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WO2024215130A1 - Nouveau dégradeur de gspt1 et son utilisation - Google Patents

Nouveau dégradeur de gspt1 et son utilisation Download PDF

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Publication number
WO2024215130A1
WO2024215130A1 PCT/KR2024/004911 KR2024004911W WO2024215130A1 WO 2024215130 A1 WO2024215130 A1 WO 2024215130A1 KR 2024004911 W KR2024004911 W KR 2024004911W WO 2024215130 A1 WO2024215130 A1 WO 2024215130A1
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Prior art keywords
compound
cancer
inhibitor
gspt1
alkyl
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Inventor
Wooseok Han
Joonwoo NAM
Min Sung Joo
Dong Hyuk KI
Eun-Jung Kim
JaeYung LEE
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Cyrus Therapeutics Inc
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Cyrus Therapeutics Inc
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Priority to CN202480025398.6A priority Critical patent/CN121002006A/zh
Priority to AU2024255772A priority patent/AU2024255772A1/en
Publication of WO2024215130A1 publication Critical patent/WO2024215130A1/fr
Priority to MX2025012045A priority patent/MX2025012045A/es
Priority to IL323841A priority patent/IL323841A/en
Anticipated expiration legal-status Critical
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/445—Non condensed piperidines, e.g. piperocaine
    • A61K31/4523—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems
    • A61K31/454—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems containing a five-membered ring with nitrogen as a ring hetero atom, e.g. pimozide, domperidone
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/535—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one oxygen as the ring hetero atoms, e.g. 1,2-oxazines
    • A61K31/5375—1,4-Oxazines, e.g. morpholine
    • A61K31/5377—1,4-Oxazines, e.g. morpholine not condensed and containing further heterocyclic rings, e.g. timolol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings directly linked by a ring-member-to-ring-member bond

Definitions

  • the present invention relates to a novel GSPT1 degrader and a use thereof. Specifically, the present invention relates to a compound represented by Formula I, a stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof; a pharmaceutical composition for treating a disorder of uncontrolled cell proliferation comprising the same; and a method of treating a disorder of uncontrolled cell proliferation by administering the same to a mammal.
  • the traditional drug development strategy is to directly regulate the activity of the protein by binding the drug to the specific active site of disease-related proteins such as enzymes, receptors, transmitters, and membrane proteins. Therefore, transcription factors, scaffold proteins, aggregates, and the like in the nucleus remain undruggable targets for drug development due to the lack of a docking pocket structure. According to the Human Protein Atlas, there are FDA-approved drugs for only about 15% of the approximately 4,500 known disease-related proteins.
  • Targeted protein degradation techniques seek to treat diseases by removing disease-related proteins using the protein degradation system existing in the body. Intracellular protein degradation is accomplished by lysosomes and proteasomes, and approximately 80% of cellular proteins are labeled with ubiquitin by the ubiquitin-proteasome system (UPS) and then degraded by proteasome.
  • UPS ubiquitin-proteasome system
  • Ubiquitin is a protein composed of 76 amino acids.
  • E1, E2, and E3 ubiquitin ligases are involved in the process of labeling ubiquitin for protein degradation by UPS (ubiquitination), and the labeled protein is degraded by 26S proteasome, which is an ATP-dependent protein degradation enzyme complex.
  • E3 ubiquitin ligase binds to both E2 ligase and substrate proteins and is responsible for recognizing substrate proteins to be labeled with ubiquitin.
  • Drugs using the TPD strategy can be classified into PROTAC (proteolysis targeting chimera) and molecular glues depending on the compound structure.
  • PROTAC proteolysis targeting chimera
  • molecular glues use the characteristic of small molecule compounds to induce the formation of specific protein complexes to promote the interaction between the target protein and E3 ligase involved in protein degradation, thereby degrading the target protein.
  • the molecular glues have a much lower molecular weight than PROTAC, and therefore have the advantage of enabling the development of therapeutic agents with excellent pharmacokinetic properties.
  • IiD immunomodulatory imide
  • thalidomide thalidomide
  • lenalidomide thalidomide
  • pomalidomide binds to cereblon (CRBN), a substrate receptor for CRL4 E3 ubiquitin ligase (Ito et al., Science 327:1345-1350 (2010)).
  • the present inventors found that when a specific substituent is present at each position of the phenylsulfonamide ring bound to the pomalidomide structure, the persistence in GSPT1 degradation and pH stability, as well as anticancer efficacy and safety, are significantly improved, and excellent anticancer efficacy could be achieved against a cancer with a neuroendocrine phenotype.
  • R 1 is halogen, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, C 1 -C 6 haloalkyl, or C 1 -C 6 haloalkoxy;
  • R 2 is C 1 -C 6 alkyl optionally substituted with R 2a ;
  • R 2a is hydroxy, halogen, C 1 -C 6 alkoxy, C 1 -C 6 haloalkoxy, or -NR'R'';
  • R' and R'' are each hydrogen or C 1 -C 6 alkyl, or R' and R'' may be taken together with the nitrogen atom to which they are attached to form a 3 to 8-membered heterocyclic ring optionally comprising one additional heteroatom selected from N, O, and S; and
  • R 3 is hydrogen or halogen.
  • a pharmaceutical composition for treating a disorder of uncontrolled cell proliferation in a mammal comprising a compound represented by Formula I, a stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof.
  • a method of regulating a cereblon activity or a GSPT1 activity in at least one cell comprising contacting the at least one cell in vitro with a compound represented by Formula I, a stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof.
  • a method of treating a disorder of uncontrolled cell proliferation in a mammal comprising administering a compound represented by Formula I, a stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof to a mammal.
  • the compound according to the present invention exhibits high selectivity and sustained degradation activity against GSPT1, excellent pH stability, and low cytotoxicity against normal cells, and thus has excellent anticancer efficacy and a high therapeutic index.
  • the compound according to the present invention can exhibit excellent anticancer activity against a cancer with a neuroendocrine phenotype, such as small cell lung cancer (SCLC), lung neuroendocrine cancer (NEC), and neuroendocrine prostate cancer (NEPC).
  • SCLC small cell lung cancer
  • NEC lung neuroendocrine cancer
  • NEPC neuroendocrine prostate cancer
  • Figure 1 illustrates a result obtained by measuring the GSPT1 degradation activity of Reference Compound 1 and example compounds on various neosubstrates by concentration.
  • Figure 2 illustrates a result obtained by observing the change in GSPT1 expression level over time when treated with Reference Compound 1 and example compounds (100 nM).
  • Figure 3 illustrates a result obtained by measuring the real time proliferation of cancer cells (NCI-H1155 cells) after treatment with Reference Compound 1 and example compounds.
  • Figure 4 illustrates a result obtained by performing TMT labeling proteomics analysis after treating the HL60 cell line with example compounds.
  • Figure 5 illustrates a result obtained by observing the GSPT1 expression level and the protein translation rate over time after treating the NCI-H1155 cells with 1 ⁇ M of Compound 3.
  • Figure 6 illustrates a result obtained by observing the GSPT1 expression level, the N-MYC expression level, and the protein translation rate (left); and a result obtained by measuring the expression levels of GSPT1, N-MYC, and ATF-4 and the degree of activation of caspase 3 (right), over time after treating the NCI-H1155 cells with 1 ⁇ M of Compound 3.
  • Figure 7 illustrates a result obtained by measuring the GSPT1 expression level, the ATF-4 expression level, and the degree of activation of caspase 3 over time after treating the HL60 cells with 0.3 ⁇ M of Compound 3.
  • Figure 8 illustrates a result obtained by observing the GSPT1 expression level and the protein translation rate over time after treating the NCI-H2023 cells with 1 ⁇ M of Reference Compound 1 and Compound 3.
  • Figure 9a illustrates a result obtained by measuring the cell viability after treating various small cell lung cancer (SCLC) and lung adenocarcinoma (LUAD) cell lines with Compound 3 and Compound 4.
  • SCLC small cell lung cancer
  • LAD lung adenocarcinoma
  • Figure 9b illustrates a result obtained by calculating the EC 50 of Compound 3 and Compound 4 for lung adenocarcinoma (LUAD), small cell lung cancer (SCLC), lung neuroendocrine cancer (NEC), and neuroendocrine prostate cancer (NEPC) cell lines.
  • LAD lung adenocarcinoma
  • SCLC small cell lung cancer
  • NEC lung neuroendocrine cancer
  • NEPC neuroendocrine prostate cancer
  • Figure 10 illustrates the therapeutic index (TI) of each compound calculated based on the cell viability of Reference Compound 1, Compound 3, and Compound 4 measured in the NCI-H1155, HL60 and HeKa cells.
  • Figure 11 illustrates the change in bioluminescence (Figure 11a) and the change in body weight (Figure 11b) over time after administration of Compound 3 in the HL-60-Luc AML animal model.
  • Figure 12a illustrates a result obtained by observing the expression levels of GSPT1 and N-MYC 6 and 24 hours after administration of Compound 4 in the NCI-H1155 lung cancer animal model ( Figure 12a); and a result obtained by measuring the change in tumor volume over time after administration of Compound 4 ( Figure 12b).
  • Figure 13 illustrates a result obtained by measuring the change in tumor volume (Figure 13a) and the change in body weight (Figure 13b) over time after administration of Compound 3 at various dosage and administration cycles.
  • R 1 is halogen, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, C 1 -C 6 haloalkyl, or C 1 -C 6 haloalkoxy;
  • R 2 is C 1 -C 6 alkyl optionally substituted with R 2a ;
  • R 2a is hydroxy, halogen, C 1 -C 6 alkoxy, C 1 -C 6 haloalkoxy, or -NR'R'';
  • R' and R'' are each hydrogen or C 1 -C 6 alkyl, or R' and R'' may be taken together with the nitrogen atom to which they are attached to form a 3 to 8-membered heterocyclic ring optionally comprising one additional heteroatom selected from N, O, and S; and
  • R 3 is hydrogen or halogen.
  • R 1 is halogen, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, C 1 -C 6 haloalkyl, or C 1 -C 6 haloalkoxy.
  • R 1 is halogen, C 1 -C 3 alkyl, C 1 -C 3 alkoxy, C 1 -C 3 haloalkyl, or C 1 -C 3 haloalkoxy.
  • R 1 may be halogen, C 1 -C 6 alkyl, C 1 -C 6 haloalkyl, or C 1 -C 6 haloalkoxy.
