RS51950B - CYCLIC PEPTIDE ANTAGONISTS CXCR4 - Google Patents

Abstract

CYCLIC PEPTIDE ANTAGONISTS CXCR4. A lactam-cyclized peptide of formula I: R1 - cyclopd - Tyr - X3 - DArg - 2Nal - Gly - X7] - X8 - X9 - X10 - R2 (SEQ ID (I) NO: 1), characterized in that: a) lactam is given is formed by forming an amide bond between the side chain of the amino group in Xi and the side chain of the carboxyl group X7, wherein X1 and X7, the pair are selected from the group consisting of (D / L) Ag1 / Glu, Dab / Glu, and Dap / Glu, and R1 is Ac or n-hexanoyl; orb) the lactam given is formed by forming an amide bond between the side chain of the carboxyl group Xt and the side chain of the amino group X7, where X1 and X7, a pair selected from the group consisting of Asp / (D / L) Agl, Asp / Dab, Asp / Dap , Glu / (D / L) Agl, Glu / Dab, Glu / Dap, Glu / DDap, and Glu / Lys, and R1 is Ac or Bz, or where Xi and X7 are a pair selected from the group consisting of succinyl / ( D / L) Agl, succinyl / Dab, succinyl / Dap, succinyl / Lys, and succinyl / Orn, and Ri is absent; ilic) the lactam given is formed by forming an amide bond between the α-amino group in Xi and the side chain of the carboxyl group X7, where X1 and X7, a pair selected from the group consisting of Ala / Glu, Ala / DGlu, DAla / Glu, DAla / DGlu, Dap (Ac) / Glu, Gly / Asp, Gly / Glu, Gly / DGIu, Leu / Glu, Leu / DGlu, Lys / DGIu, Lys (Ac) / Glu, 2Nal / Glu, Phe / Glu, Phe / DGlu, DPhe / Glu, and DPhe / DGlu, and R1 is absent; ilid) the lactam given is formed by forming an amide bond between the non-amino group of the non-side chain on X1 and the side chain of the carboxyl group X7, where X1 and X7, a pair selected from the group consisting of (3-Ala / Asp, (3- Ala / Glu, 5-amino-valeryl / Asp, 5-aminovaleryl / Glu, 4-AMB / Glu, 4-AMPA / Asp, and 4-AMPA / Glu, and R, is absent; or e) lactam is formed by formation amide bonds between the α-amino group X2 and the side chain of the carboxyl group X7, wherein X2 and X7, a pair selected from the group consisting of Tyr / Asp, Tyr / Glu, and Tyr / DGIu, and each of R1 and X1 is absent; R1 is a substituent on the a-amino group in X1 when X1 contains an n a-amino group and the a-amino group provided is not a component of the lactam amide bond, selected from the group consisting of Ac, Bz, and n-hexanoyl, or is absent, where X1 is selected from the group consisting of (D / L) Agl, Asp, Dab, Dap, and Glu; X1 is selected from the group consisting of (D / L) Agl, Ala, (3-Ala, DAla, 5-aminovaleryl, 4 -AMB, 4-AMPA, Asp, Dab, Dap, Dap (Ac), Glu, Gly, Leu, Lys, Lys (Ac), 2Nal, Phe, DPhe, and succinyl, or absent; X3 is from selected from the group consisting of Arg, Lys, Lys (iPr), and Lys (Me2); X7 is selected from the group consisting of (D / L) Agl, Asp, Dab, Dap, DDap, Glu, DGIu, Lys, and Om; X8 is selected from the group consisting of (3-Ala, Arg, DArg, Gly, Lys , Lys (iPr), and Om, or absent; X9 is selected from the group consisting of Gly, 2Nal, D2Nal, and DPhe, or absent; X10 is 2Nal, or absent; whereby, when X8 is absent, then each of X9 and X10 is absent, and when X9 is absent, then X10 is absent, iR2 is selected from the group consisting of NH2 and NHEt, Ilinjeg's pharmaceutically acceptable salt. The application contains 12 more claims.

Description

[0001] The present invention relates to novel compounds of cyclic peptides as CXCR4 antagonists and to the use and treatment of diseases in which the pathogenesis is mediated by CXCR4 and SDF-1.

[0002] CXCR4, a G-protein-coupled receptor, and its natural ligand, stromal cell-derived factor-1 (SDF-1; CXCL12), are a chemokine receptor-ligand pair. CXCR4 is successively- or overexpressed (increased)-expressed in various human cancers. SDF-1, the only known ligand of CXCR4, is highly expressed in the tumor microenvironment, as well as in the bone marrow, lungs, liver, and lymph nodes, i.e., organ sites lethally involved in tumor metastasis. CXCR4/SDF-1 interaction plays an important role in multiple stages of tumorigenesis, including tumor growth, invasion, angiogenesis, and metastasis, as well as in rheumatoid arthritis, pulmonary fibrosis, and HIV infection (Tsutsumi et al. (2006) Peptide Science 88(2):279-289).

[0003] Regarding the involvement of CXCR4/SDF-1 in these severe diseases, CXCR4 is an attractive therapeutic target.

[0004] AMD3100, a bicyclam CXCR4 antagonist, is currently in Phase III clinical trials for stem cell mobilization for stem cell transplantation in patients with multiple myeloma and non-Hodgkins lymphoma. AMD070, a next msli CXCR4 antagonist molecule, is currently in Phase II clinical trials for HIV infection. CTCE9908, a bivalent (dimeric) peptide CXCR4 antagonist, is currently in Phase 1b/II clinical trials for cancer. FC131, a cyclic pentapeptide CXCR4 antagonist, inhibits 125I-SDF-1 binding to CXCR4 transfectant with IC504 nM (Fujii et al. (2003) Angevv. Chem. Int. Ed. 42:3251-3253; Araki et al. (2003) Peptide Science. The Japanese Peptide Society (2004) 1:207-210).

[0005] There is a need for improved CXCR4 antagonists that are potent and selective, exhibit little or no activity for other chemokine receptors. The compounds of the present invention are potent and selective CXCR4 antagonists. Their high potency (strength) allows the use of low doses in therapeutic regimens, while their high selectivity minimizes non-target related adverse side effects. In addition, the compounds provided herein possess other highly desirable pharmacological properties, such as high bioavailability when administered subcutaneously, good in vivo metabolic stability, and pharmacokinetic/pharmacodynamic properties that allow convenient dosing.

[0006] Accordingly, in a first aspect, the present invention provides a lactam-cyclized peptide of the formula I: R riclopc, - Tyr - X3 - DArg - 2Nal - Gly - X7] - X8 - X9 - X10 - R2 (SEQ ID NO:1) (I)

where is:

a) give a lactam formed by amide bonding between the side chain of the amino group X^ and the side chain of the carboxyl group X7, where X and X7 are, respectively, a pair selected from the group

consist of (D/L) Agl/Glu, Dab/Glu, and Dap/Glu, and Rije Ac or n-hexanoyl; or

b) give a lactam formed by amide bonding between the side chain of the carboxyl group Xii and the side chain of the amino group X7, where Xii X7 are, respectively, a pair selected from the group

consisting of Asp/(D/L)Agl, Asp/Dab, Asp/Dap, Glu/(D/L)Agl, Glu/Dab, Glu/Dap, Glu/DDap, and Glu/Lys, and is Ac or Bz, or where X and X7 are, respectively, a pair selected from the group consisting of succinyl/(D/L)Agl, succinyl/Dab, succinyl/Dap, succinyl/Lys, and succinyl/Orn, iR^is absent; or

c) a given lactam formed by the formation of an amide bond between the a-amino group Xii of the side chain of the carboxyl group X7, where X^ and X7 are, respectively, a pair selected from the group consisting of

Ala/Glu, Ala/DGlu, DAla/Glu, DAla/DGlu, Dap(Ac)/Glu, Gly/Asp, Gly/Glu, Gly/DGIu, Leu/Glu, Leu/DGlu, Lys/DGIu, Lys(Ac)/Glu, 2Nal/Glu, Phe/Glu, Phe/DGlu, DPhe/Glu, and R, and R is omitted; or

d) give a lactam formed by the formation of an amide bond between the non-a, non-side chain of the amino group X-, and the side chain of the carboxyl group X7, where X, and X7 are, respectively, a pair selected

from the group consisting of (3-Ala/Asp, P-Ala/Glu, 5-amino-valeryl/Asp, 5-aminovaleryl/Glu, 4-AMB/Glu, 4-AMPA/Asp, and 4-AMPA/Glu, and R is omitted; or

e) give a lactam formed by the formation of an amide bond between the α-amino group X2 and the side chain of the carboxyl group X7, where X2 and X7 are, respectively, a pair selected from the group consisting of

Tyr/Asp, Tyr/Glu, and Tyr/DGIu, and Rti Xisu absent;

R-i is a substituent on the a-amino group X^wherein X contains an a-amino group and said a-amino group is not part of said lactam amide bond, selected from the group consisting of Ac, Bz, and n-hexanoyl, or is omitted, where X is selected from the group consisting of (D/L)Agl, Asp, Dab, Dap, and Glu;

Xi is selected from the group consisting of (D/L)Agl, Ala, P-Ala, DAla, 5-aminovaleryl, 4-AMB, 4-AMPA, Asp, Dab, Dap, Dap(Ac), Glu, Gly, Leu, Lys, Lys(Ac), 2Nal, Phe, DPhe, and succinyl, or is absent;

X3 is selected from the group consisting of Arg, Lys, Lys(iPr), and Lys(Me2);

X7 is selected from the group consisting of (D/L)Agl, Asp, Dab, Dap, DDap, Glu, DGIu, Lys, and Om;

X8 is selected from the group consisting of (3-Ala, Arg, DArg, Gly, Lys, Lys(iPr), and Om, or is absent;

X9 is selected from the group consisting of Gly, 2Nal, D2Nal, and DPhe, or is absent;

X10 is 2Nal, or is absent;

where when X8 is absent, then X9 and X10 are absent, and when X9 is absent, then X10 is absent, and

R 2 is selected from the group consisting of NH 2 and NHEt, or

its pharmaceutically acceptable salt.

[0007] Alternatively given, this is equivalent to the lactam-cyclized peptide of formula I: R!-cyclom-Tyr-X3-DArg-2Nal-Gly-X7]-X8-X9-X10-R2(SEQ ID (I) NO:1)

where:

Rtje substituent on the a-amino groupX^wherein X-contains an a-amino group and this a-amino group is not part of a given lactam amide bond, selected from the group consisting of Ac, Bz, and n-hexanoyl, or is absent, wherein Xi is selected from the group consisting of (D/L)Agl, Asp, Dab, Dap, and Glu;

X1 is selected from the group consisting of (D/L)Agl, Ala, (3-Ala, DAla, 5-aminovaleryl, 4-AMB, 4-AMPA, Asp, Dab, Dap, Dap(Ac), Glu, Gly, Leu, Lys, Lys(Ac), 2Nal, Phe, DPhe, and succinyl, or is absent;

X3 is selected from the group consisting of Arg, Lys, Lys(iPr), and Lys(Me2);

X7 is selected from the group consisting of (D/L)Agl, Asp, Dab, Dap, DDap, Glu, DGIu, Lys, and Om;

X8 is selected from the group consisting of (3-Ala, Arg, DArg, Gly, Lys, Lys(iPr), and Om, or is absent;

Xg is selected from the group consisting of Gly, 2Nal, D2Nal, and DPhe, or is absent;

Xi0 is 2Nal, or is absent;

where when X8 is absent, then X9 and X10 are absent, and when X9 is absent, then X10 is absent, and

R 2 is selected from the group consisting of NH 2 and NHEt,

where is:

a) give a lactam formed by the formation of an amide bond between the side chain of the amino group X-i and the side chain of the carboxyl group X7, when Xi and X7 are, respectively, a pair selected from the group consisting of (D/L)Agl/Glu, Dab/Glu, and Dap/Glu, and Rije Ac or n-hexanoyl; or b) provide a lactam formed by forming an amide bond between the side chain of the carboxyl group X^ and the side chain of the amino group X7, when X1 and X7, respectively, are a pair selected from the group consisting of Asp/(D/L)Agl, Asp/Dab, Asp/Dap, Glu/(D/L)Agl, Glu/Dab, Glu/Dap, Glu/DDap, and Glu/Lys, and R, is Ac or Bz, or when X, and X 7 , respectively, a pair selected from the group consisting of succinyl/(D/L)Agl, succinyl/Dab, succinyl/Dap, succinyl/Lys, and succinyl/Orn, and R 7 is absent; or c) provide a lactam formed by the formation of an amide bond between the α-amino group Xi and the side chain of the carboxyl group X7, where X, and X7 are, respectively, a pair selected from the group consisting of Ala/Glu, Ala/DGIU, DAla/Glu, DAla/DGlu, Dap(Ac)/Glu, Gly/Asp, Gly/Glu, Gly/DGIu, Leu/Glu, Leu/DGlu, Lys/DGIu, Lys(Ac)/Glu, 2Nal/Glu, Phe/Glu, Phe/DGlu, DPhe/Glu, and DPhe/DGlu, and Rtje absent; or d) datilactam formed by the formation of an amide bond between the non-a, non-side chains of the amino group X1 and the side chain of the carboxyl group X7, when X^ and X7, respectively,

a pair selected from the group consisting of P-Ala/Asp, (3-Ala/Glu, 5-amino-valeryl/Asp, 5-aminovaleryl/Glu, 4-AMB/Glu, 4-AMPA/Asp, and 4-AMPA/Glu, and R is absent; or

e) give a lactam formed by the formation of an amide bond between the α-amino group X2 and the side chain of the carboxyl group X7, when X2 and X7 are, respectively, a pair selected from

groups consisting of Tyr/Asp, Tyr/Glu, and Tyr/DGIu, and R^ and X are absent, or

its pharmaceutically acceptable salt.

