Compounds for radioimaging and treatment of cancer

Abstract

The present invention relates to compounds that inhibit fibroblast activation protein (FAP) and their use in treating, preventing and / or imaging cancers associated with fibroblast activation protein expression.

Description

[Technical Field]

[0001] Field The present invention relates to compounds that inhibit fibroblast activation protein (FAP) and their use in treating, preventing and / or imaging cancers associated with fibroblast activation protein expression. [Background technology]

[0002] background Fibroblast-activating protein (FAP) is a type II transmembrane serine protease with dipeptidyl peptidase and endopeptidase activity. FAP, also known as FAP-α, seprase, and α2-antiplasmin-converting enzyme, is generally expressed at low levels in healthy human tissues. In contrast, FAP is often overexpressed in fibroblasts of various cancers and proliferative disorders. FAP is thought to play a role in various tumor-promoting activities, including matrix remodeling, angiogenesis, chemotherapy resistance, and immunosuppression.

[0003] FAP is known to have enzymatic activity as a peptidase, contributing to matrix digestion and tumor microenvironment remodeling, which promotes tumor cell invasion and migration. Neuropeptide Y is a known substrate of FAP, and the cleavage products of neuropeptide Y in the presence of FAP are proangiogenic, suggesting that FAP contributes to tumor angiogenesis. In addition to its enzymatic activity, FAP also participates in cell signaling by forming complexes with other proteins.

[0004] Fibroblasts expressed by cancer cells differ from those associated with healthy cells. Because there are significant differences in FAP expression between healthy and cancerous tissues, treating cancer by administering compounds that exhibit selectivity for FAPs may offer an alternative to current cancer treatments.

[0005] However, for a therapeutic regimen to be useful in treating cancer and related disorders, the administered agent must first exhibit sufficient selectivity for FAP over other receptor sites present in other tissues. Even if a compound exhibits selectivity for FAP, the compound must be retained at the relevant site for a sufficient period of time. Compounds known to be selective for FAP often exhibit limited retention at cancer sites, making them unsuitable for use as part of a therapeutic regimen for the relevant cancer. Furthermore, when a compound selective for FAP is used in radiotherapy and / or radioimaging, the compound must be able to coordinate, retain, and transport the selected radioisotope to the desired site without significant degradation of the compound or loss of the radioisotope. Summary of the Invention [Problem to be solved by the invention]

[0006] There remains a need for compounds that are suitable for use in radiotherapy and / or radioimaging, that are selective for FAPs associated with cancer or related disorders, that have sufficient retention at cancer sites, and that have the necessary chemical stability and pharmacokinetic profile. [Means for solving the problem]

[0007] Summary of the Invention The present inventors have discovered that the compounds disclosed herein can inhibit FAP binding. The compounds disclosed herein comprise a fragment capable of binding to an FAP and a sarcofagin capable of coordinating and retaining a metal ion (e.g., a radioisotope), with the FAP-binding inhibitor and sarcofagin linked via a linker. Because the compounds of the present invention can bind to an FAP and deliver a radioisotope, the present inventors believe that the compounds of the present invention may be useful for selectively targeting malignant sites that overexpress an FAP and then delivering a dose of radioactivity provided by the attached radioisotope to the cancer site.

[0008] In a first aspect, the present invention provides a compound of formula (I), or a salt, complex, isomer, solvate or prodrug thereof: [ka] is provided, During the ceremony, R is H, OH, halogen, cyano, NO2, NH2, optionally substituted C1-C 12 Alkyl, optionally substituted amino, optionally substituted amido, optionally substituted aryl and the following structure: [ka] wherein: X and X 1 may be the same or different and independently represent O, S, NH and N—(C 1-12 alkyl).

[0009] In certain embodiments, the linker in the compound of formula (I) is Optionally substituted C1-C 12 Alkylene, where one or more alkylene groups are O, S, NH, or N—(C 1-12 alkyl); [ka] one or more amino acids; [ka] (wherein n is an integer from 1 to 10); and [ka] The compound comprises one or more moieties selected from the group consisting of:

[0010] In some embodiments, the linker in the compound of Formula (I) is a group: [ka] The compound comprises:

[0011] In some embodiments, the linker in the compound of Formula (I) is [ka] wherein a, b, and c are independently integers selected from 1 to 10.

[0012] In certain embodiments, R is H, NH, optionally substituted C 1- C 12 In a specific embodiment, R is an alkyl or an optionally substituted amido. [ka] and optionally substituted amide having the formula: In the structure, X 1 is O, S, NH or N-(C 1-12 alkyl); The linker is [ka] Optionally substituted C1-C 12 Alkylene, where one or more alkylene groups are O, S, NH, or N—(C 1-12 alkyl); one or more amino acids; [ka] (wherein n is an integer from 1 to 10); and [ka] The compound comprises one or more moieties selected from the group consisting of:

[0013] In certain embodiments, the compound of formula (I) has the structure of formula (Ia): [ka] and In the structure, X and X 1 may be the same or different and independently represent O, S, NH and N—(C 1-12 alkyl); The linker is [ka] Optionally substituted C1-C 12 Alkylene, where one or more alkylene groups are O, S, NH, or N—(C 1-12 alkyl); one or more amino acids; [ka] (wherein n is an integer from 1 to 10); and [ka] The compound comprises one or more moieties selected from the group consisting of:

[0014] In some embodiments, the linker in the compound of Formula (Ia) is a group: [ka] The compound comprises:

[0015] In some embodiments, the linkers in the compound of Formula (Ia) may be the same or different and independently [ka] wherein a, b, and c are independently integers selected from 1 to 10.

[0016] In certain embodiments, the compound of formula (I) is a Sar-FAPi and has the following structure: [ka] It has.

[0017] In certain embodiments, the compound of formula (Ia) is a Sar-bisFAPi and has the following structure: [ka] It has.

[0018] In certain embodiments, the compound of formula (I) is complexed with a metal ion.

[0019] In some embodiments, the compound of formula (I) is complexed with a radioisotope. In some embodiments, the radioisotope is a Cu radioisotope. In some embodiments, the radioisotope is 60 Cu, 61 Cu, 62 Cu, 64 Cu and 67 Cu.

[0020] We have now shown in Figures 9 and 10 that compounds of formula (I) and (Ia) can be radiolabeled with copper radioisotopes. As shown in Figures 11 and 12, the radiolabeled compounds exhibit excellent stability and radiochemical purity at ambient temperature.

[0021] Radiolabeled compounds of formula (I) and (Ia), specifically 64 Cu]Sar-FAPi and [ 64 Cu]Sar-bisFAPi was exposed to SK-MEL 187 (FAP-positive melanoma xenograft) and LNCaP cells. As shown in Figure 13, the inventors found that the compounds of the present invention as disclosed herein exhibit excellent binding affinity to cells expressing the FAP receptor.

[0022] In a second aspect, the present invention provides a composition comprising a compound according to the first aspect and a pharmaceutically acceptable excipient.

[0023] In a third aspect, the present invention provides a method for treating cancer, comprising administering to a subject in need thereof a compound of formula (I) as defined in the first aspect, or a salt, complex, isomer, solvate or prodrug thereof, wherein the compound of formula (I) contains a suitable radioisotope.

[0024] In certain embodiments, the cancer is associated with a fibroblast activation protein (FAP) receptor.

[0025] In a fourth aspect, the present invention provides a method for radioimaging of cancer, comprising administering to a subject in need thereof a compound of formula (I) as defined in the first aspect, or a salt, complex, isomer, solvate or prodrug thereof, wherein the compound of formula (I) contains a suitable radioisotope.

[0026] In certain embodiments of the third and fourth aspects, the radioisotope is a Cu radioisotope. In other embodiments, the Cu radioisotope is 60 Cu, 61 Cu, 62 Cu, 64 Cu and 67 Cu.

[0027] In certain embodiments, the cancer is selected from the group consisting of epithelial ovarian cancer, ovarian cancer, osteosarcoma, pancreatic adenocarcinoma, colorectal cancer, lung cancer, non-small cell lung cancer, gastric cancer, endometrial cancer, pancreatic adenocarcinoma, medullary thyroid carcinoma, differentiated thyroid carcinoma, breast cancer, invasive ductal carcinoma, oral squamous cell carcinoma, esophageal cancer, renal cell carcinoma, insulinoma, prostate cancer, neuroendocrine differentiated prostate cancer, pheochromocytoma, adenoid cystic carcinoma, hepatocellular carcinoma, cervical cancer, small intestine cancer, neuroendocrine tumors, anal cancer, chordoma, desmoid tumor, head and neck cancer, thymic carcinoma, pancreatic cancer, cholangiocarcinoma, esophageal cancer, salivary gland cancer, sarcoma, and cancer of unknown primary origin.

[0028] In a fourth aspect, the present invention provides the use of a compound of formula (I), or a salt, complex, isomer, solvate or prodrug thereof, in the manufacture of a medicament for the treatment of cancer.

[0029] In a sixth aspect, the present invention provides a method for producing a pharmaceutical composition comprising: i) a container comprising a compound of formula (I) as defined in the first aspect, or a pharmaceutically acceptable salt thereof; ii) a vessel comprising a solution of Cu ions; and iii) Instructions for preparing an aqueous formulation of a compound of formula (I) or a pharmaceutically acceptable salt thereof complexed with Cu ions. A kit comprising:

[0030] In certain embodiments, the Cu ions provided as part of the kit are Cu radioisotopes. In certain embodiments, the radioisotopes are 60 Cu, 61 Cu, 62 Cu, 64 Cu and 67 Cu.

[0031] In a seventh aspect, the present invention provides a process for producing a compound of formula (I) or a pharmaceutically acceptable salt thereof, comprising: i) a compound of formula A or a protected form thereof: [ka] preparing a ii) A compound of formula B or a protected form thereof: [ka] preparing a iii) coupling a compound of formula A, or a protected form thereof, with a compound of formula B, or a protected form thereof, to form a compound of formula (I), or a protected form thereof. comprising where R is H, OH, halogen, cyano, NO, NH, optionally substituted C-C 12 Alkyl, optionally substituted amino, optionally substituted amido, optionally substituted aryl and the following structure: [ka] wherein: X and X 1 may be the same or different and independently represent O, S, NH and N—(C1- 12 alkyl) A method is provided.

