.25±0.05 0.26±0.07 0.36±0.05 2 AD01416 0.38±0.05 0.5±0.04 0.68±0.13 2 AD01417 0.56±0.07 0.84±0.09 0.9±0.07 2 AD01418 0.58±0.07 0.75±0.04 0.75±0.11 2 AD01419 0.34±0.03 0.41±0.03 0.39±0.03 2 AD01420 0.52±0.04 0.88±0.1 0.89±0.26 2 AD01421 0.46±0.07 0.65±0.1 0.67±0.06 2 AD01422 0.66±0.18 0.9±0.21 0.8±0.3 2 AD01423 0.38±0.01 0.49±0.07 0.43±0.09 2

Blood samples were collected on day 8, before siRNA administration, day 15, day 22, and day 29. The results are summarized in Tables 11 and 12.

Table 11. Screening of CFB siRNAs in AAV-CFB-transduced mice with a single 2 mpk subcutaneous dose. The percent reduction of human CFB in mouse serum was normalized to CFB expression before siRNA administration and to the PBS control group. Dual chain AD# CFB remaining compared with day 8 (mean ± SEM) Dosage (mg/kg) Day 15 Day 22 Day 29 AD01399-1 0.12±0.01 0.08±0.02 0.11±0.02 2 AD01704-1 0.12±0.04 0.07±0.02 0.1±0.03 2 AD01399 0.11±0.01 0.06±0.01 0.09±0.01 2 AD01704 0.12±0.01 0.1±0.03 0.13±0.01 2 AD02226 0.33±0.04 0.26±0.04 0.39±0.02 2 AD02227 0.19±0.02 0.09±0.01 0.16±0.04 2 AD02228 0.29±0.04 0.18±0.03 0.33±0.07 2 AD02229 0.6±0.07 0.41±0.02 0.4±0.01 2 AD02230 1.38±0.38 0.89±0.23 0.95±0.27 2 AD02231 0.71±0.06 0.53±0.03 0.66±0.1 2 AD02232 0.48±0.03 0.37±0.06 0.43±0.05 2 AD02233 0.5±0.08 0.35±0.04 0.38±0.04 2 AD02234 0.32±0.07 0.28±0.05 0.4±0.05 2 AD02235 0.51±0.02 0.3±0.03 0.48±0.06 2 AD02236 0.45±0.01 0.24±0.03 0.37±0.05 2 AD02237 0.27±0.03 0.14±0.01 0.25±0.03 2 AD02238 0.54±0.07 0.38±0.07 0.46±0.07 2 AD02239 0.33±0.04 0.18±0.02 0.22±0.03 2 AD02240 0.42±0.05 0.21±0.02 0.29±0.02 2 AD02241 0.34±0.03 0.25±0.03 0.37±0.06 2

Table 12. Screening of CFB siRNA in AAV-CFB transduced mice with a single 2 mpk subcutaneous dose. The percent reduction of human CFB in mouse plasma was normalized to CFB expression before siRNA administration and to the PBS control group. Dual chain AD# CFB remaining compared with day 8 (mean ± SEM) Dosage (mg/kg) Day 15 Day 22 Day 29 AD02342 0.26±0.02 0.28±0.01 0.34±0.02 2 AD02343 0.17±0.04 0.28±0.06 0.36±0.07 2 AD02344 0.36±0.08 0.47±0.13 0.63±0.21 2 AD02345 0.24±0.04 0.26±0.03 0.34±0.03 2 AD02346 0.29±0.02 0.59±0.1 0.57±0.03 2 AD02347 0.6±0.07 1.16±0.18 0.84±0.08 2

Example 7. In vivo testing of CFB siRNA duplex in NHP PD model

This study recruited 6-year-old cynomolgus monkeys weighing 3-6 kg, 3 per group. Each monkey was subcutaneously injected with a single dose of 6 mg/kg CFB siRNA reagent or PBS on day 1 (before siRNA administration). After fasting overnight, serum was drawn on day -14 (before administration), day -7 (before administration), day 1 (before administration), day 8, day 15, day 22, day 29, day 35, day 43, day 50, day 57, day 64, and day 71; the remaining CFB protein was determined by Western blotting, and the results are shown in Table 13.

Table 13. Results of CFB protein concentration in serum measured by Western blotting. Dual Chain AD# Compared with the CFB remaining before drug administration (%) (mean ± SD) Dosage (mg/kg) Day 8 Day 15 Day 22 Day 29 Day 36 Day 43 Day 50 Day 57 Day 64 Day 71 AD01093 56.05±16.45 24.62±12.26 15.02±5.22 13.34±5.46 14.81±7.95 15.26±12.1 14.02±10.21 12.15±7.97 11.69±8.11 11.28±5 6 AD01393 79.98±11.75 38.02±8.03 26.05±12.13 20.79±8.31 27.18±3.61 21.66±7.47 20.06±2.14 28.08±11.57 21.3±1.76 38.4±12.61 6 AD01396 70.89±3.58 37.85±3.86 24.03±3.81 24.08±7.18 22.48±4.68 23.5±6.32 25.51±10.06 25.53±11.17 34.92±14.46 46.54±21.87 6 AD01399 60.47±16.8 29.19±13.91 18.8±8.44 16.75±8.76 18.28±8.53 20.84±11.75 20.98±14.01 24.26±17.94 18.87±10.36 25.1±12.43 6 AD01412 103.1±0.08 88.6±2.31 93.69±8.03 77.76±7.64 89.57±19.04 91.42±13.97 89.76±2.32 100.33±8.62 89.65±3.93 103.81±9.64 6 AD01420 90.2±6.43 67.39±8.71 54.22±10.56 51.32±16.92 56.43±18.12 53.71±32.13 62.08±25.97 66.24±33.92 64.32±40.48 72.47±29.78 6 AD01420-1 85.11±6.75 43.23±10.92 40.17±14.85 32.15±16.59 53.15±29.9 37.77±18.29 34.21±27.24 38.56±27.52 37.96±32.42 58.27±42.91 6 AD01787 120.61±11.32 102.26±9.66 113.15±8.08 115.51±6.64 122.69±11.97 120.12±10.89 110.84±20.59 123.55±33.75 112.63±31.32 129.61±21.11 6