[0008] In a further aspect, the present invention provides a lactam-cyclized peptide of the formula I: R , - cycloPd - Tyr - X3 - DArg - 2Nal - Gly - X7] - X8 - X9 - X10 - R2 (SEQ ID NO:1) (I)

or a pharmaceutically acceptable salt thereof,

where:

X is selected from the group consisting of (D/L)Agl, Ala, (3-Ala, DAla, 5-aminovaleryl, 4-AMB, 4-AMPA, Asp, Dab, Dap, Dap(Ac), Glu, Gly, Leu, Lys, Lys(Ac), 2Nal, Phe, DPhe, and succinyl, or is absent,

wherein, when X-i is (D/L)Agl, Dab, or Dap and the α-amino group of Xini is part of a lactam amide bond, then the given α-amino group substituted with Rikoi is selected from the group consisting of Ac and n-hexanoyl;

wherein, when X is Asp or Glu and the α-amino group in Xt is not an integral part of the lactam amide bond, then the given α-amino group is substituted with Rv selected from the group consisting of Ac and Bz; and

in the case where X is Ala, P-Ala, DAla, 5-aminovaleryl, 4-AMB, 4-AMPA, Dap(Ac), Gly, Leu, Lys, Lys(Ac), 2Nal, Phe, DPhe or succinyl, then it is Riodsutan;

X3 is selected from the group consisting of Arg, Lys, Lys(iPr), and Lys(Me2);

X7 is selected from the group consisting of (D/L)Agl, Asp, Dab, Dap, DDap, Glu, DGIu, Lys, and Om;

X8 is selected from the group consisting of p-Ala, Arg, DArg, Gly, Lys, Lys(iPr), and Om, or is absent;

X9 is selected from the group consisting of Gly, 2Nal, D2Nal, and DPhe, or is absent;

X10 is 2Nal, or absent,

whereby, when X8 is absent, then Xg and Xi0 are absent, and when X9 is absent, then X10 is absent; and

R 2 is selected from the group consisting of NH 2 and NHEt,

and gives, when:

provide a lactam formed by forming an amide bond between the side chain of the amino group in X-, and the side chain of the carboxyl group in X7 and Xi and X7su, respectively, a pair selected from the group consisting of (D/L)Agl/Glu, Dab/Glu, and Dap/Glu, and Rije Ac or n-hexanoyl; or

given a lactam formed by forming an amide bond between the side chain of the carboxyl group in Xii and the side chain of the amino group in X7, and Xii X7 are, respectively, a pair selected from the group consisting of Asp/(D/L)Agl, Asp/Dab, Asp/Dap, Glu/(D/L)Agl, Glu/Dab, Glu/Dap, Glu/DDap, and Glu/Lys, and Rije Ac or Bz, or wherein X, and X7, respectively, a pair selected from the group consisting of succinyl/(D/L)Agl, succinyl/Dab, succinyl/Dap, succinyl/Lys, and succinyl/Om, and R is absent; or

given a lactam formed by forming an amide bond between the α-amino group in X-, and the side chain carboxyl group in X7, iX^ and X7 are, respectively, a pair selected from the group consisting of Ala/Glu, Ala/DGIU, DAla/Glu, DAla/DGlu, Dap(Ac)/Glu, Gly/Asp, Gly/Glu, Gly/DGIu,

Leu/Glu, Leu/DGlu, Lys/DGIu, Lys(Ac)/Glu, 2Nal/Glu, Phe/Glu, Phe/DGlu, DPhe/Glu, and DPhe/DGlu, iR, is absent; or

given the lactam formed by amide bond formation between non-a, non-side chain amino group in Xii side chain carboxyl group in X7, and X, and X7 are, respectively, a pair selected from the group consisting of P-Ala/Asp, P-Ala/Glu, 5-amino-valeryl/Asp, 5-aminovaleryl/Glu, 4-AMB/Glu, 4-AMPA/Asp, and 4-AMPA/Glu, and R, is absent; or

given the lactam formed by the formation of an amide bond between the α-amino group of Tyr on X2 and the side chain of the carboxyl group in X7, and X7 is selected from the group consisting of Asp, Glu, and DGIu, iRii Xisu is absent.

[0009] The periodic sequence motif in all compounds of formula I is the presence of Tyr at position X2, DArg at position X4, 2Nal at position X5, and Gly at position X6.

[0010] In a further aspect, the present invention provides lactam-cyclized peptides or a pharmaceutically acceptable salt thereof of formula I (SEQ ID NO:1), where: Ri is selected from the group consisting of Ac and Bz, or is absent; X-, is selected from the group consisting of P-Ala, 4-AMB, 4-AMPA, Asp, Dab, Dap, Dap(Ac), Glu, 2Nal, Phe, and succinyl, or is absent;

X3 is selected from the group consisting of Arg, Lys, Lys(iPr), and Lys(Me2);

X7 is selected from the group consisting of Asp, Dab, Dap, Glu, DGIu, Lys, and Om;

X8 is selected from the group consisting of Arg and Lys, or is absent;

X9 is absent;

X10 is absent; and

R 2 is selected from the group consisting of NH 2 and NHEt.

[0011] In a further aspect, the present invention provides a lactam-cyclized peptide or its pharmaceutically acceptable salt of formula I (SEQ ID NO:1), where: Rnje is selected from the group consisting of Ac and Bz, or is absent; Xt is selected from the group consisting of DAla, 5-aminovaleryl, 4-AMPA, Asp, Glu, Leu, Lys(Ac), Phe, DPhe, and succinyl;

X3 is selected from the group consisting of Arg, Lys, Lys(iPr), and Lys(Me2);

X7 is selected from the group consisting of (D/L)Agl, Asp, Dab, Dap, DDap, Glu, and DGIu;

X8 is selected from the group consisting of Arg, DArg, and Lys, or is absent;

X9 is absent;

Xi0 is absent; and

R 2 is selected from the group consisting of NH 2 and NHEt.

[0012] In a further aspect, the present invention provides a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof of formula I (SEQ ID NO: 1), where: Ri is selected from the group consisting of Ac, Bz, and n-hexanoyl, or is absent;

X, is selected from the group consisting of (D/L)Agl, Ala, 3-Ala, Asp, Dap, Glu, Gly, Lys, and Phe;

X3 is selected from the group consisting of Arg, Lys, Lys(iPr), and Lys(Me2);

X7 is selected from the group consisting of (D/L)Agl, Asp, Dap, Glu, and DGIu;

X8 is selected from the group consisting of P-Ala, Arg, Gly, Lys, Lys(iPr), and Om, or is absent;

X9 is selected from the group consisting of Gly, 2Nal, D2Nal, and DPhe, or is absent;

X10 is 2Nal, or absent; and

R 2 is selected from the group consisting of NH 2 and NHEt.

[0013] In a further aspect, the present invention provides a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof of formula I (SEQ ID NO:1), wherein: R-i is selected from the group consisting of Ac and Bz, or is absent;

X is selected from the group consisting of Ala, 5-aminovaleryl, Asp, Glu, Gly, Phe, DPhe, and succinyl;

X3 is selected from the group consisting of Arg, Lys(iPr), and Lys(Me2);

X7 is selected from the group consisting of (D/L)Agl, Asp, Dap, Glu, and DGIu;

X8 is selected from the group consisting of P-Ala, Arg, Gly, Lys, Lys(iPr), and Om, or is absent;

X9 is selected from the group consisting of Gly, D2Nal, and DPhe, or is absent;

Xi0je 2Nal, or is absent; and

R 2 is selected from the group consisting of NH 2 and NHEt.

[0014] In a further aspect, the present invention provides a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof of formula I (SEQ ID NO:1), where: X is selected from the group consisting of Gly and Phe;

X3 is Lys(iPr); and

X7 is DGIu.

[0015] In a further aspect, the present invention provides a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof of formula I (SEQ ID NO:1), wherein: Rije is absent;

Xi is selected from the group consisting of Gly and Phe;

X3 is Lys(iPr);

X7 is DGIu;

X8 is selected from the group consisting of Arg and Lys(iPr), or is absent;

X9 is absent;

X10 is absent; and

R 2 is selected from the group consisting of NH 2 and NHEt.

[0016] In a further aspect, the present invention provides a lactam-cyclized peptide or its pharmaceutically acceptable salt of formula I (SEQ ID NO:1), where: given lactam is formed by forming an amide bond between the side chain of the amino group Xii and the side chain of the carboxyl group X7;

Pm is selected from the group consisting of Ac and n-hexanoyl;

Xi is selected from the group consisting of (U/L)Agl, Dab, and Dap;

X3 is selected from the group consisting of Arg and Lys(iPr);

X7 is Glu;

X8 is Arg;

X9 is absent;

X10 is absent; and

R2 is NH2.

[0017] In a preferred embodiment of this aspect of the pronal, Xi is (D/L)Agl or Dap.

[0018] In a further aspect, the present invention provides a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof of formula I (SEQ ID NO:1), wherein: a given lactam is formed by forming an amide bond between the side chain of the carboxyl group in Xii and the side chain of the amino group in X7;

Rtje selected from the group consisting of Ac and Bz;

X-i is selected from the group consisting of Asp and Glu;

X3 is selected from the group consisting of Arg and Lys(Me2);

X7 is selected from the group consisting of (D/L)Agl, Dab, Dap, DDap, and Lys;

X8 is Arg;

X9 is absent;

Xi0 is absent; and

R2 is NH2.

[0019] In a preferred embodiment of this aspect of the invention, X7 is (D/L)Agl, Dab, Dap, or DDap. Better yet, the X7 is (D/L)Agl or Dap.

[0020] In a further aspect, the present invention provides a lactam-cyclized peptide or its pharmaceutically acceptable salt of formula I (SEQ ID NO:1), where: given lactam is formed by forming an amide bond between the side chain of the carboxyl group in X^ and the side chain of the amino group in X7;

Ri is absent;

X is succinyl;

X3 is Arg;

X7 is selected from the group consisting of (D/L)Agl, Dab, Dap, Lys, and Om;

X8 is Arg;

X9 is absent;

Xio is absent; and

R2 is NH2.

[0021] In a preferred embodiment of this aspect of the invention, X7 is (D/L)Agl or Dap.

[0022] In a further aspect, the present invention provides a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof of formula I (SEQ ID NO:1), wherein: a given lactam is formed by forming an amide bond between the α-amino group in Xii of the side chain of the carboxyl group in X7;

Ri is absent;

Xt is selected from the group consisting of Ala, DAla, Gly, Dap(Ac), Leu, Lys, Lys(Ac), 2Nal, Phe, and DPhe;

X3 is selected from the group consisting of Arg, Lys, Lys(iPr), and Lys(Me2);

X7 is selected from the group consisting of Asp, Glu, and DGIu;

X8 is selected from the group consisting of P-Ala, Arg, Gly, Lys, Lys(iPr), and Om, or is absent;

X9 is selected from the group consisting of Gly, 2Nal, D2Nal, and DPhe, or is absent;

X10 is 2Nal, or is absent;

When X8 is absent, then X9 and X10 are absent; and

R 2 is selected from the group consisting of NH 2 and NHEt.

[0023] In a preferred embodiment of this aspect of the invention, X is Ala, DAla, Gly, Leu, Lys, Lys(Ac), Phe, or DPhe. More preferably, X is Ala, Gly, Lys, or Phe.

[0024] In a preferred embodiment of this aspect of the invention, X3 is Arg, Lys, Lys(iPr), or Lys(Me2). And better, X3 is Arg.

[0025] In a preferred embodiment of this aspect of the invention, X7 is Asp, Glu, or DGIu. Better yet, the X7 is an Asp.

[0026] In a preferred embodiment of this aspect of the invention, X8 is P-Ala, Arg, Gly, Lys, Lys(iPr), Om, or is absent. More preferably, X8 is P-Ala, Gly, Lys, Lys(iPr), Orn, or absent.

[0027] In a preferred embodiment of this aspect of the invention, X9 is Gly, 2Nal, D2Nal, DPhe, or absent. More preferably, X9 is Gly, 2Nal, D2Nal, or DPhe.

[0028] In a preferred embodiment of this aspect of the invention, X10 is 2Nal, or is absent. And better, X10 is 2Nal.

[0029] In a preferred embodiment of this aspect of the invention, R 2 is NHEt.

[0030] In a further aspect, the present invention provides a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof of formula I (SEQ ID NO:1), where: a given lactam is formed by forming an amide bond between the non-a, non-side-chain amino group in Xt and the carboxyl chain in X7;

The head is absent;

X is selected from the group consisting of p-Ala, 4-AMB, 5-aminovaleryl, and 4-AMPA;

X3 is Arg;

X7 is selected from the group consisting of Asp and Glu;

X8 is selected from the group consisting of Arg and DArg;

X9 is absent;

X10 is absent; and

R2 is NH2.

[0031] In a preferred embodiment of this aspect of the invention, X 1 is P-Ala, 5-amino-valeryl, or 4-AMPA. And better, X, is P-Ala.

[0032] In a preferred embodiment of this aspect of the invention, X7 is Asp.

[0033] In a preferred embodiment of this aspect of the invention, X 8 is Arg.

[0034] In a further aspect, the present invention provides a lactam-cyclized peptide or its pharmaceutically acceptable salt of formula I (SEQ ID NO:1), where: given lactam is formed by forming an amide bond between the α-amino group in X2 and the side chain of the carboxyl group in X7;

Ri is absent;

Xi is absent;

X3 is Arg;

X7 is selected from the group consisting of Asp, Glu, and DGIu;

X8 is Arg;

Xg is absent;

X10 is absent; and

R2 is NH2.

[0035] In a further aspect, the present invention provides a lactam-cyclized peptide of the formula:

or a pharmaceutically acceptable salt thereof. The lactam is formed by the formation of an amide bond between the α-amino group in Phe and the side chain of the carboxyl group in Glu. A pharmaceutically acceptable salt may be an acetic acid salt.

[0036] In a further aspect, the present invention provides a pharmaceutical composition, comprising a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof as previously described, and a pharmaceutically acceptable carrier, diluent, or excipient.

[0037] In a further aspect, the present invention provides a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof as described above, for use in therapy.

[0038] In a further aspect, the present invention provides a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof as described above, for the treatment of rheumatoid arthritis, pulmonary fibrosis, HIV infection, or a cancer selected from the group consisting of breast cancer, pancreatic cancer, melanoma, prostate cancer, kidney cancer, neuroblastoma, non-Hodgkin's lymphoma, lung cancer, ovarian cancer, colorectal cancer, multiple myeloma, glioblastoma multiforme, and chronic lymphocytic leukemia.

[0039] In a further aspect, the present invention provides a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof as described above, for the manufacture of a drug for the treatment of rheumatoid arthritis, pulmonary fibrosis, HIV infection, or a cancer selected from the group consisting of breast cancer, pancreatic cancer, melanoma, prostate cancer, kidney cancer, neuroblastoma, non-Hodgkin's lymphoma, lung cancer, ovarian cancer, colorectal cancer, multiple myeloma, glioblastoma. multiforme, and chronic lymphocytic leukemia.

[0040] In a further aspect, the present invention provides a method of treating rheumatoid arthritis, pulmonary fibrosis, HIV infection, or a cancer selected from the group consisting of breast cancer, pancreatic cancer, melanoma, prostate cancer, kidney cancer, neuroblastoma, non-Hodgkin's lymphoma, lung cancer, ovarian cancer, colorectal cancer, multiple myeloma, glioblastoma multiforme, and chronic lymphocytic leukemia, and which comprises administering to the patient an effective amount of a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof as previously described.

[0041] In the macrocyclic peptide compounds of the present invention (SEQ ID NO:1), amino acids X 1 to X 10 are designated herein by their usual symbols consisting of three letters, shown from left to right from the amino-terminal end to the carboxy-terminal end. D- and L- (capital letters) refer to absolute stereochemistry. In the case where there is no mark in the given formula, the L-form of the amino acid is present. It can also be a dicarboxylic acid residue, i.e., a succinyl group. Amino acid or carboxylic acid residues given in brackets "[ ]" are within the cyclic structure; groups outside the parentheses are outside the cyclic ring. In all cases, cyclization via a lactam (amide) bond between Xi (or X 2 , eg, Tyr) and X 7 , which can be formed in several different ways, depends on the structures of Xi , X 2 , and X 7 .

[0042] When the lactam bond is formed between the side chain of the amino group in X-, and the side chain of the carboxyl group of X7 (Schemes 1 and 2; Examples 1-5), then the α-amino group in

Xi is protected with Ac or n-hexanoyl.

[0043] When the lactam bond is formed by exposing the side chain of the carboxyl group Xii to the side chain of the amino group X7, then the α-amino group of X is protected with Ac or Bz (Schemes 3 and 4; Examples 6-19). X can also be a bifunctional residue next to an α-amino acid, for example one with two carboxyl groups, eg, a succinyl residue. In this case, one carboxyl group forms an amide bond with the α-amino group of Tyr, and the other forms a cyclic lactam structure via an amide bond with the side chain of the amino group X7 (Schemes 3 and 4; Examples 20-24). When X-, , is succinyl, then R, is absent.

[0044] In most of the lactam-cyclized peptides provided herein (Schemes 5-15; Examples 25-28, 32-66, and 75-89), the α-amino group of X forms the lactam structure via an amide bond to the side chain of the carboxyl group of X7, and R^ is absent. Synthesis schemes of this category include cyclic peptides containing X^substances, i.e., β-Ala, 4-AMB, 5-aminovaleryl, and 4-AMPA, wherein the amino group is a non-side chain group, non-α-amino group (Schemes 5 and 6; Examples 67-74). R^ is also omitted in these cases.