[0032] In certain embodiments, the coupling of the compound of formula A with the compound of formula B occurs in the presence of a base. [Brief explanation of the drawings]

[0033] [Figure 1A] Figure 1. HPLC trace of Sar-FAPi detected by UV-vis spectroscopy at absorbance A) 254 nm and B) 220 nm, showing the purity of the synthesized compound. [Figure 1B] Figure 1. HPLC trace of Sar-FAPi detected by UV-vis spectroscopy at absorbance A) 254 nm and B) 220 nm, showing the purity of the synthesized compound. [Figure 2] Figure 2. Fragmentation pattern of Sar-FAPi analyzed by mass spectrometry. The detected signals were consistent with the predicted molecular weight (and fragments) of Sar-FAPi. [Figure 3] Figure 3. HPLC traces of Sar-FAPi and [natCu]CuSar-FAPi detected by UV-vis at 280 nm. The difference in retention time indicates that coordination of Cu ions to Sar-FAPi has occurred. [Figure 4] Figure 4. RadioTLC of [64Cu]CuCl2 solution buffered with ammonium acetate. The presence of a single signal by TLC provides a standard retention of [64Cu]CuCl2 and allows comparison when this radioisotope is used to radiolabel Sar-FAPi. [Figure 5]Figure 5. Analysis of Sar-FAPi (1 μg) radiolabeled with 64Cu (4 MBq) by A) RadioHPLC and B: radioTLC. Analysis of the radiolabeling reaction by radioHPLC shows the appearance of new signals when compared to the radioHPLC analysis of the corresponding complex labeled with [natCu]. Analysis by radioTLC also shows the presence of the 64Cu-radiolabelled Sar-FAPi compound. [Figure 6] Figure 6. RadioHPLC trace of Sar-FAPi (10 ng) radiolabeled with 64Cu (4 MBq). Analysis of the radiolabeling reaction by radioHPLC shows the appearance of new signals when compared to analysis of the corresponding complex labeled with [natCu]. [Figure 7] Figure 7. RadioHPLC of the reaction mixture containing Sar-FAPi (1 μg) and [64Cu]CuCl2 (4 MBq) after 10 min and 23 h. Comparison of the radioHPLC traces of the reaction mixture after 10 min and 23 h is virtually identical and contains no new signals corresponding to 64Cu-containing species. This indicates that the 64Cu-radiolabeled Sar-FAPi maintains the same radiochemical purity for at least 23 h and is stable for at least that time. [Figure 8] Figure 8. HPLC traces of [64Cu]Cu-SarFAPi with either histidine or cysteine ​​added. [64Cu]Cu-SarFAPi solutions were treated with either histidine hydrochloride or cysteine ​​hydrochloride and analyzed by HPLC. There was no change in retention time in either case, indicating that the same species (i.e., [64Cu]Cu-SarFAPi) was present and no new species (e.g., copper bound to either histidine or cysteine) were formed. [Figure 9A]Figure 9. RadioTLC traces of [64Cu] buffered with A) PBS or B) NHOAc showing the retention of [64Cu]CuCl; radioTLC traces of Sar-FAPi radiolabeled with [64Cu]CuCl in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, the retention of [64Cu]CuCl was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), but the introduction of [64Cu]CuCl into the Sar-FAPi solution (either PBS or NHOAc) showed the appearance of a new species, namely, [64Cu]Sar-FAPi. In both buffer systems, unchelated copper was still present, so additional Sar-FAPi was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-SarFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 9B]Figure 9. RadioTLC traces of [64Cu] buffered with A) PBS or B) NHOAc showing the retention of [64Cu]CuCl; radioTLC traces of Sar-FAPi radiolabeled with [64Cu]CuCl in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, the retention of [64Cu]CuCl was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), but the introduction of [64Cu]CuCl into the Sar-FAPi solution (either PBS or NHOAc) showed the appearance of a new species, namely, [64Cu]Sar-FAPi. In both buffer systems, unchelated copper was still present, so additional Sar-FAPi was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-SarFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 9C]Figure 9. RadioTLC traces of [64Cu] buffered with A) PBS or B) NHOAc showing the retention of [64Cu]CuCl; radioTLC traces of Sar-FAPi radiolabeled with [64Cu]CuCl in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, the retention of [64Cu]CuCl was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), but the introduction of [64Cu]CuCl into the Sar-FAPi solution (either PBS or NHOAc) showed the appearance of a new species, namely, [64Cu]Sar-FAPi. In both buffer systems, unchelated copper was still present, so additional Sar-FAPi was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-SarFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 9D]Figure 9. RadioTLC traces of [64Cu] buffered with A) PBS or B) NHOAc showing the retention of [64Cu]CuCl; radioTLC traces of Sar-FAPi radiolabeled with [64Cu]CuCl in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, the retention of [64Cu]CuCl was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), but the introduction of [64Cu]CuCl into the Sar-FAPi solution (either PBS or NHOAc) showed the appearance of a new species, namely, [64Cu]Sar-FAPi. In both buffer systems, unchelated copper was still present, so additional Sar-FAPi was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-SarFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 9E]Figure 9. RadioTLC traces of [64Cu] buffered with A) PBS or B) NHOAc showing the retention of [64Cu]CuCl; radioTLC traces of Sar-FAPi radiolabeled with [64Cu]CuCl in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, the retention of [64Cu]CuCl was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), but the introduction of [64Cu]CuCl into the Sar-FAPi solution (either PBS or NHOAc) showed the appearance of a new species, namely, [64Cu]Sar-FAPi. In both buffer systems, unchelated copper was still present, so additional Sar-FAPi was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-SarFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 9F]Figure 9. RadioTLC traces of [64Cu] buffered with A) PBS or B) NHOAc showing the retention of [64Cu]CuCl; radioTLC traces of Sar-FAPi radiolabeled with [64Cu]CuCl in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, the retention of [64Cu]CuCl was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), but the introduction of [64Cu]CuCl into the Sar-FAPi solution (either PBS or NHOAc) showed the appearance of a new species, namely, [64Cu]Sar-FAPi. In both buffer systems, unchelated copper was still present, so additional Sar-FAPi was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-SarFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 10A]Figure 10. RadioTLC traces of [64Cu]CuCl2 buffered with A) PBS or B) NHOAc showing retention of [64Cu]CuCl2; radioTLC traces of Sar-bisFAPi radiolabeled with [64Cu]CuCl2 in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, retention of [64Cu]CuCl2 was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), whereas introduction of [64Cu]CuCl2 into the Sar-bisFAPi solution (either PBS or NHOAc) demonstrated the appearance of a new species, namely, [64Cu]Sar-bisFAPi. In both buffer systems, unchelated copper was still present, so additional Sar-bisFAPia was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-Sar-bisFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 10B]Figure 10. RadioTLC traces of [64Cu]CuCl2 buffered with A) PBS or B) NHOAc showing retention of [64Cu]CuCl2; radioTLC traces of Sar-bisFAPi radiolabeled with [64Cu]CuCl2 in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, retention of [64Cu]CuCl2 was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), whereas introduction of [64Cu]CuCl2 into the Sar-bisFAPi solution (either PBS or NHOAc) demonstrated the appearance of a new species, namely, [64Cu]Sar-bisFAPi. In both buffer systems, unchelated copper was still present, so additional Sar-bisFAPia was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-Sar-bisFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 10C]Figure 10. RadioTLC traces of [64Cu]CuCl2 buffered with A) PBS or B) NHOAc showing retention of [64Cu]CuCl2; radioTLC traces of Sar-bisFAPi radiolabeled with [64Cu]CuCl2 in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, retention of [64Cu]CuCl2 was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), whereas introduction of [64Cu]CuCl2 into the Sar-bisFAPi solution (either PBS or NHOAc) demonstrated the appearance of a new species, namely, [64Cu]Sar-bisFAPi. In both buffer systems, unchelated copper was still present, so additional Sar-bisFAPia was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-Sar-bisFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 10D]Figure 10. RadioTLC traces of [64Cu]CuCl2 buffered with A) PBS or B) NHOAc showing retention of [64Cu]CuCl2; radioTLC traces of Sar-bisFAPi radiolabeled with [64Cu]CuCl2 in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, retention of [64Cu]CuCl2 was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), whereas introduction of [64Cu]CuCl2 into the Sar-bisFAPi solution (either PBS or NHOAc) demonstrated the appearance of a new species, namely, [64Cu]Sar-bisFAPi. In both buffer systems, unchelated copper was still present, so additional Sar-bisFAPia was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-Sar-bisFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 10E]Figure 10. RadioTLC traces of [64Cu]CuCl2 buffered with A) PBS or B) NHOAc showing retention of [64Cu]CuCl2; radioTLC traces of Sar-bisFAPi radiolabeled with [64Cu]CuCl2 in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, retention of [64Cu]CuCl2 was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), whereas introduction of [64Cu]CuCl2 into the Sar-bisFAPi solution (either PBS or NHOAc) demonstrated the appearance of a new species, namely, [64Cu]Sar-bisFAPi. In both buffer systems, unchelated copper was still present, so additional Sar-bisFAPia was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-Sar-bisFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 10F]Figure 10. RadioTLC traces of [64Cu]CuCl2 buffered with A) PBS or B) NHOAc showing retention of [64Cu]CuCl2; radioTLC traces of Sar-bisFAPi radiolabeled with [64Cu]CuCl2 in C) PBS and E) NHOAc, respectively, with the additional addition of Sar-FAPi (see D and F). When EDTA (10 mM) and PBS were used as the mobile phase, retention of [64Cu]CuCl2 was comparable in both radiolabeling buffer systems (PBS and NHOAc, see A and B), whereas introduction of [64Cu]CuCl2 into the Sar-bisFAPi solution (either PBS or NHOAc) demonstrated the appearance of a new species, namely, [64Cu]Sar-bisFAPi. In both buffer systems, unchelated copper was still present, so additional Sar-bisFAPia was added and further analyzed by adioTLC (see C and E). In both buffer systems, the signal representing 64Cu-Sar-bisFAPi increased, indicating complete chelation of the 64Cu ion (i.e., disappearance of free 64Cu) in the presence of NHOAc. [Figure 11A] Figure 11. RadioTLC traces of [64Cu]Sar-FAPi at A) 1 hour and B) 24 hours and radioHPLC traces of [64Cu]Sar-FAPi at C) 1 hour and D) 24 hours. The [64Cu]Sar-FAPi solution was analyzed by radioTLC and radioHPLC at 1 hour and 24 hours after radiolabeling. The compound exhibited excellent stability, with only approximately 5% of the 64Cu radioisotope remaining unchelated after 24 hours at ambient temperature. [Figure 11B] Figure 11. RadioTLC traces of [64Cu]Sar-FAPi at A) 1 hour and B) 24 hours and radioHPLC traces of [64Cu]Sar-FAPi at C) 1 hour and D) 24 hours. The [64Cu]Sar-FAPi solution was analyzed by radioTLC and radioHPLC at 1 hour and 24 hours after radiolabeling. The compound exhibited excellent stability, with only approximately 5% of the 64Cu radioisotope remaining unchelated after 24 hours at ambient temperature. [Figure 11C] Figure 11. RadioTLC traces of [64Cu]Sar-FAPi at A) 1 hour and B) 24 hours and radioHPLC traces of [64Cu]Sar-FAPi at C) 1 hour and D) 24 hours. The [64Cu]Sar-FAPi solution was analyzed by radioTLC and radioHPLC at 1 hour and 24 hours after radiolabeling. The compound exhibited excellent stability, with only approximately 5% of the 64Cu radioisotope remaining unchelated after 24 hours at ambient temperature. [Figure 11D] Figure 11. RadioTLC traces of [64Cu]Sar-FAPi at A) 1 hour and B) 24 hours and radioHPLC traces of [64Cu]Sar-FAPi at C) 1 hour and D) 24 hours. The [64Cu]Sar-FAPi solution was analyzed by radioTLC and radioHPLC at 1 hour and 24 hours after radiolabeling. The compound exhibited excellent stability, with only approximately 5% of the 64Cu radioisotope remaining unchelated after 24 hours at ambient temperature. [Figure 12A] Figure 12. RadioTLC traces of [64Cu]Sar-bisFAPi at A) 1 hour and B) 24 hours and radioHPLC traces of [64Cu]Sar-bisFAPi at C) 1 hour and D) 24 hours. The [64Cu]Sar-bisFAPi solution was analyzed by radioTLC and radioHPLC at 1 hour and 24 hours after radiolabeling. The compound exhibited excellent stability, with little to no unchelated 64Cu radioisotope after 24 hours at ambient temperature. [Figure 12B] Figure 12. RadioTLC traces of [64Cu]Sar-bisFAPi at A) 1 hour and B) 24 hours and radioHPLC traces of [64Cu]Sar-bisFAPi at C) 1 hour and D) 24 hours. The [64Cu]Sar-bisFAPi solution was analyzed by radioTLC and radioHPLC at 1 hour and 24 hours after radiolabeling. The compound exhibited excellent stability, with little to no unchelated 64Cu radioisotope after 24 hours at ambient temperature. [Figure 12C] Figure 12. RadioTLC traces of [64Cu]Sar-bisFAPi at A) 1 hour and B) 24 hours and radioHPLC traces of [64Cu]Sar-bisFAPi at C) 1 hour and D) 24 hours. The [64Cu]Sar-bisFAPi solution was analyzed by radioTLC and radioHPLC at 1 hour and 24 hours after radiolabeling. The compound exhibited excellent stability, with little to no unchelated 64Cu radioisotope after 24 hours at ambient temperature. [Figure 12D] Figure 12. RadioTLC traces of [64Cu]Sar-bisFAPi at A) 1 hour and B) 24 hours and radioHPLC traces of [64Cu]Sar-bisFAPi at C) 1 hour and D) 24 hours. The [64Cu]Sar-bisFAPi solution was analyzed by radioTLC and radioHPLC at 1 hour and 24 hours after radiolabeling. The compound exhibited excellent stability, with little to no unchelated 64Cu radioisotope after 24 hours at ambient temperature. [Figure 13] Figure 13. Chart showing the percentage of cells that bound [64Cu]Sar-FAPi or [64Cu]Sar-bisFAPi after 1 hour of incubation. After 1 hour, approximately 91% of cells showed binding to [64Cu]Sar-bisFAPi, and at least 77% of cells showed binding to [64Cu]Sar-FAPi. DETAILED DESCRIPTION OF THE INVENTION

