Cyclic peptides as PET imaging agents for granzyme B

Abstract

Novel cyclic peptides that bind to granzyme B and may be suitable for imaging granzyme B, their salts, pharmaceutical compositions containing them, diagnostic and therapeutic uses, and methods for producing such compounds are disclosed. Additionally, compounds useful as radiotracers for positron emission tomography (PET) and / or single photon emission computed tomography (SPECT) imaging are provided. The use of the compounds as imaging agents for granzyme B is further disclosed.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to U.S. Provisional Application No. 63 / 432,012, filed December 12, 2022, the entire contents of which are incorporated herein by reference.

[0002] Reference to an electronically submitted sequence listing This application contains a Sequence Listing that has been submitted electronically in XML format, and is incorporated herein by reference in its entirety. The XML file, created on November 7, 2023, has the file name 25623_WO_PCT_SL.XML and is 361,585 bytes in size. [Background technology]

[0003] Granzyme B (GzmB) is a serine protease secreted by cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells. GzmB-induced cell death has traditionally been considered the primary mechanism used by CTLs and NK cells to eliminate harmful target cells, including allogeneic cells, virus-infected cells, and tumor cells. See U.S. Patent Application Publication No. 2019 / 0224348, Larimer, B.; et al. Cancer Res 77, 2017, 2318-2327, and Larimer, B.; et al. U. Clin Cancer Res 25, 2019, 1196-1205. Measurement of GzmB can serve as a readout of the functional status of CTLs and NKs and is of interest in cancer immunotherapy and autoimmunity. A cyclic peptide that binds to human granzyme B with high affinity has now been disclosed. These peptides can be conjugated to chelators and radiometals for use in PET imaging of granzyme B. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] US Patent Application Publication No. 2019 / 0224348 [Non-patent literature]

[0005] [Non-Patent Document 1] Larimer,B.;et al.Cancer Res 77,2017,2318-2327 [Non-patent document 2] Larimer,B.;et al.U.Clin Cancer Res 25,2019,1196-1205 Summary of the Invention [Means for solving the problem]

[0006] Provided are novel cyclic peptides, their salts, pharmaceutical compositions containing them, diagnostic and therapeutic uses, and methods for producing such compounds, which may be suitable for imaging granzyme B. Further provided are compounds useful as radiotracers for positron emission tomography (PET) and / or single photon emission computed tomography (SPECT) imaging. The present disclosure further relates to the use of the compounds as imaging agents for granzyme B. [Brief explanation of the drawings]

[0007] [Figure 1A] FIG. 1 shows representative 3D PET / CT images of a mouse injected with 18F-labeled anti-granzyme-B peptide 11a. [Figure 1B] FIG. 10 shows the corresponding SUV averages in tissues of interest induced after injection of 18F-labeled anti-granzyme-B peptide 11a. [Figure 2A] FIG. 18F-labeled scrambled peptide (negative control). [Figure 2B] FIG. 10 shows the corresponding SUV averages in tissues of interest induced after injection of 18F-labeled scrambled peptide in GvHD model mice and NOG control mice. [Figure 3]Representative human granzyme-B expression IHC images at 100 μm and 50 μm resolution from lung tissues collected from GvHD mice (top panel) and NOG control mice (bottom panel). [Figure 4A] Representative PET / CT images at different time points from a male rhesus monkey injected with 18F-labeled anti-granzyme-B peptide 11a (left). [Figure 4B] Corresponding quantitative data shown as SUV mean (right). [Figure 5A] 68Ga-labeled anti-granzyme-B peptide 19a from hPBMC donor (left). [Figure 5B] Corresponding quantitative data presented as SUV mean (right). [Figure 6A] Representative PET / CT images at different time points from a male rhesus monkey injected with 68Ga-labeled anti-granzyme-B peptide 19a (left). [Figure 6B] Corresponding quantitative data shown as SUV mean (right). DETAILED DESCRIPTION OF THE INVENTION

[0008] Compounds of Formula I, or pharmaceutically acceptable salts thereof, are described that are capable of binding to Granzyme B. In some embodiments, the compounds of Formula I, or pharmaceutically acceptable salts thereof, are irreversible binders of Granzyme B. In some embodiments, the compounds of Formula I, or pharmaceutically acceptable salts thereof, are inhibitors of Granzyme B. Additionally, in some embodiments, the compounds of Formula I, or pharmaceutically acceptable salts thereof, contain one or more imaging agents. In some embodiments, the compounds of Formula I, or pharmaceutically acceptable salts thereof, are polypeptides that bind to Granzyme B.

[0009] Formula I: [ka]

[0010] and pharmaceutically acceptable salts thereof, During the ceremony, R 1 is C 1-6 alkyl and aryl, wherein the aryl is selected from C 1-6 optionally substituted with 1 to 3 groups selected from alkyl, halogen, and hydroxyl; R 2 is hydrogen, halogen, and C 1-6 alkyl, R 3 is hydrogen, -CH2OH, C 1-7 alkyl, -CHCONH, and -CHNH; R 4 , R 5 , and R 6 are independently hydrogen, C 1-6 Alkyl, (CH2)3NHC(=NH2)NH2, (CH2)4NH2, and C 1-6 -CH2-heteroaryl, optionally substituted with 1 to 3 substituents selected from alkyl and halogen; Or, R 4 and R 6 together with the atoms to which they are attached, C 1-6 forming a nitrogen-containing 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 groups selected from alkyl and halogen; R 7 , R 8 , and R 9 are independently hydrogen, C 1-6 alkyl, CH heteroaryl, and CH aryl, wherein said alkyl is selected from R a and the heteroaryl and aryl are optionally substituted with 1 to 3 OH groups; Or, R 8 and R 9 together with the atoms to which they are attached, C 1-6forming a nitrogen-containing 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 groups selected from alkyl and halogen; R a is selected from hydroxyl, -COOH, -NH, -NHC(=NH)NH and C(O)NH; R 10 is CH2 heteroaryl and C 1-6 alkyl, wherein said heteroaryl and alkyl are selected from OH, C 1-6 optionally substituted with 1 to 3 groups selected from alkyl and halogen; R 11 and R 12 are C 1-6 is alkyl, Or, R 11 and R 12 together with the atoms to which they are attached, C 1-6 forming a 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 groups selected from alkyl, phenyl, and halogen; R 13 is -NH2, [ka]

[0011] is selected from R 14 teeth, (i) H, (ii) [ka]

[0012] NOTA-1-, represented by (iii) R0aC(O)- (wherein R0a is C 1-3 alkyl), or polyethylene glycol polymers, (iv) [ka]

[0013] RESCA-1, expressed by (v) a chelating moiety, optionally having a positron-emitting isotope as an imaging agent; (vi) [ka]

[0014] AF647, represented by (vii) Formula II: [ka]

[0015] Polyethylene glycol polymer (wherein n is an integer selected from 1 to 24), (viii) Formula III: [ka]

[0016] The polyethylene glycol polymers are selected from the group consisting of:

[0017] wherein each m is an integer independently selected from 1 to 12; R 15 is H, -C(O)(CH2)2NHC(O)OC(CH3)3, -C(O)CH2NH(NOTA-1), R 14 (ii) R 14 (iii) R 14 (iv) and R 14 (v) X is [ka]

[0018] is.

[0019] (In the formula, R 1’ is C 1-6alkyl and aryl, wherein the aryl is selected from C 1-6 optionally substituted with 1 to 3 substituents selected from alkyl, halogen, and hydroxyl; R 2’ is hydrogen, halogen, and C 1-6 alkyl, R 3’ -H, -CH2OH, C 1-7 alkyl, -CHCONH, and -CHNH; R 4’ , R 5’ , and R 6’ are independently hydrogen, C 1-6 alkyl, —(CH2)3NHC(═NH)NH2, and —(CH2)4NH2, and —CH2-heteroaryl, wherein heteroaryl is selected from C 1-6 optionally substituted with 1 to 3 substituents selected from alkyl and halogen; Or, R 4’ and R 6’ together with the atoms to which they are attached, C 1-6 forming a nitrogen-containing 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 substituents independently selected from alkyl and halogen; R 7’ , R 8’ , and R 9’ are independently hydrogen, C 1-6 alkyl, CH heteroaryl, and CH aryl, wherein the alkyl is selected from 1 to 3 R a and the heteroaryl and aryl are optionally substituted with 1 to 3 OH groups; Or, R 8’ and R 9’ together with the atoms to which they are attached, C 1-6 forming a nitrogen-containing 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 substituents independently selected from alkyl and halogen; R 10’ is CH2 heteroaryl and C 1-6alkyl, wherein said heteroaryl and alkyl are selected from OH, C 1-6 may be substituted with 1 to 3 groups selected from alkyl and halogen, and R 11’ and R 12’ are C 1-6 is alkyl, Or, R 11’ and R 12’ together with the atoms to which they are attached, C 1-6 Forms a 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 substituents selected from alkyl, phenyl, and halogen. One embodiment of Formula I of the present disclosure is realized by a compound of structural Formula I' or a pharmaceutically acceptable salt thereof: [ka]

[0020] .

[0021] One embodiment of the present disclosure is R 1 C 1-6 A subembodiment of this aspect of the disclosure is realized when R 1 is realized when is -CH(CH3)2.

[0022] Another embodiment of the present disclosure is R 1 or R 1’ But C 1-6 A subembodiment of this aspect of the disclosure is realized when R is aryl optionally substituted with 1 to 3 groups selected from alkyl, halogen, and hydroxyl. 1 or R 1’ But C 1-6 A further subembodiment is realized when R is phenyl optionally substituted with 1 to 3 groups selected from alkyl, fluorine, and hydroxyl. 1 or R 1’ is phenyl optionally para-substituted with hydroxyl, methyl, or fluoro.

[0023] Another embodiment of the present disclosure is R 2 or R 2’ is hydrogen. Another embodiment of the present disclosure is 2 or R 2’ A subembodiment of this aspect of the disclosure is realized when R 2 or R 2’ is fluorine. Another embodiment of the present disclosure is 2 or R 2’ C 1-6 A subembodiment of this aspect of the disclosure is realized when R 2 or R 2’ is methyl.

[0024] Another embodiment of the present disclosure is R 3 or R 3’ is hydrogen. Another embodiment of the present disclosure is 3 or R 3’ C 1-7 Another embodiment of the present disclosure is realized when R 3 or R 3’ Another embodiment of the present disclosure is realized when R 3 or R 3’ Another embodiment of the present disclosure is realized when R 3 or R 3’ is -CH2CONH2. Another embodiment of the present disclosure is 3 or R 3’ Another embodiment of the present disclosure is realized when R 3 or R 3’ is selected from hydrogen, —CH 2 OH, —CH 2 CONH 2 and —CH 2 NH 2 .

[0025] Another embodiment of the present disclosure is R 4 , R 5 , and R 6 and / or R 4’ , R 5’, and R 6’ Another embodiment of the present disclosure is realized when at least one of R 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ Another embodiment of the present disclosure is realized when at least two of R 5 and R 6 and / or R 5’ and R 6’ and R are hydrogen. Another embodiment of the present disclosure is 5 and / or R 5’ is hydrogen and R 4 and R 6 and / or R 4’ and R 6’ is realized when is not hydrogen.

[0026] Another embodiment of the present disclosure is R 4 , R 5 , and R 6 and / or R 4’ , R 5’ and R 6’ At least one of the 1-6 Another embodiment of the present disclosure is realized when R 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ Two of them are C 1-6 Another embodiment of the present disclosure is realized when one of R is alkyl and the other is hydrogen. 4 and R 6 and / or R 4’ and R 6’ Both are C 1-6 Another embodiment of the present disclosure is realized when R 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R6’ C 1-6 alkyl is selected from CH3, CH(CH3)2, and CH2CH3 1-6 This is realized when the group is alkyl.

[0027] Another embodiment of the present disclosure is R 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ is (CH2)3NHC(=NH2)-NH2 and the other is not. Another embodiment of the present disclosure is 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ is realized when one of R is (CH)NHC(=NH)-NH and the other is hydrogen. 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ is (CH)NH and the other is not. Another embodiment of the present disclosure is 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ is realized when one of R is (CH)NH and the other is hydrogen. 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ One of them is C 1-6 and -CH-heteroaryl, optionally substituted with 1 to 3 substituents selected from alkyl and halogen, and the others are otherwise realized. Another embodiment is -CH-heteroaryl, optionally substituted with 1 to 3 substituents selected from alkyl and halogen, and the others are otherwise realized. 4 , R 5 , and R6 and / or R 4’ , R 5’ , and R 6’ One of them is C 1-6 Another embodiment is realized when R is -CH heteroaryl, optionally substituted with 1 to 3 substituents selected from alkyl and halogen, and the others are hydrogen. 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ is -CH-indolyl, optionally substituted with methyl or fluoro, and the other is not. Another embodiment is realized when R 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ Another embodiment is realized when one of R is -CH-indolyl, optionally substituted with methyl or fluoro, and the other is hydrogen. 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ Another embodiment is realized when one of R is -CH-heteroaryl and the other is not. 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ Another embodiment is realized when one of R is -CH-heteroaryl and the other is hydrogen. 4 , R 5 , and R 6 and / or R 4’ , R 5’ , and R 6’ is -CH-indolyl and the other is not. Another embodiment is realized when one of R 4 , R 5 , and R 6and / or R 4’ , R 5’ , and R 6’ is realized when one of the groups is -CH2-indolyl and the other is hydrogen.

[0028] Another embodiment of the present disclosure is R 4 and R 6 and / or R 4’ and R 6’ together with the atoms to which they are bonded, C 1-6 Another embodiment of the present disclosure is realized when R forms a nitrogen-containing 3- to 10-membered heterocyclyl, optionally substituted with 1 to 3 groups selected from alkyl and halogen. 4 and R 6 and / or R4' and R6' together with the atoms to which they are attached are C 1-6 form a nitrogen-containing 3- to 10-membered heterocyclyl optionally substituted with 1 to 3 groups selected from alkyl and halogen, and R 5 or R 5’ is hydrogen. Another embodiment of the present disclosure is 4 and R 6 and / or R 4’ and R 6’ together with the atom to which they are attached form a nitrogen-containing 3- to 10-membered heterocyclyl selected from isoquinolinyl, pyrrolidinyl, indolyl, piperidinyl, and azetidinyl, wherein said isoquinolinyl, pyrrolidinyl, indolyl, piperidinyl, and azetidinyl are C 1-6 Another embodiment of the present disclosure is realized when R 4 and R 6 and / or R 4’ and R 6’ are taken together with the atom to which they are attached to form an optionally substituted isoquinolinyl. Another embodiment of the present disclosure is 4 and R 6 and / or R 4’ and R 6’are taken together with the atom to which they are attached to form an optionally substituted pyrrolidinyl. Another embodiment of the present disclosure is 4 and R 6 and / or R 4’ and R 6’ are taken together with the atom to which they are attached to form an optionally substituted piperidinyl. Another embodiment of the present disclosure is 4 and R 6 and / or R 4’ and R 6’ are taken together with the atom to which they are attached to form an optionally substituted azetidinyl. A subembodiment of this aspect of the disclosure is when the heterosilyl substituent is C 1-6

[0023] Another subembodiment of this disclosure is realized when the one to three heterosilyl substituents are selected from methyl and fluorine.

[0029] Another embodiment of the present disclosure is R 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ Another embodiment is realized when at least one of R 7 , R 8 , and R 9 Another embodiment is realized when two of R 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ Two of the C groups are hydrogen and the other is optionally substituted. 1-6 Another embodiment is realized when R is selected from alkyl, (CH)indolyl, CHphenyl, CHphenylOH, (CH)NH, (CH)NHC(=NH)NH, and CHimidazolyl.7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ Two of the C groups are hydrogen and the other is optionally substituted. 1-6 One aspect of this embodiment is when R 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ two of which are hydrogen and the others are selected from CH3, CH2CH(CH3)2, CH(CH3)CH2CH3, CH(CH3)CH2CH3, and CH(CH3)2; 1-6 Another embodiment is realized when R 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ is realized when two of R are hydrogen and the other is a substituted alkyl selected from CHOH, CHCOOH, CH(OH)CH, (CH)C(O)NH, —(CH)—NHC(═NH)NH, —(CH)—NH, and —(CH)C(O)OH. 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ Another embodiment is realized when two of R are hydrogen and the other is -CHOH. 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ Another embodiment of the present disclosure is realized when two of R 7 , R 8 , and R 9 and / or R 7’ , R 8’, and R 9’ is realized when two of R are hydrogen and the other is -CH(OH)CH. Another embodiment of the present disclosure is 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ is realized when two of R are hydrogen and the other is (CH)C(O)NH. 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ is realized when two of R are hydrogen and the other is -(CH)-NHC(=NH)NH. Another embodiment is 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ is realized when two of R are hydrogen and the other is —(CH)C(O)OH. Another embodiment of the present disclosure is 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ Another embodiment of the present disclosure is realized when two of R 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ is realized when two of R are hydrogen and the other is CHphenyl. Another embodiment of the present disclosure is 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ Another embodiment is realized when two of R are hydrogen and the other is -CHphenylOH. 7 , R8 , and R 9 and / or R 7’ , R 8’ , and R 9’ is realized when two of R are hydrogen and the other is -(CH)NH. 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ is realized when two of R are hydrogen and the other is -(CH)NHC(=NH)NH. Another embodiment is 7 , R 8 , and R 9 and / or R 7’ , R 8’ , and R 9’ This is achieved when two of the groups are hydrogen and the other is -CH2 imidazolyl.

[0030] Yet another embodiment of the present disclosure is R 8 and R 9 and / or R 8’ and R 9’ together with the atoms to which they are bonded, C 1-6 A subembodiment is realized when R forms a nitrogen-containing 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 groups selected from alkyl and halogen. 8 and R 9 and / or R 8’ and R 9’ A subembodiment is realized when R 8 and R 9 and / or R 8’ and R 9’ are taken together to form an optionally substituted pyrrolidinyl.

[0031] Another embodiment of the present disclosure is R 10 or R 10’is an optionally substituted CH2 indolyl. One aspect of this embodiment is 10 or R 10’ is unsubstituted CH2 indolyl. Another subembodiment is realized when R 10 or R 10’ is CH2 indolyl, and said indolyl is C 1-6 This is realized when the alkyl group is substituted with 1 to 3 groups selected from alkyl and halogen.