[0045] When R, and X are absent, the lactam structure is formed via an amide bond between the α-amino group of Tyr (X2) and the side chain of the carboxyl group of X7 (Schemes 5 and 6; Examples 29-31).

[0046] The structures of common amino acids, e.g., alanine, glycine, etc., are well known in the art. The structures of the non-standard and substituted amino acids present in the compounds of the present invention are given below.

[0047] The lactam-cyclized peptides of the present invention can be obtained as pharmaceutically acceptable salts. These salts, and the usual methodology for their preparation, are known in the art. See, eg, P. Stahl et al. (2002) Handbook of Pharmaceutical Salts: Properties, Selection and Use, VCHA/Wiley-VCH; Berge et al. (1977) "Pharmaceutical Salts," Journal of Pharmaceutical Sciences 66(1):1-19.

[0048] The subject pronalsk compounds are potent antagonists of CXCR4/SDF-1 interactions. Compounds of formula (I) and their pharmaceutically acceptable salts specifically exemplified herein exhibit an average KioD value of about 7.5 nM or less as determined by the CXCR4/<125>1-SDF-1a binding assay described below. More preferred compounds of formula (I) and their pharmaceutically acceptable salts exhibit an average Q value in the range of about 0.2 nM to about 1 nM in this assay. Particularly preferred compounds of formula (I) and their pharmaceutically acceptable salts show an average value for Ki below 0.2 nM in this assay.

[0049] Additionally, the compounds and pharmaceutically acceptable salts of the subject pronal are preferably highly selective for the CXCR4 receptor, exhibit little or no inhibitory activity for other chemokine receptors, including CCR1, CCR2, CXCR2, CXCR3, and other G-protein coupled receptors at the concentrations tested, and no significant activity for serotonin, dopamine, and opioid receptors. Also, they preferably exhibit good stability in blood and plasma, good subcutaneous bioavailability, desirable pharmacokinetic/pharmacodynamic properties, and strong in vivo efficacy (efficacy) in inhibiting tumor growth, with a wide margin of safety.

[0050] In view of these pharmacological properties, the compounds of the present invention are indicated for the treatment of disorders involving CXCR4/SDF-1 interactions, or CXCR4 receptor activity, as in HIV infections. In particular, the compounds are useful in the treatment of malignancies in which angiogenic pathways, growth pathways, survival pathways, and metastatic pathways mediated by CXCR4 and SDF-1 are implicated in the pathogenesis, including breast cancer, pancreatic cancer, melanoma, prostate cancer, kidney cancer, neuroblastoma, non-Hodgkin's lymphoma, lung cancer, ovarian cancer, colorectal cancer, multiple myeloma, glioblastoma multiforme, and chronic lymphocytic leukemia, as well as in rheumatoid arthritis, pulmonary fibrosis, and HIV infection (Tsutsumi et al. (2006) Peptide Science 88(2):279-289).

[0051] Agi (aminoglycine) is a pro-chiral building block. When this residue appears in a peptide of the formula given here, then the a-carbon becomes a chiral center on which each of the two attached a-amino groups is individually attached to different residues. In this case, the final peptide product contains two diastereomers which are not resolved, and which may be present in ratios other than 1:1. "(D/L)Agl" in the peptide formula indicates this mixture of diastereomers. "(DL)Agl" means an Ag derivative that is racemic, for example Fmoc-(DL)Agl(Boc).

[0052] Values for "Kj" were calculated using the IC50 values reported in the CXCR4/<125>1-SDF-1a binding assay described below using Equation 7.22 in Enzymes, A Practical Introduction to Structure, Mechanism, and Data Analysis, Robert A. Copeland, Wiley-VCH, New York, 1996, page 207.

[0053] The term "SDF-1" includes all isoforms, SDF-1a and SDF-1(3), which are currently thought to exhibit similar functionality.

[0054] "Treatment/Treatment" as used herein refers to the curative treatment of disorders associated with CXCR4/SDF-1 interactions or CXCR4 receptor activity. Curative treatment refers to processes that involve slowing, terminating, containing, controlling, or halting the progression of a disease, but does not necessarily include the total elimination of all symptoms associated with the disease, condition, or disorder.

[0055] The compounds of the present invention can be used as drugs in human or veterinary medicine, administered in different ways. Most preferably, these compositions are for parenteral administration. These pharmaceutical compositions can be prepared by methods known in the art. See, e.g., Remington: The Science and Practice of Pharmacv, 19th ed. (1995), A. Gennaro et al., Mack Publishing Co., and contain one or more compounds of formula (I) or a pharmaceutically acceptable salt(s) thereof and a pharmaceutically acceptable carrier, diluent, or excipient.

An effective amount of the compound ranges from about 1 mg to about 300 mg, preferably from about 1 mg to about 200 mg, more preferably from about 1 mg to about 100 mg, and most preferably from about 1 mg to about 50 mg, per day.

[0057] All lactam-cyclized peptides of the subject pronal can be synthesized either by solid phase synthesis or liquid phase synthesis, or a combination thereof, with the peptide chain on solid phase and cyclization or other modifications on resin or in solution. These methods are well known in the art.

[0058] The following abbreviations have the given meanings: Ac: acetyl; Agi: aminoglycine; AMB: aminomethyl benzoic acid; AMPA: aminomethyl phenyl acetic acid; Bn: benzyl; Boe: tert-butyloxycarbonyl; BOP: (benzotriazol-1-yloxy)-tris(dimethylamino)phosphonium hexafluorophosphate; 2-Br-Z: 2-bromobenzyloxycarbonyl; Bz: benzoyl; Bzl: benzyl; 2-CI-Z: 2-chlorobenzyloxycarbonyl; Dab: 2,4-diaminobutyric acid;Dap:2,3-diamino-propionic acid;DCC:dicyclohexyl-carbodiimide;DCM:dichloromethane;DIC:diisopropyl carbodiimide;DIEA:diisopropyl-ethylamine;DMF,:N, Ndimethyl formamide;DMSO:dimethylsulfoxide;EDT:1,2-ethanedithiol;Et:ethyl; Fm: 9-fluorenylmethyl; Fmoc: 9-fluoro-enylmethoxy carbonyl;HATU:N-[(dimethylamino)-1H-1,2,3-triazolo[4,5-b]pyridin-1-ylmethylene]-N-methylmethanaminium hexafluorophosphate N-oxide;HBTU:O-benzotriazolyl-A/,A/,A/',A/'-tetramethyluronium hexafluorophosphate;HCTU:1H-benzotriazolium 1-[bis(dimethylamino)methylene]-5-chloro-3-oxide hexafluorophosphate; HF: hydrogen fluoride; HOBt: hydroxybenzotriazole; IBCF: isobutyl chloroformate; iPr: isopropyl; IPA: isopropyl alcohol; Me: methyl;2Nal:2-naphthylalanine;NMM:N-methylmorpholine;NMP:A/-methyl-pyrrolidone;OtBu:tert-butyl ester;Pbf:2,2,4,6,7-pentamethyl-dihydrobenzofuran-5-sulfonyl;PBS:phosphate buffered saline;PyBOP:(benzotriazol-1-yloxy)-tris(pyrrolidino)-phosphonium. hexafluoro-phosphate; PyBrOP: bromotris(pyrrolidine) phosphonium hexafluorophosphate; tBu: tert-butyl; TFA: trifluoroacetic acid; THF: tetrahydrofuran; TIS: triisopropyl silane; Thos: p-toluenesulfonyl; Z: benzyloxycarbonyl; ZOSu: N-(benzyloxycarbonyl-oxy) succinimide.

[0059] The preparation of the compounds of the subject invention as described in the following examples are intended to be illustrative rather than limiting. In each of these examples, the observed molecular weight is reported as a deconvolution. The deconvolution value is taken from the formula MW(MT); observed)) = n(m/z)-n, where m/z represents the charged ion (positive form) and n is the number of charges of specific species. When multiply charged species are present in the mass spectrum, the observed molecular weight is given as a proxy.

[0060] The synthetic procedures provided in Examples 85-87, and the isotopic labeling procedures described in Example 89, for cyclic lactam peptides containing an isopropyl lysine side chain are equally applicable to other peptides described herein containing a lysine alkyl side chain, with appropriate modifications. The synthetic methods of Examples 86-88, which eliminate the need for a toxic palladium catalyst, are also equally applicable to the other peptides provided herein, with appropriate modifications.

Example 1

Ac-cyclorDap-Tvr-Arq-DArq-2Nal-Glv-Glu1-Arq-NH, (SEQ ID NO:2)

[0061] The sequence Ac-Dap(Alloc)-Tyr(tBu)-Arg(Pbf)-DArg(Pbf)-2Nal-GlyGlu(Oalyl)-Arg(Pbf) (SEQ ID NO:3) was assembled according to the standard Fmoc sequence using an ABI 431 Peptide Synthesizer (Applied Biosystems) as given in Scheme 1 below. Automated assembly was performed using the standard Applied Biosystems DCC/HOBt chemistry protocol or the FastMoc chemistry HBTU/DIEA protocol according to the client's instructions (PE Applied Biosystems Inc., Foster City, CA). The solid support is Rink amide resin (4-(2',4'-dimethoxyphenyl-Fmoc-aminomethyl)-phenoxy resin) for C-terminal amides or indole resin [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin for C-terminal ethyl amides (NovaBiochem, EMD Biosciences, Inc., San Diego, CA). The assembly (folding) of the chain in steps starts from the C-terminal end of the linear peptide and is accomplished in 9 main steps. In step 1, four equivalents of protected amino acids Fmoc-Arg(Pbf) were activated with DCC/HOBt (or HBTU/DIEA for FastMoc herniation) in NMP, and coupled to Rink Amide resin deprotected with 20% piperidine. In step 2, four equivalents of Fmoc-Glu(Oallyl) was activated and coupled to the deprotected peptide resin from step 1. Appropriate steps were performed up to step 8, Fmoc-Dap(Alloc) coupling. Next, the Fmoc at the N-terminal end was removed with 20% piperidine in DMF and acetylation of the α-amino group was performed with 5 equivalents of acetic anhydride, 10 equivalents of DIEA in dry DMF or NMP, for 1 h at room temperature.

[0062] The protecting groups of the ANI and Alloc side chains were removed using 0.1 equivalents of Pd(Ph3P)4 in the presence of 24 equivalents of phenylsilane in dichloromethane (Scheme 2). This process was repeated once more to complete the deprotection of the side chain. The deprotected carboxylic acid residue in Glu was activated with PyBOP/DIEA and cyclized to the front chain of the amino group of Dap on the resin. At the same time, protection was removed from the cyclized peptide and the peptide was separated (cleaved) from the resin using a mixture (scavenger cocktail) TF A/H20/TIS/EDT (95/2/1/2, z/z/z/z), or TFA/H20/TIS/anisole (92/2/4/2, z/z/z/z) for 2 h at room temperature. The solvents were then evaporated in vacuo, and the peptide was precipitated and washed three times with cold diethyl ether to remove the scavenger. Calculated molecular weight (MW calc.): 1142.30; MW observed (MW recorded): 1142.50.

[0063] Peptide purification was performed using standard preparative HPLC techniques. Immediately after cyclization, the peptide solution was diluted with water containing 0.1% (z/z) TFA, applied to a reversed-phase C18 HPLC column, and eluted with a gradient consisting of aqueous 0.1% trifluoroacetic acid/acetonitrile (z/z) with monitoring at 214 nm. Appropriate fractions were collected and lyophilized. Further characterization of the final product was performed using analytical HPLC and mass spectral analysis using conventional techniques. For peptides with a basic side chain, the final lyophilized product is the TFA salt.

Example 2

Ac- cyclorDab- Tvr- Ara- DArq- 2Nal- Glv- Glul- Arg- NH? (SEQ ID NO:6)

[0064] Prepare as given in Example 1, with the exception that Fmoc-Dap(Alloc) in step 8 is replaced by Fmoc-Dab(Alloc). MW calc.: 1156.33; MW note: 1156.10.

Example3

Ac-cyclorDap-Tvr-Lvs(iPr)-DArq-2Nal-Glv-Glu1-Arg-NH7 (SEQ ID NO:7)

[0065] Prepare as given in Example 1, with the exception that Fmoc-Arg(Pbf) in step 6 is replaced by Fmoc-Lys(iPr)(Boc). MVV calc.: 1156.37; MW note: 1156.78.

Example 4

n- Hexanoyl-cyclorDap- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- Glu1- Arq- NH, ( SEQ ID NO:8)

[0066] Prepare as given in Example 1, except that Fmoc-Arg(Pbf) in step 6 is replaced by Fmoc-Lys(iPr)(Boc). Additionally, the acetic anhydride in step 9 was replaced with hexanoic acid which was activated by PyBOP/DIEA. MVV calc.: 1212.47; MVV note: 1212.92.

Example 5

Ac-cyclor(D/L) Arq- Tvr- Arq- DArq- 2Nal- Glv- Glu1- Arq- NH, ( SEQ ID NO:9)

[0067] Prepare as given in Example 1, except that Fmoc-Glu(Oallyl) in step 2 is replaced by Fmoc-Glu(OtBu), and Fmoc-Dap(Alloc) in step 8 is replaced by Fmoc-(DL)Agl(Boc). After chain assembly, there is no need to treat with Pd(Ph3P)4 as no allyl protection is present. Instead, cyclization was performed in solution after cleavage of the linear peptide from the solid support (support) and deprotection. The crude linear peptide (0.25 mmol) resulting from cleavage was dried under vacuum and dissolved in 10 mL of dry DMF. This peptide solution was added to the following solution using a syringe pump over a period of 2 h: 15 mL dry dichloromethane and 15 mL dry DMF containing 1.0 mmol PyBOP and 4.0 mmol DIEA. The reaction was then allowed to proceed at room temperature for 2 h. The solvents were then evaporated in vacuo, the residue was applied to a preparative reverse phase C18 HPLC column, and the target cyclic peptide was isolated and characterized as described in Example 1. MVV calc.: 1128.27; MVV note: 1128.26.

Example 6

Ac- cyclorGlu- Tvr- Arq- DArq- 2Nal- Glv- Dap1- Arg- NH? (SEQ ID NO:10)

[0068] SequenceGlu(OtBu)-Tyr(tBu)-Arg(Pbf)-DArg(Pbf)-2Nal-Gly-Dap(Boc)-Arg(Pbf)

(SEQ ID NO:11) was assembled according to standard Fmoc herniation using an ABI 431 instrument as provided in Scheme 3 below. Automated assembly was performed according to the standard Applied Biosystems DCC/HOBt hernia protocol or the FastMoc hernia protocol (HBTU/DIEA) according to the manufacturer's instructions (PE Applied Biosystems Inc., FosterCity, CA). The solid support is Rink amide resin for C-terminal amides or indole resin [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin for C-terminal ethyl amide. The assembly (assembly) of the chain in steps starts from the C-terminal end of the linear peptide and takes place in 9 main steps. In step 1, four equivalents of protected amino acids Fmoc-Arg(Pbf) were activated with DCC/HOBt (or HBTU/DIEA for FastMoc herniation) in NMP, and coupled to deprotect the Rink Amide resin. In step 2, four equivalents of Fmoc-Dap(Boc) were activated and coupled to the deprotected resin from step 1. The corresponding steps were carried out until step 8, the Fmoc-Glu(OtBu) coupling. For step 9, the Fmoc at the N-terminal end was removed using 20% piperidine in DMF and acetylation of the α-amino group was performed offline by the addition of 5 equivalents of acetic anhydride, 10 equivalents of DIEA in dry DMF or NMP, for 1 h at room temperature. The final product is At the same time the protection was removed from the obtained peptide and the peptide was separated (cleaved) from the resin using a mixture (scavenger cocktail) TFA/H20/TIS/EDT (95/2/1/2, z/z/z/z), or TFA/H20/TIS/anisole (92/2/4/2, z/z/z/z) for 2 h at room temperature (Scheme 4). The solvents were evaporated in vacuo, and the peptide was precipitated and washed three times with cold diethyl ether to remove the scavenger. The crude product was used directly in the cyclization reaction. MW calc.: 1142.30; MVV note: 1142.83.