Example 8. In vivo testing of CFB siRNA duplex in NHP PD model

This study recruited cynomolgus monkeys (6 years old, weighing 3-6 kg, 3 per group). Each monkey was subcutaneously injected with a single 6 mg/kg CFB siRNA reagent or PBS on day 1 (before siRNA administration). After fasting overnight, serum was drawn on day -14 (before administration), day -7 (before administration), and day 1 (before administration); all groups collected samples on day -14, day -7, day 1, day 8, day 15, day 22, day 29, day 35, day 43, day 50, day 57, day 71, and day 85; Western blot (WB) was used to detect CFB protein residues, and WISLAB kits were used to detect Alternative (C5b-9) and Classical (C5b-9). The results are shown in Tables 14-16.

Table 14. Results of Western blot (WB) method to measure the CFB protein concentration in serum (normalized by Transferrin) Dual Chain AD# Compared with before drug administration, CFB remaining (%) (mean ± SEM) Dosage (mg/kg) Day 8 Day 15 Day 22 Day 29 Day 36 Day 43 Day 50 Day 57 Day 71 Day 85 AD01093-1 41.63±6.09 23.21±2.33 19.73±3.44 20.95±6.08 21.95±5.16 30.21±10.5 35.19±12.56 37.53±13.79 38.88±15.13 49.86±12.3 6 AD01393-1 53.86±7.46 24.92±6.93 18.78±6.46 17.3±7.68 19.07±8.76 21.24±9.92 27.04±13.88 28.46±10.81 41.68±16.3 51.25±15.53 6 AD01399-1 39.05±8.51 11.51±2.98 6.95±1.89 7.32±2.53 8.42±3.65 12.82±6.26 14.96±7.96 20.59±12.61 28.91±17.37 47.7±26.68 6 AD02225 51.86±1.67 35.12±3.35 24.95±4.12 26.14±6.25 27.53±6.11 30.08±6.8 36.85±4.94 39.94±7.18 46.59±7.11 49.64±2.89 6 AD01704-1 30.57±1.7 8.09±1.44 4.31±0.85 3.82±0.99 3.83±1.22 4.41±1.83 4.88±2.36 6.02±1.96 9.8±3.61 14.01±4.14 6

Table 15. Results of Alternative (C5b-9) assay in serum using WISLAB kit Dual Chain AD# Compared with before administration, the remaining complement activity (C5b-9) (%) (Mean ± SEM) Dosage (mg/kg) Day 8 Day 15 Day 22 Day 29 Day 36 Day 43 Day 50 Day 57 Day 71 Day 85 AD01093-1 6.65±2.36 1±0.81 0.32±0.33 0.42±0.18 0.75±0.19 5.91±5.77 4.42±4.25 3.3±2.57 18.37±14.68 33.39±23.87 6 AD01393-1 28±12.34 2.42±1.41 1±0.46 1.81±0.82 1.31±0.09 3.19±2.37 7.67±6.4 8.8±6.07 25.47±5.94 40.98±12.51 6 AD01399-1 12.9±5.13 0.24±0.15 0.66±0.27 0.38±0.04 0.34±0.54 0.74±0.92 1.47±1.62 5.65±5.61 20.42±15.96 27.92±18.46 6 AD02225 27±0.59 9.86±2.8 1.98±0.6 5.46±2.57 5.5±2.55 9.26±3.65 15.37±5.65 30.3±10.24 53.52±13.49 66.4±5.75 6 AD01704-1 9.18±5.2 0.18±0.6 -0.3±0.23 -0.17±0.2 -0.07±0.29 -0.2±0.15 0.03±0.53 0.16±0.52 1.44±1.24 8.29±7.81 6