[0034] Detailed Description Throughout this specification and the claims that follow, unless the context requires otherwise, the word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated integer or step or group of integers or steps, but not to the exclusion of other integers or steps or groups of integers or steps.

[0035] The terms "about" or "approximately," as used herein, mean within an acceptable error range for a particular value as determined by one of ordinary skill in the art, which error range will depend in part on how the value is measured or determined, i.e., the limitations of the measurement system.

[0036] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. For purposes of the present invention, the following terms are defined below.

[0037] The term "pharmaceutically acceptable salt" refers to a salt that retains the desired biological activity of the above-identified compounds, including pharmaceutically acceptable acid addition salts and base addition salts. Suitable pharmaceutically acceptable acid addition salts of compounds of Formula (I) can be prepared from inorganic or organic acids. Examples of such inorganic acids include hydrochloric acid, sulfuric acid, phosphoric acid, methanesulfonic acid, camphorsulfonic acid, oxalic acid, maleic acid, succinic acid, citric acid, formic acid, hydrobromic acid, benzoic acid, tartaric acid, fumaric acid, salicylic acid, mandelic acid, and carbonic acid. Suitable organic acids can be selected from aliphatic, alicyclic, aromatic, and heterocyclic carboxylic and sulfonic acid types of organic acids, including formic acid, acetic acid, propionic acid, succinic acid, glycolic acid, gluconic acid, lactic acid, malic acid, tartaric acid, citric acid, fumaric acid, maleic acid, alkylsulfonic acid, and arylsulfonic acid. Pharmaceutically acceptable salts also include those in which the main compound functions as an acid and reacts with an appropriate base to form, for example, sodium, potassium, calcium, magnesium, ammonium, and choline salts. Those skilled in the art will further recognize that acid addition salts can be prepared by reacting a compound with an appropriate inorganic or organic acid by any of a number of known methods. Alternatively, alkali metal and alkaline earth metal salts can be prepared by reacting a compound with an appropriate base by a variety of known methods. The following are further examples of acid salts which can be obtained by reaction with inorganic or organic acids: acetate, adipate, alginate, citrate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate, digluconate, cyclopentanepropionate, dodecylsulfate, ethanesulfonate, glucoheptanoate, glycerophosphate, hemisulfate, heptanoate, hexanoate, fumarate, hydrobromide, hydroiodide, 2-hydroxy-ethanesulfonate, lactate, maleate, methanesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, palmate, pectinate, persulfate, 3-phenylpropionate, picrate, pivalate, propionate, succinate, tartrate, thiocyanate, tosylate, mesylate and undecanoate.Further information regarding pharmaceutically acceptable salts can be found in Remington's Pharmaceutical Sciences, 19th Edition, Mack Publishing Co., Easton, PA 1995. For drugs that are solid, one of ordinary skill in the art will recognize that the compounds, drugs, and salts of the invention may exist in different crystalline or polymorphic forms, all of which are intended to be within the scope of the invention and the specified formula.

[0038] As used herein, the term "sarcofazine" refers to a nitrogen-containing macrocyclic ligand having the formula 3,6,10,13,16,19-hexazabicyclo[6.6.6]icosane.

[0039] As used herein, the term "optionally substituted" as used throughout the specification denotes that the group may or may not be further substituted with one or more non-hydrogen substituents or fused to one or more non-hydrogen substituents (thus forming a fused polycyclic ring system). In certain embodiments, the substituents are independently selected from halogen, ═O, ═S, —CN, —NO 2 , —CF 3 , —OCF 3 , alkyl, alkenyl, alkynyl, haloalkyl, haloalkenyl, haloalkynyl, heteroalkyl, cycloalkyl, cycloalkenyl, heterocycloalkyl, heterocycloalkenyl, aryl, heteroaryl, cycloalkylalkyl, heterocycloalkylalkyl, heteroarylalkyl, arylalkyl, cycloalkylalkenyl, heterocycloalkylalkenyl, arylalkenyl, heteroarylalkenyl, cycloalkylheteroalkyl, heterocycloalkylheteroalkyl, arylheteroalkyl, heteroarylheteroalkyl, hydroxy, hydroxyalkyl, alkyloxy, alkyloxyalkyl, alkyl. C(=O)OH, C(=O)R, alkyloxycycloalkyl, alkyloxyheterocycloalkyl, alkyloxyaryl, alkyloxyheteroaryl, alkyloxycarbonyl, alkylaminocarbonyl, alkenyloxy, alkynyloxy, cycloalkyloxy, cycloalkenyloxy, heterocycloalkyloxy, heterocycloalkenyloxy, aryloxy, phenoxy, benzyloxy, heteroaryloxy, arylalkyloxy, amino, alkylamino, acylamino, aminoalkyl, arylamino, sulfonylamino, sulfinylamino, sulfonyl, alkylsulfonyl, arylsulfonyl, aminosulfonyl, sulfinyl, alkylsulfinyl, arylsulfinyl, aminosulfinylaminoalkyl, -C(=O)OH, -C(=O)R a , -C(=O)OR a , C(=O)NR a R b , C(=NOH)R a , C(=NR a )NR b R c , N.R. a R b , N.R.a C(=O)R b , N.R. a C(=O)OR b , N.R. a C(=O)NR b R c , N.R. a C(=NR b )NR c R d , N.R. a SO2R b , -SR a , SO2NR a R b , -OR a , OC(=O)NR a R b , OC(=O)R a and acyl groups, wherein R a , R b , R c and R d are each independently H, C1-C 12 Alkyl, C1-C 12 Haloalkyl, C2-C 12 Alkenyl, C2-C 12 Alkynyl, C2-C 10 Heteroalkyl, C3-C 12 Cycloalkyl, C3-C 12 Cycloalkenyl, C2-C 12 Heterocycloalkyl, C2-C 12 Heterocycloalkenyl, C6-C 18 Aryl, C1-C 18 heteroaryl, and acyl, or R a , R b , R c and R d any two or more of these, together with the atoms to which they are attached, form a heterocyclic ring system having 3 to 12 ring atoms.

[0040] In some embodiments, each optional substituent is independently selected from the group consisting of halogen, =O, =S, -CN, -NO2, -CF3, -OCF3, alkyl, alkenyl, alkynyl, haloalkyl, haloalkenyl, haloalkynyl, heteroalkyl, cycloalkyl, cycloalkenyl, heterocycloalkyl, heterocycloalkenyl, aryl, heteroaryl, hydroxy, hydroxyalkyl, alkyloxy, alkyloxyalkyl, alkyloxyaryl, alkyloxyheteroaryl, alkenyloxy, alkynyloxy, cycloalkyloxy, cycloalkenyloxy, heterocycloalkyloxy, heterocycloalkenyloxy, aryloxy, heteroaryloxy, arylalkyl, heteroarylalkyl, arylalkyloxy, amino, alkylamino, acylamino, aminoalkyl, arylamino, sulfonyl, alkylsulfonyl, arylsulfonyl, aminoalkyl, -COOH, -SH, and acyl.

[0041] Examples of particularly suitable optional substituents include F, Cl, Br, I, CH3, CH2CH3, OH, OCH3, CF3, OCF3, NO2, NH2, COOH, COOCH3, and CN.

[0042] As used herein, unless otherwise specified, the term "alkyl" refers to a straight-chain or branched aliphatic hydrocarbon group, preferably C1-C6 12 Alkyl, more preferably C1-C 10 "C1-C6 alkyl" refers to a group or portion of a group that is alkyl, most preferably C1-C6. Examples of suitable straight chain and branched C1-C6 alkyl substituents include methyl, ethyl, n-propyl, 2-propyl, n-butyl, sec-butyl, t-butyl, hexyl, and the like.

[0043] As used herein, the term "halogen" refers to chlorine, fluorine, bromine or iodine.

[0044] As used herein, the term "heteroatom" refers to a nitrogen (N), oxygen (O), or sulfur (S) atom.

[0045] As used herein, the term "alkylene" refers to a divalent straight or branched chain aliphatic hydrocarbon group, e.g., C-C 16 An alkylene group is a divalent hydrocarbon group having 2 to 16 carbon atoms in the chain.

[0046] As used herein, the term "amine" refers to an -NH2 or -NH- group, where the valency of the group depends on the surrounding atoms. For example, when an amine group replaces an alkylene unit, the amine group becomes an -NH- group. When the amine group is in a terminal position, the amine group becomes an -NH2 group. One or more hydrogen atoms (if applicable) may be replaced with a non-hydrogen group, resulting in a substituted amine.

[0047] As used herein, the term "amide" refers to the group -NH-C(O)-. The amide group may be present in either the forward or reverse orientation, and references to an amide group are understood to encompass both forms.