  • R 1 may be halogen, C 1 -C 3 alkyl, C 1 -C 3 haloalkyl, or C 1 -C 3 haloalkoxy. In one embodiment, R 1 may be halogen or C 1 -C 6 alkyl. In one embodiment, R 1 may be halogen or C 1 -C 3 alkyl. In one embodiment, R 1 may be a halogen (F, Cl, Br, or I). For example, R 1 includes F, Cl, Br, I, -OCH 2 F, -OCHF 2 , -OCF 3 , -CH 2 F, -CHF 2 , -CF 3 , or -CH 3 , but is not limited thereto.
  • R 2 is C 1 -C 6 alkyl or C 1 -C 6 alkyl substituted with R 2a .
  • R 2 may be C 1 -C 6 alkyl.
  • R 2 may be C 1 -C 3 alkyl.
  • R 2 may be C 1 -C 6 alkyl substituted with R 2a .
  • R 2 may be C 1 -C 3 alkyl substituted with R 2a .
  • R 2a may be hydroxy, halogen, C 1 -C 6 alkoxy, or C 1 -C 6 haloalkoxy.
  • R 2a may be hydroxy, halogen, C 1 -C 3 alkoxy, or C 1 -C 3 haloalkoxy. In one embodiment, R 2a may be hydroxy or C 1 -C 6 haloalkoxy. In one embodiment, R 2a may be hydroxy or C 1 -C 3 haloalkoxy.
  • R 2 includes -CH 3 , -CH 2 CH 3 , -CH 2 OCF 3 , -CH 2 OCHF 2 , -CH 2 OCH 2 F, -CH 2 OH, or -CH 2 CH 2 OH, but is not limited thereto.
  • R 2 may be C 1 -C 6 alkyl substituted with R 2a , and R 2a may be -NR'R''. In one embodiment, R 2 may be C 1 -C 3 alkyl substituted with R 2a . R' and R'' may be each hydrogen or C 1 -C 6 alkyl. In one embodiment, said R' and R'' may be each hydrogen or C 1 -C 3 alkyl.
  • said R' and R'' may be taken together with the nitrogen atom to which they are attached to form a 3 to 8-membered heterocyclic ring optionally comprising one additional heteroatom selected from N, O, and S.
  • said R' and R'' may be taken together with the nitrogen atom to which they are attached to form a morpholinyl, thiomorpholinyl, piperazinyl, or piperidinyl ring.
  • R 2 includes -CH 2 -morpholinyl, -CH 2 -CH 2 -morpholinyl, -CH 2 -NH 2 , -CH 2 -NH(CH 3 ), -CH 2 -N(CH 3 ) 2 , -CH 2 CH 2 -NH 2 , -CH 2 -CH 2 NH(CH 3 ), or -CH 2 -CH 2 -N(CH 3 ) 2 , but is not limited thereto.
  • R 3 is hydrogen or halogen. In one embodiment, R 3 is hydrogen. In one embodiment, R 3 is halogen. For example, R 3 may be F, Cl, Br, or I. For example, R 3 may be F.
  • the present inventors have found the problem that the compounds disclosed in the prior document (WO 2022/066835) do not exhibit a sustained GSPT1 degradation efficacy, and after a certain period of time, for example, 48 hours after treatment, the GSPT1 expression level is restored and the tumor cells whose proliferation was suppressed grow again.
  • the present inventors have found that the compounds disclosed in the prior document are unstable at physiological pH (pH 7), and the present inventors believe that this pH instability of the compounds disclosed in the above prior document is one of the reasons why the GSPT1 degradation activity and tumor inhibition activity are transient.
  • the present inventors conducted extensive research on the properties of a compound in which phenylsulfonamide is bound to the pomalidomide structure by modifying its structure. As a result, it was found that a compound having an alkyl group or an alkyl group substituted with a specific substituent at the meta position based on the position at which the sulfonamide group is bonded exhibits excellent pH stability, sustained and selective degradation activity against GSPT1, and excellent tumor proliferation inhibition activity.
  • a compounds which has, in addition to the above substitution at the meta position, a halogen or an alkyl or alkoxy group optionally substituted with halogen at the adjacent ortho position, and which is unsubstituted or substituted with halogen (preferably, F) at the adjacent para position, exhibits significantly improved properties in terms of pH stability, GSPT1 degradation activity, and tumor proliferation inhibition activity.
  • the compound represented by Formula I of the present invention may have the following combination of substituents:
  • R 1 is halogen, C 1 -C 6 alkyl, C 1 -C 6 haloalkyl, or C 1 -C 6 haloalkoxy;
  • R 2 is C 1 -C 6 alkyl optionally substituted with R 2a ;
  • R 2a is hydroxy or C 1 -C 6 haloalkoxy
  • R 3 is hydrogen or halogen.
  • the compound represented by Formula I of the present invention may have the following combination of substituents:
  • R 1 is halogen or C 1 -C 6 alkyl
  • R 2 is C 1 -C 6 alkyl
  • R 3 is hydrogen or halogen.
  • R 1 may be halogen or C 1 -C 3 alkyl; R 2 may be C 1 -C 3 alkyl; and R 3 may be hydrogen or halogen.
  • R 1 may be F, Cl, Br, I, -CH 3 , or -CH 2 CH 3; R 2 may be -CH 3 or -CH 2 CH 3 ; and R 3 may be hydrogen or F.
  • the compound represented by Formula I of the present invention may have the following combination of substituents:
  • R 1 is C 1 -C 6 haloalkyl or C 1 -C 6 haloalkoxy
  • R 2 is C 1 -C 6 alkyl
  • R 3 is hydrogen or halogen.
  • R 1 may be C 1 -C 3 haloalkyl or C 1 -C 3 haloalkoxy;
  • R 2 may be C 1 -C 3 alkyl; and
  • R 3 may be hydrogen or halogen.
  • R 1 may be -OCH 2 F, -OCHF 2 , -OCF 3 , -CH 2 F, -CHF 2 , or -CF 3 ;
  • R 2 may be -CH 3 or -CH 2 CH 3 ; and
  • R 3 may be hydrogen or F.
  • the compound represented by Formula I of the present invention may have the following combination of substituents:
  • R 1 is halogen
  • R 2 is C 1 -C 6 alkyl optionally substituted with R 2a ;
  • R 2a is hydroxy or C 1 -C 6 haloalkoxy
  • R 3 is hydrogen
  • R 1 may be halogen
  • R 2 may be C 1 -C 3 alkyl optionally substituted with hydroxy or C 1 -C 3 haloalkoxy
  • R 3 may be hydrogen
  • R 1 may be Cl, Br, or I
  • R 2 may be -CH 2 OCHF 2 , -CH 2 OCH 2 F, -CH 2 OH, or -CH 2 CH 2 OH.
  • R 1 may be halogen
  • R 2 may be C 1 -C 6 alkyl
  • R 3 may be hydrogen
  • R 1 may be Cl, Br, or I
  • R 2 may be C 1 -C 3 alkyl, such as methyl or ethyl
  • R 3 may be hydrogen.
  • the compound represented by Formula I of the present invention has the following combination of substituents:
  • R 1 is halogen
  • R 2 is C 1 -C 6 alkyl substituted with NR'R'';
  • R' and R'' are each hydrogen, C 1 -C 6 alkyl, or C 1 -C 3 alkyl;
  • R 3 is hydrogen
  • R 1 may be halogen (for example, Cl); R 2 may be methylaminomethyl; and R 3 may be hydrogen.
  • the compound represented by Formula I of the present invention may have the following combination of substituents:
  • R 1 is halogen
  • R 2 is C 1 -C 6 alkyl substituted with NR'R'';
  • R' and R'' may be taken together with the nitrogen atom to which they are attached to form a 3 to 8-membered heterocyclic ring optionally comprising one additional heteroatom selected from N, O, and S;
  • R 3 is hydrogen
  • the compound represented by Formula I of the present invention may have the following combination of substituents:
  • R 1 is halogen
  • R 2 is C 1 -C 6 alkyl substituted with NR'R'';
  • R' and R'' may be taken together with the nitrogen atom to which they are attached to form a morpholinyl, thiomorpholinyl, piperazinyl, or piperidinyl ring;
  • R 3 is hydrogen
  • R 1 may be halogen (for example, Cl); R 2 may be morpholinylmethyl, and R 3 may be hydrogen.
  • the compound represented by Formula I may be a compound represented by any one of the following formulas:
  • the compound represented by Formula I according to the present invention may be a compound having any one of the following structures:
  • the numerical range indicated using the term “to” refers to a range that includes the numerical values described before and after the term “to” as the lower limit and the upper limit, respectively.
  • the term “optional” or “optionally” means that a subsequently described event or circumstance may or may not occur, and that the description includes instances in which said event or circumstance occurs and instances in which it does not occur.
  • the term “optionally substituted” means both substitution and unsubstitution with the specified substituents.
  • alkyl refers to a saturated straight chain and branched carbon chain having 1 to 6 carbon atoms, 1 to 4 carbon atoms, or 1 to 3 carbon atoms.
  • Non-limiting examples of the alkyl group include methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, iso-butyl, tert-butyl, and the like.
  • alkoxy refers to the -O-alkyl group.
  • Non-limiting examples of the alkoxy group include methoxy, ethoxy, n-propoxy, iso-propoxy, n-butoxy, sec-butoxy, iso-butoxy, tert-butoxy, and the like.
  • heteroatom means nitrogen, oxygen, or sulfur, and includes any oxidized form of nitrogen, such as N(O) (N + -O - ) and sulfur, such as S(O) and S(O) 2 , and any basic quaternized forms of nitrogen.
  • heterocycloalkyl refers to a saturated or partially unsaturated cyclic group containing one or more heteroatoms, with the remaining ring atoms being carbon.
  • the heterocycloalkyl group may contain, for example, 1 or 2 heteroatoms.
  • the heterocycloalkyl group may contain 3 to 8 ring elements, 5 to 7 ring elements, or 5 or 6 ring elements.
  • the heterocycloalkyl group includes, for example, morpholinyl, thiomorpholinyl, piperazinyl, piperidinyl, pyrrolidinyl, azetidinyl, and the like, but is not limited thereto.
  • halogen refers to an atom belonging to group 17 of the periodic table.
  • the halogen atom includes fluorine, chlorine, bromine, and iodine, etc., and may be used interchangeably with the term “halo”, which refers to a monovalent functional group composed of halogen.
  • hydroxy refers to the -OH functional group (hydroxyl group).