[0032] Another embodiment of the present disclosure is R 10 or R 10’ C optionally substituted with 1 to 3 hydroxyl groups 1-6 A subembodiment of this aspect of the disclosure is realized when R 10 or R 10’ is CH(OH)CH3.

[0033] Another embodiment of the present disclosure is R 10 or R 10’ But, OH, C 1-6 A subembodiment of this aspect is realized when R is -CH-heteroaryl, optionally substituted with 1 to 3 groups selected from alkyl and halogen. 10 or R 10’ is -CH2-indolyl, which is unsubstituted or substituted with fluoro or methyl.

[0034] Another embodiment of the present disclosure is R 11 and R 12 and / or R 11’ and R 12’ Both are C 1-6 A subembodiment is realized when R 11 and R 12 and / or R 11’ and R 12’ is independently selected from -CH3 and -CH2CH3.

[0035] Another embodiment of the present disclosure is R 11 and R 12 and / or R 11’ and R 12’ together with the atoms to which they are bonded, C 1-6 A subembodiment is realized when R forms a 3- to 10-membered heterocyclyl group that is optionally substituted with 1-3 groups of alkyl, phenyl, and halogen. 11 and R 12 and / or R 11’ and R 12’ together form a group selected from pyrrolidinyl, piperidinyl, and tetrahydroisoquinolinyl, wherein said pyrrolidinyl, piperidinyl, and tetrahydroisoquinolinyl are selected from C 1-6 A subembodiment is realized when R is optionally substituted with 1 to 3 groups of alkyl, phenyl, and halogen. 11 and R 12 and / or R 11’ and R 12’ together form a pyrrolidinyl, and said pyrrolidinyl is 1-6 A subembodiment is realized when R is optionally substituted with 1 to 3 groups of alkyl, phenyl, and halogen. 11 and R 12 and / or R 11’ and R 12’ together form piperidinyl, and the piperidinyl is C 1-6 A subembodiment is realized when R is optionally substituted with 1 to 3 groups of alkyl, phenyl, and halogen. 11 and R 12 and / or R 11’ and R 12’ together form a tetrahydroisoquinolinyl, and said tetrahydroisoquinolinyl is C 1-6 A subembodiment is realized when the substituents are selected from methyl, phenyl, and fluorine.

[0036] Another embodiment of the present disclosure is R 13 is realized when is NH2.

[0037] Another embodiment of the present disclosure is R 13 but [ka]

[0038] This is realized when

[0039] Another embodiment of the present disclosure is R 13 but [ka]

[0040] This is realized when

[0041] Another embodiment of the present disclosure is R 14 is hydrogen. Another embodiment of the present disclosure is 14 R0aC(O)- (wherein R0a is C 1-3 This is realized when the alkyl group is a polyethylene glycol polymer selected from PEG1-24.

[0042] Another embodiment of the present disclosure is R 14 but Formula II: [ka]

[0043] Polyethylene glycol polymer wherein n is an integer selected from 1 to 24, and X is as defined herein. This is realized when

[0044] Another embodiment of the present disclosure is R 14 but Formula III: [ka]

[0045] Polyethylene glycol polymer (wherein m is an integer selected from 1 to 12, and X and R 15 are as described herein) A subembodiment of Formula III is realized when R 15 is H. A subembodiment of Formula III is realized when R 15 is C(O)(CH2)2NHC(O)OC(CH3)3. Another subembodiment of Formula III is realized when R 15 is C(O)CHNH(NOTA-1). Another subembodiment of Formula III is realized when R 15 R 14 Another subembodiment of Formula III is realized when R 15 R 14 Another subembodiment of formula III is realized when R 15 R 14 Another subembodiment of formula III is realized when R 15 R 14 Another subembodiment of formula III is realized when R 15 R 14 This is realized when (v) of the above is true.

[0046] Another embodiment of the present disclosure is R 14However, deferoxamine (DFO), 1,4,7,10-tetraacetic acid (DOTA), diethylenetriaminepentaacetic acid (DTPA), ethylenediaminetetraacetic acid (EDTA), (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetra(methylenephosphonic) acid (DOTP), (1R,4R,7R,10R)-α'α"α"'-tetramethyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTMA), 1,4,8,11-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTMA), Isotetradecane-1,4,8,11-tetraacetic acid (TETA), H4 octapa, H6 phospa, H2 dedopa, H5 decapa, H2 azapa, HOPO, DO2A, 1,4,7,10-tetrakis(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane (DOTAM), 1,4,7-triazacyclononane-N,N',N"-triacetic acid (NOTA), NOTA-1,1,4,8,11-tetraazabicyclo[6.6.2]hexadecane-4,11-dicetic acid (CB TE2A), 1,4,7,10-tetraazacyclododecane (cyclen), 1,4,8,11-tetraazacyclotetradecane (cyclam), octadentate chelators, hexadentate chelators, phosphonate chelators, macrocyclic chelators, chelators containing macrocyclic terephthalamide ligands, bifunctional chelators, fusarinine C and fusarinine C derivative chelators, triacetylfusarinine C (TAFC), ferrioxamine E (FOXE), ferrioxamine B (FOXB), and ferrichrome A (FCHA), wherein the chelator is used as a positron-irradiating agent. A subembodiment is realized when the chelating agent is deferoxamine (DFO), optionally with a positron-emitting isotope. A subembodiment is realized when the chelating agent is 1,4,7,10-tetraacetic acid (DOTA), optionally with a positron-emitting isotope. A subembodiment is realized when the chelating agent is diethylenetriaminepentaacetic acid (DTPA), optionally with a positron-emitting isotope. A subembodiment is realized when the chelating agent is ethylenediaminetetraacetic acid (EDTA), optionally with a positron-emitting isotope.A subembodiment is realized when the chelating agent is (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetra(methylenephosphonic) acid (DOTP), optionally bearing a positron-emitting isotope. A subembodiment is realized when the chelating agent is (1R,4R,7R,10R)-α'α"α"'-tetramethyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTMA ... A subembodiment is realized when the chelating agent is 1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid (TETA), optionally having an emitting isotope. A subembodiment is realized when the chelating agent is 6,6'-((ethane-1,2-diylbis((carboxymethyl)azanediyl))bis(methylene))dipicolinic acid (H4 octapa), 6,6'-(2,3-bis((phosphonomethyl)amino)butane-1,4-diyl)dipicolinic acid (H6 phospa), 6,6'-((ethane-1,2-diylbis(azanediyl))bis (methylene))dipicolinic acid (H2dedopa), 6,6'-(((((carboxymethyl)azanediyl)bis(ethane-2,1-diyl))bis((carboxymethyl)azanediyl))bis(methylene))dipicolinic acid (H5decapa), and 6,6'-(2,3-bis(((1-benzyl-1H-1,2,3-triazol-4-yl)methyl)amino)butane-1,4-diyl)dipicolinic acid (H2azapa), wherein said group optionally includes a positron-emitting isotope. is realized when the chelating agent is N,N'-(butane-1,4-diyl)bis(1-hydroxy-N-(3-(1-hydroxy-6-oxo-1,6-dihydropyridine-2-carboxamido)propyl)-6-oxo-1,6-dihydropyridine-2-carboxamide) (HOPO), optionally having a positron-emitting isotope. A subembodiment is realized when the chelating agent is 1,4,7,10-tetraazacyclododecane-1,7-diacetic acid (DO2A), optionally having a positron-emitting isotope.A subembodiment is realized when the chelating agent is 1,4,7,10-tetrakis(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane (DOTAM), optionally with a positron-emitting isotope. A subembodiment is realized when the chelating agent is 1,4,7-triazacyclononane-N,N',N"-triacetic acid (NOTA), NOTA-1, optionally with a positron-emitting isotope. A subembodiment is realized when the chelating agent is 1,4,8,11-tetraazabicyclo[6.6.2]hexadecane-4,11-dicetic acid (CB TE2A). A subembodiment is realized when the chelating agent is 1,4,7,10-tetraazacyclododecane (cyclen), optionally with a positron-emitting isotope. A subembodiment is realized when the chelating agent is 1,4,8,11-tetraazacyclotetradecane (cyclam), optionally with a positron-emitting isotope. A subembodiment is realized when the chelating agent is selected from octadentate chelating agents, hexadentate chelating agents, phosphonate chelating agents, macrocyclic chelating agents, chelating agents comprising macrocyclic terephthalamide ligands, bifunctional chelating agents, Fusarinin C and Fusarinin C derivative chelating agents. , wherein the group optionally has a positron-emitting isotope. A subembodiment is realized when the chelating agent is triacetylfusarinine C (TAFC), optionally with a positron-emitting isotope. A subembodiment is realized when the chelating agent is ferrioxamine E (FOXE), optionally with a positron-emitting isotope. A subembodiment is realized when the chelating agent is ferrioxamine B (FOXB), optionally with a positron-emitting isotope. A subembodiment is realized when the chelating agent is ferrichrome A (FCHA), optionally with a positron-emitting isotope. Another embodiment is R. 14 However, DOTA, NOTA-1, and RESCA-1 68 Ga, 64 Cu and Al 18 Another subembodiment is realized when the chelating agent is a chelating agent optionally having a positron emitting isotope selected from F. 177 Lu,90 Y, and 111 This is realized when the signal is selected from In.

[0047] Another embodiment of the present disclosure is R 14 but, (ii) [ka]

[0048] This is realized when the NOTA-1 is represented by

[0049] A subembodiment of this aspect of the present disclosure is realized when the structure of (ii) optionally comprises a positron-emitting isotope. Another subembodiment is when the positron-emitting isotope is: 68 Ga and 18 F(Al 18 F), and 64 This is realized when the metal is selected from Cu.

[0050] Another embodiment of the present disclosure is R 14 (iv) [ka]

[0051] This is realized when

[0052] A subembodiment of this aspect of the present disclosure is realized when (iv) can include a positron-emitting isotope. Another subembodiment is when the positron-emitting isotope is: 68 Ga and 18 This is realized when F is selected.

[0053] Another embodiment of the present disclosure is R 14 (vi) [ka]

[0054] This is realized when

[0055] A subembodiment is wherein (vi) is 111 In and 67 This is realized when the gamma-emitting isotope is selected from Ga.

[0056] One embodiment of the present disclosure is realized when the compounds of Formula I, Formula I', and Formula Ia comprise one or more imaging agents. In another embodiment, the imaging agent is selected from the group consisting of a paramagnetic ion, an X-ray contrast agent, a fluorophore, and a radioisotope. Many suitable imaging agents are known in the art, as are methods for their attachment to antibodies (see, e.g., U.S. Pat. Nos. 5,021,236, 4,938,948, and 4,472,509, the disclosures of each of which are incorporated herein by reference in their entirety). The radiolabeled compounds of Formula I, Formula I', and Formula Ia provided herein, or pharmaceutically acceptable salts thereof, may be prepared according to methods well known in the art. For example, monoclonal antibodies may be iodinated by contact with sodium iodide and / or potassium iodide and a chemical oxidizing agent, such as sodium hypochlorite, or an enzymatic oxidizing agent, such as lactoperoxidase. In a further example, the compounds of Formula I, Formula I', and Formula Ia provided herein, or pharmaceutically acceptable salts thereof, can be chelating compounds of Formula I, Formula I', and Formula Ia to a bifunctional chelator provided herein (e.g., NOTA-1, DOTA, or NODAGA), or similar derivatives thereof, by radiometallation. 68 They may be labeled with Ga. Synthetic methods for incorporating radioisotopes into organic compounds are well known in the art, and those of skill in the art will readily recognize other methods applicable to the compounds provided herein.

[0057] In another embodiment, the imaging agent comprises one, two, or three imaging agents selected from the group consisting of a paramagnetic ion, an X-ray contrast agent, a fluorophore, and a radioisotope. In another embodiment, Formula I, Formula I', and Formula Ia comprise one imaging agent. In another embodiment, Formula I, Formula I', and Formula Ia comprise two imaging agents. In another embodiment, Formula I, Formula I', and Formula Ia comprise three imaging agents. In another embodiment, the compounds of Formula I, Formula I', and Formula Ia comprise one or more imaging agents, which may include one or more independently selected paramagnetic ions.

[0058] A subembodiment of this aspect of the disclosure is realized when each of the paramagnetic ions is independently selected from the group consisting of fluoride, chromium(III), manganese(II), iron(III), iron(II), cobalt(II), nickel(II), copper(II), neodymium(III), samarium(III), ytterbium(III), gadolinium(III), vanadium(II), terbium(III), dysprosium(III), holmium(III), and erbium(III). Another subembodiment of this aspect of the disclosure is realized when the compounds of Formula I, Formula I', and Formula Ia comprise one, two, or three independently selected paramagnetic ions. Another subembodiment of this aspect of the disclosure is realized when one or more independently selected paramagnetic ions are independently bound directly or indirectly (e.g., via a chelator) to the compounds provided herein.

[0059] Another embodiment of this aspect of the disclosure is realized when the compounds of Formula I, Formula I', and Formula Ia include one or more contrast agents that are independently selected x-ray contrast agents. A subembodiment of this aspect of the disclosure is realized when each of the x-ray contrast agents is independently selected from the group consisting of lanthanum(III), gold(III), lead(II), bismuth(III), and iodinated x-ray contrast agents (e.g., diatrizoate, ioxaglate, metrizoate, iopamidol, iohexol, ioxilan, iopromide, iodixanol, and ioversol).

[0060] Another embodiment of the present disclosure is realized when the imaging agent is a fluorophore. A subembodiment of this aspect of the present disclosure is when A is Alexa 350, Alexa 430, AMCA, BODIPY 630 / 650, BODIPY 650 / 665, BODIPY-FL, BODPY-R6G, 13BODLPY-TMR, BODLPY-TRX, Cascade Blue, Cy3, Cy5, 6-FAM, fluorescein isothiocyanate, HEX, 6-JOE, Oregon Green 488, Oregon Green 500, Oregon Green 514, quantum dot, Pacific Blue, REG, rhodamine green, rhodamine red, Renografin, ROX, TAMRA, TET, tetramethyl-rhodamine, Texas Red, AF This is achieved when the fluorophore is selected from the group consisting of 350, 405, AF532, AF488, AF647, AF680, AF750, Cy5, Cy5.5, Cy7, indocyanine green (ICG), green fluorescent protein (GFP), red fluorescent protein (RFP), dsRED, and IRdye 800.

[0061] Another embodiment of this aspect of the disclosure is realized when the compounds of Formula I, Formula I', and Formula Ia comprise one or more imaging agents comprising one or more independently selected radioisotopes. A subembodiment of this aspect of the disclosure is realized when the radioisotopes provided herein are useful as imaging agents in one or more of the methods provided herein. A subembodiment of this aspect of the disclosure is realized when one or more of the radioisotopes provided herein may also be useful in one or more therapeutic applications (e.g., when administered to a subject in a therapeutically effective amount). For example, 131 I and 64Cu may be useful as an imaging agent (e.g., as a non-toxic and / or non-therapeutic radioisotope) when administered to a subject at low concentrations (e.g., 5 mCi), and may also be useful as a therapeutic agent (i.e., as a toxic and / or therapeutic radioisotope) when administered to a subject at higher concentrations. A subembodiment of this aspect of the present disclosure is one in which each of the radioisotopes is: 3 H, 11 C. 14 C. 18 F, 32 P, 35 S, 36 C1, 51 Cr 52 Fe, 57 Co, 58 Co, 59 Fe, 64 Cu, 67 Cu, 67 Ga, 68 Ga, 75 Se, 76 Br, 77 Br, 89 Zr, 90 Y, 99m Tc 111 In 123 I 124 I 125 I 131 I 152 EU 153 Sm 166 Ho, 177 Lu, 186 Re, 188 Re, 201 Tl, 203 Pb, 212 Pb, 210 At, 211 At, 212 Bi, 213 Bi, and 225 A subembodiment of this aspect of the disclosure is realized when one or more independently radioisotopes are independently attached, directly or indirectly (e.g., via a chelator) to a compound of Formula I, Formula I', and Formula Ia.

[0062] Another embodiment of the present disclosure is realized when the compound of Formula I, Formula I', and Formula Ia comprises one or more imaging agents selected from PET (positron emission tomography), SPECT (single photon emission computed tomography), and computed tomography imaging agents. A subembodiment of this aspect of the present disclosure is realized when the imaging agent is PET. Another subembodiment of this aspect of the present disclosure is realized when the imaging agent is a SPECT. Another subembodiment of this aspect of the present disclosure is realized when the imaging agent is a computed tomography imaging agent. Another subembodiment of this aspect of the present disclosure is realized when the imaging agent is a radioisotope computed tomography imaging agent. Another subembodiment of the present disclosure is realized when the imaging agent is 11 C. 18 F, 64 Cu, 68 Ga, 76 Br, 77 Br, 89 Zr, 111 In, 123 I, 124 I, 186 Re, 188 Re, and 201 In a further aspect of the present disclosure, the imaging agent is a PET or SPECT imaging agent comprising one or more radioisotopes selected from: 68 In a further embodiment, non-limiting radioisotopes that form stable complexes with chelating moieties and have suitable physical half-lives for PET imaging purposes include: 89 Zr, 68 Ga, 64 Cu, 44 Sc, and 86 Y. In a further embodiment, non-limiting radioisotopes directly attached to the peptide are selected from: 76 Br and 124 In yet a further embodiment, non-limiting radioisotopes introduced via artificial groups include, but are not limited to, I. 18 It's F.

[0063] Another embodiment of this disclosure is realized when a compound of Formula I, Formula I', and Formula Ia, or a pharmaceutically acceptable salt thereof, is linked to one or more imaging agents via a linking group. A subembodiment of this aspect of the disclosure is realized when the linking group comprises one or more amino acid residues. Subembodiments of this aspect of the disclosure are realized when about 1 to about 100, about 1 to about 80, about 1 to about 60, about 1 to about 40, about 1 to about 20, about 1 to about 10, about 1 to about 5, about 5 to about 100, about 5 to about 80, about 5 to about 60, about 5 to about 40, about 5 to about 20, about 5 to about 10, about 10 to about 100, about 10 to about 80, about 10 to about 60, about 10 to about 40, about 10 to about 20, about 20 to about 100, about 20 to about 80, about 20 to about 60, about 20 to about 40, about 40 to about 100, about 40 to about 80, about 40 to about 60, about 60 to about 100, about 60 to about 80, or about 80 to about 100 amino acid residues are present.