Table 16. Results of determination of Classical (C5b-9) in serum using WISLAB kit Dual Chain AD# Compared with before administration, the remaining complement activity (C5b-9) (%) (Mean ± SEM) Dosage (mg/kg) Day 8 Day 15 Day 22 Day 29 Day 36 Day 43 Day 50 Day 57 Day 71 Day 85 AD01093-1 110.72±10.3 115.29±8.17 113.23±18.81 125.72±18.3 125.9±14.11 128.3±21.07 116.67±14.73 124.2±11.55 137.23±4.11 144.27±15.28 6 AD01393-1 100.03±0.78 103.27±1.25 96.03±4.63 102.31±1 98.01±4.94 100.87±5.67 95.91±2.97 98.78±3.21 115.89±2.73 120.72±3.85 6 AD01399-1 103.63±3.87 103.67±7.35 92.67±3.09 101.4±9.33 103.06±6.33 106.33±7.53 101.36±4.2 105.28±5.72 121.06±18.88 119.17±19.36 6 AD02225 96.19±3.56 95.5±6.77 96.3±11.44 92.88±5.05 96.02±4.5 98.23±7.62 94.13±5.68 104.92±7.06 137.13±3.9 133.59±9.45 6 AD01704-1 90.29±3.65 93.52±3.66 92.64±6.06 93.4±3.97 96.05±3.48 90.78±4.28 93.45±4.39 94.17±2.75 117.72±2.93 116.42±1.9 6

Example 9. In vivo testing of CFB siRNA duplex in NHP PD model

This study recruited cynomolgus monkeys (6 years old, weighing 3-6 kg, 3 per group). After fasting overnight, serum was drawn from each monkey on day -14 (before drug administration), day -7 (before drug administration), and day 1 (before drug administration). On day 1 (before siRNA administration), a single subcutaneous injection of 3 mg/kg CFB siRNA or PBS was given. The sample collection time for each group was: day -14, day -7, day 1, day 8, day 15, day 22, day 29, day 35, day 43, day 50, day 57; Western blot (WB) was used to detect the residual CFB protein. The results are shown in Table 17.

Table 17. Results of Western blot (WB) analysis of CFB protein concentration in serum. Dual chain AD# Compared with before drug administration, CFB remaining (%) (mean ± SEM) Day 8 Day 15 Day 22 Day 29 Day 36 Day 43 Day 50 Day 57 AD01093-1 73.17±5.21 46.94±1.52 41.11±4.76 38.28±5.56 42.43±6.64 45±6.63 44.08±9.05 49.95±11.73 AD01093-2 43.57±4.58 16.8±5.21 8.55±2.83 7.93±3.67 8.21±2.87 11.47±3.63 12.84±3.68 15.42±2.62

Example 10. In vivo screening of CFB siRNA duplexes.

Huh7 cells were digested with trypsin, adjusted to an appropriate density, and seeded in 96-well plates. On the second day of seeding, cells were transfected with psiCHECK(TM)-2 vector plasmid, blank vector pCNDNA3.0, siRNAs, or control siRNA complexes using Lipofectamine2000 (Invitrogen-11668-019) according to the manufacturer's recommended protocol. siRNAs were tested in triplicate at different concentrations (10 nM and 1 nM).

On day 1, trypsin was added to dissociate the Huh7 cells in the culture flask, and the cells were counted using a Vi-Cell counter. The cell density was adjusted to 1*10^5/ml and cultured in DMEM medium.

On the second day, psiCHECK(TM)-2 vector/blank vector pCDNA3.0/siRNAs/Lipofectamine 2000 were mixed for transfection.

(1) Mix the appropriate amount of Lipofectamine 2000 (Invitrogen-11668-019) with Opti-MEM® medium (Solution Mix #1). Finally, add 0.3 μl of Lipofectamine 2000 and 4.7 μl of Opti-MEM® medium to each well.

(2) Mix the appropriate psiCHECK(TM)-2 vector, blank pCNDNA 3.0 vector, and siRNAs with Opti-MEM® medium (Solution Mixture #2).

(3) Mix equal volumes of solution mixture #1 and mixture #2 (mixture #3), with a final volume of 10 μl per well. Incubate the mixture at room temperature for 15 min to allow complex formation.

(4) Remove the DMEM medium and add 10 μl of the mixed solution #3 and 90 μl of fresh DMEM medium.

(5) The no-compound control wells were defined as cells transfected with psiCHECK(TM)-2 vector and blank vector pCNDNA 3.0 and not treated with siRNA; the blank control wells were defined as wells with cells only.

Day 3, Dual-Glo® Luciferase Assay

(1) Add the reagent to the assay plate and wait 10 minutes to allow the cells to lyse.

(2) Transfer 100 μl of cell lysate to the plate and measure firefly luminescence.

(3) Add 50 μl of Dual-Glo® Stop & Glo® reagent to the assay plate and mix, wait 10 minutes, and then measure Renilla luminescence.

(4) Calculate relative expression

Data Analysis

Sample hole ratio = (sample Renilla luminescence - background blank) / (sample Firefly luminescence - background blank)

No-compound control well ratio = (control sample Renilla luminescence - background blank) / (control sample Firefly luminescence - background blank)

Inhibition rate = 100-(sample well ratio/average ratio of no compound control) × 100%