[0069] The cyclization was performed in solution after the linear peptide was separated from the solid support (support) and all side chains were deprotected (Scheme 4). The cleaved crude linear peptide (0.25 mmol) was dried under vacuum and dissolved in 10 mL of dry DMF. This peptide solution was added to the following reaction mixture using a syringe pump over a period of 2 h: 15 mL dry dichloromethane and 15 mL dry DMF containing 1.0 mmol PvBOP and 4.0 mmol DIEA. The reaction was then allowed to proceed at room temperature for 2 h. The solvents were evaporated in vacuo, the residue was applied to a preparative reverse phase C 18 HPLC column, and the target cyclic peptide was isolated and characterized as described in Example 1.

Scheme 4: Ring cyclization of the amino side chain and the carboxyl side chain

Example 7

Bz- cycloirGlu- Tvr- Arq- DArq- 2Nal- Glv- Dap1- Arq- NH? (SEQ ID NO:14)

[0070] Prepare as given in Example 6, except that the acetic anhydride in step 9 is replaced with benzoic anhydride. MVV calc.: 1204.37; MVV note: 1204.87.

Example 8

Ac-cyclorGlu-Tvr-Arg-DArq-2Nal-Glv-DDap1-Arq-NH, (SEQ ID NO:15)

[0071] Prepare as given in Example 6, except that Fmoc-Dap(Boc) in step 2 is replaced by Fmoc-DDap(Boc). MVV calc.: 1142.30; MVV note: 1142.73.

Example 9

Ac-cyclorGlu-Tvr-Arq-DArq-2Nal-Glv-Lvsl-Arq-NH, (SEQ ID NO:16)

[0072] Prepare as given in Example 6, except that Fmoc-Dap(Boc) in step 2 is replaced by Fmoc-Lvs(Boc). MVV Iraqi: 1184.38; MVV note: 1184.23.

Example 10

Ac-cyclorGlu-Tvr-Arq-DArq-2Nal-Glv-Pab1-Arq-NH, (SEQ ID NO: 17)

[0073] Prepare as in Example 6, except that Fmoc-Dap(Boc) in step 2 is replaced by Fmoc-Dab(Boc). MVV calc.: 1156.33; MVV note: 1156.07.

Example 11

Ac- cyclorGlu- Tvr- Arq- DArq- 2Nal- Glv-( D/ L) Agn- Arq- NH7 ( SEQ ID NO: 18)

[0074] Prepare as given in Example 6, except that Fmoc-Dap(Boc) in step 2 is replaced by Fmoc-(DL)Agl(Boc). MVV calc.: 1128.27; MVV note: 1128.86.

Example 12

Bz-cyclofGlu-Tvr-Arq-DArq-2Nal-Glv-(D/L)Agll-Arg-NH?(SEQ ID NO: 19)

[0075] Prepare as given in Example 6, except that Fmoc-Dap(Boc) in step 2 is replaced by Fmoc-(DL)Agl(Boc). Additionally, the anhydride in step 9 was replaced with benzoic acid anhydride. MVV calc.: 1190.34; MVV note: 1190.99.

Example 13

Bz-cyclorAsp-Tvr-Arq-DArg-2Nal-Glv-Dab1-Arq-NH7 (SEQ ID NO:20)

[0076] Prepare as in Example 6, except that Fmoc-Dap(Boc) in step 2 is replaced by Fmoc-Dab(Boc), and Fmoc-Glu(OtBu) in step 8 is replaced by Fmoc-Asp(OtBu). Additionally, acetic anhydride in step 9 was replaced with benzoic anhydride. MVV calc.: 1204.37; MVV note: 1204.87.

Example 14

Ac- cyclofAsp- Tvr- Arq- DArg- 2Nal- Glv- Dab1- Arq- NH? (SEQ ID NO:21)

[0077] Prepare as given in Example 6, except that Fmoc-Dap(Boc) in step 2 is replaced by Fmoc-Dab(Boc), and Fmoc-Glu(OtBu) in step 8 is replaced by Fmoc-Asp(OtBu). MVV calc.: 1142.30; MVV note: 1142.81.

Example 15

Ac-cyclofAsp-Tvr-Arg-PArq-2Nal-Glv-Dabl-Arq-NH, (SEQ ID NO:22)

[0078] Prepare as given in Example 6, except that Fmoc-Glu(OtBu) in step 8 is replaced by Fmoc-Asp(OtBu). MVV calc.: 1128.27; MVV note: 1128.78.

Example 16

Bz- cvclolAsp- Tvr- Ar2- DAr2- 2Nal- Glv- Dapl- Ar2- NH2 ( SEQ ID NO:23)

[0079] Prepare as given in Example 6, except that Fmoc-Glu(OtBu) in step 8 is replaced by Fmoc-Asp(OtBu). Additionally, acetic anhydride in step 9 was replaced with benzoic anhydride. MVV calc.: 1190.34; MVV note: 1190.69.

Example 17

Ac- cyclorAsp- Tvr- Arq- DAr2- 2Nal- Glv-( D/ L) Aqn- Arg- NH? (SEQ ID NO:24)

[0080] Prepare as given in Example 6, except that Fmoc-Dap(Boc) in step 2 is replaced by Fmoc-(DL)Agl(Boc), and Fmoc-Glu(OtBu) in step 8 is replaced by Fmoc-Asp(OtBu). MVV calc.: 1114.24; MVV note: 1114.85.

Example 18

Ac-cyclorAsp-Tvr-Lvs(Me7)-DArq-2Nal-Glv-Dap1-Arq-NH, (SEQ ID NO:25)

[0081] Prepare as given in Example 6, except that Fmoc-Arg(Pbf) in step 6 is replaced by Fmoc-l_ys(Me2), and Fmoc-Glu(OtBu)in in step 8 is replaced by Fmoc-Asp(OtBu). MVV calc.: 1128.31; MVV note: 1128.92.

Example 19

Bz- cyclofAsp- Tvr- Lvs( Me7)- DArg- 2Nal- Glv- Dap1- Arq- NH7 ( SEQ ID NO:26)

[0082] Prepare as given in Example 6, except that Fmoc-Arg(Pbf) in step 6 is replaced by Fmoc-Lys(Me2), and Fmoc-Glu(OtBu) in step 8 is replaced by Fmoc-Asp(OtBu). Additionally, acetic anhydride in step 9 was replaced with benzoic anhydride. MVV calc.: 1190.38; MVV note: 1191.14.

Example 20

cyclosuccinyl- Tvr- Arq- DArq- 2Nal- Glv-( D/ L) Aqn- Arq- NH7 ( SEQ ID NO:27)

[0083] Prepare as given in Example 6, except that Fmoc-Dap(Boc) in step 2 was replaced by Fmoc-(DL)Agl(Boc), Fmoc-Glu(OtBu) in step 8 was not used, and this step was omitted. Additionally, the acetic anhydride in step 9 was replaced with succinic anhydride. MVV calc.: 1057.19; MVV note: 1057.87.

Example 21

cyclosuccinyl- Tvr- Arq- DArq- 2Nal- Glv- Dap1- Arg- NH7 ( SEQ ID NO:28)

[0084] Prepare as in Example 6, except that Fmoc-Glu(OtBu) in step 8 was not used, and this step was omitted. Additionally, the acetic anhydride in step 9 was replaced with succinic anhydride. MW calc.: 1071.22; MVV note: 1071.85.

Example 22

cyclosuccinyl- Tvr- Arq- DArq- 2Nal- Glv- Dab1- Arq- NH, ( SEQ ID NO:29)

[0085] Prepare as given in Example 6, except that Fmoc-Dap(Boc) in step 2 was replaced by Fmoc-Dab(Boc), Fmoc-Glu(OtBu) in step 8 was not used, and this step was skipped. D Additionally, the acetic anhydride in step 9 was replaced with succinic anhydride. MVV calc.: 1085.25; MVV note: 1085.87.

Example 23

cyclosuccinyl- Tvr- Arq- DArg- 2Nal- Glv- Orn1- Ar,- NH? (SEQ ID NO:30)

[0086] Prepare as in Example 6, except that Fmoc-Dap(Boc) in step 2 was replaced by Fmoc-Orn(Boc), Fmoc-Glu(OtBu) in step 8 was not used, and this step was skipped. Additionally, the acetic anhydride in step 9 was replaced with succinic anhydride. MVV calc.: 1099.27; MVV note: 1100.23.

Example 24

cyclosuccinyl- Tvr- Arq- DArq- 2Nal- Glv- Lvs1- Arq- NH?( SEQ ID NO:31)

[0087] Prepare as in Example 6, except that Fmoc-Dap(Boc) in step 2 is replaced by Fmoc-Lys(Boc), Fmoc-Glu(OtBu) in step 8 is not used, and this step is omitted. Additionally, the acetic anhydride in step 9 was replaced with succinic anhydride. MVV calc.: 1113.30; MVV note: 1114.25.

Example 25

cyclorGlv- Tvr- Lvs( iPr)- DArg- 2Nal- Glv- DGIul- Arq- NH7 ( SEQ ID NO:32)

[0088] Sequence Gly-Tyr(tBu)-Lys(iPr)(Boc)-DArg(Pbf)-2Nal-Gly-DGIu(Alii)-Arg(Pbf)

(SEQ ID NO:33) was assembled according to standard Fmoc herniation using an ABI 431 instrument as provided in Scheme 5 below. Automated assembly was performed using the standard Applied Biosystems DCC/HOBt chemistry protocol or the FastMoc HBTU/DIEA chemistry protocol according to the manufacturer's instructions (PE Applied Biosystems Inc., Foster City, CA). The solid support is Rink amide resin for amides or indole resin [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin for C-terminal ethyl amides. The assembly (folding) of the chain in steps starts from the C-terminal end of the linear peptide and is carried out in 8 main steps (Scheme 5). In step 1, four equivalents of the protected amino acid Fmoc-Arg(Pbf) was activated with DCC/HOBt (or HBTU/DIEA for FastMoc herniation) in NMP, and coupled with deprotected Rink amide resin. In step 2, four equivalents of Fmoc-DGIu(Oallyl) was activated and coupled (coupled) to the peptide resin from step 1. The corresponding steps were carried out until step 8, coupling of Fmoc-Gly.

[0089] The protecting group of the allyl ester side chain was removed by adding 0.1 equivalents of Pd(Ph3P)4 in the presence of 24 equivalents of phenylsilane in dichloromethane (Scheme 6). This process was repeated once more to complete the deprotection of the side chain. The Fmoc at the N-terminal end was then removed by addition of 20% piperidine in DMF. The deprotected DGIu carboxylic acid residue was activated by the addition of PyBOP/DIEA, and cyclized with the α-amino group of glycine on the resin. At the same time, the protection was removed from the cyclized peptide and the peptide was removed from the resin using a mixture. scavenger cocktail) consisting of TFA/H20/TIS/EDT (95/2/1/2, z/z/z/z), or TFA/H20/TIS/anisole (92/2/4/2, z/z/z/z) for 2 h at room temperature. The solvents were then evaporated under vacuum, and the peptide was precipitated and washed three times with cold diethyl ether to remove the scavenger. MVV calcd.: 1085.29; MVV note: 1085.32.

Example 25a

cyclorGlv- Tvr- Lys( iPr)- DArg- 2Nal- Glv- DGIul- Arg- NH?- acetic acid salt ( SEQ ID

NO:34)

[0090] The TFA salt of the peptide from Example 25 was converted to the acetic acid salt by adsorbing the material onto a preparative C18 column of appropriate size, equilibrated with 2% acetic acid/H 2 O (z/z). The column was then washed with three to five column volumes of 2% aqueous acetic acid (w/w). The peptide was eluted with a 1:1 water/acetonitrile (z/z) solution containing 2% acetic acid (aq), and lyophilized. MVV calc.: 1085.29; MVV note: 1085.32.

Scheme 5: peptide binding of the α-amino group to the carboxyl side chain

[0091] Peptide purification was performed using standard preparative HPLC techniques. Immediately after cyclization, the peptide solution was diluted with water containing 0.1% (w/z) TFA, applied to a reversed-phase C18 HPLC column, and eluted with an aqueous 0.1% trifluoroacetic acid/acetonitrile (w/z) gradient with monitoring at 214 nm. Appropriate fractions were collected and lyophilized. Further characterization of the final product was performed using analytical HPLC and mass spectral analysis according to conventional methods. For peptides with basic side chains, the final lyophilized product is the TFA salt.

Example 26

cyclorGlv- Tvr- LvsriPr)- DArq- 2Nal- Glv- DGIu1- NH, fSEQ ID NO:37)

[0092] Prepare as given in Example 25, except that step 1 is omitted. MW calc.: 929.10; MVV note: 929.39.

Example 26a

socyclofGlv- Tvr- Lvs( iPr)- DArq.- 2Nal- Glv- DGIu1- NH7- acetic acid ( SEQ ID

NO:38)

[0093] The TFA salt of the peptide from Example 26 was converted to the acetic acid salt by adsorbing the material onto a preparative C18 column of appropriate size, equilibrated with a 2% acetic acid/H 2 O (z/z) solution. the column was then washed with three to five column volumes of 2% aqueous acetic acid (w/w). The peptide was eluted with a 1:1 water/acetonitrile (z/z) solution containing 2% acetic acid (aq), and lyophilized. MVV calc.: 929.10; MVV note: 929.39.

Example 27

cyclorGly- Tvr- Ar,- DAr,- 2Nal- Glv- DGIul- Ar,- NH, ( SEQ ID NO:391

[0094] Prepare as in Example 25, except that Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf). MVV calc.: 1071.22; MVV note: 1071.02.

Example 28

cyclofGlv- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- LvsnPr)- NH7 ( SEQ ID NO:40)

[0095] Prepare as in Example 25, except that Fmoc-Arg(Pbf) in step 1 is replaced by Fmoc-Lys(iPr)(Boc). MVV calc.: 1099.35; MVV note: 1099.91.

Example 29

cyclorTvr-Arq-DArq-2Nal-Glv-Glu1-Arq-NH, (SEQ ID NO:41)

[0096] Prepare as given in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf), Fmoc-Gly in step 8 is not used, and step 8 is omitted. MVV calc.: 1014.17; MVV note: 1014.78.

Example 30

cyclorTvr- Ar,- nAr,- 2Nal- Glv- DGIul- Ar7- NH?( SEQ ID NO:42)

[0097] Prepare as in Example 25, except that Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf), Fmoc-Gly in step 8 is not used, and step 8 is omitted. MVV calc.: 1014.17; MVV note: 1014.65.

Example 31

cyclorTvr- Arq- DAr,- 2Nal- Glv- Asp1- Ar,- NH, <SEQ ID NO:43)

[0098] Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Asp(allyl), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf), Fmoc-Gly in step 8 is not used, and step 8 is omitted. MVV calc.: 1000.14; MVV note: 1000.63.

Example 32

cyclorGlv- Tvr- Arg- DArq- 2Nal- Glv- Asp1- Ar,- NH, (SEQ ID NO:44)

[0099] Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Asp(allyl), and Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf). MVV calc.: 1057.19; MVV note: 1057.35.

Example 33

cyclorGlv- Tvr- Lvs( Me7)- DAr?- 2Nal- Glv- Asp1- Ar,- NH, (SEQ ID NO:45)

[0100] Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Asp(allyl), and Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Lys(Me 2 ). MVV calc.: 1057.23; MVV note: 1057.86.