[0048] As used herein, the term "isomer" refers to all stereoisomers of the compounds of the present invention; examples of isomers include diastereomers and enantiomers, where applicable.

[0049] As used herein, the term "amino acid" refers to a molecule containing both amino and carboxyl functional groups. The amino acid may be a natural or unnatural amino acid and may be in equilibrium with its zwitterionic form. The amino acid may contain modifications at either the amino and / or carboxyl termini, or may contain free amino or carboxyl groups. Further modifications of the amino acid side chain or additional substitutions at other portions of the amino acid are also contemplated.

[0050] As used herein, naturally occurring amino acids are the 20 amino acids commonly found in nature in either L- or D-form: glycine (Gly, G), alanine (Ala, A), valine (Val, V), leucine (Leu, L), isoleucine (Ile, I), methionine (Met, M), proline (Pro, P), phenylalanine (Phe, F), tryptophan (Trp, W), serine (Ser, S), threonine (Thr, T), asparagine (Asn, N), glutamine (Gln, Q), tyrosine (Tyr, Y), cysteine ​​(Cys, C), lysine (Lys, K), arginine (Arg, R), histidine (His, H), aspartic acid (Asp, D), and glutamic acid (Glu, E).

[0051] In certain embodiments of compounds of Formula (I), X and X 1 may be the same or different and are O, S, NH or N-(C 1-12 In some embodiments, X and X are selected from alkyl. 1 In some embodiments, X and X are the same. 1 are the same and are O. In other embodiments, in some embodiments, X and X 1 are the same and are NH. In still other embodiments, in some embodiments, X and X 1 is the same, and N-(C 1-12 In some embodiments, X and X are alkyl. 1 are the same and are N-C1 alkyl or N-Me.

[0052] The linker in the compound of formula (I) is [ka] wherein a, b, and c are independently integers selected from 1 to 10.

[0053] In some embodiments, the linker of the compound of Formula (I) has the following structure: [ka] wherein a and c are independently integers selected from 1 to 10, and one or more amino acids is phenylalanine.

[0054] In certain embodiments of the present invention, the linker in the compound of Formula (I) may comprise one or more ethylene oxide groups. In some embodiments, the linker may comprise 1 to 10 ethylene oxide groups. In some embodiments, the linker may comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 ethylene oxide groups. In certain embodiments, the linker may comprise 1, 2, 3, 4, or 5 ethylene oxide groups.

[0055] In other embodiments of the present invention, the linker in the compound of Formula (I) can comprise one or more amino acids, which can be the same or different. In some embodiments, the linker can comprise 1 to 10 amino acids. In certain embodiments, the linker can comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids, which can be the same or different. In certain embodiments, the linker can comprise 1, 2, 3, 4, or 5 amino acid groups, which can be the same or different. In some embodiments, the linker can comprise two or more amino acids, where the amino acids present are the same. In other embodiments, the linker can comprise two or more amino acids, where the amino acids present are different.

[0056] In some embodiments of the present invention, the linker in the compound of formula (I) may comprise a piperazine group, wherein one of the nitrogen atoms of the piperazine group is connected to a propylamide linker: [ka] is a nitrogen atom.

[0057] In the compounds of formula (I), the variable R represents the terminal group of sarcofagin. In the present invention, R is H, OH, halogen, cyano, NO, NH, optionally substituted C-C 12 Alkyl, optionally substituted amino, optionally substituted amido, optionally substituted aryl and groups having the structure: [ka] is a group selected from the group consisting of: X 1 are O, S, NH and N-(C 1-12 alkyl).

[0058] In certain embodiments, R is an optionally substituted C-C 12 In some embodiments, R is an unsubstituted C-C alkyl group. 12 In other embodiments, R is a substituted C-C alkyl group substituted with one or more groups selected from the group consisting of =O, =S, -CN, -NO2, -CF3, -OCF3, alkyl, haloalkyl, haloalkenyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, hydroxy, hydroxyalkyl, alkyloxy, alkyloxyalkyl, aryloxy, heteroaryloxy, arylalkyl, heteroarylalkyl, amino, alkylamino, acylamino, aminoalkyl, arylamino, sulfonyl, alkylsulfonyl, arylsulfonyl, aminosulfonyl, aminoalkyl, -COOH, -SH, and acyl. 12 In certain embodiments, R is a C-C substituted with an alkyl group. 12 In certain embodiments, R is an unsubstituted C group.

[0059] In certain embodiments of compounds of Formula (I), R has the following structure: [ka] and therefore the compound of formula (I) has the structure of a compound of formula (Ia): [ka] wherein X and X 1 may be the same or different, and O, S, NH and N—(C 1-12 alkyl).

[0060] The compound of formula (Ia) comprises two moieties capable of binding to a FAP, each moiety being linked to sarcofagin via a linker. 1 If the are the same, the compound of Formula (Ia) is symmetrical. The compound of Formula (Ia) can be considered a "dimeric" form of the compound of Formula (I). Without wishing to be bound by any particular theory, the inventors believe that by including a second group capable of binding to a targeting site (i.e., a FAP), the compound of Formula (Ia) may exhibit advantages such as better binding and retention in vivo, which in turn may result in less administered compound (and associated radioisotope), potentially leading to fewer side effects and better efficacy. The inventors also believe that the compound of Formula (Ia) comprising a second moiety capable of binding to a FAP is of a suitable size to ensure that the compound is properly metabolized.

[0061] In certain embodiments, the compound of formula (I) has the following structure: [ka] [ka] It has one of the following:

[0062] The compounds of formula (I) have one or more stereocenters. In some embodiments of the compounds of formula (I), a particular configuration of the stereocenters is preferred to impart a desired biological effect and / or binding property. For example, the compounds of formula (I) comprise a nitrile group attached to a carbon atom of a pyrrolidine ring. In a preferred embodiment, the pyrrolidine ring has the configuration shown below. The compounds of formula (I) may also comprise one or more amino acids, each having a defined stereochemistry, e.g., the stereochemistry in which the amino acid occurs naturally. In a preferred embodiment, the compounds of formula (I) have the following structure: [ka] [ka] It has.

[0063] In certain embodiments, the compound of formula (I) is a Sar-FAPi and has the following structure: [ka] It has.

[0064] In certain embodiments, the compound of formula (I) has the structure of formula (Ia): [ka] wherein: X and X 1 may be the same or different and independently represent O, S, NH and N—(C 1-12 alkyl); The linker is [ka] Optionally substituted C1-C 12 Alkylene, where one or more alkylene groups are O, S, NH, or N—(C 1-12 alkyl); one or more amino acids; [ka] (wherein n is an integer from 1 to 10); and [ka] The compound comprises one or more moieties selected from the group consisting of:

[0065] In certain embodiments, the compound of Formula (Ia) has the following structure: [ka] It has.

[0066] In certain embodiments, the compound of formula (Ia) is a Sar-bisFAPi and has the following structure: [ka] It has.

[0067] The compounds of formula (I) can be coordinated with a metal ion via the nitrogen-containing macrocycle to form the corresponding complex of formula (I). In one embodiment, the compounds of formula (I) are coordinated with a metal ion.

[0068] In one embodiment, the metal ion is an ion of Cu, Tc, Gd, Ga, In, Co, Re, Fe, Mg, Ag, Rh, Pt, Cr, Ni, V, Ir, Zn, Cd, Mn, Ru, Pd, Hg, Ti, Lu, Sc, Zr, or Pb.

[0069] The compounds have been found to be particularly useful for binding copper ions. In some embodiments, the metal ion is 60 Cu, 61 Cu, 62 Cu, 64 Cu and 67 In some embodiments, the radioisotope is selected from the group consisting of Cu. 60 In some embodiments, the radioisotope is Cu. 61In some embodiments, the radioisotope is Cu. 62 In some embodiments, the radioisotope is Cu. 64 In some embodiments, the radioisotope is Cu. 67 It is Cu.

[0070] The complexes described herein are radiolabeled with radioisotopes that undergo spontaneous decay, and the by-products of these decays are detected by various means, such as positron emission tomography (PET) or single photon emission computed tomography (SPECT). The quality of the images obtained, and therefore the reliability of diagnoses based on these images, depends on the ability of the radiolabeled complex to specifically bind to the FAP.

[0071] When the metal ion is a radioisotope and the compound of formula (I) is radiolabeled to form a complex, the complex can be administered for the purpose of radiotherapy or radioimaging.Since the compound of formula (I) (and thus the radioisotope-labeled complex) contains a group that can bind to a biological receptor, the radioisotope-labeled complex of formula (I) can be used for radiotherapy or radioimaging of cancers associated with overexpression of the target site to which the compound of formula (I) binds.

[0072] The present inventors have discovered that compounds and complexes of formula (I) containing sarcofagin and one or more biologically active moieties linked by a linker group can bind to and inhibit FAPs. The combination of these components in compounds of formula (I) allows for the administration of the corresponding complex containing a radionuclide, maintaining the stability of the complex in vivo, and allowing the complex to accumulate at its intended target, i.e., the FAP. For the biologically active moiety to bind to its intended target and deliver the coordinated radionuclide, sufficient distance must exist between the groups to prevent reaction between them. The present inventors have discovered that the linker group defined herein (i.e., at least a propylamide linker) and sarcofagin itself provide compounds in which the distance between the albumin-binding group and the group capable of binding to a biological receptor prevents such reaction. In addition to contributing to the required distance, the linker comprising the nature of the linker group may modify the lipophilicity of the compound, improving the hydrolytic stability of the compound and its various fragments.

[0073] The compounds of the present invention and their complexes with radioisotopes can be used in methods of radioimaging, diagnosis, or treatment. In some embodiments, the compounds of the present invention complexed with radionuclides can be used in methods for radioimaging, diagnosis, or treatment of cancer.

[0074] As used herein, the terms "treat," "treatment," "prevent," "prevention," and grammatical equivalents refer to any use that improves the described neuroendocrine tumor, prevents, delays, or slows the establishment of the disease, or otherwise prevents, hinders, slows, or reverses the progression of the disease. Thus, terms such as "treat" and "prevent" should be considered in their broadest context. For example, "treatment" does not necessarily mean that the patient is treated until cured. When a disease exhibits or is characterized by multiple symptoms, treatment or prevention does not necessarily improve, prevent, hinder, slow, or reverse all of the symptoms, but may prevent, hinder, slow, or reverse one or more of the symptoms.

[0075] As used herein, the term "cancer" broadly encompasses neoplastic diseases characterized by abnormal cell growth that may invade or metastasize to other parts of the body. Cancer may be benign, meaning that it does not spread to other parts of the body. Cancer may also be malignant, meaning that cancer cells may spread through the circulatory or lymphatic system. As used herein, the term includes all malignant, i.e., cancerous, conditions. Cancer may exist as a tumor. In certain embodiments, the term as used herein refers to a cancer characterized by overexpression of a FAP.

[0076] As used herein, the term "tumor" refers to any malignant cancerous or pre-cancerous cell growth. The term is particularly directed to solid tumors or carcinomas, although it may also include leukemias.