Table 18. Provides the experimental results of in vitro studies using various CFB RNAi agents to inhibit CFB expression. The duplex sequences used correspond to those shown in Table 2. Double chain AV# Average inhibition rate % (mean ± SD) 10nM 1nM AV05135 51.57±4.83 51.60±2.54 AV05136 32.31±0.69 32.34±11.14 AV05137 71.85±2.09 52.74±2.46 AV05138 53.25±5.26 44.54±3.24 AV05139 16.69±6.03 10.55±5.25 AV05140 72.53±1.26 65.88±1.76 AV05141 3.49±9.89 -0.14±1.35 AV05142 -12.92±10.03 5.19±8.18 AV05143 41.13±6.14 26.59±2.19 AV05144 41.24±2.37 36.04±4.49 AV05145 43.64±1.59 41.60±2.02 AV05146 49.94±5.88 53.33±7.09 AV05147 8.21±6.47 -2.41±5.21 AV05148 15.01±6.01 26.41±2.81 AV05149 57.70±2.98 41.88±4.99 AV05150 53.07±2.20 38.87±5.17 AV05151 23.81±4.60 17.76±4.77 AV05152 71.35±2.10 63.63±3.64 AV05153 36.66±2.35 37.38±2.22 AV05154 55.53±6.50 66.12±2.35 AV05155 13.71±15.79 -1.37±6.90 AV05156 33.91±0.86 27.81±11.82 AV05157 52.10±0.61 39.46±5.46 AV05158 50.62±1.23 45.44±6.29 AV05159 1.54±3.62 1.10±2.90 AV05160 7.48±5.49 7.84±4.84 AV05161 26.98±3.37 20.49±2.34 AV05162 43.54±2.62 31.84±4.57 AV05163 47.02±4.48 45.82±3.45 AV05164 19.99±6.09 11.83±6.63 AV05165 33.67±6.78 34.77±1.92 AV05166 53.68±1.72 48.54±0.94 AV05167 19.24±4.53 11.36±5.80 AV05168 29.33±3.39 22.61±3.48 AV05169 22.14±6.96 22.66±7.16 AV05170 20.93±10.36 19.60±5.36 AV05171 30.90±2.25 31.23±1.74 AV05172 28.21±1.88 12.81±1.58 AV05173 27.28±9.37 23.23±5.22 AV05174 49.83±4.10 40.03±6.41 AV05175 74.61±0.55 73.75±2.06 AV05176 32.49±2.21 33.65±4.89 AV05177 71.81±3.71 72.37±2.55 AV05178 55.10±1.40 53.76±1.89

Equivalent

Although several embodiments of the present invention have been described and illustrated herein, a person of ordinary skill in the art will readily conceive of various other means and/or structures for performing the functions and/or obtaining the results and/or one or more advantages described herein, and each of these variations and/or modifications is considered to be within the scope of the present invention. More generally, a person of ordinary skill in the art will readily understand that all parameters, dimensions, materials, and configurations described herein are intended to be examples, and that actual parameters, dimensions, materials, and/or configurations will depend on the specific application in which the teachings of the present invention are used. A person of ordinary skill in the art will recognize, or be able to determine using only routine experimentation, many equivalents to the specific embodiments of the present invention described herein. Therefore, it should be understood that the foregoing embodiments are presented by way of example only and are within the scope of the appended patent applications and their equivalents; the present invention may be implemented in a manner different from that specifically described and claimed. The present invention is directed to each individual feature, system, article, material and/or method described herein. In addition, any combination of two or more such features, systems, articles, materials and/or methods, if such features, systems, articles, materials and/or methods are not mutually inconsistent, are included in the scope of the present invention.

All definitions defined and used herein should be understood to control over dictionary definitions, definitions in documents incorporated by reference, and/or ordinary meanings of the defined terms.

The indefinite articles "a" and "an" used in this specification and patent application should be understood to mean "at least one" unless expressly stated otherwise.

The phrase "and/or" used in the specification and claims should be understood to mean "either or both" of the combined elements, that is, the elements are present in combination in some cases and separately in other cases. In addition to the elements expressly identified in the "and/or" clause, other elements may optionally be present, whether related or unrelated to the elements expressly identified, unless expressly stated otherwise.

All references, patents, and patent applications and publications cited or referred to in this application are incorporated herein by reference in their entirety.

TW202500169A_113124145_SEQL.xml

Claims (77)