  • amino refers to -NH 2 .
  • alkylamino refers to a group in which one of two Hs in an amino group is substituted with an alkyl group.
  • alkylamino group include, but are not limited thereto, methylamino, ethylamino, and propylamino.
  • dialkylamino refers to -N(alkyl) 2 .
  • the two alkyls may be the same or different from each other.
  • dialkylamino substituent include, but are not limited thereto, dimethylamino, diethylamino, ethylmethylamino, and dipropylamino.
  • haloalkyl refers to an alkyl group substituted by one or more halogen atoms.
  • the halogens may be the same (e.g., CHF 2 , -CF 3 ) or different (e.g., CF 2 Cl).
  • the haloalkyl group may be optionally substituted with one or more substituents other than halogen.
  • Examples of haloalkyl groups may include fluoromethyl, dichloroethyl, trifluoromethyl, trichloromethyl, pentafluoroethyl, and pentachloroethyl groups, but are not limited thereto.
  • haloalkoxy refers to an alkoxy group substituted with one or more halogen atoms, where alkoxy is as defined above.
  • Non-limiting examples of the haloalkoxy group may include fluoromethoxy, dichloroethoxy, trifluoromethoxy, trichloromethoxy, and the like.
  • hydroxyalkyl refers to an alkyl group substituted with one or more -OH groups, where alkyl is as defined above.
  • solvate may refer to a compound of the present invention or a salt thereof comprising a stoichiometric or non-stoichiometric amount of a solvent bound by non-covalent intermolecular forces.
  • Preferred solvents therefor may be solvents that are volatile, non-toxic, and/or suitable for administration to humans.
  • the solvent may be water, in which case the "solvate” is referred to as "hydrate”.
  • the compound of the present invention may exist as a hydrate, which can be obtained, for example, by crystallization from a solvent or aqueous solution.
  • one, two, three or any number of solvent or water molecules may combine with the compound according to the present disclosure to form a solvate and a hydrate.
  • the present disclosure includes all such possible solvates.
  • the compound according to the present invention may exist in different crystal polymorphic forms, which makes it possible for certain modifications to become metastable. Unless otherwise stated, the compound of the present invention includes all of these possible polymorphic forms.
  • stereoisomer may refer to a compound of the present invention or a salt thereof that has the same chemical formula or molecular formula but is optically or sterically different, and specifically, may be a diastereomer, an enantiomer, or a geometric isomer.
  • the compound of the present invention contains one or more asymmetric centers and may be in the form of a racemate, a single enantiomer, a mixture of enantiomers, a single diastereomer, a mixture of diastereomers, etc. In one embodiment, due to the nature of the asymmetric center or limited rotation, the compound of the present invention may exist in the form of an enantiomer or a diastereomer.
  • Purification of the isomers and separation of isomer mixtures can be achived by standard techniques known in the art. For example, a diastereomeric mixture can be separated into its respective diastereomers by a chromatographic process or crystallization, and a racemate can be separated into its respective enantiomers by a chromatographic process or resolution of the chiral phase.
  • the compound of the present invention may be used in the form of a pharmaceutically acceptable salt derived from an acid or a base.
  • pharmaceutically acceptable salt refers to a salt of an active pharmaceutical ingredient prepared with an acid or base that is tolerated by the biological system, or is tolerated by the subject, or is tolerated by the biological system and tolerated by the subject when administered in a therapeutically effective amount.
  • a base addition salt can be obtained by contacting the neutral form of this compound with a sufficient amount of the desired base, either in a pure solvent or in a suitable inert solvent.
  • pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amino, magnesium salts, lithium salts, strontium salts or similar salts, but are not limited thereto.
  • an acid addition salt can be obtained by contacting the neutral form of this compound with a sufficient amount of the desired acid, either in a pure solvent or in a suitable inert solvent.
  • pharmaceutically acceptable acid addition salts include those derived from inorganic acids such as hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, monohydrogen carbonic acid, phosphoric acid, monohydrogen phosphoric acid, dihydrogen phosphoric acid, sulfuric acid, monohydrogen sulfuric acid, hydroiodic acid, or phosphorous acid, as well as those derived from relatively non-toxic organic acids such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-tolylsulfonic acid, citric acid, tartaric acid, or
  • the compound according to the present invention can be easily prepared from commercially available starting materials, compounds known in the literature, or intermediates easily prepared therefrom through standard synthetic methods and procedures in the related field.
  • spectroscopic means such as nuclear magnetic resonance spectroscopy (e.g., 1 H or 13 C), infrared spectroscopy, spectrophotometry (e.g., UV-visible rays), mass spectrometry, or chromatography, such as high performance liquid chromatography (HPLC), gas chromatography (GC), gel permeation chromatography (GPC), or thin layer chromatography (TLC).
  • HPLC high performance liquid chromatography
  • GC gas chromatography
  • GPC gel permeation chromatography
  • TLC thin layer chromatography
  • the following general reaction scheme generally illustrates a representative method for preparing a compound represented by Formula I.
  • Those of ordinary skill in the art can easily prepare a compound represented by Formula I by appropriately selecting starting materials, reaction temperature, reaction conditions, catalyst, solvent, processing method, and the like, suitable for the desired compound based on the preparation method specifically disclosed in the examples herein.
  • the designation of each substituent in Formula I in the reaction scheme is the same as that of the substituent at the corresponding position in Formula I.
  • the compound represented by Formula I according to the present invention can be prepared by reacting a phenylsulfonylchloride compound substituted with R 1 , R 2 and R 3 as shown in Reaction Scheme A below:
  • the final target phenylsulfonylchloride compound substituted with R 1 , R 2 , and R 3 corresponding to the substituents R 1 , R 2 , and R 3 of a compound represented by Formula I can be purchased as a commercially available compound or can be easily manufactured based on known techniques in the field of medicinal chemistry and examples herein.
  • Step 1 the phenylsulfonylchloride compound and dimethyl 4-aminobenzene-1,2-dicarboxylate are reacted in the presence of an appropriate solvent, such as pyridine at an appropriate temperature (for example, 20 °C to 30 °C, such as about 25 °C) for a period of time (for example, about 1 hour).
  • an appropriate solvent such as pyridine
  • an appropriate temperature for example, 20 °C to 30 °C, such as about 25 °C
  • a period of time for example, about 1 hour
  • Step 2 the dicarboxylate compound obtained in Step 1 is hydrolyzed with LiOH in an appropriate solvent (for example, a mixed solution of THF, MeOH, and water) to obtain a phthalic acid compound.
  • the reaction can be performed at an appropriate temperature (for example, about 20 °C to about 80 °C, such as about 50 °C) for an appropriate period of time (for example, about 8 hours to about 15 hours, such as about 12 hours).
  • Step 3 the phthalic acid compound obtained in Step 2 can be reacted with 3-aminopiperidine-2,6-dione or an appropriate salt thereof (for example, HCl salt) in the presence of an appropriate solvent and reagent, such as acetic acid and sodium acetate to obtain a compound represented by Formula I.
  • the reaction can be performed at an appropriate temperature (for example, 80°C to 120 °C, such as 100 °C) for a certain period of time (for example, about 8 hours to about 15 hours, such as about 12 hours).
  • a pharmaceutical composition for treating a disorder of uncontrolled cell proliferation comprising the compound represented by Formula I, stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof.
  • the compound represented by Formula I, stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof is as described above.
  • treating refers to inhibiting a disease, for example, inhibiting a disease, condition or disorder in a subject who experiences or exhibits the pathology or signs of the disease, condition or disorder, i.e., preventing further development of the pathology and/or signs, or ameliorating the disease, for example, ameliorating the disease, condition or disorder in a subject who experiences or exhibits the pathology or signs of the disease, condition or disorder, i.e., reversing the pathology and/or signs, for example, reducing the severity of the disease.
  • the disorder of uncontrolled cell proliferation is cancer.
  • the cancer may be pediatric cancer, such as pediatric acute leukemia or medulloblastoma.
  • the cancer may be selected from brain cancer, lung cancer, blood cancer, bladder cancer, colon cancer, cervical cancer, endometrial cancer, ovarian cancer, squamous cell cancer, kidney cancer, peritoneal cancer, breast cancer, stomach cancer, large intestine cancer, prostate cancer, pancreatic cancer, urogenital tract cancer, lymphatic cancer, laryngeal cancer, skin cancer, malignant melanoma, large intestine cancer, endometrial carcinoma, thyroid cancer, rhabdomyosarcoma, liver cancer, and a combination thereof.
  • the cancer may be lung cancer, prostate cancer, breast cancer, ovarian cancer, endometrial cancer, bladder cancer, or skin cancer.
  • the lung cancer may be small cell lung cancer, non-small cell lung cancer, or lung neuroendocrine cancer.
  • the prostate cancer may be androgen receptor-positive prostate cancer (AR-positive prostate cancer; ARPC), castration-resistant prostate cancer (CRPC), double-negative prostate cancer (DNPC) (not expressing androgen receptor nor neuroendocrine markers), or neuroendocrine prostate cancer (NEPC).
  • the brain cancer may be glioblastoma, medulloblastoma, glioma, or a combination thereof.
  • the kidney cancer may be kidney clear cell carcinoma.
  • the bladder cancer may be bladder urothelial carcinoma.
  • the blood cancer may be selected from chronic myeloid leukemia (CML), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), acute lymphocytic leukemia (ALL), hairy cell leukemia, chronic myelomonocytic leukemia (CMML), juvenile myelomonocytic leukemia (JMML), large granular lymphocytic leukemia (LGL), acute lymphocytic leukemia, acute lymphoblastic leukemia, B-cell lymphoma, T-cell-lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, hairy cell lymphoma, Burkitt lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, and a combination thereof.
  • CML chronic myeloid leukemia
  • AML acute myeloid leukemia
  • CLL chronic lymphocytic leukemia
  • the disorder of uncontrolled cell proliferation may be related to cereblon (CRBN) dysfunction.
  • CRBN cereblon
  • cereblon and “CRBN” can be used interchangeably, and refer to a protein encoded by the human CRBN gene at the cytogenetic location of 3p26.2 and the molecular location between base pairs 3,148,489 and 3,179,716 of chromosome 3 (UCSC Genome Browser for Humans, December 2013 (GRCh38/hg38) assembly).
  • CRBN is the substrate recognition element of the DCX (DDB1-CUL4-X-box) E3 protein ligase complex that mediates ubiquitination and subsequent proteasome degradation of target proteins.