[0077] In connection with the administration of the complex of formula (I), radioimaging of cancers related to receptor expression also depends on the selection of a suitable radionuclide.For example, when the intended purpose of the complex of formula (I) is radioimaging, the selected radionuclide should have a sufficiently long half-life so that the detection of the decay of the radionuclide can obtain images of sufficient quality.This also requires that the compound of formula (I) itself, i.e., the ligand that coordinates the radionuclide, is sufficiently stable against radioactive decay.The inventors have found that the decomposition of the complex of formula (I) by radiolysis (i.e., as a result of the radioactivity of the radionuclide) is minimized, and in this respect, the complex of formula (I) generally remains intact.

[0078] Radioimaging of a subject to which a radiolabeled compound of formula (I) is administered can be by positron emission tomography (PET) or single photon emission computed tomography (SPECT). In one embodiment, the present invention provides a method for radioimaging a subject in need thereof, the method comprising administering a compound of formula (I) complexed with a radionuclide. In one embodiment, the method comprises administering a compound of formula (I) complexed with a copper radionuclide. In another embodiment, the method comprises: 64 administering a compound of formula (I) complexed with Cu.

[0079] In one embodiment, the radioimaging of the subject after administration of the radionuclide-complexed compound of Formula (I) is by PET. In another embodiment, the radioimaging of the subject after administration of the radionuclide-complexed compound of Formula (I) is by SPECT.

[0080] The term "subject," as used herein, refers to a mammal, including humans, primates, livestock (e.g., sheep, pigs, cows, horses, donkeys), clinical trial animals (e.g., mice, rabbits, rats, guinea pigs), performance and show animals (e.g., horses, livestock, dogs, cats), companion animals (e.g., dogs, cats), and domesticated wild animals. Preferably, the mammal is a human or a clinical trial animal. Even more preferably, the mammal is a human.

[0081] The compounds of the present invention complexed with a radionuclide can be administered to a subject in need thereof as a composition via a parenteral route. Administration by intravenous injection may be preferred. Alternatively, the formulations of the present invention may be administered intraarterially or via other routes for delivery to the systemic circulation. The subject to which the compound has been administered is then placed in a PET (or SPECT) scanner to obtain images showing the location of the complex and, therefore, the location of the cancer or tumor. This then allows for the diagnosis and detection of the cancer or tumor.

[0082] The compounds of the present invention and their complexes with radionuclides can be used in methods for treating diseases such as cancer. When complexed with a suitable radionuclide, the complexes of the present invention can be administered to a subject in need thereof. The methods disclosed herein comprise administering a therapeutically effective amount of a radiolabeled compound of the present invention to a subject in need thereof. In one embodiment, the present invention provides a method for treating a disease in a subject in need thereof, the method comprising administering a therapeutically effective amount of a compound of formula (I) complexed with a radionuclide.

[0083] The term "therapeutically effective amount" or "effective amount" refers to an amount sufficient to achieve a beneficial or desired clinical result. An effective amount can be administered in one or more administrations. For radioimaging purposes, an effective amount is an amount sufficient to provide an image showing the location of a compound of formula (I) administered to a subject by detection of decay products from a radioisotope complexed with the compound. For therapeutic purposes, an effective amount is generally sufficient to palliate, ameliorate, stabilize, reverse, slow and / or delay the progression of cancer.

[0084] In one embodiment, the present invention provides a method for treating cancer, the method comprising administering a compound of formula (I) complexed with a radionuclide. In one embodiment, the method comprises administering a compound of formula (I) complexed with a copper radionuclide. In another embodiment, the method comprises administering 67 In certain embodiments, the method comprises administering a compound of formula (I) complexed with Cu. 67 The method comprises administering a compound of Formula (I) complexed with Cu, wherein the cancer is associated with fibroblast activation protein (FAP). In another embodiment, the method comprises administering a compound of Formula (I) comprising a biologically active moiety that binds to a FAP. In another embodiment, the method comprises administering a compound of Formula (I) comprising a biologically active moiety that inhibits a FAP. In another embodiment, the method comprises administering a compound of Formula (I) comprising a biologically active moiety that binds to and inhibits a FAP.

[0085] Examples of cancer types treatable by administration of the compounds of the present invention include those classified as carcinomas, lymphomas, and sarcomas. Examples of carcinomas include, but are not limited to, adenocarcinoma, acinar cell adenocarcinoma, adrenocortical cell carcinoma, alveolar cell carcinoma, undifferentiated carcinoma, basaloid carcinoma, basal cell carcinoma, bronchiolar carcinoma, bronchogenic carcinoma, renaladinol carcinoma, embryonal carcinoma, anometroid carcinoma, fibrolamolar liver cell carcinoma, follicular carcinoma, giant cell carcinoma, hepatocellular carcinoma, intraepidermal carcinoma, carcinoma in situ, leptomanigio carcinoma, medullary carcinoma, melanoma, menigual carcinoma, mesometonephric carcinoma, oat cell carcinoma, squamous cell carcinoma, sweat gland carcinoma, transitional cell carcinoma, and tubular cell carcinoma. Examples of sarcomas include, but are not limited to, amelioblastic sarcoma, angiolithiasis sarcoma, botryoid sarcoma, endometrial stromal sarcoma, Ewing's sarcoma, fascicularis sarcoma, giant cell sarcoma, granulositic sarcoma, immunoblastic sarcoma, juxaccordial osteogenic sarcoma, copices sarcoma, leukocyte sarcoma (leukemia), lymphocytic sarcoma (lymphosarcoma), medullary sarcoma, myeloid sarcoma (granulocytic sarcoma), austiogenci sarcoma, periosteal sarcoma, reticulum cell sarcoma (histiocytic lymphoma), round cell sarcoma, spindle cell sarcoma, synovial sarcoma, and telangiectatic audiogenic sarcoma. Examples of lymphomas include, but are not limited to, Hodgkin's disease and lymphocytic lymphomas, such as Burkitt's lymphoma, nodular poorly differentiated lymphocytic lymphoma, nodular mixed lymphoma, nodular histocytic lymphoma, and diffuse lymphoma.

[0086] Examples of cancers that can be treated with the compounds disclosed herein include, but are not limited to, Hodgkin's disease, non-Hodgkin's lymphoma, acute lymphocytic leukemia, multiple myeloma, breast cancer, ovarian cancer, lung cancer, Wilms' tumor, testicular cancer, soft tissue sarcoma, chronic lymphocytic leukemia, primary macroglobulinemia, bladder cancer, chronic granulocytic leukemia, primary brain tumor, malignant melanoma, small cell lung cancer, gastric cancer, and ovarian cancer. These include intestinal cancer, malignant pancreatic insulinoma, malignant carcinoid cancer, malignant melanoma, choriocarcinoma, mycosis fungoides, head and neck cancer, osteogenic sarcoma, pancreatic cancer, acute granulocytic leukemia, hairy cell leukemia, rhabdomyosarcoma, Kaposi's sarcoma, genitourinary cancer, thyroid cancer, esophageal cancer, malignant hypercalcemia, renal cell carcinoma, endometrial cancer, polycythemia vera, essential thrombocytosis, adrenocortical carcinoma, skin cancer, and prostate cancer.

[0087] In certain embodiments, the cancer is lung cancer, testicular cancer, kidney cancer, bladder cancer, kidney or renal cancer, ovarian cancer, breast cancer, fallopian tube cancer, uterine leiomyoma, prostate cancer, non-Hodgkin's lymphoma, colon cancer, lipoma, basal cell skin cancer, squamous cell skin cancer, osteosarcoma, acute myeloid leukemia (AML), pancreatic cancer, prostate cancer, CNS cancer, retinoblastoma, neuroblastoma, glioblastoma, Kaposi's sarcoma, Ewing's sarcoma, rhabdomyosarcoma, hemangioma, solid tumor, blood-borne tumor, leukemia, or melanoma.

[0088] Other specific examples of cancer types include lung cancer (e.g., bronchogenic carcinoma, small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC), lung adenocarcinoma), kidney cancer (e.g., nephroblastoma or Wilms' tumor, renal cell carcinoma), acoustic neuroma, adenocarcinoma, adenocarcinoma of the adrenal gland, anal cancer, angiosarcoma (e.g., lymphangiosarcoma, lymphangioendothelial sarcoma, angiosarcoma), appendix cancer, benign monoclonal gammopathy, biliary tract cancer (e.g., cholangiocarcinoma), bladder cancer, breast cancer (e.g., adenocarcinoma of the breast, uterine ulcer, thyroid cancer ... papillary carcinoma, adenocarcinoma of the breast, medullary carcinoma of the breast), brain tumors (e.g., meningioma, glioblastoma, glioma (e.g., astrocytoma, oligodendroglioma), medulloblastoma), bronchial carcinoma, carcinoid tumor, cervical cancer (e.g., cervical adenocarcinoma), choriocarcinoma, chordoma, craniopharyngioma, colorectal cancer (e.g., colon carcinoma, rectal carcinoma, colorectal adenocarcinoma), connective tissue carcinoma, epithelial carcinoma, ependymoma, endothelial sarcoma (e.g., Kaposi's sarcoma, multiple idiopathic hemorrhagic sarcoma), endometrial cancer (e.g., uterine carcinoma, uterine sarcoma), esophageal cancer (e.g., esophageal adenocarcinoma, Barrett's adenocarcinoma), Ewing's sarcoma, eye cancer (e.g., intraocular melanoma, retinoblastoma), familial eosinophilia, gallbladder cancer, stomach cancer (e.g., gastric adenocarcinoma), gastroesophageal cancer, gastrointestinal stromal tumor (GIST), germ cell cancer, head and neck cancer (e.g., head and neck squamous cell carcinoma, oral cancer (e.g., oral squamous cell carcinoma), throat cancer (e.g., laryngeal cancer, pharyngeal cancer, nasopharyngeal cancer, oropharyngeal cancer)), H chain disease (e.g., alpha chain disease, gamma chain disease, μ chain disease, vascular blastoma, hypopharyngeal carcinoma, inflammatory myofibroblastic tumor, immune cell amyloidosis), liver cancer (e.g., hepatocellular carcinoma (HCC), malignant hepatoma, hepatocholangiocarcinoma), leiomyosarcoma (LMS), mastocytosis (e.g., systemic mastocytosis), muscle cancer, myelodysplastic syndrome (MDS), mesothelioma, myeloproliferative disorders (MPD) (e.g., polycythemia vera (PV), essential thrombocytosis (ET), primary myelofibrosis (AMM) or myelofibrosis (MF), chronic myelofibrosis (CRM),Idiopathic myelofibrosis, chronic myelocytic leukemia (CML), chronic neutrophilic leukemia (CNL), hypereosinophilic syndrome (HES), neuroblastoma, neurofibroma (e.g., type 1 or type 2 neurofibromatosis (NF), schwannomatosis), neuroendocrine carcinoma (e.g., gastrointestinal pancreatic neuroendocrine tumor (GEP-NET), carcinoid tumor), osteosarcoma (e.g., bone cancer), ovarian cancer (e.g., cystadenocarcinoma, ovarian embryonal carcinoma, ovarian adenocarcinoma), lactate adenocarcinoma, pancreatic cancer (e.g., pancreatic adenocarcinoma, intraductal papillary mucinous neoplasm (IPMN), islet cell tumor), penile cancer (e.g., Paget's disease of the penis and scrotum), pinealoma, primitive neuroectodermal tumor (PNT), plasma cell Neoplasms, paraneoplastic syndromes, intraepithelial neoplasia, rectal cancer, rhabdomyosarcoma, salivary gland cancer, skin cancer (e.g., squamous cell carcinoma (SCC), keratoacanthoma (KA), melanoma, basal cell carcinoma (BCC)), small intestine cancer (e.g., appendix cancer), soft tissue sarcomas (e.g., malignant fibrous histiocytoma (MFH), liposarcoma, malignant peripheral nerve sheath tumor (MPNST), chondrosarcoma, fibrosarcoma, myxosarcoma), sebaceous gland carcinoma, small intestine carcinoma, sweat gland carcinoma, synovial tumor; testicular cancer (e.g., seminoma, testicular embryonal carcinoma), thyroid cancer (e.g., papillary thyroid carcinoma, papillary thyroid carcinoma (PTC), medullary thyroid carcinoma), urethral cancer, vaginal cancer, and vulvar cancer (e.g., Paget's disease of the vulva).