A double-stranded ribonucleic acid (dsRNA) agent for inhibiting CFB expression, wherein the dsRNA agent includes a sense chain and an antisense chain, wherein the sense chain comprises at least 15 consecutive nucleotides that differ from the nucleotide sequence of SEQ ID NO: 1 by no more than 3 nucleotides, and the antisense chain comprises at least 15 consecutive nucleotides that differ from the nucleotide sequence of SEQ ID NO: 2 by no more than 3 nucleotides, wherein the sense chain and the antisense chain may partially, substantially, or completely complement each other. The dsRNA agent of claim 1, wherein the dsRNA agent comprises a sense strand and an antisense strand, wherein the sense strand comprises at least 15, 16, 17, 18, 19, 20 or 21 consecutive nucleotides, which differ from any one of the nucleotide sequences of nucleotides 483-513, 486-516, 491-521, 483-521, 513-543, 987-1017, 989-1019, 1317-1347, 2237-2267, 2439-2469 of SEQ ID NO1 by 0, 1, 2 or 3 nucleotides, and the antisense ... The corresponding nucleotide sequences in NO:2 differ by 0, 1, 2 or 3 nucleotides for at least 15, 16, 17, 18, 19, 20 or 21 consecutive nucleotides, wherein the sense strand and the antisense strand may partially, substantially or completely complement each other. The dsRNA agent of claim 1-2, wherein the sense strand comprises at least 15, 16, 17, 18, 19, 20 or 21 consecutive nucleotides that differ from any one of the nucleotide sequences of nucleotides 488-508, 491-511, 496-516, 488-516, 518-538, 992-1012, 994-1014, 1322-1342, 2242-2262, 2444-2464 of SEQ ID NO: 1 by 0, 1, 2 or 3 nucleotides, and the antisense strand comprises at least 15, 16, 17, 18, 19, 20 or 21 consecutive nucleotides that differ from the corresponding nucleotide sequence of SEQ ID NO: 2 by 0, 1, 2 or 3 nucleotides. The dsRNA agent of claim 1, wherein the antisense strand comprises a region complementary to the CFB RNA transcript, the complementary region comprising at least 15 consecutive nucleotides that differ by no more than 1, 2 or 3 nucleotides from any of the antisense sequences listed in any one of Tables 1-3. The dsRNA agent of claim 1, wherein the antisense strand comprises a region complementary to the CFB RNA transcript, the region comprising at least 15 consecutive nucleotides from any one of the antisense sequences listed in any one of Tables 1-3. The dsRNA agent of claim 1, wherein the double-stranded ribonucleic acid (dsRNA) agent is used to inhibit CFB expression, wherein the dsRNA agent comprises a sense strand and an antisense strand, nucleotide positions 2 to 18 in the antisense strand comprise a region complementary to the CFB RNA transcript, wherein the complementary region comprises at least 15, 16, 17, 18, 19, 20 or 21 consecutive nucleotides that differ by 0, 1, 2 or 3 nucleotides from one of the antisense sequences listed in one of Tables 1-3, and optionally comprises a targeting ligand. The dsRNA agent of claim 6, wherein the region complementary to the CFB RNA transcript comprises at least 15, 16, 17, 18 or 19 consecutive nucleotides that differ from one of the antisense sequences listed in one of Tables 1-3 by no more than 3 nucleotides. The dsRNA agent of any one of claims 1-7, wherein the antisense chain of the dsRNA is substantially or completely complementary to any one of the target regions of SEQ ID NO: 1, and preferably the dsRNA agent comprises the antisense chain sequence listed in any one of Tables 1-3. The dsRNA agent of any one of claims 1-8, wherein the sense chain sequence and the antisense chain sequence in the dsRNA agent are at least substantially complementary or completely complementary to each other, preferably, wherein the dsRNA agent comprises the sense chain sequence described in any one of Tables 1-3. The dsRNA agent of any one of claims 1 to 9, wherein the dsRNA agent comprises a sequence listed as a duplex sequence in any one of Tables 1 to 3. The dsRNA agent of any one of claims 1-10, wherein the dsRNA agent comprises at least one modified nucleotide. The dsRNA agent of any one of claims 1 to 11, wherein all or substantially all nucleotides of the sense chain and/or the antisense chain are modified nucleotides. A dsRNA agent as described in any of claims 1-12, wherein the double-stranded ribonucleic acid (dsRNA) agent is used to inhibit CFB expression, wherein the dsRNA agent includes a sense chain and an antisense chain, wherein the sense chain is complementary to the antisense chain, wherein the antisense chain comprises a region that is complementary to a portion of the CFB RNA transcript, wherein the length of each chain is about 15 to about 30 nucleotides, wherein the sense chain comprises a sequence that can be represented by formula (I): 5′-(N′ _L ) _n′ N′ _L N′ _L N′ _L N′ _L N′ L N′ _F N′ _L N′ _F N′ _L N′ _N1 N′ _N2 N′ _L N′ _L N′ _L N′ _L N′ _L (N′ _L ) _m′ -3′ (I); wherein: each N′ _F represents a 2′-fluorine-modified nucleotide; each N′ _N1 and N′ _N2 independently represents a modified or unmodified nucleotide; each _N'L independently represents a modified or unmodified nucleotide, but does not represent a 2'-fluoro-modified nucleotide, and m' and n' are each independently an integer from 0 to 7. The dsRNA agent of any one of claims 1-12, wherein the double-stranded ribonucleic acid (dsRNA) agent is used to inhibit CFB expression, wherein the dsRNA agent comprises a sense strand and an antisense strand, wherein the sense strand is complementary to the antisense strand, wherein the antisense strand comprises a region that is complementary to a portion of a CFB RNA transcript, wherein each strand is about 18 to about 30 nucleotides in length, wherein the antisense strand comprises a sequence that can