  • the DCX (DDB1-CUL4-X-box) E3 protein ligase complex is composed of at least CRBN, CUL4A, DDB1, and RBX1.
  • the CRBN protein has two isoforms generated by alternative splicing: Isoform 1 has 442 amino acids and a molecular weight of 50,546 Da; and Isoform 2 has 441 amino acids and a molecular weight of 50,475 Da.
  • the disorder of uncontrolled cell proliferation may be related to GSPT1 dysfunction.
  • the disorder of uncontrolled cell proliferation may be a disorder related to GSPT1 or caused by GSPT1.
  • the disorder of uncontrolled cell proliferation may be cancer related to GSPT1.
  • GSPT1 (G1 to S phase transition 1), also called eRF3, is a translation termination factor that binds to eRF1 and mediates stop codon recognition and release of nascent proteins from ribosomes.
  • GSPT1 plays a key role in maintaining high-fidelity protein synthesis ( Cell . 2011 Oct 14; 147(2): 396-408).
  • Ubiquitination and degradation of GSPT1 are known to induce inhibition of the expression level of cancer-inducing proteins (translationally addicted oncoproteins, for example, c-MYC, N-MYC, L-MYC, BCL-2, MCL-1, etc.) whose expression levels remain high depending on protein translation in cancer cells (Mullard, Nat Rev Drug Discov.
  • the compound represented by Formula I according to the present invention can be useful for the treatment of MYC-driven cancer, such as cancer driven by cancer-inducing proteins whose expression level is maintained at a high level depending on protein translation, such as c-MYC, n-MYC, and l-MYC.
  • MYC-driven cancer includes, for example, prostate cancer, breast cancer, liver cancer, colorectal cancer, and the like, but is not limited thereto.
  • the compound represented by Formula I according to the present invention can be useful for the treatment of cancers exhibiting high n-Myc expression level.
  • the compound represented by Formula I according to the present invention can be useful for the treatment of cancers sensitive to the integrated stress response, such as blood cancer.
  • the blood cancer includes acute myeloid leukemia (AML), chronic myeloid leukemia (CML), chronic lymphocytic leukemia (CLL), acute lymphocytic leukemia (ALL), and the like, but is not limited thereto.
  • the blood cancer is acute myeloid leukemia.
  • the compound represented by Formula I according to the present invention is a molecular glue with a structure based on pomalidomide, one of the immunomodulatory imide drugs, and can bind to CRBN protein and induce selective GSPT1 degradation.
  • a compound represented by Formula I with a specific combination of substituents can have excellent selectivity, especially for GSPT1, compared to various neosubstrates such as IKAROS, AIOLOS, and CK1 ⁇ .
  • the compound represented by Formula I of the present invention has excellent selectivity and degradation activity against GSPT1 compared to the compound disclosed in the prior document WO 2022/066835.
  • the compounds disclosed in the above prior document only exhibit temporary degradation activity against GSPT1, and then restore GSPT1 expression level and show reproliferation of tumor cells, whereas the compound represented by Formula I according to the present invention exhibits sustained and stable GSPT1 degradation activity and excellent tumor cell proliferation inhibitory activity.
  • the compound represented by Formula I according to the present invention has excellent inhibitory activity against tumor cell proliferation and significantly reduced cytotoxicity against normal cells, and, thus, it has a much larger therapeutic index than the compounds disclosed in the above prior document.
  • the cancer is a cancer with a neuroendocrine phenotype.
  • the present inventors have found that a compound represented by Formula I exhibits particularly excellent anticancer activity, especially against a cancer with a neuroendocrine phenotype, such as lung cancer or prostate cancer.
  • a cancer with a neuroendocrine phenotype have been reported to share morphology and marker-based histology, such as high nuclear-to-cytoplasm ratio, frequent mitosis characteristic, and granular chromatin (Am. Soc. Clin. Oncol. Educ. Book. 2015; 35: 92-103).
  • a cancer with a neuroendocrine phenotype essentially has TP53 and RB1 loss and/or inactivation mutations at the molecular level and expresses common neuroendocrine markers such as chromogranin A (CHGA) and synaptophysin (SYP) (Nat. Med. 2016; 22: 298-305; Lancet Oncol. 2015; 16: e435-e446).
  • CHGA chromogranin A
  • SYP synaptophysin
  • a cancer with a neuroendocrine phenotype includes neuroendocrine prostate cancer (NEPC), castration-resistant prostate cancer, and lung neuroendocrine tumor, but is not limited thereto.
  • NEPC neuroendocrine prostate cancer
  • castration-resistant prostate cancer e.g., castration-resistant prostate cancer
  • lung neuroendocrine tumor e.g., lung neuroendocrine tumor
  • the pharmaceutical composition may comprise conventional pharmaceutically acceptable carriers, excipients, or additives.
  • the pharmaceutically acceptable carrier, excipient, or additive may include one or more pharmaceutically acceptable diluents, preservatives, antioxidants, solubilizers, emulsifiers, coloring agents, releasing agents, coating agents, sweetener, flavoring agents and fragrances, and adjuvants, but is not limited thereto.
  • the disclosed pharmaceutical composition may conveniently be presented in a unit dosage form and may be prepared by any method well known in the fields of pharmaceuticals and pharmaceutical sciences.
  • the pharmaceutical composition may be formulated according to a conventional method, and may be prepared as various oral formulations such as tablets, pills, powders, capsules, syrups, emulsions, and microemulsions, or parenteral formulations such as intramuscular, intravenous or subcutaneous formulation, or formulations for topical application to the skin.
  • the pharmaceutical composition may be a single composition or separate compositions.
  • the pharmaceutical composition comprises a compound, stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt according to one aspect as an active ingredient of the pharmaceutical composition.
  • unit dosage form refers to a physically discrete unit suitable as unit dosages, wherein each unit contains a predetermined amount of an active ingredient calculated to produce the desired therapeutic effect in relation to the required pharmaceutical carrier.
  • unit dosage forms include tablets (including divided or coated tablets), capsules, or pills for oral administration; single-dose vials for solutions or suspensions for injection; suppositories for rectal administration; powder packet; wafer; and separated multiple batches thereof.
  • additives or carriers used may include cellulose, calcium silicate, corn starch, lactose, sucrose, dextrose, calcium phosphate, stearic acid, magnesium stearate, calcium stearate, gelatin, talc, surfactant, suspending agent, emulsifying agent, diluent, and the like.
  • the additive or carrier may include water, saline, aqueous glucose solution, similar aqueous sugar solution, alcohol, glycol, ether (for example, polyethylene glycol 400), oil, fatty acid, fatty acid ester, glyceride, surfactant, suspending agent, emulsifying agent, and the like.
  • the pharmaceutical composition of the present invention may be prepared in a liquid dosage form, wherein the liquid dosage form may comprise preservatives, stabilizers, buffering substances, flavor modifiers, sweeteners, coloring agents, antioxidants, and complex formers, etc.
  • the complex forming agent may include a chelating agent such as ethylenediamine tetraacetic acid, nitrilotriacetic acid, diethylenetriamine pentacetic acid, and salts thereof. It may optionally be necessary to stabilize the liquid dosage form with a physiologically acceptable base or buffer to a pH range of approximately 9 or less. To the extent possible, it is desirable to prepare stable dosage forms at neutral or slightly basic pH values (pH 8 or less).
  • the compound of the present invention having excellent stability at neutral pH may also be advantageous in terms of formulation stability.
  • co-solvents such as alcohols can improve the solubility and/or stability of the compound according to the present disclosure in a pharmaceutical composition.
  • the dosage of the pharmaceutical composition is an amount effective for treatment of a subject or patient, and may be administered orally or parenterally as desired. It may be administered in one to several divided doses to be administered in an amount of 0.01 to 1000 mg, more specifically 0.1 to 300 mg per kg of body weight daily based on the active ingredient when administered orally, or in an amount of 0.01 to 100 mg, more specifically 0.1 to 50 mg per kg of body weight daily based on the active ingredient when administered parenterally.
  • the dose to be administered to a specific subject or patient should be determined in light of several related factors such as body weight, age, sex, and health condition of the patient, the diet, the administration time, the administration method, the severity of the disease, and the like, and it should be understood that it may be appropriately increased or decreased by a specialist.
  • the above dosage is not intended to limit the scope of the present invention in any way.
  • a physician or veterinarian of ordinary skill in the art can readily determine and prescribe the required effective amount of the pharmaceutical composition.
  • a dose of the compound of the present invention used in a pharmaceutical composition may start at a level lower than that required to achieve the desired therapeutic effect, and may gradually increase until the desired effect is achieved.
  • the pharmaceutical composition includes within its scope a pharmaceutical composition comprising, as an active ingredient, a therapeutically effective amount of at least one of the disclosed compounds, alone or in combination with a pharmaceutically acceptable carrier.
  • a pharmaceutical composition comprising, as an active ingredient, a therapeutically effective amount of at least one of the disclosed compounds, alone or in combination with a pharmaceutically acceptable carrier.
  • therapeutically effective amount or “effective amount” refers to an amount sufficient to produce a beneficial or desired clinical result, for example, an amount sufficient to alleviate, ameliorate, stabilize, reverse, slow or delay the progression of a disease.
  • the pharmaceutical composition comprises 0.05 to 99% by weight, preferably 0.1 to 70% by weight, more preferably 0.1 to 50% by weight of an active ingredient, and 1 to 99.95% by weight, preferably 30 to 99.9% by weight, more preferably 50 to 99.9% by weight of a pharmaceutically acceptable carrier, with all percentages based on the total weight of the composition.
  • the pharmaceutical composition may further comprise at least one agent known to treat cancer.
  • the at least one agent may be selected from uracil mustard, chlormethine, cyclophosphamide, ifosfamide, melphalan, chlorambucil, pipobroman, triethylenemelamine, triethylenethiophosphoramine, busulfan, carmustine, lomustine, streptozocin, dacarbazine, temozolomide, thiotepa, altretamine, methotrexate, 5-fluorouracil, floxuridine, cytarabine, 6-mercaptopurine, 6-thioguanine, fludarabine phosphate, pentostatin, bortezomib, vinblastine, vincristine, vinorelbine, vindesine, bleomycin, dactinomycin, daunorubicin, doxorubicin, epirubicin, dexamethasone, clofarabine, cladribine, pemetrexed, idarubicin,
  • the at least one agent may be selected from a DNA methyltransferase inhibitor, a HDAC-inhibitor, a glucocorticoid, a mTOR inhibitor, a cytotoxic agent, or a combination thereof.