[0089] Examples of specific types of breast cancer include lobular carcinoma in situ (LCIS), ductal carcinoma in situ (DCIS), invasive ductal carcinoma (IDC), inflammatory breast cancer, Paget's disease of the nipple, Phyllodes' tumor, angiosarcoma, adenoid cystic carcinoma, low-grade adenosquamous carcinoma, medullary carcinoma, mucinous carcinoma, papillary carcinoma, tubular carcinoma, metastatic carcinoma, micropapillary carcinoma, mixed carcinoma, or other breast cancers, including triple-negative (TNBC), HER-positive, neoadjuvant HER2-negative, estrogen receptor-positive, progesterone receptor-positive, HER·estrogen receptor-positive, HER·progesterone receptor-positive, estrogen·progesterone receptor-positive, and HER·estrogen·progesterone receptor-positive.

[0090] Examples of specific types of ovarian cancer include epithelial ovarian carcinoma (EOC), mature teratoma, dysblastoma, endodermal sinus tumor, granulosa tumor, Sertoli-Leydig cell tumor, primary peritoneal carcinoma, small cell carcinoma of the ovary (SCCO), ovarian teratoma, ovarian sex cord-stromal carcinoma, ovarian dysgerminoma, choriocarcinoma, carcinosarcoma, adenosarcoma, leiomyosarcoma, fibrosarcoma, and Krukenberg tumor.

[0091] Examples of specific types of pancreatic cancer include tumors affecting the exocrine glands, exocrine tumors, endocrine tumors, islet cell tumors, neuroendocrine tumors, cystic tumors, acinar cell carcinoma, insulinoma, somatostatinoma, gastrinoma, glucagonoma, pancreatic adenocarcinoma, pancreatic blastoma, pancreatic sarcoma, adenosquamous carcinoma, colloid carcinoma, hepatocarcinoid, intraductal papillary mucinous neoplasm, mucinous cystic neoplasm, pancreatic intraepithelial neoplasm, pancreatoblastoma, serous cystadenoma, signet ring cell carcinoma, solid pseudopapillary neoplasm, and undifferentiated carcinoma with osteoclast-like giant cells.

[0092] Examples of specific types of prostate cancer include prostate carcinoma, acinar adenocarcinoma, tubular adenocarcinoma, transitional cell (or urothelial) carcinoma, squamous cell carcinoma, small cell prostate carcinoma, carcinoid, sarcoma, small cell carcinoma, neuroendocrine tumor, and transitional cell carcinoma.

[0093] In certain embodiments, the cancer is selected from the group consisting of epithelial ovarian cancer, ovarian cancer, osteosarcoma, pancreatic adenocarcinoma, colorectal cancer, lung cancer, non-small cell lung cancer, gastric cancer, endometrial cancer, pancreatic adenocarcinoma, medullary thyroid carcinoma, differentiated thyroid carcinoma, breast cancer, invasive ductal carcinoma, oral squamous cell carcinoma, esophageal cancer, renal cell carcinoma, insulinoma, prostate cancer, neuroendocrine differentiated prostate cancer, pheochromocytoma, adenoid cystic carcinoma, hepatocellular carcinoma, cervical cancer, small intestine cancer, neuroendocrine tumors, anal cancer, chordoma, desmoid tumor, head and neck cancer, thymic carcinoma, pancreatic cancer, cholangiocarcinoma, esophageal cancer, salivary gland cancer, sarcoma, and cancer of unknown primary origin.

[0094] The compounds and complexes of the present invention can be administered alone or in combination with a pharmaceutically acceptable carrier, diluent, or excipient in the form of a pharmaceutical composition. The compounds of the present invention, although effective in themselves, are generally formulated and administered in the form of their pharmaceutically acceptable salts, as these forms are generally more stable, more prone to crystallization, and more soluble.

[0095] The compounds of the present invention are generally used in the form of pharmaceutical compositions formulated according to the desired mode of administration, and these compositions are prepared in a manner well known in the art.

[0096] When using the compounds of the present invention, they can be administered in any form or manner that makes the compound available for the desired application (imaging or radiotherapy).Those skilled in the art of preparing this type of formulation can easily select the appropriate form and administration mode depending on the specific characteristics of the selected compound, the pathology to be treated, the stage of the pathology to be treated and other relevant circumstances.For further information, please refer to Remington's Pharmaceutical Sciences, 19th edition, Mack Publishing Co. (1995).In certain embodiments, the compound of formula (I) is administered by injection.In other embodiments, the compound of formula (I) is administered intravenously.In other embodiments, the compound of formula (I) is administered directly to the site of cancer.

[0097] In another embodiment, the present invention provides a pharmaceutical pack or kit comprising one or more containers filled with one or more of the components of the pharmaceutical composition of the present invention. In such a pack or kit, at least one container containing a unit dose of the drug can be found. For convenience, these kits can provide single doses in sterile vials, allowing the clinician to use the vials directly, containing the desired amounts and concentrations of compound and radionucleotide, which can be mixed before use. Such containers can be accompanied by various documents, such as instructions for use or notices in a format prescribed by a government agency regulating the manufacture, use, or sale of pharmaceuticals, imaging agents, or biological agents, reflecting approval by the agency for manufacture, use, or sale for human administration.

[0098] In one embodiment, the present invention provides a composition comprising a compound as described above, together with one or more pharmaceutically acceptable excipients.

[0099] Pharmaceutical compositions of the present invention for parenteral injection include pharmaceutically acceptable sterile aqueous or non-aqueous solutions, dispersions, suspensions, or emulsions, as well as sterile powders for reconstitution into sterile injectable solutions or dispersions immediately prior to injection. Examples of suitable aqueous and non-aqueous carriers, diluents, solvents, or vehicles include water, ethanol, polyols (e.g., glycerol, propylene glycol, polyethylene glycol, etc.), and suitable mixtures thereof, vegetable oils (e.g., olive oil), and injectable organic esters such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of coatings such as lecithin, the maintenance of the required particle size in the case of dispersions, or the use of surfactants. Proper fluidity can be maintained, for example, by the use of coatings such as lecithin, the maintenance of the required particle size in the case of dispersions, or the use of surfactants.

[0100] These compositions may also contain adjuvants such as preservatives, wetting agents, emulsifiers, and dispersants. Prevention of microbial action can be ensured by including various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol sorbic acid, etc. In addition, it is also preferable to include isotonic agents such as sugars and sodium chloride. The inclusion of agents that delay absorption, such as aluminum monostearate and gelatin, can prolong the absorption of injectable pharmaceutical forms.

[0101] If desired, and for more effective distribution, the compounds can be incorporated into slow-release or targeted-delivery systems such as polymer matrices, liposomes, and microspheres.

[0102] Injectable preparations can be sterilized, for example, by filtration through a bacterial-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium immediately before use.

[0103] The present invention also provides a method for synthesizing or preparing the compounds of the present invention. The inventors have found that the established procedures that can be used to prepare the compounds of the present invention by various coupling procedures and conditions cannot obtain the desired compounds. This is mainly due to incompatibility between functional groups, solubility problems of reagents, and general problems of reactivity.

[0104] The present inventors have found that compounds of formula (I) and (Ia) can be synthesized by various routes, for example, following the schemes disclosed in WO2019 / 154886 and WO2019 / 154859 (the disclosures of which are incorporated herein by reference). A simplified route to compounds of formula (I) is shown in Scheme 1, in which an isoquinolone (or N-acylated aminoacylpyrrolidine-substituted isoquinoline) group is sequentially coupled to a linker and a sarcofagin moiety. [ka]

[0105] Scheme 1. i) BBr3; ii) HBTU / HOBt, DIPEA, H-Gly-Pro-FF-CN; iii) 1-Bromo-3-chloropropane, Cs2CO3, DMF; iv) NaN3, DMF; v) PPh3, MeCN; vi) tBoc4MeCOSar-NHS, DIPEA, DMF; vii) TFA.

[0106] Alternatively, the compounds of formula (I), difluoropyrrolidine-substituted isoquinolones, can be prepared in several steps from simple precursor molecules as shown in Schemes 2-4.

[0107] Scheme 2 illustrates the synthesis of embodiments of compounds of formula (I) comprising specific linkers, where the difluoropyrrolidine group is linked to the precursor: [ka] It is prepared from

[0108] Scheme 2. Synthesis of compounds of formula (I): i) BBr3; ii) 1-bromo-3-chloropropane, Cs2CO3, DMF; iii) NaN3, DMF; iv) PPh3, MeCN; v) t-Boc2O, MeCN; vi) HBTU / HOBt, DIPEA, H-Gly-Pro-FF-CN, DMF; vii) TFA; viiii) tBoc4MeCOSar-NHS, DIPEA, DMF; ix) TFA.

[0109] Scheme 3 illustrates the synthesis of another embodiment of a compound of formula (I) having a linker comprising one or more amino acids, where the difluoropyrrolidine group is prepared from a precursor. [ka]

[0110] Scheme 3. Synthesis of compounds of formula (I) comprising an amino acid linker: i) BBr3; ii) 1-bromo-3-chloropropane, Cs2CO3, DMF; iii) NaN3, DMF; iv) PPh3, MeCN; v) t-Boc-Phe-Phe-NHS, MeCN; vi) HBTU / HOBt, DIPEA, H-Gly-Pro-FF-CN, DMF; vii) TFA; viii) tBoc4bisCOSar-NHS2, DIPEA, DMF; ix) TFA.

[0111] Scheme 4 illustrates the synthesis of an embodiment of a compound of Formula (Ia) comprising sarcofagin and two difluoropyrrolidine groups prepared from precursors, where each moiety is joined by a linker. [ka]

[0112] Scheme 4. Synthesis of compounds of formula (Ia) comprising two linkers and two biologically active FAP inhibitor moieties: i) BBr; ii) 1-bromo-3-chloropropane, CsCO, DMF; iii) NaN, DMF; iv) PPh, MeCN; v) t-BocO, MeCN; vi) HBTU / HOBt, DIPEA, H-Gly-Pro-FF-CN, DMF; vii) TFA; viiii) tBocbisCOSar-NHS, DIPEA, DMF; ix) TFA.