be represented by formula (II): ₃ ′-( _NL ) _{nNM1N LNM2N L N FN L} N _{M3N M4N L N L N L N M5N} L N _{M6N L} N _L N _{FN L} -5′ (II); wherein: each _NF represents a 2′-fluorine-modified nucleotide; each _NM1 , _NM2 , _NM3 , _NM4 , _NM5 and N _M6 independently represents a modified or unmodified nucleotide; each _NL independently represents a modified or unmodified nucleotide but not a 2'-fluorine-modified nucleotide, and n is an integer from 0 to 7. The dsRNA agent of any one of claims 1 to 12, wherein the double-stranded ribonucleic acid (dsRNA) agent is used to inhibit CFB expression, wherein the dsRNA agent comprises a sense chain and an antisense chain, wherein the sense chain and the antisense chain form a dsRNA duplex, wherein the sense chain is complementary to the antisense chain, wherein the antisense chain comprises a region complementary to the CFB RNA transcript, wherein the complementary region comprises at least 15 consecutive nucleotides, wherein the dsRNA comprises a duplex represented by formula (III): Sense chain: 5′-(N′ _L ) _n′ N′ _{L N′ L N} ′ _L N′ _L N′ _F N′ _L N′ _F N′ _L N′ _N1 N′ _N2 N′ L N′ _L N′ _L N′ _{L N} ′ _L (N′ _L ) _m′ -3′ Antisense chain: 3′-( _{NL ) nNM1NLNM2NLNFNLNM3NM4NLNLNLNM5NLNM6NLNLNFNL - 5 ′} ( _III ); wherein: the length of each chain is about ₁₈ to about ₃₀ nucleotides; each _NF and _N'F independently _represents a _2' - _{fluorine -} modified nucleotide; _NM1 , _NM2 , _NM3 , _NM4 , _NM5 , N'N1, and _{N'N2 each independently represent a modified or unmodified nucleotide; N'N1 and N'N2 contain only one 2' - fluorine} -modified nucleotide; _NM1 , _NM2 , _NM3 , _NM4 , _{NM5 and NM6} have _only three 2'-fluorine-modified nucleotides _; each _NL and _N ' _L independently represents a modified or unmodified nucleotide, but does not represent a 2'-fluoro-modified nucleotide, and m', n' and n are each independently an integer from 0 to 7. The dsRNA agent of any one of claims 11-15, wherein the one or more modified nucleotides are independently selected from: 2'-O-methyl nucleotides, 2'-fluoro nucleotides, 2'-deoxy nucleotides, 2'3'-seco nucleotide mimetics, locked nucleotides, unblocked nucleic acid nucleotides (UNA), glycol nucleic acid nucleotides (GNA), 2'-F-arabinonucleotides, 2'-methoxyethyl nucleotides, debasing nucleotides, ribitol, inverted nucleotides, inverted debasing nucleotides, isomannoside nucleotides, inverted 2'-OMe nucleotides, inverted 2'-deoxynucleotides, 2'-amino modified nucleotides, 2'-alkyl-modified nucleotides, morpholino nucleotides, 3'-OMe nucleotides, nucleotides containing a 5'-phosphorothioate group, terminal nucleotides linked to a cholesterol derivative or dodecanoic acid bisdecylamide group, 2'-amino modified nucleotides, phosphoramidates, or nucleotides containing an unnatural base. The dsRNA agent of any one of claims 1-16, wherein the 5' end of the guide strand comprises an E-vinylphosphonate nucleotide. The dsRNA agent of any one of claims 1-17, wherein the dsRNA agent comprises at least one phosphorothioate nucleoside interlinkage. The dsRNA agent of any one of claims 1-17, wherein the sense strand comprises at least one phosphorothioate nucleoside bond. The dsRNA agent of any one of claims 1-17, wherein the antisense strand comprises at least one phosphorothioate nucleoside bond. The dsRNA agent of any one of claims 1-17, wherein the sense strand comprises 1, 2, 3, 4, 5 or 6 phosphorothioate internucleoside bonds. The dsRNA agent of any one of claims 1-17, wherein the antisense strand comprises 1, 2, 3, 4, 5 or 6 phosphorothioate nucleoside bonds. The dsRNA agent of any one of claims 1-22, wherein the modified sense chain is a modified sense chain sequence shown in one of Tables 2-3. The dsRNA agent of any one of claims 1-22, wherein the modified antisense chain is a modified antisense chain sequence shown in one of Tables 2-3. The dsRNA agent of any one of claims 1-24, wherein the sense strand is complementary or substantially complementary to the antisense strand, and the length of the complementary region is between 16-23 nucleotides. The dsRNA agent of any one of claims 1-25, wherein the complementary region is 19-21 nucleotides in length. The dsRNA agent of any one of claims 1-26, wherein the length of each strand does not exceed 30 nucleotides. The dsRNA agent of any one of claims 1-26, wherein the length of each strand does not exceed 25 nucleotides. The dsRNA agent of any one of claims 1-26, wherein the length of each strand does not exceed 23 nucleotides. The dsRNA agent of any one of claims 1-29, wherein the dsRNA agent comprises at least one modified nucleotide and further comprises one or more targeting groups or linking groups. The dsRNA agent of claim 30, wherein one or more targeting groups or linking groups are conjugated to the sense chain. The dsRNA agent of claim 30 or 31, wherein the targeting group or linking group comprises N-acetylgalactosamine (GalNAc). The dsRNA agent of any one of claims 30-32, wherein the targeting group has the following structure: , n'' is independently 1 or 2. The dsRNA agent of any one of claims 30-33, wherein the targeting group has the following structure: GLO-1 GLS-1* GLO-2 GLS-2* GLO-3 GLS-3* GLO-4 GLS-4* GLO-5 GLS-5* GLO-6 GLS-6* GLO-7 GLS-7* GLO-8 GLS-8* GLO-9 GLS-9* GLO-10 GLS-10* GLO-11 GLS-11* GLO-12 GLS-12* GLO-13 GLS-13* GLO-14 GLS-14* GLO-15 GLS-15* GLO-16 GLS-16* . The dsRNA agent of any one of claims 1-34, wherein the dsRNA agent comprises a targeting group linked to the 5'-end of the sense strand. The dsRNA agent of any one of claims 1-34, wherein the dsRNA agent comprises a targeting group linked to the 3' end of the sense strand. The dsRNA agent of any one of claims 1-34, wherein the