  • the DNA methyltransferase inhibitor may be 5-aza-2'-deoxycytidine, 5-azacitidine, zebularine, epigallocatechin-3-gallate, procaine, or a combination thereof.
  • the HDAC-inhibitor may be vorinostat, entinostat, panobinostat, trichostatin A, mocetinostat, belinostat, dacinostat, givinostat, tubastatin A, pracinostat, droxinostat, quisinostat, romidepsin, valproic acid, AR-42 (OSU-HDAC42), tacedinaline, ricolinostat, apicidin, or a combination thereof.
  • the glucocorticoid may be dexamethasone, prednisolone, methylprednisolone, betamethasone, triamcinolone, fludrocortisone, beclomethasone, or a combination thereof.
  • the mTor inhibitor may be BEZ235, everolimus, sirolimus, temsirolimus, rapamycin, AZD8055, or a combination thereof.
  • the cytotoxic agent may be selected from an alkylating agent, an antimetabolite, an antitumor antibiotic, a mitosis inhibitor, or a chemotherapeutic agent selected from anthracycline, cytarabine, purine analog, sorafenib, gemtuzumab ozogamicin, rituximab, or a combination thereof.
  • the alkylating agent may be selected from carboplatin, cisplatin, cyclophosphamide, chlorambucil, melphalan, carmustine, busulfan, lomustine, dacarbazine, oxaliplatin, ifosfamide, mechlorethamine, temozolomide, thiotepa, bendamustine, and streptozocin.
  • the antimetabolite may be selected from gemcitabine, 5-fluorouracil, capecitabine, hydroxyurea, mercaptopurine, pemetrexed, fludarabine, nelarabine, cladribine, clofarabine, cytarabine, decitabine, pralatrexate, floxuridine, methotrexate, and thioguanine.
  • the antitumor antibiotic may be selected from doxorubicin, mitoxantrone, bleomycin, daunorubicin, dactinomycin, epirubicin, idarubicin, plicamycin, mitomycin, pentostatin, and valrubicin.
  • the mitosis inhibitor may be selected from irinotecan, topotecan, rubitecan, cabazitaxel, docetaxel, paclitaxel, etoposide, vincristine, ixabepilone, vinorelbine, vinblastine, and teniposide.
  • the at least one agent may be a BCL2 inhibitor, a FLT3 inhibitor, an IDH 1/2 inhibitor, a CDK (cyclin-dependent kinase) inhibitor, a transcription inhibitor, a HSP inhibitor, or a combination thereof.
  • the BCL2 (B-cell lymphoma 2) inhibitor may include venetoclax, navitoclax, obatoclax mesylate, sabutoclax, lisaftoclax, and the like.
  • the FLT3 (FMS-like tyrosine kinase 3) inhibitor may include midostaurin, quizartinib, gilteritinib, sorafenib, crenolanib, pexidartinib, and the like.
  • the IDH (isocitrate dehydrogenase) 1/2 inhibitor may include ivosidenib, enasidenib, vorasidenib, olutasidenib, AGI-6780, AGI-5198, GSK321, and the like.
  • the CDK inhibitor may include a CDK7 inhibitor, a CDK9 inhibitor, a CDK 12 inhibitor, and the like, and may include, for example, samuraciclib, alvocidib, fadraciclib, seliciclib, zotiraciclib, atuveciclib, enitociclib, voruciclib, SY5609, XL201, Q-901, KRLS-017, GTAEXS-617, TGN-1062, THZ1, THZ2, SY-1365, YKL-5-124, ICEC0942, LY3405105, LDC4297, BS-181, SNS-32, AT-7519, AZD-4573, KB-0742, AU-07, BTXA-51, GFH-009, JS-101, PRT-2527, QHRD-107, TP-1287, SYHX-1903, CTX-439, KIN-004, SY-12882, THZ-531
  • the transcription inhibitor includes any agent capable of inhibiting the transcription of a cancer-inducing protein.
  • the compound of the present invention can exhibit anticancer effects by inhibiting the translation process from RNA to protein based on GSPT1 degradation activity. Therefore, combined use with a transcription inhibitor that inhibits the transcription process from DNA to RNA can exhibit enhanced anticancer effects.
  • Such transcription inhibitors may include, for example, lurbinectedin, which is known to inhibit transcription by covalently binding to residues present in the minor groove of DNA.
  • the HSP (heat shock protein) inhibitor includes a HSP70 inhibitor, a HSP90 inhibitor, and the like, and may include, for example, pimitespib, luminespib, tanespimycin, alvespimycin, ganetespib, onalespib, geldanamycin, rocaglamide, and the like.
  • the at least one agent may be co-packaged, co-formulated, and/or co-transferred with the compound represented by Formula I according to the present invention, a stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof.
  • the at least one agent may be provided in the form of a kit comprising the compound represented by Formula I according to the present invention, a stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof.
  • a kit comprising the compound represented by Formula I according to the present invention, a stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof.
  • two or more ingredients which may be active or inactive ingredients, carriers, diluents, etc., may be provided along with instructions for the preparation of the actual dosage form by the patient or the individual administering the drug to the patient.
  • a method of regulating a cereblon activity or a GSPT1 activity in at least one cell comprising contacting the at least one cell in vitro with a compound represented by Formula I, a stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof.
  • a compound represented by Formula I a stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof.
  • the term "contacting" refers to bringing together the disclosed compound or pharmaceutical composition in close proximity to a cell, target protein, or other biological entity, in a manner that allows the disclosed compound or pharmaceutical composition to directly affect the activity of the cell, target protein, or other biological entity; that is, by interacting with the cell, target protein, or other biological entity itself, or in a manner that allows the disclosed compound or pharmaceutical composition to indirectly affect the activity of the cell, target protein, or other biological entity; that is, by interacting with other molecules, cofactors, factors, or proteins on which the activity of the cell, target protein, or other biological entity itself depends.
  • the compound represented by Formula I according to the present invention or a pharmaceutical composition comprising the same has activity as a modulator of cereblon protein.
  • the compound represented by Formula I according to the present invention or a pharmaceutical composition comprising the same has activity as a modulator of GSPT1 expression and/or activity.
  • the compound represented by Formula I according to the present invention or a pharmaceutical composition comprising the same has activity as a cell proliferation inhibitor.
  • the compound represented by Formula I according to the present invention can be used to evaluate new compounds.
  • the test compound can be evaluated by comparing the analytical values obtained by performing a biological analysis with the test compound and the compound of the present invention, respectively.
  • the biological analysis includes cereblon binding analysis, GSPT1 degradation activity analysis, or cell proliferation test, etc., but is not limited thereto.
  • a method of treating a disorder of uncontrolled cell proliferation comprising administering a compound represented by Formula I, a stereoisomer, hydrate, solvate, or pharmaceutically acceptable salt thereof to a subject.
  • the term "subject” refers to a subject in need of treatment for a disease, and more specifically means a mammal such as a human or non-human primate, a mouse, a dog, a cat, a horse, and a cow.
  • a disorder of uncontrolled cell proliferation such as cancer
  • a use of the compound represented by Formula I, stereoisomer, hydrate, or solvate thereof for manufacturing a medicament for the treatment of a disorder of uncontrolled cell proliferation, such as cancer.
  • Reaction Scheme 1 Synthesis of 2-chloro -N -(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)-3-(morpholinomethyl)benzenesulfonamide
  • Step 1 tert-butyl((2-chloro-3-((4-methoxybenzyl)thio)benzyl)oxy)diphenylsilane
  • Step 2 3-(((tert-butyldiphenylsilyl)oxy)methyl)-2-chlorobenzenesulfonylchloride ( Intermediate C1 )
  • Step 3 Dimethyl 4-((3-(((tert-butyldimethylsilyl)oxo)methyl)-2-chlorophenyl)sulfinamido)phthalate
  • Step 4 Dimethyl 4-((2-chloro-3-(hydroxymethyl)phenyl)sulfonamide)phthalate ( Intermediate D1 )
  • Step 5 Dimethyl 4-[(2-chloro-3-formyl-phenyl)sulfonamido]benzene-1,2-dicarboxylic acid
  • Step 6 Dimethyl 4-((2-chloro-3-(morpholinomethyl)phenyl)sulfinamido)phthalate
  • Step 7 4-((2-chloro-3-(morpholinomethyl)phenyl)sulfinamido)phthalic acid
  • Step 8 2-chloro -N -(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)-3-(morpholinomethyl)benzenesulfonamide ( Compound 1 )
  • Step 1 3-(difluoromethoxy)-1-fluoro-4-[(4-methoxyphenyl)methylsulfanyl]-2-methyl-benzene
  • Step 3 2-(difluoromethoxy)-N-[2-(2,6-dioxo-3-piperidyl)-1, 3-dioxoisoindolin-5-yl]-4-fluoro-3-methyl-benzenesulfonamide (Compound 2 )
  • Step 2 2-chloro- N -[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindolin-5-yl]-3-methyl-benzenesulfonamide (Compound 3 )
  • Step 1 4-[(2-chloro-3-methyl-phenyl)sulfonamino]benzene-1,2-dicarboxylic acid
  • Step 3 2-chloro- N -[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindolin-5-yl]-3-methyl-benzenesulfonamide (Compound 3 )
  • Step 1 2-chloro- N -[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindolin-5-yl]-3-methyl-benzenesulfonamide
  • the reaction mixture was concentrated, and then water (100 mL) was poured, and extracted with EtOAc (200 mL x 3). The combined organic layer was washed with brine (300 mL), dried over Na 2 SO 4 , filtered, and then concentrated under reduced pressure to obtain a crude product.
  • Compound 3 was obtained by reacting 2-chloro-3-methyl-benzenesulfonyl chloride in the same manner as in Steps 1 to 3 of Method 2.
  • Step 2 Dimethyl 4-[(2-bromo-3-methyl-phenyl)sulfonylamino]benzene-1,2-dicarboxylate ( Intermediate D2 )
  • Step 4 2-bromo- N -[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindolin-5-yl]-3-methyl-benzenesulfonamide (Compound 4 )
  • reaction mixture was concentrated under reduced pressure to obtain a crude product, which was purified by prep -HPLC (column: Phenomenex luna C18 150*25 mm* 10 um; mobile phase: [water (FA)-ACN]; B%: 30%-60%, 10 minutes) to obtain Compound 4 (38.94 mg, 63% yield, 98.71% purity) as a gray solid.