[0113] The inventors have also found that the routes described in Schemes 5 and 6 can also provide for obtaining compounds of Formula (I) or Formula (Ia). In contrast to the routes described in Schemes 1-4, the route detailed in Scheme 5 does not use an intermediate azide functional group, which is associated with various health and safety issues. Furthermore, the routes of Schemes 5 and 6 require fewer steps due to the reduced number of functional group transformations required. Without wishing to be bound by any particular theory, the inventors believe that the synthetic routes disclosed herein provide a variety of methods for obtaining compounds of the present invention.

[0114] Scheme 5 shows a modified route to compounds of formula (I), in which the components for inhibiting FAP are prepared in three steps from pyridine precursors similar to those used in Schemes 1-4. The TFA-protected form of the inhibitor component is then coupled under standard peptide coupling conditions to give compounds of formula (I). In Scheme 6, MeCOSar (i.e., methyl-substituted sarcofagin) is replaced with bisCOSar. [ka]

[0115] Scheme 5. Synthesis of compounds of formula (I). i) SOCl2 / MeOH; ii) Br(CH2)2NHBoc, Cs2CO3, DMF, Cs2CO3, DMF; iii) LiOH, THF; iv) Boc4MeCOSar-NHS, Et3N, DMF; vi) TFA. [ka]

[0116] Scheme 6. Synthesis of compounds of formula (Ia). iv) BisCOSar(NHS)2, Et3N, DMF; vi) TFA.

[0117] Thus, in a further aspect, the present invention provides a process for preparing a compound of formula (I) or a pharmaceutically acceptable salt thereof, the process comprising: i) a compound of formula A or a protected form thereof: [ka] preparing a ii) A compound of formula B or a protected form thereof: [ka] preparing a iii) coupling a compound of formula A, or a protected form thereof, with a compound of formula B, or a protected form thereof, to form a compound of formula (I), or a protected form thereof. wherein R is H, OH, halogen, cyano, NO2, NH2, optionally substituted C1-C 12 Alkyl, optionally substituted amino, optionally substituted amido, optionally substituted aryl and the following structure: [ka] wherein: X and X 1 may be the same or different and independently represent O, S, NH and N—(C 1-12 alkyl).

[0118] In one embodiment, the protected form of the compound of formula A contains a nitrogen protecting group or an oxygen protecting group. In another embodiment, the protected form of the compound of formula B contains a nitrogen protecting group or an oxygen protecting group.

[0119] As used herein, the term "oxygen protecting group" refers to a group that can prevent reaction of the oxygen moiety during further derivatization of the protected compound and can be easily removed when desired. In one embodiment, the protecting group is removable by natural metabolic processes under physiological conditions. Examples of oxygen protecting groups include acyl groups (e.g., acetyl), ethers (e.g., methoxymethyl ether (MOM), α-methoxyethoxymethyl ether (MEM), p-methoxybenzyl ether (PMB), methylthiomethyl ether, pivaloyl (Piv), tetrahydropyran (THP)), and silyl ethers (e.g., trimethylsilyl (TMS), tert-butyldimethylsilyl (TBDMS), and triisopropylsilyl (TIPS) groups).

[0120] As used herein, the term "nitrogen protecting group" refers to a group that can prevent reaction of the nitrogen moiety during further derivatization of the protected compound and that can be easily removed when desired. In one embodiment, the protecting group is removable by natural metabolic processes in physiological situations; essentially, the protected compound acts as a prodrug of the active, unprotected species.Examples of suitable nitrogen protecting groups that can be used include formyl, trityl, phthalimido, acetyl, trichloroacetyl, chloroacetyl, bromoacetyl, iodoacetyl; benzyloxycarbonyl (CBz), 4-phenylbenzyloxycarbonyl, 2-methylbenzyloxycarbonyl, 4-methoxybenzyloxycarbonyl, 4-fluorobenzyloxycarbonyl, 4-chlorobenzyloxycarbonyl, 3-chlorobenzyloxycarbonyl, 2-chlorobenzyloxycarbonyl, 2,4-dichlorobenzyloxycarbonyl, and the like. oxycarbonyl, 4-bromobenzyloxycarbonyl, 3-bromobenzyloxycarbonyl, 4-nitrobenzyloxycarbonyl, 4-cyanobenzyloxycarbonyl, t-butoxycarbonyl (tBoc), 2-(4-xenyl)-isopropoxycarbonyl, 1,1-diphenyleth-1-yloxycarbonyl, 1,1-diphenylprop-1-yloxycarbonyl, 2-phenylprop-2-yloxycarbonyl, 2-(p-toluyl)-prop-2-yloxycarbonyl, cyclopentanyloxycarbonyl oxycarbonyl, 1-methylcyclopentanyloxycarbonyl, cyclohexanyloxycarbonyl, 1-methylcyclohexanyloxycarbonyl, 2-methylcyclohexanyloxycarbonyl, 2-(4-tolylsulfono)-ethoxycarbonyl, 2-(methylsulfono)ethoxycarbonyl, 2-(triphenylphosphino)-ethoxycarbonyl, fluorenylmethoxycarbonyl (Fmoc), 2-(trimethylsilyl)ethoxycarbonyl, allyloxycarbonyl, 1-(trimethylsilylmethyl) Urethane-type blocking groups such as prop-1-enyloxycarbonyl, 5-benzisoxalylmethoxycarbonyl, 4-acetoxybenzyloxycarbonyl, 2,2,2-trichloroethoxycarbonyl, 2-ethynyl-2-propoxycarbonyl, cyclopropylmethoxycarbonyl, 4-(decyloxy)benzyloxycarbonyl, isobornyloxycarbonyl, and 1-piperidyloxycarbonyl; benzoylmethylsulfono group, 2-nitrophenylsulfenyl, and diphenylphosphine oxide are mentioned.The actual nitrogen protecting group employed is not critical, so long as the derivatized nitrogen group is stable to the conditions of the subsequent reaction and can be selectively removed as needed without substantially destroying the remainder of the molecule, including other nitrogen protecting groups. Further examples of these groups can be found in Greene, TW and Wuts, PGM, Protective Groups in Organic Synthesis, 2nd ed.; Wiley-Interscience: 1991; Chapter 7; McOmie, JFW (ed.), Protective Groups in Organic Chemistry, Plenum Press, 1973; and Kocienski, PJ, Protecting Groups, 2nd ed., Thieme Medical Pub., 2000.

[0121] Reference herein to any prior publication (or information derived therefrom) or public knowledge is not an acknowledgement or admission, or in any way intended to suggest, that the prior publication (or information derived therefrom) or public knowledge forms part of the common general knowledge in the field of endeavor to which this specification pertains.

[0122] Those skilled in the art will appreciate that the invention described herein is susceptible to variations and modifications other than those specifically described. It is to be understood that the invention includes all such variations and modifications within its spirit and scope. The invention also includes all steps, features, compositions, and compounds referred to or shown in this specification, individually or collectively, and any combination of any two or more of said steps or features. [Example]

[0123] The following examples are illustrative of the present disclosure and should not be construed as limiting in any way the general nature of the disclosure of the description throughout this specification.

[0124] Synthesis of Compounds of the Invention The agents of various embodiments can be prepared using the reaction pathways and synthetic schemes described herein, using readily available starting materials, and employing techniques available in the art. The preparation of certain compounds of these embodiments is described in detail in the Examples below, but those skilled in the art will recognize that the described chemical reactions can be easily adapted to prepare numerous other agents of various embodiments. For example, the synthesis of non-exemplified compounds can be successfully carried out by modifications obvious to those skilled in the art, such as by appropriately protecting interfering groups, by substituting other suitable reagents known in the art, or by routine modification of reaction conditions. A list of suitable protecting groups in organic synthesis can be found in T.W. Greene's Protective Groups in Organic Synthesis, 3rd Edition, John Wiley & Sons, 1991. Alternatively, it will be recognized that other reactions disclosed herein or known in the art have applicability for preparing other compounds of various embodiments. Reagents useful for synthesizing the compounds can be obtained or prepared according to techniques known in the art.

[0125] device Mass spectra were collected using a Thermo Scientific Exactive Plus OrbiTrap LC / MS (Thermo Fisher Scientific, MA, USA) and calibrated against an internal standard.

[0126] Copper-64G was supplied by the University of Queensland as a 0.1 M HCl solution. The copper was used approximately 48 h (4 half-lives) after preparation. An aliquot of copper-64 (74 μL, 121 MBq) was buffered with NaOH solution (0.1 M, 60 μL) and then with NHOAc buffer (1 M, pH 5.6, 54 μL) to a final pH of 5–6.

[0127] NMR spectra were obtained using an Agilent MR400 NMR (California, USA) (1 H, 400 MHz) at 297 K and referenced to solvent residues.

[0128] Analytical RP-HPLC traces were recorded using either: i) An Agilent 1200 HPLC system equipped with an Alltech Hypersil BDS C18 analytical HPLC column (4.6 × 150 mm, 5 μm) was used (flow rate 1 mL / min, UV absorbance was recorded at 214 and 254 nm). Retention time (R t / min) recorded using a gradient elution of 5 to 100% B in A in 30 min (A = 0.1% TFA, B = MeCN containing 0.1% TFA); or ii) Shimadzu LC-20AT system equipped with an SPD-20A UV detector and a Lab Logic Flow-RAM Radio HPLC detector. Samples were run on a Phenomenex Luna C18 5 μm 4.6 × 150 mm column at a flow rate of 1 mL / min using a solvent gradient of 5–100% B in 15 min (solvent A: 0.1% TFA in HO, solvent B: 0.1% TFA in acetonitrile).

[0129] Semi-preparative HPLC was performed on an Agilent 1200 HPLC system using buffers A=0.1% TFA and B=0.1% TFA in MeCN with UV detection at 214 nm.

[0130] Microwave synthesis was carried out using a Biotage (Uppsala, Sweden) Initator+ microwave system.

[0131] Radio-iTLC analysis was performed on a Lab Logic Scan-RAM PET / SPECT radio-TLC scanner using silica-injected glass microfiber iTLC plates developed with a mobile phase of 10 mM Na2EDTA in Dulbecco's phosphate-buffered saline, with an origin position of 10 mm and a solvent front of 110 mm (total measurement of 120 mm).

[0132] Mass spectrometry was performed using an Orbitrap Exactive Plus and a Dionex UltiMate® 3000 (ThermoFisher Scientific).

[0133] Example 1 - Mass spectrometry of Sar-FAPi Mass spectrometry was performed on Sar-FAPi. Calculated values: [M+H]+ m / z = 827.4850, [M+2H]2+ m / z = 414.2462, [M+3H]3+ m / z = 276.4999. Found values: 827.4856, 414.2464, 276.5004.