antisense chain comprises an inverted debasing residue at the 3' end. The dsRNA agent of any one of claims 1 to 34, wherein the sense strand comprises one or two inverted debasing residues or imann residues at the 3' or/and 5' ends. The dsRNA agent of any one of claims 1-38, wherein the dsRNA agent has two blunt ends. The dsRNA agent of any one of claims 1-38, wherein at least one strand comprises a 3' overhang of at least 1 nucleotide. The dsRNA agent of any one of claims 1-38, wherein at least one strand comprises a 3' overhang of at least 2 nucleotides. The dsRNA agent of any one of claims 1-41, wherein the CFB RNA transcript is SEQ ID NO: 1. A composition comprising the dsRNA agent of any one of claims 1-42. The composition as described in claim 43 further comprises a pharmaceutically acceptable carrier. The composition of claim 44, further comprising one or more additional therapeutic agents. The composition of claim 45, wherein the composition is packaged in a kit, container, package, dispenser, prefilled syringe or vial. The composition of any one of claims 43-46, wherein the composition is formulated for subcutaneous administration or formulated for intravenous (IV) administration. A cell comprising a dsRNA agent as described in any one of claims 1-42, optionally, the cell is a mammalian cell, optionally a human cell. A method for inhibiting CFB gene expression in a cell, wherein the method comprises: (i) preparing a cell containing an effective amount of a double-stranded RNA (dsRNA) agent as described in any one of claims 1-42 or a composition as described in any one of claims 43-47. The method as described in claim 49, further comprising: (ii) maintaining the cells prepared in claim 49(i) for a sufficient time to obtain degradation of the mRNA transcript of the CFB gene, thereby inhibiting the expression of the CFB gene in the cells. The method of any one of claims 49-50, wherein the cell is located in a subject and the dsRNA agent is administered subcutaneously to the subject. The method of any of claims 49-50, wherein the cell is located in a subject and the dsRNA agent is administered to the subject by IV administration. A method as described in claim 51 or 52, further comprising evaluating the suppression of the CFB gene after administering a dsRNA agent to a subject, wherein the means for evaluating comprises: (i) determining one or more physiological characteristics of a CFB-related disease or condition in the subject, and (ii) comparing the determined physiological characteristics with baseline pre-treatment physiological characteristics of the CFB-related disease or condition and/or with control physiological characteristics of the CFB-related disease or condition; wherein the comparison indicates one or more of the presence or absence of suppression of CFB gene expression in the subject. A method as described in claim 53, wherein the determined physiological characteristic is one or more of: CFB mRNA level, CFB protein level, or CH50 activity, AH50, lactate dehydrogenase (LDH), hemoglobin level in the subject; any one or more levels of C3, C9, C5, C5a, C5b and soluble C5b-9 complex. A method as described in claim 54, wherein one or more of the CFB mRNA level in the subject, the CFB protein level in the subject is reduced; and/or one or more of the CH50 activity, AH50, lactate dehydrogenase (LDH), hemoglobin level, the level of any one or more of C3/C9/C5/C5a/C5b/soluble C5b-9 complex, and lipid levels in the blood or serum are reduced, indicating a reduction in CFB gene expression in the subject. A method for inhibiting CFB gene expression in a subject, wherein the method comprises administering to the subject an effective amount of a double-stranded RNA (dsRNA) agent as described in any one of claims 1 to 42 or a composition as described in any one of claims 43 to 47. The method of claim 56, wherein the dsRNA agent is administered subcutaneously to the subject. The method of claim 56, wherein the dsRNA agent is administered to the subject via IV administration. A method as in any of claims 56-58, further comprising evaluating suppression of a CFB gene after administration of the dsRNA agent, wherein the means for evaluating comprises: (i) determining one or more physiological characteristics of a CFB-related disease or condition in the subject, and (ii) comparing the determined physiological characteristics to baseline pre-treatment physiological characteristics of the CFB-related disease or condition and/or to control physiological characteristics of a CFB-related disease or condition; wherein the comparison indicates one or more of the presence or absence of suppression of CFB gene expression in the subject. A method as described in claim 60, wherein the determined physiological characteristics are one or more of: CFB mRNA level, CFB protein level, or CH50 activity, AH50, lactate dehydrogenase (LDH), hemoglobin level in the subject; the level of any one or more of C3, C9, C5, C5a, C5b, soluble C5b-9 complex and lipid levels in blood or serum. A method as described in claim 60, wherein a reduction in one or more of the subject's CFB mRNA level, the subject's CFB protein level; and/or a reduction in CH50 activity, AH50, lactate dehydrogenase (LDH), hemoglobin level, the level of any one or more of C3, C9, C5, C5a, C5b, soluble C5b-9 complex, and one or more of lipid levels in the blood or serum indicates a reduction in the subject's CFB gene expression. A method for treating a disease or condition associated with CFB protein, the method comprising administering to a subject an effective amount of a double-stranded RNA (dsRNA) agent as described in any one of claims 1 to 42 or a composition as described in any one