  • Step 1 Dimethyl 4-[(2-iodo-3-methyl-phenyl)sulfonylamino]benzene-1,2-dicarboxylate ( Intermediate D3 )
  • Step 3 N -[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindolin-5-yl]-2-iodo-3-methyl-benzenesulfonamide (Compound 5 )
  • reaction mixture was concentrated under reduced pressure to obtain a crude product, which was purified by prep -HPLC (formic acid addition, column: Phenomenex luna C18 150*25 mm* 10 um; mobile phase: [water (FA)-ACN]; B%: 36%-56%, 10 minutes) to obtain Compound 5 (35.46 mg, 62% yield, 98.40% purity) as a green solid.
  • prep -HPLC formic acid addition, column: Phenomenex luna C18 150*25 mm* 10 um; mobile phase: [water (FA)-ACN]; B%: 36%-56%, 10 minutes
  • Step 1 Dimethyl 4-[(2-chloro-3-formyl-phenyl)sulfonylamino]benzene-1,2-dicarboxylate
  • Step 2 Dimethyl 4-[[2-chloro-3-(methylaminomethyl)phenyl]sulfonylamino]benzene-1,2-dicarboxylate ( Intermediate D4 )
  • Step 3 Dimethyl 4-[[3-[[ tert -butoxycarbonyl(methyl)amino]methyl]-2-chloro-phenyl]sulfonylamino]benzene-1,2-dicarboxylate
  • Step 4 4-[[3-[[ tert -butoxycarbonyl(methyl)amino]methyl]-2-chloro-phenyl]sulfonylamino]phthalic acid
  • Step 5 tert -butyl N -[[2-chloro-3-[[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindolin-5-yl]sulfamoyl]phenyl]methyl]- N -methyl-carbamate
  • Step 6 2-chloro- N -[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindolin-5-yl]-3-(methylaminomethyl)benzenesulfonamide (Compound 6 )
  • reaction mixture was concentrated under reduced pressure and then purified by prep -HPLC (hydrochloric acid addition, column: YMC Triart C18 150*25 mm*5 um; mobile phase: [water (hydrochloric acid)-ACN]; gradient: 13%-43% B over 10 min) to obtain Compound 6 (39.07 mg, 85% yield, 99.44% purity, hydrochloride) as an off-white solid.
  • Step 1 Dimethyl 4-[[2-chloro-3-(trifluoromethoxymethyl)phenyl]sulfonylamino]benzene-1,2-dicarboxylate
  • Step 2 4-[[2-chloro-3-(trifluoromethoxymethyl)phenyl]sulfonylamino]phthalic acid
  • Step 3 2-chloro- N -[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindolin-5-yl]-3-(trifluoromethoxymethyl)benzenesulfonamide (Compound 7 )
  • reaction mixture was concentrated under reduced pressure to obtain a crude product, which was purified by prep -HPLC (formic acid addition, column: Phenomenex luna C18 150*25 mm* 10 um; mobile phase: [water (FA)-ACN]; gradient: 36%-66% B over 10 min) to obtain Compound 7 (3.09 mg, 18% yield, 98.88% purity) as a green solid.
  • Step 3 dimethyl 4-((2-(difluoromethoxy)-3-methylphenyl)sulfonamido)phthalate ( Intermediate D5 )
  • Step 4 4-[[2-(difluoromethoxy)-3-methyl-phenyl]sulfonylamino]phthalic acid
  • Step 5 2-(difluoromethoxy)- N -[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindolin-5-yl]-3-methyl-benzenesulfonamide (Compound 8 )
  • Step 2 dimethyl 4-[(3-bromo-2-chloro-phenyl)sulfonylamino]benzene-1,2-dicarboxylate ( Intermediate D6 )
  • Step 3 4-[(3-bromo-2-chloro-phenyl)sulfonylamino]phthalic acid
  • Step 4 3-bromo-2-chloro- N -(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)benzenesulfonamide
  • Step 5 2-chloro- N -(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)-3-(hydroxymethyl)benzenesulfonamide (Compound 9 )
  • the NCI-H1155 cell line was purchased from the American Type Culture Collection (ATCC, Manassas, VA) and maintained in RPMI 1640 (Roswell Park Memorial Institute Medium 1640; Cytiva) supplemented with 10% FBS, 2 mM L-glutamine, and 100 unit/mL streptomycin-penicillin.
  • the HL-60 cell line was purchased from the Korean Cell Line Bank (KCLB, Seoul, Korea) and maintained in RPMI 1640 supplemented with 10% FBS, 2 mM L-glutamine, and 100 unit/mL streptomycin-penicillin.
  • the NCI-H2023 cell line was purchased from the ATCC and maintained in Dulbecco's Modified Eagle Medium (DMEM / F12 1:1 medium, Cytiva) mixed 1:1 with F12 supplemented with 5% FBS, 0.005 mg/mL insulin, 0.01 mg/mL transferrin, 30 nM sodium selenite, 10 nM hydrocortisone, 10 nM beta-estradiol, and 100 unit/mL streptomycin-penicillin.
  • DMEM / F12 1:1 medium, Cytiva Dulbecco's Modified Eagle Medium
  • the HEKa cell line was purchased from the ATCC and maintained in Keratinocyte Growth Kit (ATCC) supplemented with 6 mM L-glutamine, 0.4% bovine pituitary extract, 0.5 ng/mL rh TGF-a, 100 ng/mL hydrocortisone hemisuccinate, 5mg/mL rh insulin, 1 mM epinephrine, and 5 mg/mL apo-transferrin and Dermal Cell Basal Medium (ATCC) supplemented with 100 unit/mL streptomycin-penicillin.
  • ATCC Keratinocyte Growth Kit
  • the HEK-293 hGSPT1 HiBiT tag cell line was purchased from Synthego Corporation (Redwood City, CA) and maintained in DMEM medium supplemented with 10% FBS, 2 mM L-glutamine, and 100 unit/mL streptomycin-penicillin.
  • CRBN binding ability analysis was commissioned by Eurofins and analyzed using the E3scan TM platform.
  • the experimental method used is as follows.
  • the CRBN-DDB1 protein complex was produced using the HEK-293 cell line, and then DNA for qPCR detection was labeled.
  • Streptavidin-coated magnetic beads and a biotinylated small molecule ligand that binds to CRBN were reacted for 30 minutes to produce affinity beads with the small molecule ligand immobilized.
  • the affinity beads with the ligand immobilized were blocked with an excess of biotin and washed with the blocking buffer (SeaBlock (Pierce), 1% BSA, 0.05% Tween 20, 1 mM DTT) to remove unbound ligand and prevent non-specific binding.
  • the binding reaction was performed by mixing the CRBN-DDB1 ligase, the affinity beads, and the test compound in 1x binding buffer (20% SeaBlock, 0.17x PBS, 0.05% Tween 20, 6 mM DTT).
  • the test compound was prepared in a 100% DMSO solution at a concentration 111 times higher than the final concentration.
  • the solution of the test compound was diluted to to 11 concentrations in 3-fold serial dilution.
  • the test compound was dispensed with 100% DMSO using a non-contact dispensing method. Thereafter, the compounds were diluted directly into solutions for experiments to a final concentration of DMSO of 0.9%. All reactions were performed in a polypropylene 384 well plate. The final volume of each reaction solution was set to 0.02 mL. The experimental plate was reacted at room temperature with shaking for about 1 hour, and the affinity beads were washed with the washing buffer (1x PBS, 0.05% Tween 20). Thereafter, the beads were redispersed in the elution buffer (1x PBS, 0.05% Tween 20, 0.5 ⁇ M non-biotinylated affinity ligand) and then reacted at room temperature with shaking for about 30 minutes. CRBN-DDB1 concentration in the eluate was measured by qPCR. Kd was calculated by analyzing the solutions of the test compounds at the 11 concentrations and 3 control groups (DMSO).
  • Reference Compound 1 and Reference Compound 2 are compounds having the following structures, described as “Compound 1" and “Compound 5" in International Publication No. WO 2022/066835.
  • the example compounds showed equal or higher binding ability (lower Kd) to CRBN-DDB1 compared to Reference Compound 1 and Reference Compound 2. However, since the binding ability to CRBN-DDB1 does not necessarily lead to GSPT1 degradation activity, GSPT1 degradation activity was also evaluated below.
  • the HEK293 cell line expressing HiBiT-GSPT1 was produced by Synthego (California, USA).
  • DNA donor capable of expressing the HiBiT protein fragment consisting of 11 amino acids (VSGWRLFKKIS (SEQ ID NO. 1)(CCACTCCTCTCCGGCCGGGCGCCCCTGCCTCCATTTCCCGCTCTCTGTCCACCACACACACGGCCCCCCCGATAATG GTGAGCGGCTGGCGGCTGTTCAAGAAGATTAGCG GCGGTAGCGATCCGGGCAGTGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGCGGGAGCAGCAGCGGCAGCAGCAGCAGCGACTCGGC (SEQ ID NO.
  • the underlined part is the HiBiT nucleotide sequence
  • the underlined part is the HiBiT nucleotide sequence
  • a single clone was obtained through single cell sorting.
  • the GSPT1 gene sequence edited from the obtained single clone was amplified using the following two primers [Forward (5'-3'): TTGGCGTTGACGTTGAGTTG (SEQ ID NO. 3), Reverse (5'-3'): ACACGAGGAGGAGGGTTGAG (SEQ ID NO. 4)], and the edited HiBiT-GSPT1 gene sequence was determined by Sanger sequencing method.
  • the HEK-293 hGSPT1 HiBiT tag cells were seeded in triplicate in a white 96-well analysis plate at a density of 5,000 cells per well. After overnight incubation, a medium mixed with the 3-fold concentration of the test compound was used to treat the cells with the test compound at final concentrations of 1 nM, 3 nM, 10 nM, 30 nM, 100 nM, 1 ⁇ M, 3 ⁇ M, and 10 ⁇ M. After 72 hours of incubation, the level of the GSPT1 HiBiT tag protein was evaluated using the Nano-Glo® HiBiT Lytic Detection System (Promega) according to the manufacturer's instructions. The luminescence signals were measured using a Varioskan LUX multimode microplate reader.
  • the GSPT1 degradation activity was evaluated in three aspects: EC 50 , DC 50 , and D max .
  • Dmax (%) represents the maximum value for degrading GSPT1 at the highest concentration.
  • DC 50 was calculated by defining it as the concentration required to cause the absolute degradation of GSPT1 by 50%.