[0134] Example 2 - Radiolabeling An aliquot of copper-64 (90 μL, approximately 60 MBq) buffered with either PBS or ammonium acetate was added to an aliquot of either Sar-MonoFAPi or Sar-BisFAPi (300 ng, 3 μL from a 10 μg / mL stock solution in MilliQ HO prepared immediately before use). The mixture was allowed to stand at ambient temperature, and iTLC was performed on a small sample of the reaction mixture. An additional 6 μg (0.6 μL from a 0.1 mg / mL stock solution in MilliQ HO prepared immediately before use) was added, and quantitative labeling was confirmed by TLC after an additional 5 min. HPLC analysis was then performed immediately thereafter.

[0135] Example 3 - Stability Study Samples were analyzed by HPLC and iTLC as described in Example 2. The radiolabeled solution was stored at ambient temperature without further dilution and analyzed again by HPLC and iTLC after 24 hours.

[0136] Example 4 - Challenge Test To study the binding of the ligand to copper ions, the radiolabeled complexes were mixed with cysteine ​​or histidine, biologically relevant chelators of copper that compete with Sar-FAPi for copper ions. Stock solutions of cysteine ​​hydrochloride and histidine hydrochloride (50 mM in ammonium acetate buffer) were added to [ 64CuSarFAPi to a final concentration of 10 mM cysteine ​​or histidine. The reaction mixture was left at ambient temperature for 1 hour and then analyzed by HPLC.

[0137] When either cysteine ​​or histidine was added, [ 64 No change was observed in Cu]Cu-Sar-FAPi, indicating that it was not affected by the presence of a competitive chelator. 64 It is shown that the Cu radioisotope remains chelated to the Sar-FAPi ligand.

[0138] Example 5 - Cell Binding Studies SK-MEL-187 human melanoma cells were cultured at 30 × 10 cells / 2 mL screw-cap Eppendorf tube in medium (RPMI + 10% fetal bovine serum + 10% DMSO). 6 Cells were frozen in aliquots at the following cell density: 0.5 x 10 cells / 300 μl. Cells were thawed at 37°C, resuspended in PBS (2 mL), and transferred to 10 mL tubes. Cells were spun down (2000 rpm for 2 minutes) and resuspended at 20 million cells / 300 μl. Cells were diluted to the required concentration, and all samples were at the following cell density: 0.5 x 10 cells / 300 μl. 6 , 1×10 6 , 2 × 10 6 , 5×10 6 , 10×10 6 and 20×10 6 The medium was diluted to 300 μl with PBS. Approximately 5 kBq of radioactive ligand was added to this medium, with activity diluted so that 200 μl was added to the assay. Samples were incubated on a rotating wheel for 1 hour, then centrifuged (2000 rpm, 2 minutes) to pellet the cells and excess activity (supernatant) was removed. Samples were washed with 500 μl of PBS and centrifuged again (2000 rpm, 2 minutes) to pellet the cells. Samples were resuspended in 500 μl of PBS and the cell pellets were counted in a gamma counter.

[0139] LNCaP C42 human prostate cancer cells were used as a negative control following the same protocol. LNCaP C42 cells were frozen in medium (DMEM / F12 + 10% fetal bovine serum + 10% DMSO).

[0140] All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. Any examples provided herein, or the use of exemplary language (e.g., "such as," "for example"), are intended merely to better describe exemplary embodiments and do not pose a limitation on the scope of the claimed invention unless otherwise asserted. No language in the specification should be construed as indicating any non-claimed element as essential.

[0141] The description provided herein is of several embodiments that may share common properties and characteristics. It should be understood that one or more features of one embodiment may be combined with one or more features of other embodiments. Furthermore, a single feature or combination of features of an embodiment may constitute an additional embodiment.

[0142] Those skilled in the art will understand that the invention described herein is susceptible to variations and modifications other than those specifically described. It is to be understood that the invention includes all such variations and modifications. The invention also includes all steps, features, compositions, and compounds referred to or shown in this specification, individually or collectively, and any combination of any two or more of the steps or features.

[0143] Although the present invention has been described in detail herein for purposes of clarity and understanding, it will be apparent to those skilled in the art that various modifications and variations can be made to the embodiments and methods described herein without departing from the scope of the inventive concepts disclosed herein.

[0144] Throughout this specification and the claims that follow, unless the context requires otherwise, the word "comprise" and variations such as "comprises" or "comprising" will be understood to imply the inclusion of a stated integer or step or group of integers or steps, but not to the exclusion of other integers or steps or groups of integers or steps.

[0145] Reference herein to any prior publication (or information derived therefrom) or public knowledge is not an acknowledgement or admission, or in any way intended to suggest, that the prior publication (or information derived therefrom) or public knowledge forms part of the common general knowledge in the field of endeavor to which this specification pertains.

Claims

1. A compound of formula (I) or a salt, complex, isomer, solvate or prodrug thereof: 【Chemistry 1】 [In the formula, R is H, OH, halogen, cyano, NO ２ , N.H. ２ , optionally substituted C １ -C １２ Alkyl, optionally substituted amino, optionally substituted amido, optionally substituted aryl and groups having the structure: 【Chemistry 2】 is a group selected from the group consisting of: X and X １ may be the same or different and independently represent O, S, NH and N—(C １－１２ alkyl).

2. The linker is Optionally substituted C １ -C １２ Alkylene, where one or more alkylene groups are O, S, NH, or N—(C １－１２ alkyl); 【Transformation 3】 one or more amino acids; 【Chemistry 4】 wherein n is an integer from 1 to 10; and 【Transformation 5】 2. The compound of formula (I) of claim 1, comprising one or more moieties selected from the group consisting of:

3. The linker may be a group: 【Transformation 6】 3. A compound of formula (I) according to claim 1 or 2, comprising:

4. The linker is 【Transformation 7】 wherein a, b and c are integers independently selected from 1 to 10.

5. R has the following structure: 【Transformation 8】 and optionally substituted amide having the formula: In the structure, X １ is O, S, NH or N-(C １－１２ alkyl); The linker is 【Chemistry 9】 Optionally substituted C １ -C １２ Alkylene, where one or more alkylene groups are O, S, NH, or N—(C １－１２ alkyl); one or more amino acids; 【Chemistry 10】 wherein n is an integer from 1 to 10; and 【Chemistry 11】 A compound of formula (I) according to any one of claims 1 to 4, comprising one or more moieties selected from the group consisting of:

6. The compound of formula (I) has the structure of formula (Ia): 【Chemistry 12】 and In the structure, X and X １ may be the same or different and independently represent O, S, NH, and N—(C １－１２ alkyl); The linker is 【Chemistry 13】 Optionally substituted C １ -C １２ Alkylene, where one or more alkylene groups are O, S, NH, or N—(C １－１２ alkyl); one or more amino acids; 【Chemistry 14】 wherein n is an integer from 1 to 10; and 【Chemistry 15】 A compound of formula (I) according to any one of claims 1 to 5, comprising one or more moieties selected from the group consisting of:

7. Each linker in the compound is a group: 【Chemistry 16】 7. The compound of formula (Ia) according to claim 6, comprising:

8. Each linker may be the same or different and independently 【Chemistry 17】 and a, b and c are integers independently selected from 1 to 10.

9. The following structure: 【Chemistry 18-1】 【Chemistry 18-2】 【Chemistry 18-3】 The compound of formula (I) according to any one of claims 1 to 8, wherein

10. The compound of formula (I) is Sar-FAPi and has the following structure: 【Chemistry 19】 The compound according to any one of claims 1 to 8, having the formula:

11. The compound of formula (Ia) is Sar-bisFAPi and has the following structure: 【Chemistry 20】 The compound according to any one of claims 1 to 8, having the formula:

12. A compound of formula (I) according to any one of claims 1 to 11, wherein the compound is complexed with a metal ion.

13. 13. The compound of formula (I) according to claim 12, wherein the metal ion is a Cu radioisotope.

14. The radioisotope ６０ Cu, ６１ Cu, ６２ Cu, ６４ Cu and ６７ 14. The compound of formula (I) according to claim 13, wherein the compound is selected from the group consisting of Cu.

15. A composition comprising a compound according to any one of claims 1 to 14 and a pharmaceutically acceptable excipient.

16. A method for the treatment of cancer, comprising administering to a subject in need thereof a compound of formula (I) according to any one of claims 1 to 14, or a salt, complex, isomer, solvate or prodrug thereof, wherein the compound of formula (I) contains a suitable radioisotope.

17. 19. A method for radioimaging of cancer, comprising administering to a subject in need thereof a compound of formula (I) according to any one of claims 1 to 14, or a salt, complex, isomer, solvate or prodrug thereof, wherein the compound of formula (I) contains a suitable radioisotope.

18. The radioisotope ６０ Cu, ６１ Cu, ６２ Cu, ６４ Cu and ６７ 16. The method of claim 14 or 15, wherein the metal is selected from the group consisting of Cu.

19. The method of any one of claims 16 to 18, wherein the cancer is selected from the group consisting of epithelial ovarian cancer, ovarian cancer, osteosarcoma, pancreatic adenocarcinoma, colorectal cancer, lung cancer, non-small cell lung cancer, gastric cancer, endometrial cancer, pancreatic adenocarcinoma, medullary thyroid carcinoma, differentiated thyroid carcinoma, breast cancer, invasive ductal carcinoma, oral squamous cell carcinoma, esophageal cancer, renal cell carcinoma, insulinoma, prostate cancer, neuroendocrine differentiated prostate cancer, pheochromocytoma, adenoid cystic carcinoma, hepatocellular carcinoma, cervical cancer, small intestine cancer, neuroendocrine tumor, anal cancer, chordoma, desmoid tumor, head and neck cancer, thymic carcinoma, pancreatic cancer, cholangiocarcinoma, esophageal cancer, salivary gland cancer, sarcoma, and cancer of unknown primary origin.

20. 15. Use of a compound of formula (I) according to any one of claims 1 to 14, or a salt, complex, isomer, solvate or prodrug thereof, in the manufacture of a medicament for the treatment of cancer.

21. i) a container comprising a compound of formula (I) according to any one of claims 1 to 14, or a pharmaceutically acceptable salt thereof; ii) a vessel comprising a solution of Cu ions; and iii) Instructions for preparing an aqueous formulation of a compound of formula (I) or a pharmaceutically acceptable salt thereof complexed with Cu ions. A kit comprising:

22. 22. The kit of claim 21, wherein the Cu ions are Cu radioisotopes.

23. The radioisotope ６０ Cu, ６１ Cu, ６２ Cu, ６４ Cu and ６７ 23. The kit of claim 22, wherein the metal is selected from the group consisting of Cu.

24. A process for producing a compound of formula (I) or a pharmaceutically acceptable salt thereof, comprising: i) a compound of formula A or a protected form thereof: 【Chemistry 21】 preparing ii) A compound of formula B or a protected form thereof: 【Chemistry 22】 preparing iii) coupling a compound of formula A, or a protected form thereof, with a compound of formula B, or a protected form thereof, to form a compound of formula (I), or a protected form thereof. wherein R is H, OH, halogen, cyano, NO ２ , N.H. ２ , optionally substituted C １ -C １２ Alkyl, optionally substituted amino, optionally substituted amido, optionally substituted aryl and the following structure: 【Chemistry 23】 wherein: X and X １ may be the same or different and independently represent O, S, NH, and N—(C １－１２ alkyl) ,method.

Images