[0064] Another embodiment of this aspect of the disclosure is realized when the linking group comprises one or more alkylene groups, one or more amine groups, one or more amide groups, one or more alkyleneoxy groups, one or more thiol groups, one or more carbohydrate groups, or any combination thereof. A subembodiment of this aspect of the disclosure is when the linking group comprises one or more C 1-50 an alkylene group, one or more amine groups, one or more amide groups, one or more C 1-50 Alkyleneoxy group, one or more C 1-50 Another subembodiment of this aspect of the disclosure is realized when the linking group comprises one or more -(OCH2CH2) p-group, and p is an integer of, for example, about 1 to about 100, about 1 to about 80, about 1 to about 60, about 1 to about 40, about 1 to about 20, about 1 to about 10, about 1 to about 5, about 5 to about 100, about 5 to about 80, about 5 to about 60, about 5 to about 40, about 5 to about 20, about 5 to about 10, about 10 to about 100, about 10 to about 80, about 10 to about 60, about 10 to about 40, about 10 to about 20, about 20 to about 100, about 20 to about 80, about 20 to about 60, about 20 to about 40, about 40 to about 100, about 40 to about 80, about 40 to about 60, about 60 to about 100, about 60 to about 80, or about 80 to about 100 amino acid residues. In some embodiments, p is an integer from about 10 to about 40. In some embodiments, p is an integer from about 20 to about 40. In some embodiments, p is an integer from about 25 to about 35.

[0065] Another embodiment of the present disclosure is realized when a compound of Formula I, Formula I', and Formula Ia, or a pharmaceutically acceptable salt thereof, is linked to a chelating moiety that contains one or more of the imaging agents described herein.

[0066] Another embodiment of the present disclosure of Formula I, Formula I′ and Formula Ia is R 1 and / or R 1’ is optionally substituted phenyl or C 1-6 alkyl, and R 2 and / or R 2’ But hydrogen, C 1-6 alkyl, and fluorine; R 3 and / or R 3’ is selected from hydrogen, —CH hydroxyl, —CH NH , and —CH C(O)NH ; R 4 and R 5 and / or R 4’ and R 5’ One of the groups is hydrogen or C 1-6 alkyl and the other is C 1-6 alkyl, (CH2)3NHC(=NH2)NH2, and (CH2)2NH2, and R 6 and / or R 6’ is hydrogen or C 1-6 alkyl, and R 10and / or R 10’ optionally substituted CH2 indolyl or C 1-6 alkyl, and R 11 and R 12 and / or R 11’ and R 12’ taken together to form an optionally substituted pyrrolidinyl, tetrahydroisoquinolinyl, or piperidinyl.

[0067] Another embodiment of the present disclosure of Formula I, Formula I′ and Formula Ia is R 1 and / or R 1’ is optionally substituted phenyl or C 1-6 alkyl, and R 2 and / or R 2’ But hydrogen, C 1-6 alkyl, and fluorine; R 3 and / or R 3’ is selected from hydrogen, CH2 hydroxyl, CH2NH2, and CH2C(O)NH2; R 4 and R 6 and / or R 4’ and R 6’ taken together form optionally substituted pyrrolidinyl, indolyl, piperidinyl, azetidinyl, morpholinyl, and isoquinolinyl; R 5 and / or R 5’ is hydrogen and R 10 and / or R 10’ optionally substituted CH2 indolyl or C 1-6 alkyl, and R 11 and R 12 and / or R 11’ and R 12’ taken together to form an optionally substituted pyrrolidinyl, tetrahydroisoquinolinyl, or piperidinyl.

[0068] Another embodiment of the present disclosure of Formula I, Formula I′ and Formula Ia is R 1 and / or R 1’ is optionally substituted phenyl or C1-6 alkyl, and R 2 and / or R 2’ But hydrogen, C 1-6 alkyl, and fluorine; R 3 and / or R 3’ is selected from hydrogen, CH2 hydroxyl, CH2NH2, and CH2C(O)NH2; R 4 and R 6 and / or R 4’ and R 6’ taken together form an optionally substituted pyrrolidinyl, indolyl, piperidinyl, azetidinyl, morpholinyl, or isoquinolinyl, and R 5 and / or R 5’ is hydrogen and R 10 and / or R 10’ is an optionally substituted CH2 indolyl, and R 11 and R 12 and / or R 11’ and R 12’ together form an optionally substituted pyrrolidinyl, R 13 and / or R 13 'But NH2, [ka]

[0069] A subembodiment is realized when R 7 and / or R 7’ is hydrogen and R 8 and R 9 and / or R 8’ and R 9’ taken together to form an unsubstituted pyrrolidinyl or a fluorine-substituted pyrrolidinyl. Another subembodiment is realized when R 7 and R 8 and / or R 7’ and R 8’is hydrogen and the other is selected from CH3, CH2CH(CH3)2, CH(CH3)CH2CH3, CH(CH3)CH2CH3, CH(CH3)2, )CH2OH, -CH2COOH, -CH(OH)CH3, -(CH2)2C(O)NH2, -(CH2)indolyl, -CH2phenyl, -CH2phenylOH, -(CH2)4NH2, -(CH2)3NHC(=NH)NH2, and -CH2imidazolyl; 9 and / or R 9’ is hydrogen. Another subembodiment is realized when R 7 and R 8 and / or R 7’ and R 9’ is hydrogen and the other is selected from -CH, -CHCH(CH), -CHOH, -CHCOOH, and -CH(OH)CH. Another subembodiment is realized when R 7 and R 8 and / or R 7’ and R 8’ is hydrogen and the other is selected from -CHCOOH and -CH(OH)CH. Another subembodiment is realized when R 3 and / or R 3’ Another subembodiment is realized when R 1 and / or R 1’ is —CH(CH3)2. Another subembodiment is realized when R 1 and / or R 1’ is optionally substituted phenyl. Another subembodiment is realized when R 4 and R 6 and / or R 4’ and R 6’ and R are taken together to form an optionally substituted pyrrolidinyl. Another subembodiment is realized when R 4 and R 6 and / or R 4’ and R 6’ and R are taken together to form an optionally substituted indolyl. 4 and R6 and / or R 4’ and R 6’ and R are taken together to form an optionally substituted piperidinyl. 4 and R 6 and / or R 4’ and R 6’ and R are taken together to form an optionally substituted azetidinyl. 4 and R 6 and / or R 4’ and R 6’ and R are taken together to form an optionally substituted morpholinyl. 4 and R 6 and / or R 4’ and R 6’ and R are taken together to form an optionally substituted isoquinolinyl. 4 and R 6 and / or R 4’ and R 6’ Another subembodiment is realized when the substituents on the pyrrolidinyl, indolyl, piperidinyl, azetidinyl, morpholinyl, and isoquinolinyl of R are selected from methyl and fluorine. 11 and R 12 and / or R 11’ and R 12’ and R are taken together to form an optionally substituted pyrrolidinyl. Another subembodiment is realized when R 11 and R 12 and / or R 11’ and R 12’ and R are taken together to form an optionally substituted tetrahydroisoquinolinyl. 11 and R 12 and / or R 11’ and R 12’ and R are taken together to form an optionally substituted piperidinyl. 11 and R 12and R 11’ and R 12’ Both are C 1-6 Another subembodiment is realized when R 13 is NH2. Another subembodiment is realized when R 13 and / or R 13’ but [ka]

[0070] Another subembodiment of this aspect of the disclosure is realized when R 13 and / or R 13’ but [ka]

[0071] This is realized when

[0072] In one embodiment of the present disclosure of Formula I, Formula I′, and Formula Ia, R 1 and / or R 1’ is optionally substituted phenyl or C 1-6 alkyl, and R 2 and / or R 2’ But hydrogen, C 1-6 alkyl, and fluorine; R 3 and / or R 3’ is selected from hydrogen, —CH hydroxyl, —CH NH , and —CH C(O)NH ; R 4 and R 5 and / or R 4’ and R 5’ One of the groups is hydrogen or C 1-6 alkyl and the other is C 1-6 alkyl, (CH2)3NHC(=NH2)NH2, and (CH2)2NH2, and R 6 and / or R 6’ is hydrogen or C 1-6 alkyl, and R 10 and / or R 10’is an optionally substituted CH2 indolyl, and R 11 and R 12 and / or R 11’ and R 12’ taken together form an optionally substituted tetrahydroisoquinolinyl, pyrrolidinyl, or piperidinyl, and R 13 and / or R 13’ But NH2, [ka]

[0073] Another subembodiment of this aspect of the disclosure is realized when R 7 and / or R7' is hydrogen and R 8 and R 9 and / or R 8’ and R 9’ taken together to form an unsubstituted pyrrolidinyl or a fluorine-substituted pyrrolidinyl. Another subembodiment is realized when R 7 and R 8 and / or R 7’ and R 8’ is hydrogen and the other is selected from —CH, —CHCH(CH), —CH(CH)CHCH, —CH(CH)CHCH, —CH(CH)CH, —CH(CH)—CHOH, —CHCOOH, —CH(OH)CH, —(CH)C(O)NH, —(CH)indolyl, —CHphenyl, —CHphenylOH, —(CH)NH, —(CH)NHC(═NH)NH, and —CHimidazolyl; 9 and / or R 9’ is hydrogen. Another subembodiment is realized when R 7 and R 8 and / or R 7’ and R 8’ is hydrogen and the other is selected from -CH, -CHCH(CH), -CHOH, -CHCOOH, and -CH(OH)CH. Another subembodiment is realized when R 7 and R 8and / or R 7’ and R 8’ is hydrogen and the other is -CHCOOH and -CH(OH)CH. Another subembodiment is realized when R 3 and / or R 3’ is CHOH. Another subembodiment is realized when R 1 and / or R 1’ is —CH(CH3)2. Another subembodiment is realized when R 1 and / or R 1’ is optionally substituted phenyl. Another subembodiment is realized when R 11 and R 12 and / or R 11’ and R 12’ and R are taken together to form an optionally substituted pyrrolidinyl. Another subembodiment is realized when R 11 and R 12 and / or R 11’ and R 12’ and R are taken together to form an optionally substituted tetrahydroisoquinolinyl. 11 and R 12 and / or R 11’ and R 12’ and R are taken together to form an optionally substituted piperidinyl. 13 is NH2. Another subembodiment is realized when R 13 and / or R 13’ but [ka]

[0074] Another subembodiment is realized when R 13 and / or R 13’ but [ka]

[0075] This is realized when

[0076] Another embodiment of the present disclosure of Formula I is represented by structural formula Ia [ka]

[0077] (In the formula, R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 11 , R 12 and R 13 is as described herein, and R x and R y are independently hydrogen, hydroxyl, C 1-6 alkyl and halogen) This is achieved by:

[0078] A subembodiment of the present disclosure of Formula Ia is realized when it includes one or more positron-emitting imaging agents.

[0079] Another subembodiment of the present disclosure of formula Ia is R x and R y is realized when independently is methyl or fluorine. a One embodiment of the present disclosure is R 2 But hydrogen, C 1-6 alkyl, and fluorine; R 3 is selected from hydrogen, —CHOH, —CHNH, and —CHC(O)NH; R 4 and R 5 One of the groups is hydrogen or C 1-6 alkyl and the other is C 1-6 alkyl, —(CH2)3NHC(═NH)NH2, and —(CH2)2NH2; R 6 is hydrogen or C 1-6 alkyl, and R 11 and R 12are taken together to form optionally substituted pyrrolidinyl, tetrahydroisoquinolinyl, and piperidinyl. Another subembodiment of the present disclosure of formula Ia is realized when R 7 and R 8 is hydrogen and the other is selected from -CH, -CHCH(CH), -CH(CH)CHCH, -CH(CH)CHCH, -CH(CH)CH, -CH(CH), -CHOH, -CHCOOH, -CH(OH)CH, -(CH)C(O)NH, -(CH)indolyl, -CHphenyl, -CHphenylOH, -(CH)NH, -(CH)NHC(=NH)NH, and -CHimidazolyl. Another subembodiment of the present disclosure of Formula Ia is realized when R 7 and R 8 is hydrogen and the other is selected from -CH, -CHCH(CH), -CHOH, -CHCOOH, and -CH(OH)CH. Another subembodiment of the present disclosure of Formula Ia is realized when R 3 Another subembodiment of this aspect of the disclosure of Formula Ia is realized when R 13 is —NH or —CHNH. Yet another subembodiment of Formula Ia is realized when R 13 but [ka]

[0080] Yet another subembodiment of this aspect of the disclosure of Formula Ia is realized when R 13 but [ka]

[0081] Another subembodiment of formula Ia is realized when R 14 R 14 (i). Another subembodiment of formula Ia is realized when R 14 R 14 Another subembodiment of formula Ia is realized when R14 R 14 (iii). Another subembodiment of formula Ia is realized when R 14 R 14 (iv) Another subembodiment of formula Ia is realized when R 14 R 14 (v) Another subembodiment of formula Ia is realized when R 14 R 14 (vi). Another subembodiment of formula Ia is realized when R 14 R 14 (vii). Another subembodiment of formula Ia is realized when R 14 R 14 (viii). Yet another subembodiment of formula Ia is realized when R 14 R 14 (vii) or (viii), and is realized when X is as described herein.

[0082] Another subembodiment of the present disclosure of formula Ia is R 2 and / or R 2’ But hydrogen, C 1-6 alkyl, and fluorine; R 3 and / or R 3’ is selected from hydrogen, —CHOH, —CHNH, and —CHC(O)NH; R 4 and R 6 and / or R 4’ and R 6’ taken together form optionally substituted pyrrolidinyl, indolyl, piperidinyl, azetidinyl, morpholinyl, and isoquinolinyl; R 5 and / or R5' is hydrogen and R 11 and R 12 and / or R 11’ and R 12’ are taken together to form optionally substituted pyrrolidinyl, tetrahydroisoquinolinyl, and piperidinyl. A subembodiment of this aspect of the disclosure of formula Ia is realized when R 7 and / or R 7’ is hydrogen and R8 and R 9 and / or R 8’ and R 9’ and R are taken together to form an optionally substituted pyrrolidinyl. Another subembodiment of this aspect of the disclosure is 7 and R 8 and / or R 7’ and R 8’ is hydrogen and the other is selected from -CH3, CH2CH(CH3)2, -CH(CH3)CH2CH3, -CH(CH3)CH2CH3, -CH(CH3)2, -CH2OH, -CH2COOH, -CH(OH)CH3, -(CH2)2C(O)NH2, -(CH2)indolyl, -CH2phenyl, -CH2phenylOH, -(CH2)4NH2, -(CH2)3NHC(=NH)NH2, and -CH2imidazolyl; 9 and / or R 9’ Another subembodiment of this aspect of the disclosure of Formula Ia is realized when R 7 and R 8 and / or R 7’ and R 8’ is hydrogen and the other is selected from -CH, -CHCH(CH), -CHOH, -CHCOOH, and -CH(OH)CH. Another subembodiment of this aspect of the disclosure of Formula Ia is realized when R 7 and R 8 and / or R 7’ and R 8’ is hydrogen and the other is -CHCOOH and -CH(OH)CH. Another subembodiment of the present disclosure of formula Ia is realized when R 3 and / or R 3’ Another subembodiment of the present disclosure of formula Ia is realized when R 4 and R 6 and / or R 4’ and R 6’ Another subembodiment of the present disclosure of formula Ia is realized when R 4 and R 6and / or R 4’ and R 6’ Another subembodiment of the present disclosure of formula Ia is realized when R 4 and R 6 and / or R and R taken together form an optionally substituted piperidinyl. Another subembodiment of the present disclosure of formula Ia is realized when R 4 and R 6 and / or R 4’ and R 6’ Another subembodiment of the present disclosure of formula Ia is realized when R 4 and R 6 and / or R 4’ and R 6’ Another subembodiment of the present disclosure of formula Ia is realized when R 4 and R 6 and / or R 4’ and R 6’ Another subembodiment of this aspect of the disclosure of formula Ia is realized when R 4 and R 6 and / or R 4’ and R 6’ Another subembodiment of the present disclosure of formula Ia is realized when the substituents on pyrrolidinyl, indolyl, piperidinyl, azetidinyl, morpholinyl, and isoquinolinyl formed from the combination of: are selected from methyl and fluorine. 11 and R 12 and / or R 11’ and R 12’ Another subembodiment of the present disclosure of formula Ia is realized when R 11 and R 12 and / or R 11’ and R 12’Another subembodiment of the present disclosure of formula Ia is realized when R 11 and R 12 and / or R 11’ and R 12’ Another subembodiment of this aspect of the disclosure of formula Ia is realized when R 13 Yet another subembodiment of this aspect of the disclosure of Formula Ia is realized when R 13 but [ka]

[0083] Yet another subembodiment of this aspect of the disclosure of Formula Ia is realized when R 13 but [ka]

[0084] Another subembodiment is realized when R 14 R 14 (i). Another subembodiment is realized when R 14 R 14 (ii). Another subembodiment is realized when R 14 R 14 (iii). Another subembodiment is realized when R 14 R 14 (iv). Another subembodiment is realized when R 14 R 14 (v). Another subembodiment is realized when R 14 R 14 (vi). Another subembodiment is realized when R 14 R 14 (vii). Another subembodiment is realized when R 14 R 14 (viii). Yet another subembodiment is realized when R14 R 14 (vii) or (viii) and is realized when X is as described herein. Another subembodiment of this aspect of formula Ia is R 15 is H, -C(O)(CH2)2NHC(O)OC(CH3)3, -C(O)CH2NH(NOTA-1), and R 14 Another subembodiment of this aspect of formula Ia is realized when R 4 and R 6 and / or R 4’ and R 6’ are taken together to form optionally substituted pyrrolidinyl, indolyl, piperidinyl, azetidinyl, morpholinyl, and isoquinolinyl. Another subembodiment of this aspect of Formula Ia is realized when R 3 and / or R 3’ is CH2OH.

[0085] The compounds of the present disclosure may contain one or more asymmetric centers and therefore may exist as racemates and racemic mixtures, single enantiomers, diastereomeric mixtures, and individual diastereomers. Depending on the nature of the various substituents on the molecule, additional asymmetric centers may exist. Each such asymmetric center independently produces two optical isomers, and all possible optical isomers and diastereomers, in mixtures and as pure or partially purified compounds, are intended to be included within the scope of the present disclosure. Unless a specific stereochemistry is indicated, the present disclosure is intended to encompass all such isomeric forms of these compounds.