of claims 43 to 47 to inhibit CFB gene expression. The method of claim 62, wherein the disease or condition is one or more of the following: an autoimmune disease; a dysfunction of the complement system, including abnormal upregulation of complement components, such as CFB, C3 glomerulopathy (C3G), systemic lupus erythematosus (SLE), lupus nephritis; an Ig-mediated renal disease, such as IgA nephropathy and primary membranous nephropathy, nephropathy, diabetic nephropathy, polycystic nephropathy, membranous nephropathy; age-related macular degeneration (AMD), including dry AMD and geographic atrophy, typical or infectious hemolytic uremic syndrome (tHUS), atypical uremic hemolytic syndrome (aHUS), asthma, psoriasis, thrombotic microangiopathy, ischemia-reperfusion injury, paroxysmal nocturnal hemoglobinuria (PNH), rheumatic diseases, rheumatoid arthritis, multiple sclerosis (MS), neuromyelitis optica (NMO), immune complex-mediated glomerulonephritis (IC-mediated GN), post-infectious glomerulonephritis (PIGN), antineutrophil cytoplasmic autoantibody-associated vasculitis (ANCA-AV), antiphospholipid antibody syndrome (APS), dysbacteriosis Periodontal disease, malaria-induced anemia, dermatomyositis with sphaeroidal ecchymositis, Shiga toxin E-associated hemolytic uremic syndrome, myasthenia gravis (MG), neuromyelitis optica (NMO), dense deposits, coronary artery disease, dermatomyositis, Graves' disease, atherosclerosis, Alzheimer's disease, systemic inflammatory response sepsis, septic shock, spinal cord injury, glomerulonephritis, Hashimoto's thyroiditis, type I diabetes, psoriasis, sphaeroidal ecchymoses, autoimmune hemolytic anemia (AIHA), cold agglutinin disease, humoral and vascular transplant rejection, graft dysfunction, myocardial infarction, graft allergy, hyperlipidemia, and sepsis. The method of claim 63, further comprising administering an additional treatment regimen to the subject. The method of claim 64, wherein the additional treatment regimen comprises: administering to the subject one or more CFB antisense polynucleotides of the present invention, administering to the subject a non-CFB dsRNA therapeutic agent, and modifying the subject's behavior. A method as described in claim 65, wherein the non-CFB dsRNA therapeutic agent is one or more of the following: a C5 inhibitor, such as an anti-complement component C5 antibody or an antigen-binding fragment thereof (e.g., eculizumab, ravulizumab-cwvz or pozelimab (REGN3918)) or a C5 peptide inhibitor (e.g., zilucoplan); a C3 peptide inhibitor (e.g., compstatin). The method of any one of claims 62-66, wherein the dsRNA agent is administered subcutaneously to the subject. The method of any one of claims 62-66, wherein the dsRNA agent is administered to the subject by IV administration. The method of any of claims 62-69, further comprising determining the efficacy of the administered double-stranded RNA (dsRNA) agent in the subject. A method as described in claim 69, wherein the method of determining the efficacy of a treatment in a subject comprises: (i) determining one or more physiological characteristics of a CFB-related disease or condition in the subject, and (ii) comparing the determined physiological characteristics to a baseline pre-treatment physiological characteristic of the CFB-related disease or condition, wherein the comparison indicates one or more of the presence, absence, and efficacy level of a double-stranded RNA (dsRNA) agent administered to the subject. A method as described in claim 70, wherein the physiological characteristics determined are: the subject's CFB mRNA level, CFB protein level, or CH50 activity, AH50, lactate dehydrogenase (LDH), hemoglobin level; the level of one or more of C3, C9, C5, C5a, C5b, soluble C5b-9 complex and lipid levels in blood or serum. A method as described in claim 70, wherein a reduction in one or more of CFB mRNA levels, CFB protein levels, and/or a reduction in CH50 activity, AH50, lactate dehydrogenase (LDH), hemoglobin levels, levels of one or more of C3, C9, C5, C5a, C5b, soluble C5b-9 complex, and lipid levels in the blood or serum of the subject indicates that administration of a double-stranded RNA (dsRNA) agent to the subject has efficacy. A method for reducing the level of CFB protein in a subject compared to the baseline pre-treatment level of CFB protein in the subject, the method comprising administering to the subject an effective amount of a double-stranded RNA (dsRNA) agent as described in any one of claims 1-42 or a composition as described in any one of claims 43-47 to reduce the level of CFB gene expression. The method of claim 73, wherein the dsRNA agent is administered to the subject subcutaneously or by IV administration. A method for altering the physiological characteristics of a CFB-related disease or condition in a subject as compared to a baseline, pre-treatment physiological characteristic of the CFB-related disease or condition in the subject, the method comprising administering to the subject an effective amount of a double-stranded RNA (dsRNA) agent as described in any one of claims 1-42 or a composition as described in any one of claims 43-47 to alter the physiological characteristics of the CFB-related disease or condition in the subject. The method of claim 75, wherein the dsRNA agent is administered to the subject subcutaneously or by IV administration. A method as described in any of claims 75-76, wherein the physiological characteristics are one or more of: CFB mRNA level, CFB protein level, CH50 activity, AH50, lactate dehydrogenase (LDH), hemoglobin level in the subject; the level of any one or more of C3, C9, C5, C5a, C5b, soluble C5b-9 complex and lipid level in blood or serum.