Example 34

cyclorGlv- Tvr- LvrnPr)- DArq- 2Nal- Glv- Asp1- Arg- NH, (SEQ ID NO:46)

[0101] Prepare as given in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Asp(allyl). MVV calc.: 1071.26; MVV note: 1071.76.

Example 35

cyclorGlv- Tvr- LvsnPr)- DArg- 2Nal- Glv- Asp1- NH? (SEQ ID NO:47)

[0102] Prepare as given in Example 25, except that step 1 is omitted, and Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Asp(allyl). MVV calc.: 915.07; MVV note: 915.38.

Example 36

cyclorGlv- Tvr- LvsfiPr)- DArq- 2Nal- Glv- Asp1- Lvs( iPr)- NH, (SEQ ID NO:48)

[0103] Prepare as given in Example 25, except that Fmoc-Arg(Pbf) in step 1 is replaced by Fmoc-l_ys(iPr)(Boc), and Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Asp(allyl). MVV calc.: 1085.33; MVV note: 1085.78.

Example 37

cyclorGly-Tvr-Lvs(Me?)-nArq-2Nal-Glv-Glul-Arq-NH, (SEQ ID NO:49)

[0104] Prepare as given in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), and Fmoc-Lys(iPr)(Boc) is replaced by Fmoc-Lys(Me2) in step 6. MVV calc.: 1071.26; MVV note: 1071.05.

Example 38

cyclorGlv- Tvr- Arq- DArq- 2Nal- Clv- Glu1- Arq- NH, ( SEQ ID NO:50)

[0105] Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), and Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf). MVV calc.: 1071.22; MVV note: 1071.50.

Example 39

cyclorGlv- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- Glu1- Arg- NH, ( SEO ID NO:51)

[0106] Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl). MW calc.: 1085.29; MW note: 1085.91.

Example 40

cyclorGlv. Tvr- Lvs( iPr)- DArsi- 2Nal- Glv- Glu1- NH7 ( SEQ ID NO:52)

[0107] Prepare as in Example 25, except that step 1 is omitted, and Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl). MVV cal. ; 929.10; MVV note: 929.39.

Example 41

cyclorGlv- Tvr- LvsfiPr)- DArq- 2Nal- Glv- Asp1- NHEt ( SEQ ID NO:53)

[0108] Prepare as in Example 25, except that the Rink amide resin was replaced with [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin, step 1 was omitted, and Fmoc-DGIu(Oallyl) in step 2 was replaced with Fmoc-Asp(allyl). MVV calc.: 943.13; MVV note: 943.36.

Example 42

cyclorGlv- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- Glu1- NHEt ( SEQ ID NO:54)

[0109] Prepare as in Example 25, except that the Rink amide resin was replaced with [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin, step 1 was omitted, and Fmoc-DGIu(Oallyl) in step 2 was replaced with Fmoc-Glu(Oallyl). MVV calc.: 957.15; MVV note: 957.50.

Example 43

cyclofGlv- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- Glu1- NHEt ( SEQ ID NO:55)

[0110] Prepare as given in Example 25, except that the Rink amide resin is replaced with [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin, and step 1 is omitted. MVV calc.: 957.15; MVV note: 957.46.

Example 44

cycloFGIv- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- Arg- NHEt ( SEQ ID NO:56)

[0111] Prepare as given in Example 25, except that the Rink amide resin is replaced with [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin. MVV calc.: 1113.34; MVV note: 1113.81.

Example 44a

So cyclorGlv- Tvr- Lvs( iPr)- DAr2- 2Nal- Gly- DGIu1- Arg- NHEt acetic acid ( SEQ ID

NO: 57)

[0112] The peptide TFA salt of Example 44 was converted to the acetic acid salt by adsorbing the material onto a preparative C18 column of appropriate size, equilibrated with 2% acetic acid/H 2 O (z/z). the column was then washed with three to five column volumes of 2% aqueous acetic acid (w/w). The peptide was eluted with a 1:1 water/acetonitrile (z/z) solution containing 2% acetic acid (aq), and lyophilized. MVV calc.: 1113.34; MVV note: 1113.81.

Example 45

cyclorGlv- Tvr- Lvs( iPr)- PArg- 2N al- Glv- DGlu1- Lvs( iPr)- NHEt ( SEQ ID NO:58)

[0113] Prepare as given in Example 25, except that Rink amide resin is replaced with [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin, and Fmoc-Arg(Pbf) in step 1 is replaced with Fmoc-Lys(iPr)(Boc). MVV calc.: 1127.41; MVV note: 1127.35.

Example 46

cyclofLvs( Ac)- Tvr- Lvs( Me,)- DArq- 2Nat- Glv- Glu1- Arg- NH, (SEQ ID NO:59)

[0114] Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(OtBu), Fmoc-l_ys(iPr)(Boc) in step 6 is replaced by Fmoc-Lys(Me2), and Fmoc-Gly in step 8 is replaced by Boc-Lys(Fmoc). After chain assembly, the Fmoc on the N-terminal Lys side chain was removed using 20% piperidine in DMF and the Lys side chain was acetylated with 10 equivalents of acetic anhydride/DlEA at room temperature for one hour. All protecting groups on the side chain were removed and the linear peptide was cleaved from the solid support with a mixture of TFA/water/TIS/anisole (90/5/2.5/2.5, z/z/z/z) for 2 h at room temperature. Cyclization of the crude linear peptide was performed in solution. The cleaved crude linear peptide (-0.25 mmol) was dried under vacuum and dissolved in 10 mL of dry DMF. This peptide solution was added to the following reaction mixture using an injection pump over 2 h: 15 mL dry dichloromethane and 15 mL dry DMF containing 1.0 mmol PyBOP and 4.0 mmol DIEA. The reaction was then allowed to proceed at room temperature for 2 h. Solvents were evaporated in vacuo, the residue was applied to a reverse-phase C18 preparative HPLC column, and the cyanocyclic peptide was isolated and characterized as described in Example 1. MVV calcd.: 1184.42; MVV note: 1184.06.

Example 47

cyclorDap( Ac>- Tvr- Lvs( Me,)- DArq- 2Nal- Glv- Glul- NH7( SEQ ID NO:60)

[0115] Prepare as given in Example 25, except that column 1 is omitted, Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(OtBu), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Lys(Me2), and Fmoc-Gly in step 8 is replaced by Boc-Dap(Fmoc). After chain assembly, the Fmoc on the N-terminal Dap side chain was removed using 20% piperidine in DMF, and the Dap side chain was then acetylated with 10 equivalents of acetic anhydride/DIEA at room temperature for one hour. All side chain protecting groups were removed and the linear peptide was cleaved from the solid support using a mixture of TFA/water/TIS/anisole (90/5/2.5/2.5, z/z/z/z) for 2 h at room temperature. Cyclization of the crude linear peptide was carried out in solution. The cleaved crude linear peptide (~0.25 mmol) was dried under vacuum and dissolved in 10 mL of dry DMF. This peptide solution was added to the following reaction mixture using an injection pump over a period of 2 h: 15 mL dry dichloromethane and 15 mL dry DMF with 1.0 mmol PyBOP and 4.0 mmol DIEA. The reaction was then allowed to proceed at room temperature for 2 h. The solvents were then evaporated in vacuo, the residue was applied to a preparative HPLC column, and the target cyclic peptide was isolated and characterized as described in Example 1. MVV calcd.: 986.15; MVV note: 985.97.

Example 48

cycloirAla- Tvr- Lvs( iPh- DArg- 2Nal- Glv- Glu1- NH7 ( SEQ ID NO:61)

[0116] Prepare as in Example 25, except that step 1 is omitted, Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), and Fmoc-Gly in step 8 is replaced by Fmoc-Ala. MVV calc.: 943.13; MVV note: 942.92.

Example 49

cyclorDAIa- Tvr- Lvs( iPr)- DArg- 2Nal- Glv- Glul- NH7 ( SEQ ID NO:62)

[0117] Prepare as given in Example 25, except that step 1 is omitted, Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), and Fmoc-Gly in step 8 is replaced by Fmoc-DAla. MVV calc.: 943.13; MVV note: 943.44.

Example 50

cyclorDAIa- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- NH7 ( SEQ ID NO:63)

[0118] Prepare as in Example 25, except that step 1 is omitted, and Fmoc-Gly in step 8 is replaced by Fmoc-DAla. MVV calc.: 943.13; MVV note: 943.42.

Example 51

cyclorAla- Tvr- Lvs( iPr)- DArg- 2Nal- Glv- DGIu1- NH7 ( SEQ ID NO:64)

[0119] Prepare as in Example 25, except that step 1 is omitted, and Fmoc-Gly in step 8 is replaced by Fmoc-Ala. MVV calc.: 943.13; MVV note: 943.48.

Example 52

cyclorLeu- Tvr- Lvs( iPr)- DArg- 2Nal- Glv- DGIu1- NH, ( SEQ ID NO:65)

[0120] Prepare as in Example 25, except that step 1 is omitted, and Fmoc-Gly in step 8 is replaced with Fmoc-Leu. MVV calc.: 985.21; MVV note: 985.56.

Example 53

cyclorLeu- Tvr- Lvs( iPh- DArg- 2Nal- Glv- Glul- NH7 ( SEQ ID NO:66)

[0121] Prepare as in Example 25, except that step 1 is omitted, Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), and Fmoc-Gly in step 8 is replaced by Fmoc-Leu. MVV calc.: 985.21; MVV note: 985.49.

Example 54

cyclorDPhe- Tvr- LvsfiPr>- DArq- 2Nal- Glv- Glu1- NH?( SEQ ID NO:67)

[0122] Prepare as in Example 25, except that step 1 is omitted, Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), and Fmoc-Gly in step 8 is replaced by Fmoc-DPhe. MVV calc.: 1019.22; MVV note: 1019.52.

Example 55

cyclofPhe- Tvr- LvsnPr)- DArg- 2Nal- Glv- Glul- NH7-( SEQ ID NO:68)

[0123] Prepare as given in Example 25, except that step 1 is omitted, Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), and Fmoc-Gly in step 8 is replaced by Fmoc-Phe. MVV calc.: 1019.22; MVV note: 1019.53.

Example 56

cycloirDPhe- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- NH, ( SEQ ID NO:69)

[0124] Prepare as in Example 25, except that step 1 is omitted, and Fmoc-Gly in step 8 is replaced by Fmoc-DPhe. MVV calc.: 1019.22; MVV note: 1019.50.

Example 57

cyclorPhe- Tvr- Lvr( iPr)- DArq- 2Nal- Glv- DGIu1- Lvs( iPr)- NH?( SEQ IDNO:701

[0125] Prepare as in Example 25, except that Fmoc-Arg(Pbf) in step 1 is replaced by Fmoc-l_ys(iPr)(Boc), and Fmoc-Gly in step 8 is replaced by Fmoc-Phe. MVV calc.: 1189.48; MVV note: 1189.92.

Example 57a

with acetic acid cycloPhe- Tyr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- Lvs( iPr)- NH?-( SEQ ID NO:71)

[0126] The TFA salt of the peptide from Example 57 was converted to the acetic acid salt by adsorbing the material onto a preparative C18 column of appropriate size, equilibrated with 2% acetic acid/H 2 O (z/z). the column was then washed with three to five column volumes of 2% aqueous acetic acid (w/w). The peptide was eluted with 1:1 water/acetonitrile (z/z) containing 2% acetic acid (aq) and lyophilized. MVV calc.: 1189.48; MVV note: 1189.92.

Example 58

cycloPhe- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- Arg- NH? (SEQ ID NO:72)

[0127] Prepare as in Example 25, except that Fmoc-Gly in step 8 is replaced by Fmoc-Phe. MVV calc.: 1175.41; MVV note: 1175.81.

Example 59

cycloPhe- Tvr- Lvs( iPr)- DArg- 2Nal- Glv- DGIu1- NH, ( SEQ ID NO:73)

[0128] Prepare as in Example 25, except that step 1 is omitted, and Fmoc-Gly in step 8 is replaced by Fmoc-Phe. MVV calc.: 1019.22; MVV note: 1019.56.

Example 60

cyclofPhe- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- Lvs( iPr)- NHEt ( SEQ ID NO:74)

[0129] Prepare as in Example 25, except that Rink amide resin is replaced with [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin, Fmoc-Arg(Pbf) in step 1 is replaced with Fmoc-l_ys(iPr)(Boc), and Fmoc-Gly in step 8 is replaced with Fmoc-Phe. MVV calc.: 1217.53; MVV note: 1217.97.

Example 60a

Acetic acid salt cycloPhe- Tyr- LvsfiPr)- DArg- 2Nal- Glv- DGIu1- Lvs( iPr)- NHEt-( SEQ ID NO:75)

[0130] The TFA salt of the peptide from Example 60 was converted to the acetic acid salt by adsorbing the material onto a preparative C18 column of appropriate size, equilibrated with 2% acetic acid/H 2 O (z/z). The column was then washed with three to five column volumes of 2% aqueous acetic acid (w/w). The peptide was eluted with 1:1 water/acetonitrile (z/z) containing 2% acetic acid (aq) and lyophilized. MVV calc.: 1217.53; MVV note: 1217.97.

Example 61

cyclorDAIa- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- GluVNHEt ( SEQ ID NO. 76)

[0131] Prepare as in Example 25, except that Rink amide resin is replaced with [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin, step 1 is omitted, Fmoc-DGIu(Oallyl) in step 2 is replaced with Fmoc-Glu(Oallyl), and Fmoc-Gly in step 8 is replaced with Fmoc-DAla. MVV calc.: 971.18; MVV note: 971.49.

Example 62

cyclor2Nal- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- Glu1- NHEt ( SEQ ID NO:77)

[0132] Prepare as given in Example 25, except that Rink amide resin is replaced with [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin, step 1 is omitted, Fmoc-DGIu(Oallyl) in step 2 is replaced with Fmoc-Glu(Oallyl), and Fmoc-Gly in step 8 is replaced with Fmoc-2Nal. MVV calc.: 1097.34; MVV note: 1097.53.

Example 63

cyclorDPhe- Tvr- LvsfiPh- DArq- 2Nal- Glv- Glu1- NHEt ( SEQ ID NO:78)

[0133] Prepare as given in Example 25, except that Rink amide resin is replaced with [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin, step 1 is omitted, Fmoc-DGIu(Oallyl) in step 2 is replaced with Fmoc-Glu(Oallyl), and Fmoc- in step 8 is replaced with Fmoc-DPhe. MVV calc.: 1047.28; MVV note: 1047.51.

Example 64

cycloPhe- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- Glu1- NHEt ( SEQ ID NO:79)

[0134] Prepare as given in Example 25, except that Rink amide resin is replaced with [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin, step 1 is omitted, Fmoc-DGIu(Oallyl) in step 2 is replaced with Fmoc-Glu(Oallyl), and Fmoc-Gly in step 8 is replaced with Fmoc-Phe. MVV calc.: 1047.28; MVV note: 1047.57.

Example 65

cyclorGlv- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIuVGIv- 2Nal- NH, ( SEQ ID NO:80)

[0135] Prepare as in Example 25, except that Fmoc-Arg(Pbf) in step 1 was replaced by Fmoc-2Nal, and one step was added between steps 1 and 2 where Fmoc-Gly was used. MVV calc.: 1183.39; MVV note: 1183.26.

Example 66

cyclorGlv- Tvr- Lvs( iPr)- DArg- 2Nal- Glv- Glv- B- Ala- D2Nal- NH, ( SEQ ID NO:81)

[0136] Prepared as in Example 25, except that Fmoc-Arg(Pbf) in step 1 was replaced by Fmoc-D2Nal, one step was added between steps 1 and 2 using Fmoc-(3-Ala), and Fmoc-DGIu(Oallyl) in step 2 was replaced by Fmoc-Glu(Oallyl). MVV calc.: 1197.40; MVV obv.: 1196.70.