[0086] The independent syntheses of these diastereomers or their chromatographic separations may be achieved as known in the art by appropriate modification of the methodology disclosed herein. Their absolute stereochemistry may be determined, among other methods, by X-ray crystallography of crystalline products or crystalline intermediates which are derivatized, if necessary, with a reagent containing an asymmetric center of known absolute configuration.

[0087] If desired, a racemic mixture of a compound can be separated so that individual enantiomers are isolated. Separation can be carried out by methods well known in the art, for example, by coupling a racemic mixture of a compound with an enantiomerically pure compound to form a diastereomeric mixture, and then separating the individual diastereomers by standard methods such as fractional crystallization or chromatography. The coupling reaction is often the formation of a salt with an enantiomerically pure acid or base. The diastereomeric derivative can then be converted to a pure enantiomer by cleaving the added chiral residue. A racemic mixture of a compound can also be directly separated by chromatographic methods using chiral stationary phases, which are well known in the art.

[0088] Alternatively, any enantiomer of a compound may be obtained by stereoselective synthesis using optically pure starting materials or reagents of known configuration by methods well known in the art.

[0089] In compounds of Formula I or Ia, atoms may exhibit their natural isotopic abundance, or one or more of the atoms may be artificially enriched in a particular isotope having the same atomic number but an atomic mass or mass number different from that predominantly found in nature. The present disclosure may include all suitable isotopic variations of compounds of general Formula I or Ia. For example, different isotopic forms of hydrogen (H) include protium ( 1 H) and deuterium ( 2H). Protium is the predominant hydrogen isotope found in nature. Enrichment with deuterium may provide certain therapeutic advantages (e.g., increased in vivo half-life or reduced dosage requirements) or may result in compounds useful as standards for characterizing biological samples. For purposes of this disclosure, when a compound is referred to as "non-deuterated," it means that it is not enriched in deuterium above background conditions. Isotopically enriched compounds within general formula I or Ia may be prepared without undue experimentation by conventional techniques well known to those skilled in the art, or by methods similar to those described in the schemes and examples herein using appropriate isotopically enriched reagents and / or intermediates.

[0090] When compounds of the present disclosure can form tautomers, all such tautomeric forms are included within the scope of the present disclosure. For example, compounds containing a carbonyl -CHC(O)- group (keto form) can undergo tautomerization to form a hydroxyl -CH=C(OH)- group (enol form). When present, both the keto and enol forms are included within the scope of the present disclosure.

[0091] Any variable (e.g., R 5 When a group (e.g., aryl, aryl, aryl) occurs more than one time, its definition on each occurrence is independent at every other occurrence. Also, combinations of substituents and variables are permissible only if such combinations result in stable compounds. Lines drawn from substituents into ring systems indicate that the indicated bond may be attached to any of the substitutable ring atoms. When the ring system is bicyclic, it is intended that the bond be attached to any of the suitable atoms on either ring of the bicyclic moiety.

[0092] It is understood by those skilled in the art that one or more silicon (Si) atoms can be incorporated into the compounds of the present disclosure instead of one or more carbon atoms, in order to provide compounds that are chemically stable and can be easily synthesized from readily available starting materials by techniques known in the art.Carbon and silicon have different covalent bond radii, and as a result, when comparing the bonds of similar C and Si elements, the bond distance and steric configuration are different.These differences result in slight changes in the size and shape of silicon-containing compounds compared with carbon.Those skilled in the art will understand that the difference in size and shape can result in slight or dramatic changes in potency, solubility, lack of off-target activity, packaging properties, etc. (Diass, JO et al. Organometallics (2006) 5:1188-1198; Showell, GA et al. Bioorganic & Medicinal Chemistry Letters (2006) 16:2555-2558).

[0093] It is understood that the substituents and substitution patterns on the compounds of the present disclosure can be selected by those skilled in the art to provide compounds that are chemically stable and can be easily synthesized from readily available starting materials by techniques known in the art and the methods described below.When a substituent itself is substituted with two or more groups, it is understood that these multiple groups can be on the same carbon or different carbons, as long as a stable structure is obtained.The phrase "optionally substituted with one or more substituents" should be understood to mean that the group in question is unsubstituted or can be substituted with one or more substituents.

[0094] Absolute stereochemistry is indicated by the use of hashed and solid wedge bonds, as shown in Illus-I and Illus-II. Thus, the methyl group in Illus-I rises from the page, the ethyl group in Illus-II falls into the page, and the cyclohexene ring is within the plane of the paper. The hydrogen on the same carbon as the methyl group in Illus-I falls into the page, and the hydrogen on the same carbon as the ethyl group in Illus-II rises from the page. The convention remains the same, with both hashed and solid rectangles attached to the same carbon in Illus-III, with the methyl group rising from the page, the ethyl group falling into the page, and the cyclohexene ring remaining within the plane of the paper. [ka]

[0095] As is conventional, unless otherwise indicated in the accompanying text, a regular "stick" or "wavy" bond indicates that all possible stereochemistries are represented, including pure compounds, mixtures of isomers, and racemic mixtures.

[0096] As used herein, unless otherwise specified, the following terms have the following meanings:

[0097] The phrase "at least one" when used in reference to the number of components comprising a composition, e.g., "at least one pharmaceutical excipient," means that one member of a particular group is present in the composition, and more than one may be present. The components of the composition are typically aliquots of isolated, pure materials that are added to the composition, and the purity level of the isolated material that is added to the composition is the purity level normally accepted for that type of reagent.

[0098] The phrase "one or more," whether used in reference to substituents on a compound or component of a pharmaceutical composition, means the same as "at least one."

[0099] "Effective amount" or "therapeutically effective amount" is meant to describe providing an amount of at least one active compound or pharmaceutical agent of the present disclosure, or a composition comprising at least one compound or pharmaceutical agent of the present disclosure, that is effective in eliciting the desired biological or medical response in a tissue, system, animal, individual, or human, or in treating or inhibiting a disease or condition described herein, thereby resulting in the desired therapeutic, ameliorative, inhibitory, or prophylactic effect. For example, when treating a central nervous system disease or disorder with one or more of the compounds described herein, an "effective amount" (or "therapeutically effective amount") means providing an amount of at least one compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt thereof that results in a therapeutic response in a patient suffering from a central nervous system disease or disorder ("condition") (including, for example, a response suitable for managing, alleviating, ameliorating, or treating the condition, or alleviating, ameliorating, reducing, or eradicating the condition and / or one or more symptoms resulting from long-term stabilization of the condition, as determined by analysis of pharmacodynamic markers or clinical evaluation of the patient suffering from the condition).

[0100] "Patient" and "subject" mean an animal, such as a mammal (e.g., a human), preferably a human.

[0101] "Prodrug" means a compound that is rapidly converted in vivo to the parent compound, for example, by hydrolysis in blood; for example, conversion of a prodrug of Formula I to Formula IV to a compound of Formula I, Formula I', Formula II, Formula III, or Formula IV, or a salt thereof, is included; thorough discussions are provided in T. Higuchi and V. Stella, Prodrugs as Novel Delivery Systems, Vol. 14 of the ACS Symposium Series, and Edward B. Roche, ed., Bioreversible Carriers in Drug Design, American Pharmaceutical Association and Pergamon Press, 1987 (both of which are incorporated herein by reference); the scope of the present disclosure includes prodrugs of the novel compounds of the present disclosure.

[0102] The term "substituted" means that one or more of the listed substituents can occupy one or more of the bonding positions on the substrate typically occupied by "-H," provided that such substitution does not exceed the normal valence rules for atoms in the bonding configuration presented on the substrate, and the substitution ultimately provides a stable compound; i.e., such substitution does not provide the compound with mutually reactive substituents located geminally or adjacent to one another. The substitution provides a compound that is sufficiently robust to withstand isolation from a reaction mixture to a useful degree of purity.

[0103] When optional substitution of a moiety is described (e.g., "optionally substituted"), the term means that, if substituents are present, one or more of the listed substituents for a particular substrate may be present on the substrate at the bonding position normally occupied by the default substituent normally occupying that position. For example, the default substituent on a carbon atom of an alkyl moiety is a hydrogen atom, and any substituent can replace the default substituent.

[0104] As used herein, unless otherwise specified, the following terms used to describe moieties, whether including the overall definition of a variable portion of a structural representation of a compound of the disclosure or a substituent attached to a variable portion of a structural representation of a group of a compound of the disclosure, have the following meanings, and unless otherwise specified, the definition of each term (i.e., moiety or substituent) applies when that term is used individually or as a component of another term (e.g., the definition of aryl is the same for aryl, and aryl moieties such as arylalkyl, alkylaryl, and arylalkynyl moieties). Moieties are equivalently described herein by structure, typographical designation, or chemical terminology without any intended distinction in meaning, e.g., an "acyl" substituent is referred to by the term "acyl," the typographical designation "R'-(C=O)-" or "R'-C(O)-," or by the structural designation: [ka]

[0105] and the like, and no distinction is implied by the use of any or all of these designations.

[0106] The term "alkyl" (including the alkyl portions of other moieties such as trifluoromethyl-alkyl- and alkoxy-) means a straight- or branched-chain aliphatic hydrocarbon moiety containing up to about 20 carbon atoms (e.g., "C 1-20 (The term "C-alkyl" refers to an aliphatic hydrocarbon moiety of 1 to 20 carbon atoms.) In some embodiments, alkyl preferably contains up to about 10 carbon atoms; for example, alkyl moieties of 1 up to 8 carbon atoms are referred to herein as "C-alkyl" unless the term is modified by an indication that a shorter chain is contemplated. 1-8 When the term "alkyl" is indicated with two hyphens (i.e., "-alkyl-"), it indicates that the alkyl moiety is attached to connect the substituents on either side of it, e.g., "-alkyl-OH" indicates an alkyl moiety connecting a hydroxyl moiety to a substrate.

[0107] Unless otherwise specified, "natural amino acid" refers to any one of the 20 amino acids commonly found in nature and synthesized peptides and known by the one-letter abbreviations A, R, N, C, D, Q, E, G, H, I, L, K, M, F, P, S, T, W, Y and V.

[0108] The term "unnatural amino acid" refers to a molecule that is structurally similar to an amino acid and can be substituted for an amino acid in the formation of a macrocycle. Unnatural analogs include, but are not limited to, compounds that are structurally identical to the amino acids defined herein except that they contain one or more additional methylene groups between the amino and carboxyl groups (e.g., α-amino acids, β-carboxy acids), or replace the amino or carboxy group with a similarly reactive group (e.g., replacing a primary amine with a secondary or tertiary amine, or replacing a carboxy group with an ester).

[0109] Substitutions for specific amino acids can be conservative, i.e., substitutions of amino acids with other amino acids have similar characteristics (e.g., charge, side chain size, hydrophobicity / hydrophilicity, main chain conformation and rigidity, etc.), so that changes can frequently be made without changing the biological activity of the protein. Those skilled in the art generally recognize that single amino acid substitutions in non-essential regions of a polypeptide do not substantially change biological activity (see, for example, Watson et al., Molecular Biology of the Gene, The Benjamin / Cummings Pub. Co., p. 224 (4th Ed.) (1987)). Furthermore, substitutions of structurally or functionally similar amino acids are unlikely to destroy biological activity. Exemplary conservative substitutions are shown in Table Y. [Table 1]

[0110] "Dose," "dosage," "unit dose," "unit dosage," "effective dose," and related terms refer to a physically discrete unit containing a predetermined quantity of active ingredient (e.g., peptidomimetic macrocycle) calculated to produce a desired therapeutic effect (e.g., cancer cell death). These terms are synonymous with therapeutically effective amount and amount sufficient to achieve the stated objectives of the methods disclosed herein.

[0111] The term "cycloalkyl" refers to a moiety having a main hydrocarbon chain forming a monocyclic or bicyclic aliphatic moiety containing at least 3 carbon atoms (the minimum number necessary to provide a monocyclic moiety) up to the specified maximum number of carbon atoms (generally 8 for monocyclic moieties and 10 for bicyclic moieties, including spirocyclic moieties). Examples of cycloalkyl moieties include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl. The term "cycloalkyl" also generally includes non-aromatic fused polycyclic ring systems containing up to 20 carbon atoms, which may be substituted as defined herein for "alkyl." Suitable polycyclic cycloalkyls include, but are not limited to, 1-decalin, norbornyl, adamantly, and the like.

[0112] As used herein, the term "alkylene" refers to a saturated, straight-chain or branched-chain aliphatic hydrocarbon group having two residues derived from the removal of two hydrogen atoms from the same carbon atom or two different carbon atoms of a parent alkane. Alkylene is a straight-chain or branched-chain group having 1 to 20 carbon atoms, preferably 1 to 12 carbon atoms, and more preferably 1 to 6 carbon atoms. Non-limiting examples include methylene, ethylene, propylene, butylene, pentylene, etc.

[0113] As used herein, when the term "alkyl" is modified by "substituted" or "optionally substituted," it means that one or more C-H bonds in an alkyl moiety are replaced or optionally substituted by a substituent attached to the alkyl group that is invoked in defining the moiety.

[0114] The term "cycloalkyl" refers to a saturated or partially unsaturated monocyclic or polycyclic hydrocarbon substituent having 3 to 20 carbon atoms, preferably 3 to 12 carbon atoms, more preferably 3 to 10 carbon atoms, and most preferably 3 to 8 carbon atoms. Non-limiting examples of monocyclic cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatrienyl, cyclooctyl, and the like. Polycyclic cycloalkyls include cycloalkyls having spirocyclic, fused, or bridged rings.

[0115] When a structural formula represents a bond between a moiety and a substrate using a bond line that terminates in the middle of the structure, for example, the following notation: [ka]

[0116] indicates that, regardless of whether the structure is numbered or not, unless otherwise defined, the moiety may be attached to the substrate via any available ring atom, e.g., the numbered atom of the exemplary moiety.

[0117] The term "aryl" refers to a 6- to 14-membered all-carbon monocyclic or fused polycyclic ring (i.e., each ring in the system shares adjacent pairs of carbon atoms with another ring in the system) having a conjugated electron system, preferably a 6- to 10-membered aryl, such as phenyl and naphthyl, preferably phenyl.

[0118] The term "heteroaryl" refers to an aromatic 5- to 8-membered monocyclic, 8- to 12-membered bicyclic, or 11- to 14-membered tricyclic ring system having 1 to 3 heteroatoms for monocyclic, 1 to 6 heteroatoms for bicyclic, or 1 to 9 heteroatoms for tricyclic, wherein the heteroatoms are selected from O, N, or S (e.g., carbon atoms and 1 to 3, 1 to 6, or 1 to 9 heteroatoms of N, O, or S, respectively, for monocyclic, bicyclic, or tricyclic rings). Non-limiting examples of heteroaryl are imidazolyl, pyridyl, pyrazolyl, pyrimidinyl, furanyl, oxazolyl, triazolyl, oxadiazolyl, and thiophenyl. Heteroaryl groups described herein may also contain fused rings sharing a common carbon-carbon bond, such as indolyl.

[0119] The term "heterocyclyl" (or heterocycloalkyl) means a non-aromatic saturated monocyclic or polycyclic ring system containing 3 to 10 ring atoms, preferably 5 to 10 ring atoms, in which one or more of the atoms in the ring system is an element other than carbon, for example, nitrogen (e.g., azetidinyl, piperidyl, pyrrolidinyl, tetrahydroisoquinolinyl), oxygen (e.g., furanyl and tetrahydropyranyl), or sulfur (e.g., tetrahydrothiopheneyl and tetrahydrothiopyranyl), and the heteroatoms can be present alone or in combination, provided the moiety does not include adjacent oxygen and / or sulfur atoms present in the ring system. A heterocyclyl is optionally substituted by one or more independently selected substituents.

[0120] The nitrogen or sulfur atom of the heterocyclyl can be optionally oxidized to the corresponding N-oxide, S-oxide, or S,S-dioxide (SO2). Non-limiting examples of suitable monocyclic heterocyclyl rings include azetidinyl, piperidyl, pyrrolidinyl, piperazinyl, morpholinyl- [ka]

[0121] (Unless otherwise specified, the moiety is attached to the substrate through any of the ring carbon atoms C2, C3, C5, or C6), bicyclic rings such as thiomorpholinyl, thiazolidinyl, 1,3-dioxolanyl, 1,4-dioxanyl, tetrahydrofuranyl, tetrahydrothiophenyl, tetrahydrothiopyranyl, etc., and tetrahydroisoquinolinyl.

[0122] The term "solvate" refers to a pharmaceutically acceptable solvate formed by a compound of the present disclosure with one or more solvent molecules. Non-limiting examples of solvent molecules include water, ethanol, acetonitrile, isopropanol, DMSO, and ethyl acetate.

[0123] The term "halogen" means fluorine, chlorine, bromine, or iodine; preferred halogens are fluorine, chlorine, and bromine unless otherwise specified, when this term is used; a substituent that is a halogen atom means -F, -Cl, -Br, or -I; "halo" means a fluoro, chloro, bromo, or iodo substituent bonded to a defined moiety; for example, "haloalkyl" means an alkyl as defined above, where one or more of the bonding positions on the alkyl moiety typically occupied by a hydrogen atom are instead occupied by a halo group; and perhaloalkyl (or "fully halogenated" alkyl) means that all bonding positions not involved in bonding the alkyl substituent to the substrate are occupied by halogens; for example, if the alkyl is selected to be methyl, the term perfluoroalkyl means -CF.

[0124] The terms "hydroxyl" and "hydroxy" refer to the group HO-, and "hydroxyalkyl" refers to a substituent of the formula "HO-alkyl-", where the alkyl group is attached to a substrate and can be substituted or unsubstituted as defined above, with preferred hydroxyalkyl moieties comprising lower alkyls. Non-limiting examples of suitable hydroxyalkyl groups include hydroxymethyl and 2-hydroxyethyl.

[0125] Bond sequences are indicated by hyphens and moieties are represented by letters, e.g., -alkyl indicates a single bond between the substrate and the alkyl moiety, -alkyl-X indicates that the alkyl group attaches the "X" substituent to the substrate, and in structural representations, bond sequences are indicated by wavy lines terminating the bond representation, e.g., [ka]

[0126] indicates that the methylphenyl moiety is attached to the substrate through the carbon atom ortho to the methyl substituent, whereas a bond designation drawn to a structure terminating in a wavy line and where the attached atom is not specifically shown indicates that the moiety may be attached to the substrate through any of the atoms of the moiety available for attachment, as described in the example above.