  • the calculated EC 50 , DC 50 , and D max values are shown in Table 2 below.
  • Reference Compound 2 Dmax was 42.8% and DC 50 was greater than 10 ⁇ M.
  • GSPT1 HiBiT test In order to determine whether the results of the GSPT1 HiBiT test, a test system artificially created for high-throughput screening of compounds, are reproduced in cancer cell lines and determine the selectivity of the compounds of the present invention for the degradation of various neosubstrates, the NCI-H1155 cell line, a lung cancer cell line, was treated with the example compounds, and then the expression levels of GSPT1 and other known neosubstrates were determined through immunoblot experiments.
  • the NCI-H1155 cells were seeded in a 6 well plate (2 x 10 6 cells per well). After overnight incubation, the cells were treated with Reference Compound 1, Compound 3, Compound 4, and Compound 5 at concentrations of 0 nM, 3 nM, 10 nM, 30 nM, 100 nM, 300 nM, and 1000 nM, respectively. After 6 hours, the cells were harvested. In addition, in order to confirm the change in the amount of GSPT1 degradation over time, the cells were treated with Reference Compound 1, Compound 2, Compound 3, Compound 4, Compound 8, and Compound 11 at a concentration of 100 nM, respectively. After 0, 2, 4, 8, 24, 48, and 72 hours, the cells were harvested.
  • the harvested cells were spun down, washed with PBS, lysed with the RIPA buffer supplemented with Protease Inhibitor Cocktail (Roche) and Phosphatase Inhibitor Cocktail (Roche), and then frozen overnight in a -80 °C cryogenic freezer.
  • the samples thawed on ice were centrifuged, and 1x LDS loading buffer and 1x reducing agent were added, and the samples were heated at 95 °C.
  • the prepared samples were loaded on NuPAGE 4-12% Bis-Tris protein gel and transferred to a nitrocellulose membrane. The membrane was blocked with 5% bovine serum albumin and incubated with primary antibodies overnight, and then the corresponding protein signals were detected using HRP-linked secondary antibodies and an iBright CL1500 imaging system.
  • the antibodies used were as follows:
  • Compound 3, Compound 4, and Compound 5 all showed equivalent or improved GSPT1 degradation activity compared to Reference Compound 1 (see Figure 1).
  • all the example compounds evaluated had no significant effect on the expression level of other neosubstrates even at the highest concentration. As a result, it was confirmed that the compounds of the present invention exhibit selective degradation activity against GSPT1.
  • the cancer cells were treated with Reference Compound 1 and the example compounds, and then the real time cancer cell growth was measured using an Incucyte S3 live cell analysis system (Sartorius, Ann Arbor, MI, USA).
  • Compound 3 Compound 4, Compound 5, Compound 15, and Compound 19 showed sustained cancer cell growth inhibition at a concentration of 100 nM (marked as ⁇ in Figure 3). These compounds did not show any rebound in cell growth or showed a minimal rebound compared to Reference Compound 1 at 100 nM, even at a lower concentration of 30 nM (marked as ⁇ in Figure 3). Overall, it was confirmed that the compounds of the present invention can induce the GSPT1 degradation and cancer cell growth inhibition more consistently and stably than Reference Compound 1.
  • Reference Compound 1 As shown in Table 5, the residual amounts of Reference Compound 1 were only 87%, 86.5%, and 79.2%, respectively, after 24 hours in buffer solutions of pH 4.4, 5.2, and 6.0. Furthermore, Reference Compound 1 began to show instability with a residual amount of 87.4%, at 4 hours in the neutral condition of pH 7.1, and after 24 hours, the residual amount was only 54.2%, indicating that it was very unstable (Table 5). From the above results, it can be seen that Reference Compound 1 is unstable at physiological pH (neutral pH) and will not show sustained drug efficacy. This is consistent with the results showing that the expression level of GSPT1 was restored 48 hours after treatment of the cancer cell line with Reference Compound 1 ( Figure 2) and that cell proliferation occurred again (Figure 3).
  • the cell lysis solution was prepared by dissolving a tablet containing a protease inhibitor and a phosphatase inhibitor in a mixed solution of 8 M urea, 1% SDS, and 50 mM Tris pH 8.5.
  • the HL60 cell line was treated with 3 samples of DMSO, 4 samples of Compound 3, and 4 samples of Compound 4 for 4 hours, respectively, and then the cells were lysed with the cell lysis solution.
  • the lysed cell solution was treated with an ultrasonic homogenizer three times for 10 seconds on ice and centrifuged at 14,000 g for 15 minutes at 4 °C. Only the supernatant was separated, and proteomics analysis was performed at the Korea Basic Science Institute (KBSI).
  • GSPT1 is an enzyme that mediates the termination step of protein translation, and degradation of GSPT1 slows down protein translation rate. Accordingly, the degradation of GSPT1 is known to induce inhibition of the expression level of cancer-inducing proteins (for example, c-MYC, N-MYC, L-MYC, BCL-2, MCL-1, and the like) whose expression level is maintained at a high level depending on protein translation in cancer cells (Mullard, Nat Rev Drug Discov. 2022, 21: 865-867). Meanwhile, the degradation of GSPT1 is known to activate a cell death mechanism called "integrated stress response" (Surka et al., Blood 2021, 137(5): 661-677). This response is known to be mediated by ATF-4 and ultimately cause cell death in a caspase 3-dependent manner.
  • integrated stress response a cell death mechanism
  • the protein translation rate was measured using puromycin.
  • Puromycin is a substance commonly used as a means to measure protein translation rate because it can be inserted into the protein sequence instead of tyrosyl-tRNA during the protein translation process.
  • the NCI-H1155 cell line was purchased from the American Type Culture Collection (ATCC, Manassas, VA). The cells were cultured in RPMI 1640 medium (Cytiva) according to recommendations.
  • the HL-60 cell line was purchased from the Korean Cell Line Bank (KCLB, Seoul, Korea) and cultured in RPMI 1640 (Cytiva) supplemented with 10% FBS, 2 mM L-glutamine, and 100 unit/mL streptomycin-penicillin.
  • the NCI-H2023 cell line was purchased from the American Type Culture Collection (ATCC, Manassas, VA). The cells were cultured in DMEM / F12 1:1 medium (Cytiva) according to recommendations.
  • the NCI-H1155 cells, the HL-60 cells, and the NCI-H2023 cells were each seeded in a 6-well plate (2 x 10 6 cells per well). After overnight incubation, the following experiment was performed.
  • the NCI-H1155 cells were treated with 1 ⁇ M of Compound 3. 6 hours after the compound treatment, the cells were treated with 1 ⁇ M puromycin. The cells were harvested 0, 10, 20, 30, and 60 minutes after the puromycin treatment, respectively. The NCI-H2023 cells were treated with 1 ⁇ M of Compound 3 and Reference Compound 1, respectively, in a similar manner. 48 hours after the compound treatment, the cells were treated with 1 ⁇ M puromycin. The cells were harvested 0, 30, 60, and 90 minutes after the puromycin treatment, respectively. The DMSO treated group was used as a control group.
  • the NCI-H1155 cells were treated with 1 ⁇ M of Compound 3. Protein translation was labeled by treatment with 1 ⁇ M puromycin at 0, 2, 4, 8, and 24 hours after the compound treatment. The cells were harvested 30 minutes after the puromycin treatment.
  • the NCI-H1155 cells and the HL60 cells were treated with 0.3 ⁇ M and 1 ⁇ M of Compound 3, respectively. After 0, 2, 4, 6, 8, and 24 hours, the cells were harvested.
  • the cells harvested at each time point were spun down, washed with PBS, lysed with the RIPA buffer supplemented with Protease Inhibitor Cocktail (Roche) and Phosphatase Inhibitor Cocktail (Roche), and then frozen overnight in a -80 °C cryogenic freezer.
  • the samples thawed on ice were centrifuged, and 1x LDS loading buffer and 1x reducing agent were added, and the samples were heated at 95 °C.
  • the prepared samples were loaded on NuPAGE 4-12% Bis-Tris protein gel and transferred to a nitrocellulose membrane. The membrane was blocked with 5% bovine serum albumin and incubated with primary antibodies overnight, and then the corresponding protein signals were detected using HRP-linked secondary antibodies and an iBright CL1500 imaging system.
  • the antibodies used were as follows.
  • mice anti-Puromcyin MABE343, sigma
  • rabbit anti-human eRF3/GSPT1 [EPR22908-103]
  • rabbit anti-human ATF-4 D4B8
  • rabbit anti-human Cleaved Caspase-3 Asp175)
  • mouse anti- ⁇ -Actin-Peroxidase antibody A3854, Sigma
  • the inhibitory efficacy of the compounds of the present invention on cell proliferation was observed using the NCI-H1155 cell line, a lung cancer cell line, and the HL60, MOLM13, MOLM14, MV-4-11, and U937 cells, which are acute myeloid leukemia (AML) cell lines.
  • AML acute myeloid leukemia
  • the NCI-H1155 cell line was purchased from the American Type Culture Collection (ATCC, Manassas, VA). The cells were cultured in RPMI 1640 medium (Cytiva) according to recommendations. The exponentially growing NCI-H1155 cells were plated at 6,000 cells per well in a SPL 33596 white 96-well analysis plate and cultured overnight at 37 °C in a humidified 5% CO 2 incubator. Using a medium mixed with the 3-fold concentration of the test compound, the cells were finally treated at concentrations of 1.5 nM, 4.6 nM, 13.7 nM, 41.2 nM, 123.5 nM, 370.4 nM, 1.111 ⁇ M, 3.333 ⁇ M, and 10 ⁇ M.
  • the HL-60 cell line was purchased from the Korean Cell Line Bank (KCLB, Seoul, Korea). The cells were cultured in RPMI 1640 medium (Cytiva) according to recommendations. The exponentially growing HL-60 cells were plated at 4,000 cells per well in a SPL 33596 white 96-well analysis plate and cultured overnight at 37 °C in a humidified 5% CO 2 incubator. Using a medium mixed with the 3-fold concentration of the test compound, the cells were finally treated at concentrations of 1.5 nM, 4.6 nM, 13.7 nM, 41.2 nM, 123.5 nM, 370.4 nM, 1.111 ⁇ M, 3.333 ⁇ M, and 10 ⁇ M.
  • the cytotoxicity was measured after 72 hours of incubation using the Promega Cell Titer Glo reagent according to the manufacturer's recommendations.