Example 67

cyclor6- Ala- Tvr- Arg- DArq- 2Nal- Glv- Glu1- Arq- NH? (SEQ ID NO:82)

[0137] Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf), and Fmoc-Gly in step 8 is replaced by Fmoc-p-Ala. MVV calc.: 1085.25; MVV note: 1085.05.

Example 68

cyclorB- Ala- Tvr- Arq- DArg- 2Nal- Glv- Aspl- Ara- NH, (SEQ ID NO:83)

[0138] Prepare as in Example 25, except that FmOC-DGIu(Oallyl) in step 2 is replaced by Fmoc-Asp(Oallyl), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf), and Fmoc-Gly in step 8 is replaced by Fmoc-p-Ala. MVV calc.: 1071.22; MVV note: 1071.15.

Example 69

cyclor5- aminovaleryl- Tvr- Arg- DArq- 2Nal- Glv- Glul- Arg- NH?( SEQ ID NO:84)

[0139] Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf), and Fmoc-Gly in step 8 is replaced by Fmoc-5-aminovaleric acid. MVV calc.: 1113.30; MVV note: 1113.40.

Example 70

cyclof5- aminovaleryl- Tvr- Arq- DArq- 2Nal- Glv- Asp1- Arq- NH? (SEQ ID NO:85)

[0140] Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Asp(Oallyl), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf), and Fmoc-Gly in step 8 is replaced by Fmoc-5-amino valeric acid. MVV calc.: 1099.27; MVV note: 1100.25.

Example 71

cyclo(4- AMPA)- Tvr- Arq- DArq- 2Nal- Glv- AspT- Arg- NH, (SEQ ID NO:86)

Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Asp(Oallyl), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf), and Fmoc-Gly in step 8 is replaced by Fmoc-4-aminomethyl phenylacetic acid (4-AMPA). MVV calc.: 1147.32; MVV note: 1148.20.

Example 72

cyclor( 4- AMPA)- Tvr- Arq- DArq- 2Nal- Glv- Glu1- Arq- NH? (SEQ ID NO:87)

Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf), and Fmoc-Gly in step 8 is replaced by Fmoc-4-aminomethyl phenylacetic acid (4-AMPA). MVV calc.: 1161.34; MVV note: 1161.99.

Example 73

cyclo(4- AMPA)- Tvr- Arq- DArq- 2Nal- Glv- Glu1- DArg- NH7 ( SEQ ID NO:88)

[0143] Prepare as in Example 25, except that Fmoc-Arg(Pbf) in step 1 is replaced by Fmoc-DArg(Pbf), Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf), and Fmoc-Gly in step 8 is replaced. Fmoc-4-aminomethyl phenylacetic acid (4-AMPA). MVV calc.: 1161.34; MVV note: 1161.83.

Example 74

cyclo(4- AMB)- Tvr- Arg- DArq- 2Nal- Glv- Glu1- Arq- NH, (SEQ ID NO. 89)

[0144] Prepare as in Example 25, except that Fmoc-DGIu(Oallyl) in step 2 is replaced by Fmoc-Glu(Oallyl), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Arg(Pbf), and Fmoc-Gly in step 8 is replaced by Fmoc-4-aminomethyl benzoic acid (4-AMB). MVV calc.: 1147.32; MVV note: 1147.66.

Example 75

cyclorGlv- Tvr- Lvs( iPr)- DArg- 2Nal- Glv- DGIu1- Lvs( iPr)- Glv- 2Nal- NH, ( SEQ ID NO:90)

[0145] Prepare as given in Example 25, except that step 1 is replaced by three sequential coupling residues: first Fmoc-2Nal, then Fmoc-Gly, and then Fmoc-Lys(iPr)(Boc). MVV calc.: 1353.69; MVV note: 1354.03.

Example 76

cyclorPhe- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- Lvs( iPr)- Glv- 2Nal- NH, ( SEO ID NO:91)

[0146] Prepare as given in Example 25, except that step 1 is replaced by three sequential coupling residues: first Fmoc-2Nal, then Fmoc-Gly, and finally Fmoc-Lys(iPr)(Boc). Additionally, Fmoc-Gly in step 8 was replaced by Fmoc-Phe. MVV calc.: 1443.82; MVV note: 1444.13.

Example 77

cyclorGlv- Tvr- Lvs( iPrt- DArq- 2Nal- Glv- DGIu1- Glv- DPhe- NH, ( SEQ ID NO:92)

[0147] Prepare as given in Example 25, except that step 1 is replaced by two sequential coupling residues: first Fmoc-DPhe, then Fmoc-Gly. MVV calc.: 1133.36; MVV note: 1133.73.

Example 78

cyclorGlv- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- Lvs( iPr)- DPhe- NH7 ( SEQ ID NO:93)

[0148] Prepare as in Example 25, except that step 1 is except that step is Fmoc-DPhe, then Fmoc-Lys(iPr)(Boc). MW calc.: 1246.58; MVV note: 1246.88.

Example 79

cyclorLvs- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIul- Lvs( iPr)- NH7 ( SEQ ID NO:94)

[0149] Prepare as in Example 25, except that Fmoc-Arg(Pbf) in step 1 is replaced by Fmoc-Lys(iPr)(Boc), and Fmoc-Gly in step 8 is replaced by Fmoc-Lys(Boc). MVV calc.: 1170.50; MVV note: 1169.80.

Example 80

cycloPhe- Tvr- Lvr- DArq- 2Nal- Glv- DGIu1- LvsfiPr)- NH, (SEQ ID NO:95)

[0150] Prepare as in Example 25, except that Fmoc-Arg(Pbf) in step 1 is replaced by Fmoc-Lys(iPr)(Boc), Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Lys(Boc), and Fmoc-Gly in step 8 is replaced by Fmoc-Phe. MVV calc.: 1147.40; MVV note: 1146.70.

Example 81

cycloPhe- Tvr- LvsfiPh- DArq- 2Nal- Glv- DGIul- Lvs- NH, ( SEQ ID NO:96)

[0151] Prepare as in Example 25, except that Fmoc-Arg(Pbf) in step 1 is replaced by Fmoc-Lys(Boc), and Fmoc-Gly in step 8 is replaced by Fmoc-Phe. MVV calc.: 1147.40; MVV note: 1146.70.

Example 82

cyclofPhe- Tvr- Lvs( iPh- DArq- 2Nal- Glv- DGIu1- Orn- NH, ( SEQ ID NO:97)

[0152] Prepare as in Example 25, except that Fmoc-Arg(Pbf) in step 1 is replaced by Fmoc-Orn(Boc), and Fmoc-Gly in step 8 is replaced by Fmoc-Phe. MVV calc.: 1133.40; MVV note: 1132.70.

Example 83

cycloPhe- Tvr- Lvs- DArq- 2Nal- Glv- nGlu1- Lvs- NH7 ( SEQ ID NO:98)

[0153] Prepare as in Example 25, except that each of Fmoc-Arg(Pbf) in step 1 and Fmoc-Lys(iPr)(Boc) in step 6 is replaced by Fmoc-Lys(Boc), and Fmoc-Gly in step 8 is replaced by Fmoc-Phe. MVV calc.: 1105.37; MVV note: 1105.40.

Example 84

cycloPhe- Tvr- Lvs- DArq- 2Nal- Glv- DGIu1- Lvs- NHEt ( SEQ ID NO:99)

[0154] Prepare as in Example 25, except that Rink resin is replaced with [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin, each of Fmoc-Arg(Pbf) in step 1 and Fmoc-Lys(iPr)(Boc) in step 6 is replaced with Fmoc-Lys(Boc), and Fmoc-Gly in step 8 is replaced with Fmoc-Phe. MW calc.: 1133.36; MVV note: 1133.82.

Example 85

Alternative synthesis of I cyclorPhe- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- LvsnPr)- NH,

(SEO ID NO:70)

[0155] Example 57 described in the synthesis of SEQ ID NO:70 via heimian peptide synthesis on Fmoc solid phase using the commercially available building blocks Fmoc-Lys(iPr)Boc, which is expensive and difficult to obtain in large quantities. The process described in this example enables the synthesis of SEQ ID NO:70 by using the less expensive Fmoc-Lys(Boc), a cyclization solvent, and alkylating the lysine via reductive amination using sodium cyanoborohydride, thus providing a more economical route to the final product. Additional advantages are that the reaction medium (acetic acid, acetone, and methanol) are relatively inexpensive, the reaction conditions are easily controlled, the solvent ratio can be varied significantly without affecting the alkylation reaction, and the yield is 90% or more.

[0156] The sequence Phe-Tyr(tBu)-Lys(Boc)-DArg(Pbf)-2Nal-Gly-DGIu(Oalyl)-Lys(Boc) (SEQ ID NO:100) was assembled on Rink Amide resin according to standard Fmoc herniation using an ABI 431 Peptide Synthesizer as given in Scheme 7. Automated assembly was performed according to Applied Biosystems standard. DCC/HOBt chemistry protocol or FastMoc chemistry (HBTU/DIEA) protocol according to the supplier's instructions (PE Applied Biosystems Inc., Foster City, CA). The scheme of protecting groups on the side chain is: Lys(Boc), DGIu(Oalyl), DArg(Pbf), Tyr(tBu). Chain assembly in steps starts from the C-terminal end of the linear peptide and is performed in 8 steps. In step 1, four equivalents of the protected amino acid Fmoc-Lys(Boc) was activated with DCC/HOBt (or HBTU/DIEA for FastMoc herniation) in NMP, and coupled to a deprotected Rink amide resin. In step 2, four equivalents of Fmoc-DGIu(Oallyl) was activated and coupled to the deprotected peptide resin from step 1. These steps are repeats up to step 8, Fmoc-Phe coupling.

[0157] The protecting group of the allyl ester side chain was removed using 0.1 equivalents of Pd(Ph3P)4 in the presence of 24 equivalents of phenylsilane in dichloromethane (Scheme 8). This process was repeated once to complete the oiling of the protection from the side chain. The Fmoc at the N-terminal end was then removed with 20% piperidine in DMF. The deprotected DGIu carboxylic acid residue was activated with PyBOP/DIEA and cyclized with the α-amino group of Phe on the resin. Simultaneously, the cyclic peptide was deprotected and the peptide was cleaved from the resin using scavenger cocktails TFA/H2G7TIS/EDT (95/2/1/2, z/z/z/z) or TFA/H20/TIS/anisole (92/2/4/2, z/z/z/z) for 2 h at room temperature. The solvents were then evaporated in vacuo, and the peptide was settled and washed three times with cold diethyl ether in order to remove the scavenger cocktail.

Scheme 7: Assembly (assembly) of the peptide chain on the solid phase

[0158] Purification of the cyclic peptide precursor was achieved according to standard preparative HPLC techniques. The crude cleaved product was dissolved in a minimal amount of DMSO, applied to a reverse-phase C18 HPLC column, and eluted with a 0.1% aqueous trifluoroacetic acid/acetonitrile (z/z) gradient with monitoring at 214 nm. The releasing fractions were collected and lyophilized. Further characterization of the cyclic peptide intermediate precursor was performed by analytical HPLC and mass spectral analysis according to conventional techniques.

[0159] The lyophilized cyclic peptide precursor was then alkylated in a solution of acetic acid/acetone/methanol (1:1:4, z/z/z) via reductive amination using sodium cyanoborohydride (Scheme 9). The peptide concentration is around 10mg/mL, and can vary significantly without affecting the results. Three to 5 equivalents of the reducing agent sodium cyanoborohydride were used, and the reaction was normally completed within 2 h at room temperature. The yield is 90% or more. For example, 20 mg of cyclic peptide precursor was dissolved in 2 mL of methanol, to which 0.5 mL of acetic acid and 0.5 mL of acetone were added, mixed, and then 5.6 mg of sodium cyanoborohydride (2.5 equivalents in methanol) was added with stirring. The reaction mixture was stirred at room temperature for 30 min, after which another 5.6 mg of sodium cyanoborohydride was added. The reaction was monitored with HPLC and mass spectral analysis. After the completion of the reaction, removing the salt from the reaction mixture and lyophilization gave 18.9 mg of the final product (SEQ ID NO:70) with a purity of 97.5%. MVV calc.: 1189.48; MVV note: 1189.6.

[0160] The compound from Example 60 (SEQ ID NO:74) can also be obtained in an analogous way, with appropriate modifications based on the amino acids included in the composition, using [3-({ethyl-Fmoc-amino}-methyl)-indol-1-yl]-acetyl AM resin. First, the precursor peptide (SEQ ID NO:99) was prepared as described in Example 84, and then reductive amination was performed as shown in Scheme 9. This gave the final cyclic C-terminal ethyl amide peptide with a purity of 99.16%. MVV Iraqi: 1217.53; MVV note: 1217.84.

Example 86

Alternative synthesis of HcyclofPhe- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- Lvs( iPr)- NH,

(SEQ ID NO. 701

[0161] Example 85 describes the inexpensive synthesis of SEQ ID NO:70 via solid phase synthesis of Fmoc peptides using relatively inexpensive Fmoc-Lvs(Boc) and alkylation of lysine via reductive amination with sodium cyanoborohydride in relatively inexpensive solvents (acetic acid, acetone, and methanol). However, this process involves the use of palladium as a heavy metal catalyst in order to remove the protecting groups from the allyl ester side chains. Palladium is highly toxic, and quality control to ensure complete removal of this element is complicated and difficult. The Boe solid phase peptide synthesis process with cyclization on resin described in this example allows the preparation of SEQ ID NO:70 using the relatively inexpensive Boc-Lys(2-CI-Z) without the need for a toxic and expensive palladium catalyst, providing an even more economical, less toxic way to obtain an easily upgradable end product that requires a simpler quality control process.

[0162] The sequence Fmoc-Phe-Tyr(2-Br-Z)-Lys(2-CI-Z)-DArg(Tos)-2Nal-GlyDGIu(OFm)-Lys(2-CI-Z) (SEQ ID NO:102) was manually assembled on MBHA (4-methyl-benzhydryl-amine) resin (Cat. No. D-2095, BaChem Inc., Torrance, California). CA) according to established peptide synthesis herniation on Boe solid phase (Schnolzer et al. (1992) Int. J. Pept. Protein Res. 40:180-193) as given in Scheme 10. Chain assembly was performed according to the in situ neutralization/HBTU/DlEA activation process procedure as described in ref. The scheme of side chain protecting groups is: Lys(2-CI-Z), DGIu(OFm), DArg(Tos), and Tyr(2-Br-Z). The alpha-amino group of all constituent amino acid blocks is protected with tert-butoxycarbonyl (Boe) except for the N-terminal Phe residue, which is protected with Fmoc for synthesis efficiency. Chain assembly in steps starts from the C-terminal end of the linear peptide and is accomplished in 8 steps. In step 1, five equivalents of the protected amino acids Boc-Lys(2-Cl-Z) were activated with HBTU/DIEA in DMF, and coupled with MBHA resin. In step 2, five equivalents of Boc-DGIu(OFm) were activated and coupled to the deprotected peptide resin from step 1 with neat TFA. These steps were repeated as needed until step 8, coupling of Fmoc-Phe.

[0163] The protecting group for the Fm side chain in DGI, along with the Fmoc at the N-terminal end, was removed with 20% piperidine in DMF. The deprotected carboxylic acid residue in DGI is activated with PyBOP/DIEA, HCTU/DIEA, or other suitable activation reagents, and cyclized with the α-amino group of Phe on the resin. At the same time, the cyclic peptide was deprotected and the peptide was cleaved from the resin using HF with 5% m-cresol ilip-/frezol as scavenger for 1 h at 0 °C. The solvents were then evaporated and the crude peptide was settled and washed three times with cold diethyl ether.