[0127] The line -, as a bond, generally indicates a mixture of or either of the possible isomers, including, for example, (R)- and (S)-stereochemical configurations.

[0128] Additionally, non-wedged bold or non-wedged hashed lines are used in structures containing multiple stereocenters to indicate known relative configurations. For example, [ka]

[0129] on the other hand, [ka] .

[0130] In all cases, the compound name(s) accompany the drawn structure and are intended to capture each of the possible stereochemical permutations for a given structural isomer based on the synthetic procedures used in its preparation. A list of separate stereoisomers attached or indicating that the presented compound (e.g., "Example Number") was isolated as a single stereoisomer and that the identity of that stereoisomer corresponds to one of the possible configurations listed. A list of separate stereoisomers attached or indicating that the presented compound was isolated as a racemic or diastereomeric mixture.

[0131] Specific absolute configurations are indicated by the use of a bold wedge or a hashed wedge. Unless a specific absolute configuration is indicated, the present disclosure is meant to encompass all such stereoisomeric forms of these compounds.

[0132] As used herein, when there are multiple oxygen and / or sulfur atoms present in a ring system, there cannot be any adjacent oxygen and / or sulfur atoms present in said ring system.

[0133] As is well known in the art, unless otherwise specified, a bond drawn from a particular atom without a moiety shown at the end of the bond indicates a methyl group attached to the atom through that bond. For example, [ka]

[0134] Any unsatisfied valences in the text, schemes, examples, structures, and any tables herein are assumed to have the hydrogen atom or atoms sufficient to satisfy the valences.

[0135] One or more compounds of the present disclosure may also exist as or be converted into a solvate. The preparation of solvates is generally known. Thus, for example, see M. Caira et al., J. Pharmaceutical Sci., 93(3), 601-611 (2004) describes the preparation of solvates of the antifungal fluconazole in ethyl acetate and water. Similar preparations of solvates and hemisolvates, including hydrates (where the solvent is water or an aqueous system), are described in EC van Tonder et al., AAPS PharmSciTech., 5(1), article 12 (2004) and ALBingham et al., Chem. Commun., 603-604 (2001). A typical, non-limiting method involves dissolving a compound of the invention in a desired amount of a desired solvent (e.g., an organic solvent, an aqueous solvent, water, or a mixture of two or more thereof) at above ambient temperature and cooling the solution at a rate sufficient to form crystals, which are isolated by standard methods, with or without an antisolvent present. Analytical techniques, such as IR spectroscopy, reveal the presence of the solvent (including water) in the crystals as a solvate (or hydrate, if water is incorporated into the crystalline form).

[0136] The present disclosure also includes the compounds of the present disclosure in isolated and purified form obtained by routine techniques.The polymorphic forms of the compounds of Formula I and Formula Ia, and the salts, solvates and prodrugs of the compounds of Formula I and Formula Ia, are intended to be included in the present disclosure.Certain compounds of the present disclosure may exist in different isomeric forms (e.g., enantiomers, diastereoisomers, atropisomers).The compounds of the present invention include all isomeric forms thereof, both in pure form and in mixtures of two or more, including racemic mixtures.

[0137] Similarly, unless otherwise indicated, the representation of a structure of any tautomeric form of a compound that exhibits tautomerism is meant to include all such tautomeric forms of the compound. Thus, when the compounds of the present disclosure, their salts, and their solvates and prodrugs can exist in different tautomeric forms or in equilibrium between such forms, all such forms of the compound are encompassed by and within the scope of the present disclosure. Examples of such tautomers include, but are not limited to, ketone / enol tautomeric forms, imine-enamine tautomeric forms, and heteroaromatic forms, such as, for example, the following moieties: [ka] .

[0138] The phrase "pharmaceutically acceptable" is used herein to refer to compounds, materials, compositions, and / or dosage forms that are suitable, within the scope of sound medical judgment, for use in contact with the tissues of human beings and animals without undue toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio.

[0139] As used herein, "pharmaceutically acceptable salts" refers to derivatives in which the parent compound is modified by making its acid or base salt. Salts in solid form may exist in two or more crystal structures and may also be in the form of hydrates. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines, alkali or organic salts of acidic residues such as carboxylic acids, and the like. Pharmaceutically acceptable salts include conventional non-toxic salts or quaternary ammonium salts of the parent compound formed, for example, from non-toxic inorganic or organic acids. For example, such conventional non-toxic salts include those prepared from inorganic acids such as formic acid, hydrochloric acid, hydrobromic acid, sulfuric acid, sulfamic acid, phosphoric acid, nitric acid, and the like, and from organic acids such as acetic acid, propionic acid, succinic acid, glycolic acid, stearic acid, lactic acid, malic acid, tartaric acid, citric acid, ascorbic acid, pamoic acid, maleic acid, hydroxymaleic acid, phenylacetic acid, glutamic acid, benzoic acid, salicylic acid, sulfanilic acid, 2-acetoxybenzoic acid, fumaric acid, toluenesulfonic acid, methanesulfonic acid, ethanedisulfonic acid, oxalic acid, isethionic acid, and the like. Salts derived from inorganic bases include aluminum, ammonium, calcium, copper, ferric acid, ferrous acid, lithium, magnesium, manganic acid, manganous, potassium, sodium, zinc, and the like.

[0140] When the compound of the present disclosure is basic, salts can be prepared from pharmaceutically acceptable non-toxic acids, including inorganic and organic acids. Such acids include acetic acid, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, citric acid, ethanesulfonic acid, fumaric acid, gluconic acid, glutamic acid, hydrobromic acid, hydrochloric acid, isethionic acid, lactic acid, maleic acid, malic acid, mandelic acid, methanesulfonic acid, mucic acid, nitric acid, pamoic acid, pantothenic acid, phosphoric acid, succinic acid, sulfuric acid, tartaric acid, p-toluenesulfonic acid, etc. In one embodiment of the present disclosure, the salts are citric acid, hydrobromic acid, hydrochloric acid, maleic acid, phosphoric acid, sulfuric acid, fumaric acid, and tartaric acid. Similarly, salts of acidic compounds can be formed by reaction with a suitable inorganic or organic base.

[0141] As used herein, the terms "treating" or "treatment" (e.g., of a disease, disorder, or condition or associated symptoms, which may together or individually be referred to as an "indication") include inhibiting the disease, disorder, or condition, i.e., halting or reducing the onset of the disease or its biological process or its progression or clinical symptoms, or alleviating the disease, i.e., causing regression of the disease or its biological process or progression and / or its clinical symptoms. As used herein, "treatment" also refers to controlling, ameliorating, or reducing the risk to a subject suffering from a disease, disorder, or condition involving a tumor. As used herein, the terms "preventing" a disease, disorder, or condition or "prevention" or "prophylaxis" include, for example, impeding the onset or progression of clinical symptoms of the disease, disorder, or condition in a mammal that may be exposed to or susceptible to the disease, disorder, or condition, but that has not yet experienced or displayed symptoms of the disease.

[0142] As will be apparent to one of skill in the art, the subjects treated by the methods described herein are generally mammals, including humans and non-human animals (e.g., laboratory animals and companion animals). The term "therapeutically effective amount" refers to that amount of the subject compound that elicits the biological or medical response in a tissue, system, animal, or human that is desired by a researcher, veterinarian, physician, or other clinician.

[0143] As used herein, the term "composition" is intended to encompass a product comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, together with a specified amount of one or more additional specified ingredients, as well as any product resulting directly or indirectly from the combination of the specified amounts of the specified ingredients. Such terms, with respect to pharmaceutical compositions, are intended to encompass products comprising an active ingredient(s), including a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, optionally together with one or more additional active ingredients and one or more inactive ingredients that constitute the carrier, as well as any product resulting directly or indirectly from the combination, complexation, or aggregation of any two or more of the ingredients, or from the dissociation of one or more of the ingredients, or from any other type of reaction or interaction of one or more of the ingredients. Thus, a pharmaceutical composition of the present disclosure encompasses any composition made by mixing a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, with a pharmaceutically acceptable carrier. "Pharmaceutically acceptable" means that the carrier, diluent, or excipient must be compatible with the other ingredients of the formulation and not deleterious to the recipient thereof.

[0144] In some embodiments, the compound of Formula I is a polypeptide that binds to Granzyme B, wherein the polypeptide comprises an amino acid sequence having at least 90% sequence identity to the sequence of Compound 1 in Table 1. In some embodiments, the compound of Formula I is a polypeptide that binds to Granzyme B, wherein the polypeptide comprises an amino acid sequence having at least 95% sequence identity to the sequence of Compound 1 in Table 1. In some embodiments, the compound of Formula I is a polypeptide that binds to Granzyme B, wherein the polypeptide comprises an amino acid sequence having at least 98% sequence identity to the sequence of Compound 1 in Table 1. In some embodiments, the compound of Formula I is a polypeptide that binds to Granzyme B, wherein the polypeptide comprises an amino acid sequence having at least 99% sequence identity to the sequence of Compound 1 in Table 1. In some embodiments, the compound of Formula I is a polypeptide that binds to Granzyme B, wherein the polypeptide comprises an amino acid sequence having at least 99.5% sequence identity to the sequence of Compound 1 in Table 1.

[0145] As noted above, additional embodiments of the present disclosure each relate to a method for treating a disease, disorder, or condition, or one or more symptoms thereof ("indications"), comprising administering to a subject in need of such treatment a therapeutically effective amount of a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising said compound or salt thereof. Accordingly, one embodiment of the present disclosure is a method for treating a disease, disorder, or condition, or one or more symptoms thereof ("indications") in which granzyme B is involved.

[0146] Another embodiment of this aspect of the disclosure is realized when the disease is selected from an autoimmune disorder, an inflammatory disorder, a skin disorder, cancer, and a cardiovascular disorder. A subembodiment of this aspect of the disclosure relates to the disease is a cancer selected from breast cancer, ovarian cancer, cervical cancer, uterine cancer, prostate cancer, kidney cancer, urethral cancer, bladder cancer, liver cancer, gastric cancer, endometrial cancer, salivary gland cancer, esophageal cancer, melanoma, glioma, neuroblastoma, sarcoma, lung cancer (e.g., small cell lung cancer and non-small cell lung cancer), colon cancer, rectal cancer, colorectal cancer, leukemia (e.g., acute lymphocytic leukemia, acute myeloid leukemia, acute promyelocytic leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia), bone cancer, skin cancer, thyroid cancer, pancreatic cancer, and lymphoma (e.g., Hodgkin's lymphoma, non-Hodgkin's lymphoma, or recurrent anaplastic large cell lymphoma). Another subembodiment of this aspect of the present disclosure relates to a method of treating or preventing cancer in a subject in need thereof, comprising administering to a subject in need thereof a therapeutically effective amount of a compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition comprising said compound, salt or solvate thereof, hi one such embodiment, the subject is a human.

[0147] Another embodiment of the present disclosure relates to a method for imaging Granzyme B. A subembodiment of this aspect of the present disclosure relates to a method in which imaging is performed on a cell, tissue, cell sample, tissue sample, or subject. As used herein, the term "subject" refers to any animal, including mammals (e.g., humans, domestic animals, livestock, etc.) and invertebrates (e.g., fish). Another embodiment of the present disclosure relates to a method for imaging Granzyme B in a cell or tissue, comprising contacting the cell or tissue with a compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt thereof, and imaging the cell or tissue using a suitable imaging technique to image Granzyme B in the cell or tissue. A subembodiment of this aspect of the present disclosure is realized when the compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt thereof comprises an imaging agent.

[0148] Another embodiment of the present disclosure relates to a method of imaging an immune response in a subject, comprising administering to the subject a compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt thereof, and imaging the subject using a suitable imaging technique to image Granzyme B in a cell or tissue. A subembodiment of this aspect of the present disclosure is realized when the compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt thereof comprises an imaging agent.

[0149] Another embodiment of the present disclosure relates to a method of monitoring an immune response in the treatment of a disease in a subject, comprising administering to the subject a compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt thereof, and imaging the subject using a suitable imaging technique to image granzyme B in a cell or tissue. A subembodiment of this aspect of the present disclosure is realized when the compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt thereof comprises an imaging agent.

[0150] Another embodiment of the present disclosure is one in which the compound is effective in treating graft versus host disease, rheumatoid arthritis, systemic lupus erythematosus, Hashimoto's thyroiditis, multiple sclerosis, myasthenia gravis, type 1 diabetes, uveitis, posterior uveitis, allergic encephalomyelitis, glomerulonephritis, rheumatic fever, post-infectious glomerulonephritis, psoriasis, atopic dermatitis, contact dermatitis, eczematous dermatitis, seborrheic dermatitis, lichen planus, pemphigus, bullous pemphigoid, epidermolysis bullosa, urticaria, angioedema, vasculitis, erythema, cutaneous eosinophilia, lupus erythematosus, acne, alopecia areata, keratoconjunctivitis, vernal keratoconjunctivitis, uveitis associated with Behcet's disease, keratitis, herpetic keratitis, keratoconus, corneal epithelial dystrophy, corneal leukoplakia, ocular pemphigus, Mooren's ulcer, scleritis, Graves' ophthalmopathy, falk's disease, glaucoma ... Tono-Koyanagi-Harada syndrome, sarcoidosis, pollen allergy, reversible obstructive airway disease, bronchial asthma, allergic asthma, intrinsic asthma, extrinsic asthma, dust asthma, chronic or refractory asthma, late-onset asthma and airway hyperresponsiveness, bronchitis, gastric ulcer, vascular damage caused by ischemic disease and thrombosis, ischemic bowel disease, inflammatory bowel disease, necrotizing enterocolitis, intestinal lesions associated with burns, celiac disease, proctitis, eosinophilic gastroenteritis, mastocytosis, Crohn's disease, ulcerative colitis, migraine, rhinitis, eczema, interstitial nephritis, Goodpasture's syndrome, hemolytic uremic syndrome, diabetic nephropathy, polymyositis, Guillain-Barré syndrome, Meniere's disease, polyneuritis, multiple neuritis, mononeuritis, radiculopathy, hyperthyroidism, Graves' disease, pure red cell aplasia, aplastic anemia, hypoplastic anemia, idiopathic thrombocytopenic purpura, autoimmune hemolytic anemia, agranulocytosis, pernicious anemia, megaloblastic anemia, red blood cell hypoplasia, osteoporosis, sarcoidosis, fibrotic lung, idiopathic interstitial pneumonia, dermatomyositis, vitiligo vulgaris, ichthyosis vulgaris, photoallergic sensitivity, cutaneous T-cell lymphoma, arteriosclerosis, atherosclerosis, aortitis syndrome, polyarteritis nodosa, cardiomyopathy, scleroderma, Wegener's granulomatosis, Sjögren's syndrome, steatosis, eosinophilic fasciitisfascitis), lesions of the gums, periodontium, alveolar bone, dental cementum, glomerulonephritis, male pattern baldness, age-related alopecia by preventing hair loss, age-related alopecia by providing hair germination and promoting hair development and growth, muscular dystrophy, pyoderma, Sézary syndrome, Addison's disease, organ ischemia-reperfusion injury, transplant disease, ischemic disease, endotoxin shock, pseudomembranous colitis, colitis caused by drugs or radiation, ischemic acute renal failure, chronic renal failure, pulmonary oxygen or drug-induced intoxication, lung cancer, emphysema, cataracts, iron deposition, retinitis pigmentosa, senile macular degeneration, vitreous scarring, corneal alkalinity, dermatitis erythema multiforme, linear IgA ballous dermatitis and cement dermatitis, gingivitis, periodontitis, sepsis, pancreatitis, aging, carcinogenesis, metastasis of carcinoma and altitude sickness, histamine or leukotriene-C4 release-related diseases, Behcet's disease, autoimmune hepatitis, primary biliary cirrhosis, sclerosing cholangitis, partial hepatectomy, acute liver necrosis, necrosis caused by toxins, viral hepatitis, shock, anoxia, type B viral hepatitis, non-A / non-B hepatitis, liver cirrhosis, alcoholic liver cirrhosis, liver failure, fulminant liver failure, delayed liver failure, acute exacerbation of chronic liver failure, cytomegalovirus infection, HCMV infection, AIDS, senile dementia, trauma, chronic bacterial infection, malignant tumors of lymphatic origin, acute lymphocytic leukemia, chronic lymphocytic leukemia, acute lymphocytic lymphoma, and chronic lymphocytic lymphoma.

[0151] Another embodiment of the present disclosure is realized when the disease is selected from systemic lupus erythematosus, rheumatoid arthritis, type I diabetes, inflammatory bowel disease, biliary cirrhosis, uveitis, multiple sclerosis, Crohn's disease, ulcerative colitis, bullous pemphigoid, sarcoidosis, psoriasis, autoimmune myositis, Wegener's granulomatosis, ichthyosis, Graves' ophthalmopathy, asthma, schleroderma, and Sjogren's syndrome. Another embodiment of the present disclosure is realized when the disease is selected from bone marrow rejection, organ transplant rejection, and graft-versus-host disease.

[0152] Another aspect of the present disclosure relates to uses and methods for producing such compounds, which may be suitable for imaging granzyme B.

[0153] The present disclosure includes within its scope prodrugs of the compounds of the present disclosure. Generally, such prodrugs will be functional derivatives of the compounds of the present disclosure that are readily convertible in vivo to the required compound. Thus, in the methods of treatment of the present disclosure, the terms "administration of" a compound or "administering" a compound are intended to encompass treatment of the various conditions described with a specifically disclosed compound or a compound that may not be specifically disclosed but that converts to the specified compound in vivo after administration to a patient. Conventional procedures for the selection and preparation of suitable prodrug derivatives are described, for example, in "Design of Prodrugs," ed. H. Bundgaard, Elsevier, 1985. Metabolites of these compounds include active species produced upon introduction of the compounds of the present disclosure into a biological milieu.

[0154] The compounds described herein, or pharmaceutically acceptable salts and / or solvates thereof, may be administered alone, in combination with other compounds of the present disclosure, and / or in cocktails in combination with other therapeutic agents. The choice of therapeutic agent that can be co-administered with the compounds of the present disclosure will depend in part on the condition being treated.