  • the luminescence signals were measured using a Varioskan LUX multimode microplate reader.
  • Relative IC 50 and absolute IC 50 were calculated in the same manner as in the experiment in "3. Evaluation of GSPT1 degradation ability" described above.
  • Experiments using MOLM13, MOLM14, MV-4-11, and U937 cell lines were commissioned by Seoul National University and performed using WST-8 reagent under the drug treatment condition for 72 hours.
  • the exponentially growing MOLM13, MOLM14, MV-4-11, and U937 cells were plated at 4,000 cells per well in a SPL 30096 transparent 96-well analysis plate, respectively, and cultured overnight at 37 °C in a humidified 5% CO 2 incubator. Using a medium mixed with the 2-fold concentration of the test compound, the cells were finally treated at concentrations of 1 nM, 3 nM, 10 nM, 30 nM, 100 nM, 300 nM, 1 ⁇ M, 3 ⁇ M, and 10 ⁇ M. The cytotoxicity was measured after 72 hours of incubation using the CELLOMAX TM WST-8 reagent (PRECAREGENE, CM-VA0500) according to the manufacturer's recommendations. The absorbance of light at a wavelength of 450 nm was measured using a SpectraMAX i3x microplate reader (Molecular Devices, SpectraMAX i3x).
  • Compounds 1 to 20 generally showed improved anticancer efficacy compared to Reference Compounds 1 and 2 in the lung cancer cell lines and AML cell lines tested.
  • SCLC small cell lung cancer
  • LAD lung adenocarcinoma
  • SCLC Small cell lung cancer
  • Lung adenocarcinoma NCI-H1975, A549, NCI-H358, NCI-H460
  • the exponentially growing cells were each plated in a Greiner CELLSTAR #655090 black 96-well analysis plate at the number of cells per well under conditions established by Wuxi Apptec and cultured overnight at 37 °C in a humidified 5% CO 2 incubator. Using a medium mixed with the 10-fold higher concentration of the test compound than the target concentrations, the cells were finally treated at concentrations of 1.5 nM, 4.6 nM, 13.7 nM, 41.2 nM, 123.5 nM, 370.4 nM, 1.111 ⁇ M, 3.333 ⁇ M, and 10 ⁇ M. The cytotoxicity was measured after 72 hours of incubation using the Promega Cell Titer Glo reagent according to the manufacturer's recommendations. The luminescence signals were measured using a 2104 EnVision microplate reader (PerkinElmer, 2104 EnVision Multilabel Reader).
  • Compound 3 and Compound 4 showed minimal cell proliferation inhibition efficacy on the LUAD cell line, while showed overall excellent anticancer effect on the SCLC cell line.
  • SCLC is mostly known as a carcinoma with a neuroendocrine phenotype (Nat Rev Dis Primers. 2021 Jan 14;7(1):3.).
  • lung neuroendocrine cancer Lung NEC
  • NSCLC non-small cell lung cancers
  • NEPC neuroendocrine prostate cancer
  • Lung neuroendocrine cancer (Lung NEC): NCI-H1770, NCI-H2106
  • Neuroendocrine prostate cancer NCI-H660
  • the compound of the present invention exhibited safety against normal cells and the improved therapeutic range accordingly.
  • the cytotoxicity of compounds was compared and evaluated using the HEKa cells, skin primary cells donated from healthy adults.
  • the HEKa cell line was purchased from the American Type Culture Collection (ATCC, Manassas, VA). The cells were cultured in Dermal Cell Basal Medium (ATCC) according to recommendations. The HEKa cells were plated at 10,000 cells per well in a SPL 33596 white 96-well analysis plate and cultured overnight at 37 °C in a humidified 5% CO 2 incubator. Using a medium mixed with the 3-fold concentration of the test compound, the cells were treated at concentrations of 1.5 nM, 4.6 nM, 13.7 nM, 41.2 nM, 123.5 nM, 370.4 nM, 1.111 ⁇ M, 3.333 ⁇ M, and 10 ⁇ M.
  • ATCC American Type Culture Collection
  • VA Dermal Cell Basal Medium
  • the cytotoxicity was measured after 72 hours of incubation using the Promega Cell Titer Glo reagent according to the manufacturer's recommendations.
  • the luminescence signals were measured using a Varioskan LUX multimode microplate reader. Absolute IC 50 was calculated in the same manner as in the experiment in "3. Evaluation of GSPT1 degradation ability" described above.
  • Table 8 The results of comparing the cytotoxicity of the compounds against normal cells are shown in Table 8 below.
  • the therapeutic index in Table 8 was calculated using the IC 50 values of NCI-H1155 and HL60 listed in Table 7.
  • the compounds of the present invention were confirmed to be safer compounds with a wider therapeutic range due to their overall lower cytotoxicity against normal cells (HEKa cells), compared to Reference Compound 1.
  • Reference Compound 1 did not show a significant difference between the concentration at which it exhibited anticancer activity against NCI-H1155 or HL60 and the concentration at which it exhibited cytotoxicity against HEKa cells (i.e., it exhibited a low therapeutic index).
  • Compound 3 and Compound 4 showed a much wider therapeutic window (much higher therapeutic index) compared to Reference Compound 1.
  • Compound 3 showed little cytotoxicity to the HEKa cells, showing a very wide therapeutic window (Figure 10, bottom).
  • HL60-Luc cell line (5x10 6 cells/0.2 mL PBS), which is HL60 cells, an acute myeloid leukemia cell line, labeled with luciferase, were transplanted by injection into the tail vein of 6 to 8-week-old female NCG mice (Charles River), which were completely immune deficient. Bioluminescence was quantified to measure the progression of acute myeloid leukemia (AML). Bioluminescence was recorded from 4 to 10 days after transplantation.
  • FIG. 11a The results of evaluating the drug efficacy of the compound in the HL-60-Luc AML animal model are shown in Figure 11a.
  • Compound 3 inhibited the progression of AML when administered orally at 5 mg/kg daily. Furthermore, at a dose of 15 mg/kg once a day or more, Compound 3 showed tumor remission in addition to the inhibition of the progression of AML.
  • Compound 3 was administered at a dose of 5 mg/kg, 15 mg/kg or 30 mg/kg once a day, at a dose of 15 mg/kg twice a day, or at a dose of 30 mg/kg for 5 days with a 9-day break taken.
  • Compound 4 was administered orally at a dose of 3 mg/kg and 10 mg/kg once a day for 6 days. At 6 and 24 hours after the last dose, cancer tissue was removed and pharmacodynamic changes were observed.
  • Compound 3 significantly reduced tumor proliferation without significant body weight loss at all doses evaluated (Figure 13). Furthermore, similar to the results in the AML animal model, at a dose of 15 mg/kg once a day or more, Compound 3 inhibited the growth of lung cancer tissue and further showed tumor complete regression. In addition, follow-up was conducted until the 40th day of the experiment to confirm whether recurrence of lung cancer was still inhibited even after discontinuation of medication. As a result, when Compound 3 was administered at a dose of 15 mg/kg twice a day, no recurrence of lung cancer was observed, except for 1 out of 9 animals, even after about 1 month after discontinuation of medication.

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Abstract

La présente divulgation concerne un nouveau dégradeur de GSPT1 et son utilisation. Plus particulièrement, la présente divulgation concerne un composé représenté par la formule I, un stéréoisomère, un hydrate, un solvate ou un sel pharmaceutiquement acceptable de celui-ci, une composition pharmaceutique pour le traitement d'un trouble de la prolifération cellulaire incontrôlée le comprenant, et une méthode de traitement d'un trouble de la prolifération cellulaire incontrôlée par administration de celui-ci à un mammifère. Le composé selon la présente divulgation présente une sélectivité élevée et une activité de dégradation prolongée contre GSPT1, une excellente stabilité de pH et une faible cytotoxicité contre des cellules normales, et a ainsi une excellente efficacité anticancéreuse et un indice thérapeutique élevé. De plus, le composé selon la présente divulgation peut présenter une excellente activité anticancéreuse contre un cancer avec un phénotype neuroendocrinien, tel qu'un cancer du poumon à petites cellules (SCLC), un cancer neuroendocrinien pulmonaire (NEC), et un cancer de la prostate neuroendocrinien (NEPC).
PCT/KR2024/004911 2023-04-13 2024-04-12 Nouveau dégradeur de gspt1 et son utilisation Pending WO2024215130A1 (fr)

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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20090060314A (ko) * 2006-08-30 2009-06-11 셀진 코포레이션 5-치환 이소인돌린 화합물
KR20180095094A (ko) * 2016-01-08 2018-08-24 셀진 코포레이션 암을 치료하기 위한 방법 및 치료요법에 대한 임상 감수성의 예측변수로서 바이오마커의 용도
KR20220080003A (ko) * 2020-09-23 2022-06-14 세인트 쥬드 칠드런즈 리써치 호스피탈, 인코포레이티드 세레브론 단백질의 조절제로서의 치환된 n-(2-(2,6-디옥소피페리딘-3-일)-1,3-디옥소이소인돌린-5-일)아릴설폰아미드 유사체
KR102570883B1 (ko) * 2023-04-13 2023-08-29 (주) 사이러스테라퓨틱스 신규한 gspt1 분해제 및 이의 용도

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20090060314A (ko) * 2006-08-30 2009-06-11 셀진 코포레이션 5-치환 이소인돌린 화합물
KR20180095094A (ko) * 2016-01-08 2018-08-24 셀진 코포레이션 암을 치료하기 위한 방법 및 치료요법에 대한 임상 감수성의 예측변수로서 바이오마커의 용도
KR20220080003A (ko) * 2020-09-23 2022-06-14 세인트 쥬드 칠드런즈 리써치 호스피탈, 인코포레이티드 세레브론 단백질의 조절제로서의 치환된 n-(2-(2,6-디옥소피페리딘-3-일)-1,3-디옥소이소인돌린-5-일)아릴설폰아미드 유사체
KR102570883B1 (ko) * 2023-04-13 2023-08-29 (주) 사이러스테라퓨틱스 신규한 gspt1 분해제 및 이의 용도

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
NISHIGUCHI, G. ET AL.: "Identification of potent, selective, and orally bioavailable small-molecule GSPT1/2 degraders from a focused library of cereblon modulators", JOURNAL OF MEDICINAL CHEMISTRY, vol. 64, 2021, pages 7296 - 7311, XP055907311, DOI: 10.1021/acs.jmedchem.0c01313 *

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