[0164] Purification of the cyclic peptide precursor (SEQ ID NO:98 as a snapshot is provided in Scheme 10) was achieved using standard preparative HPLC techniques. The crude cleaved product was dissolved in a minimal amount of DMSO, applied to a reverse phase C18 HPLC column, and eluted with an aqueous 0.1% trifluoroacetic acid/acetonitrile (z/z) gradient with monitoring at 214 nm. Appropriate fractions were collected and lyophilized. Further characterization of the cyclic peptide intermediate precursor was performed by analytical HPLC and mass spectral analysis using conventional techniques. For SEQ

IDNO:98, MVV calc.: 1105.29; MVV note: 1105.4.

[0165] The lyophilized cyclic peptide precursor (SEQ ID NO:98) was then alkylated in a solution of acetic acid/acetone/methanol (1:1:4, z/z/z) via reductive amination using sodium cyanoborohydride as given in Scheme 9. The concentration of the peptide is about 10mg/ml_, and can vary significantly without affecting the results. Five equivalents of the reducing agent sodium cyanoborohydride were used, and the reaction was normally completed in 2 h at room temperature. The yield is 90% or more. For example, 6.6 mg of cyclic peptide precursor was dissolved in 0.8 mL of methanol, to which 0.2 mL of acetic acid and 0.2 mL of acetone were added, mixed well, and then 1.9 mg of sodium cyanoborohydride (2.5 equivalents in methanol) was added in two equal portions with stirring. The reaction mixture was stirred at room temperature for 30 min, after which another 1.9 mg of sodium cyanoborohydride was added. The reaction was monitored by HPLC and mass spectral analysis. When the reaction was finished, the salt was removed from the reaction mixture and the final product (SEQ ID NO:70) with a purity of 96.5% was obtained by lyophilization. MVV calc.: 1189.45; MVV note: 1189.6.

Example 87

Alternative synthesis of III cyclorPhe- Tvr- Lvs( iPr)- DArg- 2Nal- Glv- DGIul- Lys( iPr)- NH?

(SEQIDNO:70)

[0166] The compound of Example 57 (SEQ ID NO:70) can also be obtained without the use of a palladium catalyst via a cyclization solution, improved, as follows.

[0167] The sequence Boc-Phe-Tyr(2-Br-Z)-Lys(Fmoc)-DArg(Tos)-2Nal-GlyDGIu(OBzl)-Lys(Fmoc) (SEQ ID NO:103) was manually assembled on MBHA resin by Boe hernia solid phase peptide synthesis (Schnolzer et al. (1992) Int. J. Pept. Protein Res. 40:180-193) as indicated in Scheme 11. Chain assembly was performed using an in situ neutralization/HBTU/DlEA activation procedure as described by Schnolzer et al. The side chain protecting group scheme is as follows: Lys(Fmoc), DGIu(OBzl), DArg(Tos), Tyr(2-Br-Z). The alpha-amino group of all amino acids as building blocks are protected with tert-butoxycarbonyl (Boe). Stepwise chain assembly starts from the C-terminal end of the linear peptide and is accomplished in 8 steps as shown in Scheme 11. In step 1, five equivalents of the protected amino acids Boc-Lys(2-CI-Z) were activated with HBTU (4 eq) /DIEA (10 eq) in DMF, and coupled with MBHA resin. In step 2, five equivalents of Boc-DGIu(OBzl) were activated and coupled to the peptide resin from step 1 which was deprotected by addition of pure TFA. These steps are repeated an adequate number of times until step 8, the Boc-Phe coupling. The Boe protecting group was removed by addition of neat TFA, the resin was neutralized with DIEA, and washed with DMF and methanol and air-dried before cleavage with HF. At the same time, protection was removed from the linear peptide and the peptide was cleaved from the resin using HF with 5% m-cresol or p-cresol as scavenger for 1 h at 0 °C. The solvents were then evaporated and the crude peptide was settled and washed three times with cold diethyl ether.

[0168] Purification of the linear precursor peptide (SEQ ID NO: 104) was achieved using standard preparative HPLC techniques. The crude product that was cleaved was dissolved in a minimal amount of DMSO, applied to a reverse phase C18 HPLC column, and eluted with a gradient of 0.1% aqueous trifluoroacetic acid/acetonitrile (z/z) with monitoring at 214 nm. Appropriate fractions were collected and lyophilized. Further characterization of the cyclic peptide intermediate precursor was performed using analytical HPLC and mass spectral analysis using conventional techniques. For SEQ ID NO: 104, MVV calc.: 1567.78; MVV note: 1567.6.

[0169] Cyclization of the lyophilized linear peptide precursor (SEQ ID NO: 104) was performed in solution (Scheme 12). The linear peptide was dissolved in a small amount of dry DMF (-10 mg/mL). This peptide solution was slowly added via injection pump to a reaction mixture of PyBOP (2 eq., or other suitable activation reagents, such as HCTU, BOP, HBTU, etc.) and DIEA (10 eq.) in dry DMF with stirring on a magnetic stirrer. This reaction was allowed to proceed at room temperature for 2 h. Then pure piperidine was added to the reaction mixture to a final concentration of 25% (w/w). The reaction mixture was stirred for another 20 min until the Fmoc protection was completely removed. Solvents were evaporated in vacuo and the residue was applied to a preparative reverse phase C18 HPLC column, and eluted with a gradient of 0.1% aqueous trifluoroacetic acid/acetonitrile (z/z) with monitoring at 214 nm. Appropriate fractions were collected and lyophilized, and the cyclic precursor peptides (SEQ ID NO:98) were obtained. Further characterization of the intermediate cyclic peptide precursor was performed using analytical HPLC and mass spectral analysis using conventional techniques. For SEQ ID NO:98, MVV calc.: 1105.29; MVV note: 1105.4.

[0170] Alkylation of the cyclic peptide SEQ ID NO:98 was carried out in a solution of acetic acid/acetone/methanol (1:1:4, z/z/z) through reductive amination with sodium cyanoborohydride as in Scheme 9 to obtain the final product (SEQ ID NO:70). MVV calc.: 1189.45; MVV note: 1189.6.

Example 88

Alternative synthesis of IV cyclorPhe- Tvr- Lvs( iPr)- DArq- 2Nal- Glv- DGIu1- Lvs( iPrW

NH,,(SEQ ID NO:70)

[0171] The compound of Example 57 (SEQ ID NO:70) can also be prepared without the use of a palladium catalyst by the synthesis process given in Scheme 13 below.

[0172] The dipeptide Fmoc-DGIu-Lys(iPr,Z)-NH2 was first obtained in solution with the Dglutamic acid side chain exposed. The dipeptide was bound to a hyper-acid labile CTC (2-chlorotritylchloride PS resin, 1% DVB (100-200 mix) resin (Senn Chemicals USA Inc., San Diego, CA; catalog number 40207), and the terminal peptide product was then synthesized on this resin via standard Fmoc synthesis as previously described. Selective removal of the peptide from the CTC resin allows only the Dglutamic acid side chain to react with N-terminus of the peptide in solution and produce a cyclic peptide product.The remaining side chains are then cleaved with 95% TFA or another strong acid.

[0173] The dipeptide Fmoc-DGIu-Lys(iPr,Z)-NH2 was first obtained as given below (Scheme 14):

[0174] Boc-Lys(iPr,Z)-OH was reacted with NMM and IBCF in THF. After addition of NH 4 OH, the solvents were evaporated on a rotary evaporator and ethyl acetate was added to the product. The ethyl acetate phase was washed with 5% NaHCO 3 and then with 0.1 N HCl, and then dried over anhydrous sodium sulfate. The sodium sulfate was evaporated by filtration and the ethyl acetate was removed by evaporation under reduced pressure. The resulting Boc-Lys(iPr,Z)-NH2 was dissolved in DCM, and TFA was added. Upon completion of the reaction, the solvents were evaporated on a rotary evaporator. H-Lys(iPr,Z)-NH2 was then dissolved in DMF. The pH was adjusted to 8 with DIEA. In a separate vessel, Fmoc-DGIu(OtBu)-OH, HBTU, and HOBt were dissolved in DMF; DIEA was added to adjust the pH to 8. The two solutions were mixed, and the reaction was monitored by C18 reverse phase HPLC. The pH was monitored and adjusted, if necessary, with DIEA. The solvents were evaporated on a rotary evaporator, and the product was dissolved in ethyl acetate. The ethyl acetate phase was washed thoroughly with 5% NaHCO 3 and then with 0.1 N HCl, and then dried over anhydrous sodium sulfate. The sodium sulfate was filtered off and the ethyl acetate was evaporated under reduced pressure. Evaporation on a rotary evaporator was continued until a dry residue was formed. The resulting Fmoc-DGIu(OtBu)-Lys(iPr,Z)-NH2 was dissolved in DCM, and TFA was added. upon completion of the reaction, the solvents were evaporated on a rotary evaporator. The solid product was obtained by trituration with diethyl ether. After washing the precipitate with ether, the product was dried in a vacuum oven.

[0175] The solid phase peptide synthesis of the final product was performed as follows, Fmoc-DGIu(OtBu)-Lys(iPr,Z)-NH2 was dissolved in DCM and reacted with CTC resin in the presence of DIEA in a reaction vessel. After 3 h, the peptide-resin was washed from the reagent with DCM and added to Z-OS. The pH was monitored and adjusted to pH 8-9 with the addition of DIEA as needed. After 8 h, the peptide-resin was washed from the reagents with DCM, transferred to a polypropylene dish, and dried in a vacuum oven.

[0176] The protected peptide resin was assembled using Fmoc-chemistry as follows. The following coupling cycle was applied: 1) De-blocking: reaction with 25% piperidine in DMF; 2) washing cycles with DMF, IPA and again DMF; 3) Ninhydrin test (qualitative: if positive, go to Step 4 of coupling); 4) coupling with 2 equivalents of Fmoc-amino acid in the presence of HOBt/DIC in DMF; 5) DMF washing cycles; 6) Ninhydrin test (qualitative: if negative, go to the next de-blocking/coupling cycle; if positive, continue re-coupling Step 7; if slightly positive, go to step 10 of acetylation); 7) Re-coupling (if necessary), with 1 equivalent of Fmoc-amino acid in the presence of HOBt, HBTU/DIEA in DMF; 8) Washing cycles with DMF; 9) Ninhydrin test (qualitative: if negative, go to the next de-blocking/coupling cycle; if positive, go to Step 10 of acetylation); 10) Acetylation (if needed) with 2% acetic anhydride in 4% DIEA in DMF; 11) Wash cycles (with DMF, IPA, and again DMF); 12) Ninhydrin test (quantitative: if positive, continue the next unblocking/coupling cycle). After the final coupling cycle, the peptide resin (SEQ ID NO:105) was washed with ether and dried under vacuum.

[0177] The protected peptide resin was washed with DCM. Separation (cleavage) of the fully protected linear peptide from the resin was performed with 2% TFA in DCM and then profile titrated. The solvents were evaporated on a rotary evaporator, and the fully protected linear peptide (SEQ ID NO: 106) was precipitated by trituration with ether. Fully shielded linear

The peptide was transferred to a polypropylene container and dried in a vacuum oven.

[0178] The fully protected linear peptide was cyclized in the presence of PyBOP, HOBt, and DIEA in DMF. The pH was maintained between pH 7-8 with the addition of DIEA, as needed. After completion of the reaction, the solvents were evaporated in a rotary evaporator, and ethyl acetate was added to the product. The ethyl acetate phase was washed sequentially with 5% NaHCO 3 and then with 0.1 N HCl and saturated NaCl solution. It was then dried over anhydrous sodium sulfate. The sodium sulfate was removed by filtration and the ethyl acetate was evaporated on a rotary evaporator under reduced pressure. The protected cyclic peptide (SEQ ID NO:107) was precipitated by trituration with ether and dried in a vacuum oven.

[0179] Deprotection was performed in TFA:H 2 O:TIS. When the reaction was complete, the solvents were evaporated in a rotary evaporator and the cyclic peptide (SEQ ID NO:70.) was precipitated by trituration with ether and dried in a vacuum oven. MVV calc.: 1189.48; MVV note: 1189.50.

Example 89

Incorporation of isotopic markers: Synthesis of cyclorPhe- Tvr- Lys( iPr- dK)- DArg- 2Nal- Glv-PGIul- Lvs( iPr- cU- NH, ( SEQ iD NO:108)

[0180] Starting from isotopically labeled (marked) acetone such as 13C-, 14C-, deuterium-, or tritium-labeled acetones as given below, the processes given in Examples 85-87 enable site-specific isotopic labeling of cyclic peptide CXCR4 antagonists for various pharmacological and imaging studies. imaging) examinations. Isotopically labeled acetones are commercially available from various sources. The example given below uses acetone-d6 to produce a peptide containing 12 deuterium atoms. The resulting compound, which differs in molecular weight by 12 Da compared to the unlabeled copy, is easily distinguished in the mass spectrum and shows receptor affinity for the identical target.

[0181] Using any of the methods of Examples 85-87, one skilled in the art can prepare and purify cyclic peptide precursors (SEQ ID NO:98, SEQ ID NO:99, etc.). Alkylation was carried out in acetic acid/acetone/methanol (1:1:4, z/z/z) via reductive amination using sodium cyanoborohydride as in Scheme 9, except that the standard acetone was replaced with the desired isotopically labeled acetone. In this example, deuterium acetone-d6 was used.

[0182] Peptide (97 mg) was dissolved in 15 mL of acetic acid/acetone-d6/methanol (1:1:4, z/z/z). The peptide concentration can vary significantly without affecting the results. Five equivalents of sodium cyanoborohydride were used, and the reaction was normally completed within 2 h at room temperature. The reaction was monitored with HPLC and mass spectral analysis. After the completion of the reaction, salt removal from the reaction mixture and lyophilization yielded 90.5 mg of the final product (SEQ ID NO: 108) with a purity of 99.9%. MVV calc.: 1201.48; MVV note: 1201.7.

[0183] The use of isotopically labeled sodium cyanoborohydride (such as NaBD3CN, NaBT3CN, etc.) allows the incorporation of additional variants of site-specific labeling methods.

[0184] The pharmacological properties of these compounds can be determined using the tests described below.

Human CXCR4/<125>1-SDF-1a binding inhibition assay

[0185] The binding of SDF-1 to CXCR4 is the first step in the activation of the CXCR4 intracellular signaling pathway. To determine the ability of Compound to block the interaction of SDF-1 and CXCR4, human leukemic CCRF-CEM cells (ATCC CCL 119) expressing endogenous CXCR4 were used in a 125 I -labeled SDF-1a binding assay. The test was performed on an untreated 96-well U-bottom polystyrene plate (Corning Incorporated, Costar, No. 3632). Binding assay buffer was prepared with RPMI 1640 medium (Gibco, Grand Island, NY) containing 10 mM HEPES, pH 7.5, and 0.2% BSA. Briefly, 200 pL of a reaction mixture containing 300 pM SDF ligand (60 pM<125>l-SDF-1a (Perkin Elmer) and 240 pM cold SDF-1a (R&D Systems), various concentrations of test compounds in buffer, 100,000 human CCRF-CEM cells, and 0.5 mg SPA beads (VVheatgerm agglutinin beads; Amersham) was incubated at room temperature for 2 hr. Plates were then read using a 1450 Microbeta Luminescence Counter (VVallac) in a dose-dependent manner. CXCR4 antagonists reduced bound radioactivity in a dose-dependent manner.

[0186] All of the previously described compounds have an average K value of about 7.5 nM or less in this assay. For example, the compound of Example 1 shows an average K value of 3.45 nM in this assay. Many of these compounds show an average K value between 0.2 nM and about 1 nM. For example, the compound of Example 50 shows an average K value of 0.285 nM in this assay. Other compounds show an average K value of less than about 0.2 nM. For example, the compound of Example 75 exhibits an average K value of 0.096 nM in this assay.