[0155] The compounds of the present disclosure may be administered by oral, parenteral (e.g., intramuscular, intraperitoneal, intravenous, ICV, intracisternal injection or infusion, subcutaneous injection, or implant), inhalation spray, nasal, vaginal, rectal, sublingual, buccal, or topical routes of administration, and may be formulated, alone or together, in suitable dosage unit formulations containing conventional non-toxic pharmaceutically acceptable carriers, adjuvants, and vehicles appropriate for each route of administration. In addition to the treatment of warm-blooded animals, the compounds of the present disclosure are effective for use in humans.

[0156] Pharmaceutical compositions for administering the compounds of the present disclosure may be conveniently provided in unit dosage form and may be prepared by any method well known in the art of pharmacy. All methods include the step of bringing the active ingredient into association with the carrier, which constitutes one or more accessory ingredients. In general, pharmaceutical compositions are prepared by uniformly and intimately associating the active ingredient with a liquid carrier, or a finely divided solid carrier, or both, and then, if necessary, shaping the product into the desired formulation. In pharmaceutical compositions, the active compound is contained in an amount sufficient to produce the desired effect on the process or condition of a disease. As used herein, the term "composition" is intended to encompass a product containing a specified amount of the specified ingredient, and any product resulting directly or indirectly from the combination of the specified amounts of the specified ingredients.

[0157] Pharmaceutical compositions containing the active ingredient may be in a form suitable for oral use, such as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, solutions, hard or soft capsules, or syrups or elixirs. Compositions intended for oral use may be prepared according to any method known in the art for the manufacture of pharmaceutical compositions, and such compositions may contain one or more agents selected from the group consisting of sweeteners, flavoring agents, coloring agents, and preservatives to provide pharmaceutically elegant and palatable preparations. Tablets contain the active ingredient mixed with non-toxic pharmaceutically acceptable excipients suitable for tablet manufacture. These excipients may include, for example, inert diluents such as calcium carbonate, sodium carbonate, lactose, calcium phosphate, or sodium phosphate; granulating disintegrants such as cornstarch or alginic acid; binders such as starch, gelatin, or acacia; and lubricants such as magnesium stearate, stearic acid, or talc. Tablets may be uncoated or may be coated by known techniques to delay disintegration and absorption in the gastrointestinal tract, thereby providing a sustained action over a longer period. For example, a time-delay material such as glyceryl monostearate or glyceryl distearate may be used. Tablets may also be coated by the techniques described in U.S. Patent Nos. 4,256,108, 4,166,452, and 4,265,874 to form osmotic therapeutic tablets for controlled release. Oral tablets may also be formulated for immediate release, such as fast-dissolving tablets or wafers, fast-dissolving tablets, or fast-dissolving films.

[0158] Formulations for oral use may also be presented as hard gelatin capsules in which the active ingredient is mixed with an inert solid diluent, such as calcium carbonate, calcium phosphate, or kaolin, or as soft gelatin capsules in which the active ingredient is mixed with water or an oil medium, such as peanut oil, liquid paraffin, or olive oil.

[0159] Aqueous suspensions contain the active material mixed with excipients suitable for the manufacture of aqueous suspensions.Such excipients are suspending agents, for example, sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose, sodium alginate, polyvinylpyrrolidone, tragacanth gum and gum arabic.Dispersing agents or wetting agents can be naturally occurring phosphatides, such as lecithin, or condensation products of alkylene oxides with fatty acids, for example, polyoxyethylene stearate, or condensation products of ethylene oxide with long-chain aliphatic alcohols, for example, heptadecaethyleneoxycetanol, or condensation products of ethylene oxide with fatty acids and hexitol-derived partial esters, for example, polyoxyethylene sorbitol monooleate, or condensation products of ethylene oxide with fatty acids and hexitol anhydrides, for example, polyethylene sorbitan monooleate. The aqueous suspensions may also contain one or more preservatives, for example ethyl, or n-propyl, p-hydroxybenzoate, one or more coloring agents, one or more flavoring agents, and one or more sweetening agents, such as sucrose or saccharin.

[0160] Oily suspensions can be prepared by suspending active ingredients in vegetable oils, such as peanut oil, olive oil, sesame oil or coconut oil, or mineral oils such as liquid paraffin.Oily suspensions can contain thickening agents, such as beeswax, hard paraffin or acetyl alcohol.Sweeteners and flavoring agents as mentioned above can be added to provide a palatable oral preparation.These compositions can be preserved by adding antioxidants such as ascorbic acid.

[0161] Dispersible powders and granules suitable for preparation of an aqueous suspension by adding water provide the active ingredient mixed with a dispersing or wetting agent, a suspending agent, and one or more preservatives. Suitable dispersing or wetting agents and suspending agents are exemplified by those already mentioned above. Additional excipients, such as sweeteners, flavorings, and coloring agents, may also be present.

[0162] The pharmaceutical composition of the present disclosure may be in the form of an oil-in-water emulsion.The oily phase may be vegetable oil, such as olive oil or peanut oil, or mineral oil, such as liquid paraffin, or their mixture.Suitable emulsifiers may be naturally occurring gums, such as gum arabic or gum tragacanth, naturally occurring phosphatides, such as soybeans, lecithin, and esters or partial esters derived from fatty acids and hexitol anhydrides, such as sorbitan monooleate, and the condensation products of the partial esters with ethylene oxide, such as polyoxyethylene sorbitan monooleate.Emulsions may also contain sweeteners and flavoring agents.

[0163] Syrups and elixirs may be formulated with sweetening agents, for example glycerol, propylene glycol, sorbitol or sucrose, and such formulations may also contain a demulcent, a preservative and flavoring and coloring agents.

[0164] The pharmaceutical compositions may be in the form of a sterile injectable aqueous or oleaginous suspension. This suspension may be formulated according to known techniques using suitable dispersing or wetting agents and suspending agents as described above. Sterile injectable preparations may also be sterile injectable solutions or suspensions in non-toxic parenterally acceptable diluents or solvents, such as solutions in 1,3-butanediol. Among the acceptable vehicles and solvents that may be used are water, Ringer's solution, and isotonic sodium chloride solution. Additionally, sterile fixed oils are conventionally used as solvents or suspending media. For this purpose, any non-irritating fixed oil, including synthetic mono- or diglycerides, may be used. Additionally, fatty acids, such as oleic acid, are used in the preparation of injectables.

[0165] The compound of the present disclosure can also be administered in the form of suppositories for rectal administration of drugs.These compositions can be prepared by mixing the drug with suitable non-irritating excipients that are solid at room temperature but liquid at rectal temperature, and therefore melt in the rectum to release the drug.Such materials are cocoa butter and polyethylene glycol.

[0166] For topical use, creams, ointments, jellies, solutions or suspensions, etc., containing the compounds of the present disclosure are employed. Likewise, transdermal patches may also be used for topical administration.

[0167] The pharmaceutical compositions and methods of the present disclosure may further comprise other therapeutically active compounds as described herein that are typically applied in the treatment of the above-mentioned pathological conditions.

[0168] In treating, preventing, controlling, ameliorating, or reducing the risk of a condition disclosed herein, the phrase "therapeutically effective amount" refers to that amount of an active compound or pharmaceutical agent that elicits the desired biological or medical response in a tissue, system, animal, individual, or human. In some embodiments, the dosage of the compound or a pharmaceutically acceptable salt thereof administered to a subject or individual is about 1 μg to about 2 g, about 1 μg to about 1000 mg, about 1 μg to about 500 mg, about 1 μg to about 100 mg, about 1 μg to about 50 mg, about 1 μg to about 1 mg, about 1 μg to about 500 μg, about 1 μg to about 100 μg, about 1 μg to about 10 μg, about 10 μg to about 2 g, etc. The compound may be administered on a regimen of 1 to 4 times daily, or may be administered once or twice daily.

[0169] Another embodiment of the present disclosure is realized where a compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt thereof, can be administered in combination with one or more of the additional therapeutic agents provided herein. A subembodiment of this aspect of the present disclosure is realized where the additional therapeutic agent includes, but is not limited to, an anti-inflammatory agent, a steroid, an immunotherapeutic agent, a chemotherapeutic agent, and a therapeutic antibody.

[0170] Another embodiment of the present disclosure is realized when administration of a therapeutic agent induces an immune response in a cell or tissue sample or subject. A subembodiment of this aspect of the present disclosure is realized when the therapeutic agent is a compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt thereof. A subembodiment of this aspect of the present disclosure is realized when the therapeutic agent is a compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt thereof, comprising a radioisotope (e.g., a therapeutic radioisotope). Another subembodiment of this aspect of the present disclosure is realized when the therapeutic agent is a compound of Formula I, Formula Ia, or a pharmaceutically acceptable salt thereof, comprising a toxic radioisotope. Another subembodiment of this aspect of the present disclosure is realized when the toxic radioisotope is an alpha emitter ( 211 At, 212 Pb, 212 Bi, 213 Bi, 225 Ac, 227 Th) and beta emitters (e.g., 90 Y, 131 I and 177 Lu, 161 This is realized when the eigenvalue is selected from the following:

[0171] However, it will be understood that the specific dose level and frequency of administration for any particular patient may vary and will depend upon a variety of factors including the activity of the particular compound used, the metabolic stability and length of action of that compound, age, body weight, general health, sex, diet, mode and time of administration, rate of excretion, drug combination, severity of the particular condition, and the host being treated.

[0172] Methods for preparing compounds of this disclosure are illustrated in the following schemes and examples. Starting materials are made according to procedures known in the art or as illustrated herein.

[0173] It will be understood by those skilled in the art that the described methods are not the exclusive means by which the compounds provided herein can be synthesized, and that a wide repertoire of synthetic organic reactions is available for potential use in synthesizing the compounds provided herein. The compounds of the present disclosure can be prepared according to the following exemplary schemes and specific examples, or modifications thereof, using readily available starting materials, reagents, and conventional synthetic procedures. It is also possible to use variants that are known per se to those skilled in the art but are not specifically mentioned. The general procedures for producing the compounds claimed in this disclosure can be easily understood by those skilled in the art from viewing the following schemes and descriptions. Abbreviations used in the experiments may include, but are not limited to, the following: [Table 2] TIFF2026500238000040.tif57166

[0174] Table A lists unnatural amino acids with their abbreviations and structures. [Table 3] TIFF2026500238000042.tif222165 TIFF2026500238000043.tif232165 TIFF2026500238000044.tif236165 TIFF2026500238000045.tif113166 [Table 4] TIFF2026500238000047.tif136165.

[0175] General Peptide Synthesis Procedure All reagents and solvents were purchased from commercial sources and used as received unless otherwise noted. Reaction progress and synthesis intermediate analysis were assessed by LCMS (UV detection with ESI, APCI, or other mass detection) using MeCN / water gradients with either TFA, formic acid, or NH4HCO3 modifiers, where applicable. Silica gel and reversed-phase flash column chromatography were performed using commercially available pre-packed columns. Reverse-phase preparative HPLC purification was performed on a preparative HPLC instrument with UV and MS detection using MeCN / water gradients with either TFA, formic acid, or NH4OH modifiers. All K presented in the tables were used unless otherwise noted. D Data refers to surface plasmon resonance assays described in the biological assays section.

[0176] Peptides were synthesized using standard solid-phase synthesis using Fmoc / tert-Bu chemistry as exemplified in Chan, W.C.; White, P.D. "Fmoc Solid-Phase Synthesis: a Practical Approach," Oxford University Press, Oxford, 2000; Steward, J.; Young, J. "Solid Phase Peptide Synthesis," Pierce Chemical Company, Rockford, 1984; N.L. Benoiton, "Chemistry of Peptide Synthesis," CRC Press, New York, 2006; and Lloyd-Williams, P.; Albericio, F. "Chemical Approaches to the Synthesis of Peptides and Proteins," CRC Press, New York, 1997.

[0177] Unless otherwise indicated, peptide synthesis was completed using Protocol A or B outlined below. The following protected natural amino acids were used: Fmoc-Ala-OH, Fmoc-Arg(Pbf)-OH, Fmoc-Asn(Trt)-OH, Fmoc-Cys(Trt)-OH, Fmoc-Gln(Trt)-OH, Fmoc-Gly-OH, Fmoc-Glu(OtBu)-OH, Fmoc-His(Trt)-OH, Fmoc-Ile-OH, Fmoc-Leu-OH, Fmoc-Lys(Boc)-OH, Fmoc-Phe-OH, Fmoc-Pro-OH, Fmoc-Ser(t-Bu)-OH, Fmoc-Thr(t-Bu)-OH, Fmoc-Trp-OH, Fmoc-Tyr(t-Bu)-OH, and Fmoc-Val-OH. All unnatural amino acids in the sequence contained Fmoc-α-N-protected amines and the relevant side chains were protected with Boc- or tBu-protecting groups.

[0178] Protocol A: Step 1 - Synthesis of Linear Peptides: N α -Fmoc-protected Rink Amide MBHA resin (0.7 mmol, 1.0 g) was swelled in DCM (15 mL) in a fritted glass sintered vessel for 1.0 hour. The mixture was filtered, and 20% piperidine in DMF (15 mL) was added to the resin. The mixture was shaken on an orbital shaker at room temperature for 5 minutes. The solution was drained, and the resin was again treated with 20% piperidine / DMF solution on an orbital shaker for 20 minutes. The mixture was filtered, and the peptidyl resin was then washed with DMF (5 x 15 mL), then DCM (5 x 15 mL), then DMF (5 x 15 mL). A Kaiser test was performed on the beads to confirm the presence of free amines. The resin was then washed with 20% piperidine / DMF for 20 minutes. 1) Fmoc-protected amino acid (3.5 mmol), DMF (10 mL), N-methylmorpholine (7 mmol), and HBTU (3.5 mmol) or 2) Fmoc-protected amino acid (3.5 mmol), DMF (10 mL), HBTU (7 mmol), and HOBt (3.5 mmol), and N,N-diisopropylethylamine (7 mmol) or 3) Fmoc-protected amino acid (3.5 mmol), DMF (10 mL), N,N'-diisopropylcarbodiimide (7 mmol), and HOBt (3.5 mmol) The mixture was treated with a solution containing

[0179] The resulting suspension was maintained on an orbital shaker at room temperature for 2 hours. After the Kaiser test indicated the coupling reaction was complete, the mixture was filtered, and the peptidyl resin was washed with DMF (5 × 15 mL), DCM (5 × 15 mL), and DMF (5 × 15 mL). The subsequent Fmoc deprotection and amino acid coupling steps were repeated as described above. N-terminal chloroacetylation was performed using 10% chloroacetic anhydride / DCM in the presence of 6 equivalents (relative to resin substitution) of N,N-diisopropylethylamine. After synthesis, the resin was washed with DMF and DCM (3 × 15 mL), followed by methanol (20 mL) and diethyl ether (2 × 20 mL). Finally, the resin was dried overnight under vacuum.

[0180] Step 2—Cleavage and Deprotection: Cleavage of the peptide from the solid support was achieved by treating the peptide-resin (2.5 g) with a reagent cocktail (95% TFA / 2.5% TIS (triisopropylsilane) / 2.5% water / 2.5% DODT, 10 mL / g peptidyl-resin) at room temperature for 3 hours. The cleavage mixture was collected by filtration, and the resin was washed with TFA. For peptide precipitation, 5 mL of the cleavage mixture containing the peptide was precipitated into 45 mL of cold (0° C.) anhydrous ether. The precipitated peptide was centrifuged (3800 rpm), and the supernatant ether was removed. Fresh ether was added to the peptide and recentrifuged. This process was repeated three times. The precipitated peptide was then lyophilized overnight under high vacuum to obtain a linear crude peptide solid.

[0181] Step 3 - Peptide cyclization: The crude linear peptide was dissolved in DMSO to a concentration of 5 mM, and the pH was adjusted to 8-9 by adding triethylamine or N,N'-diisopropylethylamine (monitored with water-moistened pH paper). The reaction was stirred at room temperature for 16 hours and then neutralized to a pH of approximately 7 by adding TFA. The reaction solution was then directly taken on for purification.

[0182] Step 4—Peptide Purification: The crude cyclized material was purified using preparative HPLC using a Phenomenex Luna C18 5u 100A 250×21.2 mm column and a linear gradient of acetonitrile in water, both buffered with 0.1% TFA. The HPLC fractions containing the pure peptide product were pooled and lyophilized.

[0183] Protocol B: Step 1 - Linear Peptide Synthesis: The peptide sequence was assembled by solid-phase synthesis using a Microwave Liberty Blue synthesizer (CEM, Matthews, NC). Synthesis was initiated using 100 μmol of Novabiochem Rink Amide AM resin LL, 100-200 mesh, 0.29 mmol / g (CEM). Each amino acid was coupled in a 9-fold excess as a 0.2 M solution in DMF, which was activated using a 9-fold excess of 0.5 M diisopropylcarbodiimide (DIC) and 1 M Oxyma in DMF. The Fmoc deprotection step was performed using 20% ​​pyrrolidine in DMF containing 0.1 M HOBT at 90 °C for 60 s. Single and double couplings were performed at 90 °C with a 5 min coupling time, except for Fmoc-His(Trt)-OH, which was coupled at 50 °C. Acylation of secondary amino acids in the sequence was performed by double coupling. Aspartic acid was coupled as Fmoc-Asp(OBno)-OH. For the subsequent coupling cycles, the Fmoc protecting group was deprotected at room temperature. N-terminal chloroacetylation was performed by treating the resin twice with chloroacetic anhydride (10 equivalents) in NMP for 15 minutes at ambient temperature.

[0184] Step 2—Cleavage and Deprotection: Cleavage of the peptide from the solid support was achieved by treating the peptide-resin with a reagent cocktail (87.5% TFA / 5% water / 2.5% TIS (triisopropylsilane) / 2.5% phenol, 15 mL / g of peptidyl-resin) for 1–2 h at room temperature or (60% TFA / 5% TIS (triisopropylsilane) / 35% DCM, 5 mL / g of peptidyl-resin) for 45 min at room temperature. The cleavage mixture was collected by filtration, and the resin was washed with TFA. The peptide was precipitated in cold methyl tert-butyl ether, centrifuged (3800 rpm), and the supernatant ether was removed. The peptide pellet was resuspended in cold methyl tert-butyl ether and centrifuged twice more. The precipitated peptide was then lyophilized overnight under high vacuum to yield the crude linear peptide.

[0185] Step 3 - Peptide cyclization: The crude linear peptide was dissolved in MeCN / HO (50:50) at a concentration of 1 mg / mL, and DIPEA (10 equiv.) was added. The mixture was stirred at ambient temperature for 20 min. The pH was adjusted to approximately 5 with TFA, and the mixture was lyophilized.

[0186] Step 4—Peptide Purification: The crude cyclized peptide was purified using preparative HPLC using a C4 reverse-phase column (Reprosil Gold, 120 Å, 5 μm) and a linear gradient of acetonitrile in water, both buffered with 0.1% TFA. The HPLC fractions containing the pure peptide product were pooled and lyophilized.

[0187] Examples of cyclic peptide sequences and structures The macrocyclic peptide contains a thioether bond between the cysteine ​​side chain and the N-terminal -CHC(O) bond. Representative cyclic peptides of the present disclosure are described by a period-delimited sequence of amino acids and a structural representation, as shown in Table 1. In the period-delimited sequence, substitutions on the amino acid side chain are indicated in parentheses immediately following the substituted amino acid (see, for example, compounds 2a, 3a, 4a, 7a, 8a, 9a, 10a, 11a, and 16a in Table 1 below). Reference macrocyclic compound 1 in Table 1 can be used as a reference to determine the structure of compounds listed only by sequence as compounds in Tables 2, 3, 5, 6, 7, and 7a. An "*" indicates the position of the cyclization residue. [Table 5] TIFF2026500238000049.tif219165 TIFF2026500238000050.tif236165 TIFF2026500238000051.tif239165 TIFF2026500238000052.tif217165 TIFF2026500238000053.tif226165 TIFF2026500238000054.tif215166 TIFF2026500238000055.tif220165 TIFF2026500238000056.tif196165 TIFF2026500238000057.tif85166

[0188] Example 1a Preparation of Compound 1a (SEQ ID NO: 1) [ka]

[0189] Step A: Synthesis of 1" Peptides were synthesized using General Protocol B. The peptide sequence was assembled by solid-phase synthesis on a Microwave Liberty Blue synthesizer (CEM, Matthews, NC). Synthesis began with 250 μmol of Rink Amide MBHA resin (0.2 mmol, 0.35 g). Each amino acid was coupled in a 4-fold excess as a 0.2 M solution in DMF, which was activated with a 4-fold excess of 1.0 M diisopropylcarbodiimide (DIC) and 1 M Oxyma in DMF. The Fmoc deprotection step was carried out using 20% ​​piperidine in DMF containing 0.1 M HOBT at 90°C for 60 seconds. Single and double couplings were performed at 90°C with a 2-minute coupling time. Acylation of secondary amino acids in the sequence was carried out by double coupling. N-terminal chloroacetylation was carried out by treating the resin twice with chloroacetic anhydride (10 equivalents) in NMP for 15 minutes at ambient temperature. After solid-phase synthesis, the peptide was cleaved from the solid support by treating the peptide-resin with a reagent cocktail (92.5% TFA / 2.5% water / 2.5% TIS (triisopropylsilane) / 2.5% DODT, 10 mL / g of peptidyl-resin) for 3 hours at room temperature. The cleavage mixture was collected by filtration, and the resin was washed with TFA. The peptide was precipitated in cold methyl tert-butyl ether, centrifuged (3800 rpm), and the supernatant ether was removed. The peptide pellet was resuspended in cold methyl tert-butyl ether and centrifuged two more times. The precipitated peptide was then lyophilized overnight under high vacuum to give 1". MS: m / z = 911.7 [M + 2H] / 2 [ka]

[0190] Compound 1" (1.0 g, 0.55 mmol) was dissolved in DMSO to a concentration of 5 mM, and the pH was adjusted to 8-9 with DIPEA (monitored with water-moistened pH paper). The mixture was stirred at ambient temperature for 16 h. The pH was adjusted to approximately 7 with TFA. The resulting material was purified using preparative HPLC using a C18 reverse-phase column (Phenomenex Luna Gold, 100 Å, 5 μm, 21 mm × 250 mm) and 5-40% acetonitrile in water, both buffered with 0.1% TFA. HPLC fractions containing the pure peptide product were pooled and lyophilized to give 1a. MS: m / z = 893.5 [M + 2H] / 2 [Example 2″] (SEQ ID NO: 58) [ka]

[0191] Step A: Synthesis of 2" Peptides were synthesized using General Protocol B. Peptide sequences were assembled by solid-phase synthesis on a Microwave Liberty Blue synthesizer (CEM, Matthews, NC). Synthesis was initiated with 250 μmol of Novabiochem Rink Amide AM resin LL, 100-200 mesh, 0.29 mmol / g (CEM).

[0192] Each amino acid was coupled in a 9-fold excess as a 0.2 M solution in DMF, which was activated with 0.5 M diisopropylcarbodiimide (DIC) and 1 M Oxyma in a 9-fold excess of DMF. The Fmoc deprotection step was carried out using 20% ​​pyrrolidine in DMF containing 0.1 M HOBT at 90 °C for 60 seconds. Single and double couplings were performed at 90 °C with a 5-minute coupling time. Acylation of secondary amino acids in the sequence was carried out by double coupling. Aspartic acid was coupled as Fmoc-Asp(OBno)-OH. For the subsequent coupling cycle of Asp-(OBno)-OH, the Fmoc protecting group was deprotected at room temperature. N-terminal chloroacetylation was carried out by treating the resin twice with chloroacetic anhydride (10 equivalents) in NMP for 15 minutes at ambient temperature. After solid-phase synthesis, the peptide was cleaved from the solid support by treating the peptide-resin with a reagent cocktail (87.5% TFA / 5% water / 2.5% TIS (triisopropylsilane) / 2.5% phenol, 15 mL / g of peptidyl-resin) for 1 hour at room temperature. The cleavage mixture was collected by filtration, and the resin was washed with TFA. The peptide was precipitated in cold methyl tert-butyl ether, centrifuged (3800 rpm), and the supernatant ether was removed. The peptide pellet was resuspended in cold methyl tert-butyl ether and centrifuged two more times. The precipitated peptide was then lyophilized overnight under high vacuum to give 2". MS: m / z = 1003.6 [M + 2H] / 2 Step B: Synthesis of 2b (SEQ ID NOS: 58 and 135, respectively, in order of appearance) [ka]

[0193] Compound 2" (130 mg, 0.063 mmol) was dissolved in MeCN / HO (50:50) at a concentration of 1 mg / mL, and DIPEA (10 equiv.) was added. The mixture was stirred at ambient temperature for 20 min. The pH was adjusted to approximately 5 with TFA, and the mixture was lyophilized. The resulting material was purified using preparative HPLC using a C4 reverse-phase column (Reprosil Gold, 120 Å, 5 μm) and 15–35% acetonitrile in water, both buffered with 0.1% TFA. The HPLC fractions containing the pure peptide product were pooled and lyophilized to give 2b. MS: m / z = 985.6 [M+2H] / 2 Step C: Synthesis of 2a Preparation of Compound 2a (SEQ ID NOs: 135 and 2, respectively, in order of appearance) [ka]

[0194] A solution of commercially available 2,2',2"-(2-(4-isothiocyanatobenzyl)-1,4,7-triazonane-1,4,7-triyl)triacetic acid (40 mg, 0.039 mmol) in DMSO (1.5 ml) and DIPEA (14 uL, 0.78 mmol) was added to a mixture of peptide 2b (60 mg, 0.030 mmol) and DIPEA (14 uL, 0.78 mmol) in DMSO (1.5 ml). After stirring at ambient temperature for 4 h, the pH was adjusted to approximately 5 with TFA. The crude material was purified by column chromatography on a C4 reversed-phase column (Reprosil Gold, 120 Å, 5 μm) or Phenomenex Luna C18 5u 100A column, both buffered with 0.1% TFA. Purification was performed using preparative HPLC using a 250 x 21.2 mm column and a linear gradient of acetonitrile in water. The HPLC fractions containing the pure peptide product were pooled and lyophilized. MS: m / z = 1210.8 [M + 2H] / 2.

[0195] The following compounds 12a and 13a were made using the methods described in Example 2, substituting the appropriate reactants and / or reagents. [Table 6]

[0196] Example 3a Preparation of Compound 3a (SEQ ID NO: 3) [ka]

[0197] A solution of commercially available 2,2'-(((1R,2R)-2-((carboxymethyl)(4-(2-oxo-2-(2,3,5,6-tetrafluorophenoxy)ethyl)benzyl)amino)cyclohexyl)azanediyl)diacetic acid (75.7 mg, 0.13 mmol) in DMSO (0.5 mL) was added to a solution of peptide 2b (0.212 g, 0.108 mmol) in DMSO (2.5 mL) and DIPEA (188 μL, 1.1 mmol). After stirring at room temperature for 15 min, the pH was adjusted to approximately 5 with TFA. The crude material was purified using preparative HPLC using a C4 reverse-phase column (Reprosil Gold, 120 Å, 5 μm) or a Phenomenex Luna C18 5 μL 100A 250 × 21.2 mm column, both buffered with 0.1% TFA, and a linear gradient of acetonitrile in water. HPLC fractions containing the pure peptide product were pooled and lyophilized. MS: m / z=1195.0 [M+2H] / 2.

[0198] Example 4a Preparation of Compound 4a (SEQ ID NO: 4) [ka]

[0199] To a solution of propionic acid (1.3 mg, 0.017 mmol) in DMF (0.5 mL) was added HATU (5.0 mg, 0.013 mmol) and DIPEA (4.5 μL, 0.026 mmol). After 5 min, the mixture was added to a solution of peptide 2b (20 mg, 0.010 mmol) in DMF (2.5 mL) and stirred at ambient temperature for 15 min. The pH was adjusted to approximately 5 with TFA. The crude material was purified using preparative HPLC using a C4 reverse-phase column (Reprosil Gold, 120 Å, 5 μm) or a Phenomenex Luna C18 5u 100A 250 × 21.2 mm column, both buffered with 0.1% TFA, and a linear gradient of acetonitrile in water. HPLC fractions containing the pure peptide product were pooled and lyophilized. MS: m / z = 1013.8 [M + 2H] / 2.

[0200] The following compounds 14a-16a were made using the method described in Example 4a, substituting the appropriate reactants and / or reagents. [Table 7]

[0201] [Example 5a] (SEQ ID NOs: 120 and 121, respectively, in order of appearance) [ka]

[0202] A solution of commercially available bis(2,5-dioxopyrrolidin-1-yl) 3,3'-(ethane-1,2-diylbis(oxy))dipropionate (5.2 mg, 0.013 mmol) in DMSO (0.5 mL) was added to a solution of 2b (55.4 mg, 0.028 mmol) and DIPEA (10 equiv.) in DMSO (2.5 mL). After stirring at ambient temperature for 15 min, the pH was adjusted to approximately 5 with TFA. The crude material was purified using preparative HPLC on a C4 reverse-phase column (Reprosil Gold, 120 Å, 5 μm) or a Phenomenex Luna C18 5u 100A 250 × 21.2 mm column, both buffered with 0.1% TFA, using a linear gradient of acetonitrile in water. HPLC fractions containing the pure peptide product were pooled and lyophilized. MS: m / z = 1370.6 [M+3H] / 3.

[0203] The following compounds 17a-18a were made using the method described in Example 5a, substituting the appropriate reactants and / or reagents. [Table 8] TIFF2026500238000069.tif243166

[0204] [Example 6] [ka]

[0205] Step A: Synthesis of 6b To a solution of commercially available NH-bis(PEG4-acid) hydrochloride (0.065 g, 0.118 mmol) in DMF (0.6 mL) and triethylamine (0.025 mL, 0.177 mmol), 2,5-dioxopyrrolidin-1-yl 3-((tert-butoxycarbonyl)amino)propanoate (0.051 g, 0.177 mmol) was added, and the mixture was stirred at ambient temperature for 1 h. The mixture was diluted with 50% MeCN / water and lyophilized to give 6b.

[0206] Step B: Synthesis of 6 To a solution of 6b in DMF (0.6 mL) and triethylamine (0.147 mL, 1.065 mmol) was added N,N-disuccinimidyl carbonate (0.273 g, 1.065 mmol). The mixture was stirred for 4 h, then diluted with 50% MeCN / water and lyophilized. The resulting material was dissolved in DMSO (0.5 mL) and purified on a Biotage Isolera automated reverse-phase flash column chromatography system (linear gradient: 5% to 50% acetonitrile in HO, 0.1% TFA, over 12 CV, flow rate: 19 mL / min, column: Luknova SuperSep C18 (5.5 g) cartridge) to give 6c (30 mg). MS: m / z = 879.7 [M+H].

[0207] Example 7a Preparation of Compound 7a (SEQ ID NOs: 122 and 123, respectively, in order of appearance) [ka]

[0208] A solution of intermediate 6 (0.032 g, 0.037 mmol) in DMSO (0.5 mL) was added to a solution of 2b (0.16 g, 0.081 mmol) and DIPEA (10 equiv.) in DMSO (2.5 mL). After stirring at ambient temperature for 15 minutes, the pH was adjusted to approximately 5 with TFA. The crude material was purified using preparative HPLC using a Delta-Pak double cartridge C4, 25 × 100 mm, 15 um, 300A, both buffered with 0.1% TFA, and a linear gradient of acetonitrile in water. HPLC fractions containing the pure peptide product were pooled and lyophilized. MS: m / z = 1530.0 [M + 3H] / 3.

[0209] Example 8a Preparation of Compound 8a (SEQ ID NOs: 124 and 125, respectively, in order of appearance) [ka]

[0210] Compound 7a (60 mg, 0.013 mmol) was treated with 10% water in TFA (3 mL) for 30 min and then lyophilized from 50% MeCN / water. To the resulting solid was added a solution of 2,2',2"-(2-(4-isothiocyanatobenzyl)-1,4,7-triazonane-1,4,7-triyl)triacetic acid (8.88 mg, 0.020 mmol) in DMSO (3.0 ml) and DIPEA (6.85 uL, 0.039 mmol). The reaction was stirred at ambient temperature for 4 hours, then the pH was adjusted to 5 with TFA. The crude material was purified using preparative HPLC using a C4 reverse-phase column (Reprosil Gold, 120 Å, 5 μm) or a Phenomenex Luna C18 5u 100A 250 × 21.2 mm column and a linear gradient of acetonitrile in water, both buffered with 0.1% TFA. HPLC fractions containing the pure peptide product were pooled and lyophilized. MS: m / z = 1646.7 [M + 3H] / 3.

[0211] [Example 9] Preparation of Compound 9a (SEQ ID NO: 11) [ka]

[0212] A solution of commercially available Alexa Fluor 647 NHS ester (1.8 equiv.) in DMSO (0.5 mL) was added to a mixture of purified cyclic peptide 2b (0.025 mmol) and DIPEA (10 equiv.) in DMSO (2.5 mL). After stirring at ambient temperature for 15 min, the pH was adjusted to approximately 5 with TFA. The crude material was purified using preparative HPLC on a C4 reverse-phase column (Reprosil Gold, 120 Å, 5 μm) or a Phenomenex Luna C18 5u 100A 250 × 21.2 mm column, both buffered with 0.1% TFA, using a linear gradient of acetonitrile in water. HPLC fractions containing the peptide product were pooled and lyophilized. MS: m / z = 1410.4 [M + 2H] / 2.

[0213] Example 10a Preparation of Compound 10a (SEQ ID NOs: 2 and 12, respectively, in order of appearance) [ka]

[0214] [ 68 [Ga]GaCl was eluted from an IGG100 gallium-68 generator (Eckert & Ziegler) with 5 ml of 0.1 M HCl and passed through a Phenomenex Strata-XC cartridge. [Ga]GaCl was then eluted with 0.5 ml of 98% acetone (in 0.02 M HCl). 68 For labeling, 10A (50 μg, 20 nmol) was dissolved in 500 μL of HEPES buffer (0.1 M, pH 4.1) and 0.26 GBq of [Ga]GaCl was eluted from the cartridge. 68 Ga]GaCl3 were mixed and incubated for 15 min at 70° C. Radiochemical purity was monitored by radio-iTLC or radio-HPLC on an Onyx Monolithic C18 column (100 mm × 3 mm) eluted with a gradient from 5% aqueous acetonitrile (0.1% HC02H2) to 90% acetonitrile over 7 min at a flow rate of 1.5 ml / min.

[0215] The following compounds 19a-20a were made using the method described in Example 10a, substituting the appropriate reactants and / or reagents. [Table 9]

[0216] Example 11a Preparation of Compound 11a (SEQ ID NOs: 2 and 126, respectively, in order of appearance) [ka]

[0217] 0.4 mL of KHCO3 (0.4 M) was used to transfer the solution from a commercial QMA cartridge to a vial containing 125 μL of sodium acetate (0.1 M, pH 4), 25 μL of AlCl3 (2 mM in 0.1 M sodium acetate) and 10 μL of acetic acid. 18 F (7.4 GBq) was released. The mixture was kept at room temperature for 2 min. Compound 11a (0.4 mg, 0.16 μmol) dissolved in a mixture of 75 μL of 0.1 M sodium acetate (pH 4) and 75 μL of acetonitrile was then added to the above mixture. After heating at 100 °C for 15 min, 700 μL of deionized (DI) water was added to the reaction solution, which was then loaded onto an HPLC column for purification. For this purpose, a Phenomenex Gemini C18 column (10 mm i.d. × 150 mm) was eluted with a gradient consisting of 0.1% TFA in both water and acetonitrile at a flow rate of 5 mL / min. The proportion of acetonitrile was linearly increased from 20% to 70% over 15 min. The solvent in the pooled HPLC fractions containing the radiolabeled peptide was partially concentrated under reduced pressure. Radiochemical purity was monitored by radio-iTLC or radio-HPLC on an Onyx Monolithic C18 column (100 mm × 3 mm) eluted with a gradient from 5% aqueous acetonitrile (0.1% HC02H2) to 90% acetonitrile over 7 min at a flow rate of 1.5 ml / min.

[0218] The following compounds 21a-22a were made using the method described in Example 11a, substituting the appropriate reactants and / or reagents. [Table 10]

[0219] Cyclic peptides of the present disclosure were prepared according to Protocol A or Protocol B outlined and / or exemplified in this disclosure.