Chemotaxis test

[0187] CXCR4/SDF-1 interaction regulates migration (chemotaxis) of cells bearing CXCR4 on their surface. To determine the antagonistic and cellular activity of the test compounds, a chemotaxis assay was applied using human histocytic leukemia cells U937 (ATCC CRL 1593) expressing endogenous CXCR4. Briefly, U937 cells, grown in DMEM medium (Gibco, Grand Island, NY) containing 10% FBS, 1% MEM sodium pyruvate (Gibco), 1% MEM nonessential amino acids (Gibco), and 1% GlutaMAX 1

(Gibco), were collected and washed once with assay buffer prepared with 1x RPMI medium (Gibco) containing 10 mM HEPES, pH 7.5, and 0.3% BSA. After washing, cells were resuspended in assay buffer at a concentration of 5x10<6> cells/mL. The assay was performed on a 96-well ChemoTx plate (NeuroProbe) according to the manufacturer's instructions. Generally, 50 µL of cell mixture with or without test compound was applied to the upper chamber, and 30 µL of SDF-1a (R&D Systems, 10 ng/mL) prepared in 1x buffer was added to the lower chamber. After assembly, the plate was incubated for 2.5 hr at 37°C in 5% CO 2 . After incubation, 5 µL of CellTiter 96 AQ (Promega, Madison, WI) was added to the lower chamber. The plate was then incubated for 60 min at 37°C, and migrated cells were detected by measuring absorbance at 492 nm using a Liquid Spectrafluor Plus Microplate Reader (Salzburg, Austria). CXCR4 antagonists inhibit cell migration, reducing absorbance readings. The inhibitory potency (IC50) of the test compounds in this assay was calculated using GraphPad Prism software, based on the dose-dependent decrease in absorbance at 492 nm.

[0188] Most of the compounds described above have an average IC 50 value of about 60 nM or less in this assay. A number of compounds have average IC 50 values of about 6 nM or less, eg, the compounds of Example 19 have an average IC 50 value of 2.05 nM in this assay. A number of compounds have average IC 50 values of about 0.6 nM or less, eg, the compounds of Example 50 have average IC 50 values of 0.171 nM in this assay.

Chemokine receptor binding selectivity assay

[0189] The binding selectivity of the subject compounds to CXCR4 receptors compared to other chemokine receptors, such as human CCR1, CCR2, CXCR2, or CXCR3,

and other G-protein-coupled receptors, can be determined in cells transfected with nucleic acids that encode and express these receptors, or in cells in which these receptors are endogenously expressed. Whole cells or membrane fragments can be used to determine the competition of test compounds with appropriate ligands for these receptors in a manner similar to that previously described for the CXCR4/<125>I-SDF-1 binding inhibition assay.

[0190] For example, the compound of Example 57a exhibits K values greater than 73,000 nM in a ligand binding assay using the human chemokine receptor CXCR2.

Compound-induced white blood cell and neutrophil mobilization in C57BL/6

mice

[0191] Stem cells in the bone marrow actively maintain the continuous production of these mature blood cell lines throughout life. The bone marrow is the primary site for the production of white blood cells (WBC)/neutrophils and their release into the bloodstream. The CXCR4/SDF-1 axis is critical for the retention and release of VVBCs, neutrophils, and hematopoietic stem cells in the bone marrow, and disruption of the CXCR4/SDF-1 interaction in the bone marrow results in increased numbers of these cells in the peripheral blood. A short-term VVBC/neutrophil mobilization model in mice can be applied to determine the in vivo target-modulating activity of test compounds. Briefly, pathogen-free female C57BL/6 (Taconic) age 5-6 mice were housed for at least one week prior to testing. The animals were given access to sterilized rodent food and acidified water. Groups of 5 mice were injected subcutaneously with the test compound in saline, or saline as a control, and were then asphyxiated with CO 2 and cervical dislocation was performed at various time intervals after compound administration. Peripheral blood was collected by cardiac puncture using EDTA-coated syringes and pipettes. Complete blood analysis was performed using a Hemavet Mascot hematology analyzer (Drew Scientific Group, Dallas, TX). Recorded: VVBCs, neutrophils, and lymphocytes in peripheral blood. Effective CXCR4 antagonists administered subcutaneously to mice increase the number of peripheral blood neutrophils and WBCs compared to controls.

[0192] A significant number of the above compounds have an average neutrophil ratio (ratio of neutrophil increase in treated group vs. neutrophil increase in control group (saline)), measured 3 h after compound administration, greater than about 2 in this assay. For example, the compound of Example 39 has an average neutrophil ratio of 4.6 at a dose of 5 mg/kg in this trial.

Anti-tumor activity in the SCID/Namalwa xenograft model

[0193] SDF-1/CXCR4 interaction plays an important role in numerous stages of tumorigenesis, including tumor growth, invasion, angiogenesis, and metastasis. To determine the in vivo anti-tumor activity of the test compounds, a tumor xenograft model on NOD/SCID mice (Jackson Laboratories) and human non-Hodgkin's lymphoma Namalwa cells (ATCC CRL 1432) were used. Briefly, 200,000 Namalwa cells mixed with matrigel (1:1) were subcutaneously implanted on the hind flank of the animals. The implanted tumor cells grow as solid tumors, the dimensions of which can be continuously monitored and measured using a vernier caliper. To determine the in vivo efficacy of the test compound in this model, the expert can treat the animals (1 O/group) with different doses of the test compound dissolved in saline or PBS, starting 48 hours after tumor cell implantation. Compounds were dosed subcutaneously, and tumor volumes and body weights were determined every 2 or 3 days. Examinations generally last 3-4 weeks, depending on tumor growth. The anti-tumor growth (development) activity of the test compound was determined by the percent reduction in tumor volume in the treated groups compared to the tumor volume in the control groups treated with smao vehicle.

[0194] Several of the previously described compounds, for example the compound of Example 26, significantly inhibited tumor growth in this assay when administered at a dose of 1 mg/kg BID.

[0195] Pharmacological properties such as compound bioavailability, in vivo metabolic stability, and pharmacokinetic/pharmacodynamic properties can be determined by methods known in the art of drug development. Preferred compounds of the present invention exhibit high bioavailability when administered subcutaneously. Some of the compounds described herein have a bioavailability of about 100% in rats, for example the compound of Example 44. Preferred compounds also exhibit good in vivo metabolic stability. For example, no metabolites were observed in the plasma and urine of dogs and monkeys, up to 24 h after administration of the compound of Example 57a. Preferred compounds also exhibit favorable pharmacokinetic/pharmacodynamic properties that allow for convenient dosing. For example, in mice, the half-life (T1/2) of the compound of Example 58 is about 3 h. In terms of pharmacodynamic properties, preferred compounds induce prolonged mobilization of neutrophils and white cells in mice. For example, the compound of Example 25 induces a significant increase in peripheral blood neutrophils and white blood cells in mice for at least 6 h after a single dose of 5 mg/kg administered subcutaneously.

Claims (13)

1. Lactam-cyclized peptide of formula I: Ri-cyclo[Xi-Tyr-X3-DArg-2Nal-Gly-X7]-X8-X9-X10-R2 (SEQ ID (I) NO:1) characterized by the fact that: a) given lactam is formed by forming an amide bond between the side chain of the amino group in Xti of the side chain of the carboxyl group X7, where Xi'\X7, a pair selected from the group consisting of (D/L)Agl/Glu, Dab/Glu, and Dap/Glu, i.e. Ac or n-hexanoyl; or b) a given lactam is formed by forming an amide bond between the side chain of the carboxyl group Xi and the side chain of the amino group X7, where X, and X7 are a pair selected from the group consisting of Asp/(D/L)Agl, Asp/Dab, Asp/Dap, Glu/(D/L)Agl, Glu/Dab, Glu/Dap, Glu/DDap, and Glu/Lys, and Rije are Ac or Bz, or where Xi and X7 are a pair selected from the group consisting of succinyl/(D/L)Agl, succinyl/Dab, succinyl/Dap, succinyl/Lys, and succinyl/Om, iRije absent; or c) the given lactam is formed by the formation of an amide bond between the a-amino group and the side chain of the carboxyl group X7, where Xi and X7 are a pair selected from the group consisting of Ala/Glu, Ala/DGlu, DAla/Glu, DAla/DGlu, Dap(Ac)/Glu, Gly/Asp, Gly/Glu, Gly/DGIu, Leu/Glu, Leu/DGlu, Lys/DGIu, Lys(Ac)/Glu, 2Nal/Glu, Phe/Glu, Phe/DGlu, DPhe/Glu, and DPhe/DGlu, and R: is absent; or d) a given lactam is formed by the formation of an amide bond between the non-a amino group of the non-side chain of NaX^ and the side chain of the carboxyl group of X7, where Xii X7, a pair selected from the group consisting of P-Ala/Asp, p-Ala/Glu, 5-amino-valeryl/Asp, 5-aminovaleryl/Glu, 4-AMB/Glu, 4-AMPA/Asp, and 4-AMPA/Glu, and R, is absent; or e) the given lactam is formed by the formation of an amide bond between the a-amino group of X2 and the side chain of the carboxyl group of X7, where X2 and X7 are a pair selected from the group consisting of Tyr/Asp, Tyr/Glu, and Tyr/DGIu, and each of Rii Xi is absent; the substituent on the a-amino group of X when X contains n a-amino group and the given a-amino group is not an integral part of the lactam amide bond, selected from the group consisting of Ac, Bz, and n-hexanoyl, or is absent, wherein Xiodabran is from the group consisting of (D/L)Agl, Asp, Dab, Dap, and Glu; X is selected from the group consisting of (D/L)Agl, Ala, P-Ala, DAla, 5-aminovaleryl, 4-AMB, 4-AMPA, Asp, Dab, Dap, Dap(Ac), Glu, Gly, Leu, Lys, Lys(Ac), 2Nal, Phe, DPhe, and succinyl, or is absent; X3 is selected from the group consisting of Arg, Lys, Lys(iPr), and Lys(Me2); X7 is selected from the group consisting of (D/L)Agl, Asp, Dab, Dap, DDap, Glu, DGIu, Lys, and Om; X8 is selected from the group consisting of P-Ala, Arg, DArg, Gly, Lys, Lys(iPr), and Om, or is absent; X9 is selected from the group consisting of Gly, 2Nal, D2Nal, and DPhe, or is absent; X10 is 2Nal, or absent; wherein, when X8 is absent, then each of Xg and X10 is absent, and when X9 is absent, then X10 is absent, and R2 is selected from the group consisting of NH2 and NHEt, or a pharmaceutically acceptable salt thereof. 2. Lactam-cyclized peptide or its pharmaceutically acceptable salt from claim 1, characterized in that: It is selected from the group consisting of Ac and Bz, or is absent; Xi is selected from the group consisting of P-Ala, 4-AMB, 4-AMPA, Asp, Dab, Dap, Dap(Ac), Glu, 2Nal, Phe, and succinyl, or is absent; X3 is selected from the group consisting of Arg, Lys, Lys(iPr), and Lys(Me2); X7 is selected from the group consisting of Asp, Dab, Dap, Glu, DGIu, Lys, and Orn; X8 is selected from the group consisting of Arg and Lys, or is absent; X9 is absent; X10 is absent; and R 2 is selected from the group consisting of NH 2 and NHEt. 3. Lactam-cyclized peptide or its pharmaceutically acceptable salt from claim 1, characterized in that: It is selected from the group consisting of Ac and Bz, or is absent; X is selected from the group consisting of DAla, 5-aminovaleryl, 4-AMPA, Asp, Glu, Leu, Lys(Ac), Phe, DPhe, and succinyl; X3 is selected from the group consisting of Arg, Lys, Lys(iPr), and Lys(Me2); X7 is selected from the group consisting of (D/L)Agl, Asp, Dab, Dap, DDap, Glu, and Glu; X8 is selected from the group consisting of Arg, DArg, and Lys, or is absent; X9 is absent; Xio is absent; and R 2 is selected from the group consisting of NH 2 and NHEt. 4. Lactam-cyclized peptide or its pharmaceutically acceptable salt from claim 1, characterized in that: It is selected from the group consisting of Ac, Bz, and n-hexanoyl, or is absent; X-, is selected from the group consisting of (D/L)Agl, Ala, P-Ala, Asp, Dap, Glu, Gly, Lys, and Phe; X3 is selected from the group consisting of Arg, Lys, Lys(iPr), and Lys(Me2); X7 is selected from the group consisting of D/L)Agl, Asp, Dap, Glu, and DGIu; X8 is selected from the group consisting of p-Ala, Arg, Gly, Lys, Lys(iPr), and Om, or is absent; X9 is selected from the group consisting of Gly, 2Nal, D2Nal, and DPhe, or is absent; X10 is 2Nal, or absent; and R 2 is selected from the group consisting of NH 2 and NHEt. 5. Lactam-cyclized peptide or its pharmaceutically acceptable salt from claim 1, characterized in that: It is selected from the group consisting of Ac and Bz, or is absent; X is selected from the group consisting of Ala, 5-aminovaleryl, Asp, Glu, Gly, Phe, DPhe, and succinyl; X3 is selected from the group consisting of Arg, Lys(iPr), and Lys(Me2); X7 is selected from the group consisting of (D/L)Agl, Asp, Dap, Glu, and Glu; X8 is selected from the group consisting of P-Ala, Arg, Gly, Lys, Lys(iPr), and Om, or is absent; X9 is selected from the group consisting of Gly, D2Nal, and DPhe, or is absent; X10 is 2Nal, or absent; and R 2 is selected from the group consisting of NH 2 and NHEt. 6. The lactam-cyclized peptide or its pharmaceutically acceptable salt according to claim 1, 4, or 5, characterized in that: Xi is selected from the group consisting of Gly and Phe; X3 is Lys(iPr); and X7 is DGIu. 7. The lactam-cyclized peptide or its pharmaceutically acceptable salt according to claim 5, characterized in that: Ri is absent; Xi is selected from the group consisting of Gly and Phe; X3 is Lys(iPr); X7 is DGIu; X8 is selected from the group consisting of Arg and Lys(iPr), or is absent; X9 is absent; X10 is absent; and R 2 is selected from the group consisting of NH 2 and NHEt. 8. Lactam-cyclized peptide of the formula: or a pharmaceutically acceptable salt thereof. 9. The lactam-cyclized peptide of claim 8, characterized in that it is a pharmaceutically acceptable salt, an acetic acid salt. 10. A pharmaceutical composition characterized in that it contains a lactam-cyclized peptide or its pharmaceutically acceptable salt according to one of claims 1-9, and a pharmaceutically acceptable carrier, diluent, or excipient. 11. A lactam-cyclized peptide or a pharmaceutically acceptable salt thereof according to one of claims 1-9, for use in treatment. 12. Lactam-cyclized peptide or its pharmaceutically acceptable salt according to one of claims 1-9, for use in the treatment of rheumatoid arthritis, pulmonary fibrosis, HIV infection, or cancer selected from the group consisting of breast cancer, pancreatic cancer, melanoma, prostate cancer, kidney cancer, neuroblastoma, non-Hodgkin's lymphoma, lung cancer, ovarian cancer, colorectal cancer, multiple myeloma, glioblastoma multiforme, and chronic lymphocytic leukemia. 13. Use of a lactam-cyclized peptide or a pharmaceutically acceptable salt thereof according to one of claims 1-9, for the production of a drug for the treatment of rheumatoid arthritis, pulmonary fibrosis, HIV infection, or a cancer selected from the group consisting of breast cancer, pancreatic cancer, melanoma, prostate cancer, kidney cancer, neuroblastoma, non-Hodgkin's lymphoma, lung cancer, ovarian cancer, colorectal cancer, multiple myeloma, glioblastoma multiforme, and chronic lymphocytic leukemia.