[0220] The amino acid linear sequences of the parent peptides 1-5 and 7-87 are listed in Table 7. [Table 11] TIFF2026500238000079.tif237165 TIFF2026500238000080.tif239166 TIFF2026500238000081.tif51166

[0221] The amino acid sequences of macrocyclic peptides 1a-5a and 7a-87a are listed in Table 7a. An asterisk (*) indicates where cyclization (i.e., the point of attachment of amino acids into the molecule) occurs: between the amino acid leucine (L), phenylalanine (F), or tyrosine (Y) and the sulfur on the amino acid cysteine ​​(C). See, for example, compound 1a shown below. [ka] [Table 12] TIFF2026500238000084.tif240165 TIFF2026500238000085.tif241166 TIFF2026500238000086.tif88166

[0222] Biological assays: Surface Plasmon Resonance (SPR) The kinetics of test peptide binding to human granzyme B (hGranzyme B) was determined by SPR. All experiments were performed at 25°C using a Biacore 8k+ instrument (Cytiva, Marlborough, MA). The data were fitted with a 1:1 binding model using Biacore 8k+Insight Evaluation Software version 3.0 to determine the association rate constant, k a (M -1 s -1 , where "M" equals moles and "s" equals seconds) and the dissociation rate constant k d (s -1 ) was determined. These rate constants were used to calculate the equilibrium dissociation constant, K D (M)=kd / k a was calculated.

[0223] To measure the binding kinetics of peptides of interest, surfaces were prepared using a series S streptavidin (SA) chip capture method using 1x HBS-EP+ running buffer (Cytiva). Granzyme B was diluted to 10 μg / mL in 1x HBS-EP+ and flowed over spot 2 of each flow cell on the chip at a flow rate of 10 μl / min for 20 seconds, achieving a capture level of approximately 1,100 RU. Unmodified spot 1 of each flow cell was used as a matrix-binding control or reference surface. Binding kinetics were measured using single-cycle kinetics at 25 °C by injecting five concentrations using 3-fold serial dilutions of the test peptide from 0.01 μM to 1 μM. Compound dilutions were performed manually in running buffer containing 1x HBS-EP+ (Cytiva) and a final concentration of 2% DMSO. Interaction analysis for granzyme B was performed using an association time of 120 seconds and a dissociation time of 900 seconds. All experiments were performed at a flow rate of 50 μL / min with a data collection rate of 10 Hz.

[0224] Surface plasmon resonance data analysis The peptide dilution series included two zero concentration points (no compound), which were averaged and subtracted from the peptide concentration injections as the DMSO control response. Binding data from the reference surface (matrix-bound control) was subtracted from the binding data of the hGranzyme B surface on the chip to determine specific binding. The association phase data were then solvent-corrected to account for signal variations across experiments due to slight DMSO concentration differences. The corrected response data were analyzed using Biacore 8k+ Evaluation Software version 3.0 according to the manufacturer. Table 8 shows the SPR data for representative compounds of the present disclosure. [Table 13] TIFF2026500238000088.tif239165 TIFF2026500238000089.tif121165

[0225] Granzyme B biochemical inhibition assay Compounds to be tested as inhibitors were manually prepared in two-fold serial dilutions in DMSO ranging from 200 μM to 0.39 μM in Labcyte Echo-qualified low-dead-volume 384-well plates. 125 nL of each compound was then dispensed from the 384-well plate into a PerkinElmer Proxiplate-384 Plus F assay plate using the ECHO acoustic liquid handler. For the positive inhibitor control, (S)-3-((3S,6S)-3-((2S,3S)-2-acetamido-3-methylpentanamido)-4-oxo-1,2,3,4,6,7-hexahydroazepino[3,2,1-hi]indole-6-carboxamido)-4-oxobutanoic acid (see CA Willoughby et al. / Bioorg.Med.Chem.Lett. 12 (2002) 2197-2200 2199) was used at a 2 uM assay concentration. DMSO was used as a no inhibitor control. Each compound was run in duplicate.

[0226] Once the compounds were dispensed into the plate, the enzyme and substrate mixtures were prepared. The enzyme mixture was prepared by combining 300 μL of granzyme B (final concentration 8 nM) with 10.8 mL of assay buffer (from the Sigma kit). The substrate mixture was prepared by combining 500 μL of substrate (from the Sigma kit) with 3 mL of assay buffer (from the Sigma kit). The enzyme mixture was loaded into a BioRaptr and 9.25 μL was added to each well of the compound-containing assay plate. The plate was incubated at room temperature for 30 minutes under a metal gasket cover. Subsequently, 3.125 μL of substrate mixture was added to each well using the BioRaptr to reach a final assay volume of 12.5 μL. The plate was monitored in fluorescence mode for 30 minutes using a Pherastar FS, taking one reading per minute. The excitation wavelength was 400 nm and the emission wavelength was 510 nm.

[0227] Data analysis was first performed in Excel, and the slope of the measured fluorescence from each sample was determined using BMG Labtech's MARS software. These percentages were then multiplied by 10,000 for visualization purposes. Percentages multiplied by 10,000 were then resectively normalized to 0% and 100% activity based on the average values ​​of samples with or without 2 μM inhibitor (S)-3-((3S,6S)-3-((2S,3S)-2-acetamido-3-methylpentanamido)-4-oxo-1,2,3,4,6,7-hexahydroazepino[3,2,1-hi]indole-6-carboxamide)-4-oxobutanoic acid, respectively. All samples were then normalized to these values ​​using the formula = {100-[(sample-100% inhib) / 0% inhib]} x 100 to achieve a normalized percent inhibition. The normalized percent inhibition versus inhibitor concentration was then plotted in Graphpad Prism version 9.0.0, and a variable slope (four parameter) fit was calculated for each inhibitor. Because the concentration of granzyme B in the assay was 8 nM, the IC was calculated by Prism to be less than 8 nM. 50 Values ​​(see Table 9 for inhibition data for representative compounds of the present disclosure) were reported as <8 nM. [Table 14]

[0228] To determine granzyme B-positive signals in PBMC and NOG mice, immunohistochemistry (IHC) analysis of lung, liver, kidney, and spleen tissues was performed. For IHC analysis, tissues were collected after in vivo PET imaging. Mice were euthanized by CO2 and cervical dislocation according to IACUC regulations. Lung, liver, kidney, and spleen tissues were then collected and immersed in 45 mL of 10% normal-buffered formalin (HistoPrep, Fisherbrand) for approximately 20–24 hours with gentle agitation at room temperature. The tissue samples were then immersed in 45 mL of 70% EtOH for processing.

[0229] Formalin-fixed tissue samples were then processed at 37°C using a Shandon Excelsior ES. Briefly, tissue samples were subjected to 70% ethanol (EtOH, HistoPrep, Fisherbrand) overnight, followed by increasing grades of EtOH (2 times 70% EtOH for 30 minutes; 1 time 80% EtOH for 30 minutes; 1 time 95% EtOH for 30 minutes; 3 times 100% EtOH for 30 minutes). Tissue samples were then subjected to xylene (HistoPrep, Fisherbrand) for 30 minutes three times, followed by paraffin (Paraplast Plus, Sigma) for 30 minutes three times. Tissue samples were then immediately embedded in paraffin using a Shandon Histocentre 2 and then stored at 4°C. Embedded tissue samples were sectioned at 5 μm using an HM355S automated microtome and an MB35 Premier microtome blade (34° / 80 mm). The microtome transfer system was set to 43 °C. The microtome transfer system controls the water bath temperature. Tissue sections were placed on superfrost microscope slides and allowed to air dry at room temperature before IHC.

[0230] For IHC, all reagents were brought to room temperature before staining. Formalin-fixed, paraffin-embedded (FFPE) sections were baked at 60°C for 45 minutes. Immediately after deparaffinization, FFPE sections were immersed in 1x Target Retrieval Solution, pH 6.1 (Agilent) and subjected to heat-induced epitope retrieval using a digital decoking chamber (BioCare Medical) set at 120°C for 4 minutes at 10–15 PSI. After epitope retrieval, FFPE sections were immersed in diH2O and then in 1x Tris-buffered saline with Tween (TBST, Agilent) for 2 minutes each. FFPE sections were then immersed in 3% peroxidase block (FisherScientific) for 10 minutes, washed in TBST for 3 minutes each, and then treated with primary antibody (MAB2906, R&D Systems) for 60 minutes at room temperature. The primary antibody was diluted (Antibody Diluent, Agilent) to a final working solution of 1 μg / mL. The FFPE sections were then washed with TBST (5 times, 3 minutes), and the secondary antibody (Envision Mouse Horseradish Peroxidase-HRP, Agilent) was added for 30 minutes at room temperature. The FFPE sections were then washed with TBST (5 times, 3 minutes) and immersed in a solution of 3,3'-diaminobenzidine (DAB, Agilent). The FFPE sections were then washed with diH2O (5 times, 2 minutes), immersed in Gill's Hematoxylin Solution No. 1 (Sigma), rinsed with diH2O, and immersed in Bluing Reagent S Series (Expredia). The FFPE sections were then rinsed with diH2O and baked at 60°C until dry. The FFPE sections were then coverslipped using DPX mounting medium (Sigma) for histology. FFPE slides (see Figure 3) were scanned using an Axios slide scanner, and the images were then analyzed using HALO software.

[0231] In vivo preclinical imaging Disease Model: A graft-versus-host disease (GvHD) model was selected to evaluate radiolabeled granzyme-B peptides due to the presence of human granzyme-B after disease onset. Ten million human peripheral blood mononuclear cells (PBMCs) were transplanted into immunocompromised NOG (NOD / Shi-scid / IL-2Rγ) mice. Body weight was monitored regularly, and PET / CT scans were performed when mice began to lose weight (an indicator of disease onset). To determine target specificity, NOG mice without human PBMCs were used as controls. Additionally, a scrambled peptide with minimal or no binding to human granzyme-B was used as an additional control. Mice were injected with approximately 150 uCi (35 Ci / mmol). 18 F-labeled peptides were injected intravenously, and a 45-minute whole-body static PET / CT scan was performed 1 hour after injection. Attenuation correction was performed using low-dose CT scans. Regions of interest (ROIs) were manually extracted on the tissues of interest, and mean standardized uptake values ​​(SUVs) were determined.

[0232] hPBMC donor derived 18 PET imaging performed with F-labeled anti-granzyme-B peptide 11a revealed significantly higher uptake in the lung, liver, and bone in the GvHD model than in the NOG control (Figure 1:A). Figure 1:B shows 18 The corresponding SUV averages in the tissues of interest induced after injection of F-labeled anti-granzyme B peptide 11a are shown. The kidney was the primary tissue through which the peptide was excreted. No significant differences were observed between the kidney and the bladder. 18 When F-labeled scrambled peptide (negative control) was used, no significant differences were observed between the lung, liver, and bone in the GvHD model and NOG control mice (Figure 2: A). Figure 2: B shows the results in the GvHD model mice and NOG control mice. 18 The corresponding SUV averages in tissues of interest induced after injection of F-labeled scrambled peptide are shown. For purposes of this disclosure, scrambled peptide is meant to be *FQWQASNEDDTPF*CGGK(NOTA-1).NH2).

[0233] hPBMC derived from a different donor than that performed in 11a 68 PET imaging studies performed with Ga-labeled anti-granzyme B peptide 19a revealed significantly higher uptake in the lung and liver in the GvHD model than in the NOG control (Figure 5A and Figure 5B).

[0234] Granzyme-B expression in infiltrating cells was confirmed by IHC performed on tissues collected from GvHD mice and NOG control mice. Representative human granzyme-B IHC images at 100 μm and 50 μm resolution from lung tissues collected from GvHD mice (upper panel) and NOG control mice (lower panel) are shown in Figure 3. Granzyme-B staining is indicated by arrows.

[0235] Healthy non-human primates: A 3-hour dynamic PET / CT scan was performed in healthy male rhesus monkeys to determine whole-body biodistribution and clearance characteristics in additional species. 18 Animals were intravenously injected with F-labeled anti-granzyme B peptide. Attenuation correction was performed using low-dose CT scans. Regions of interest (ROIs) were manually extracted on the tissues of interest, and mean standardized uptake values ​​(SUVs) were determined.

[0236] PET imaging in healthy rhesus monkeys revealed rapid blood pool clearance and high uptake in the kidney within the first 20 minutes of injection (Figure 4(A) and Figure 4(B)). As shown in Figure 4(A), 18 F-labeled anti-granzyme B peptide 11a was primarily excreted via the renal route. 平均 The corresponding quantitative data, denoted as , are shown (right).

[0237] In normal rhesus monkeys, as shown in Figure 6(A), 68 Ga-labeled anti-granzyme B peptide 19a was primarily excreted via renal excretion, with the highest uptake observed in the urine / bladder and kidney. Figure 6(B) shows the SUV 平均The corresponding quantitative data, denoted as , are shown (right).

Claims

1. Formula I: 【Chemistry 1】 [In the formula, R １ is C １－６ selected from alkyl and aryl, said aryl being C １－６ optionally substituted with 1 to 3 groups selected from alkyl, halogen, and hydroxyl; R ２ is hydrogen, halogen, and C １－６ alkyl, R ３ is hydrogen, -CH ２ OH, C １－７ Alkyl, —CH ２ CONH ２ , and -CH ２ NH ２ is selected from R ４ , R ５ , and R ６ are independently hydrogen, C １－６ Alkyl, (CH ２ ) ３ NHC (=NH ２ ) NH ２ , (CH ２ ) ４ NH ２ , and C １－６ -CH optionally substituted with 1 to 3 substituents selected from alkyl and halogen ２ -heteroaryl, Or, R ４ and R ６ together with the atoms to which they are attached, １－６ forming a nitrogen-containing 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 groups selected from alkyl and halogen; R ７ , R ８ , and R ９ are independently hydrogen, C １－６ Alkyl, CH ２ Heteroaryl, and CH ２ aryl, wherein the alkyl is selected from R ａ wherein said heteroaryl and aryl are optionally substituted with 1 to 3 OH groups; Or, R ８ and R ９ together with the atoms to which they are attached, １－６ forming a nitrogen-containing 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 groups selected from alkyl and halogen; R ａ is hydroxyl, -COOH, -NH ２ , -NHC(=NH ２ ) NH ２ and C(O)NH ２ is selected from R １０ is CH ２ Heteroaryl and C １－６ alkyl, wherein said heteroaryl and alkyl are selected from OH, C １－６ optionally substituted with 1 to 3 groups selected from alkyl and halogen; R １１ and R １２ are both C １－６ alkyl or R １１ and R １２ together with the atoms to which they are attached, １－６ forming a 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 groups selected from alkyl, phenyl, and halogen; R １３ is -NH ２ , 【Chemistry 2】 is selected from R １４ teeth, (i) H, (ii) 【Transformation 3】 , wherein (ii) may comprise a positron-emitting isotope; (iii) R0aC(O)- (wherein R0a is C １－３ alkyl), or a polyethylene glycol polymer, (iv) 【Chemistry 4】 , wherein (iv) may comprise a positron-emitting isotope; (v) a chelating moiety, which may include a positron-emitting isotope; (vi) 【Transformation 5】 、 (vii) Formula II: 【Transformation 6】 wherein n is an integer selected from 1 to 24. a polyethylene glycol polymer of (viii) Formula III: 【Transformation 7】 wherein each m is an integer independently selected from 1 to 12; R １５ is H, —C(O)(CH ２ ) ２ NHC(O)OC(CH ３ ) ３ , —C(O)CH ２ NH (NOTA-1), R １４ (ii) R １４ (iii) R １４ (iv) and R １４ (v) selected from Polyethylene glycol polymer is selected from X is 【Transformation 8】 (In the formula, R １’ is C １－６ selected from alkyl and aryl, said aryl being C １－６ optionally substituted with 1 to 3 substituents selected from alkyl, halogen, and hydroxyl; R ２’ is hydrogen, halogen, and C １－６ alkyl, R ３’ is H, -CH ２ OH, C １－７ Alkyl, —CH ２ CONH ２ , and -CH ２ NH ２ is selected from R ４’ , R ５’ , and R ６’ are independently hydrogen, C １－６ Alkyl, -(CH ２ ) ３ NHC (=NH)NH ２ , and -(CH ２ ) ４ NH ２ , and -CH ２ -heteroaryl, said heteroaryl being selected from C １－６ optionally substituted with 1 to 3 substituents selected from alkyl and halogen; Or, R ４’ and R ６’ together with the atoms to which they are attached, １－６ forming a nitrogen-containing 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 substituents independently selected from alkyl and halogen; R ７’ , R ８’ , and R ９’ are independently hydrogen, C １－６ Alkyl, CH ２ Heteroaryl and CH ２ aryl, wherein said alkyl is selected from 1 to 3 R ａ and the heteroaryl and aryl are optionally substituted with 1 to 3 OH groups; Or, R ８’ and R ９’ together with the atoms to which they are attached, １－６ forming a nitrogen-containing 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 substituents independently selected from alkyl and halogen; R １０’ is -CH ２ Heteroaryl and C １－６ alkyl, wherein said heteroaryl and alkyl are selected from OH, C １－６ optionally substituted with 1 to 3 groups selected from alkyl and halogen; and R １１’ and R １２’ are C １－６ is alkyl, Or, R １１’ and R １２’ together with the atoms to which they are attached, １－６ Forms a 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 substituents selected from alkyl, phenyl, and halogen. It is.] The macrocyclic peptides of the formula (I) and pharmaceutically acceptable salts thereof.

2. R １ But C １－６ alkyl and phenyl, wherein said alkyl and phenyl are selected from C １－６ 2. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, optionally substituted with 1 to 3 groups selected from alkyl, halogen, and hydroxyl.

3. R ３ is hydrogen, CH ２ OH, CH ２ CONH ２ and C.H. ２ NH ２ The compound according to any one of claims 1 to 2, or a pharmaceutically acceptable salt thereof, selected from:

4. R ４ , R ５ , and R ６ each independently represents hydrogen, C １－６ Alkyl, (CH ２ ) ３ NHC (=NH)NH ２ , (CH ２ ) ４ NH ２ , and C.H. ２ The compound according to any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, selected from indolyl.

5. R ４ and R ６ together with the atoms to which they are bonded, １－６ The compound according to any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, which forms a nitrogen-containing 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 groups selected from alkyl and halogen.

6. R ７ , R ８ , and R ９ are independently hydrogen, C １－６ Alkyl, —CH ２ OH, -CH ２ COOH, -CH(OH)CH ３ , -(CH ２ ) ２ C(O)NH ２ , -(CH ２ ) indolyl, -CH ２ Phenyl, —CH ２ PhenylOH, -(CH ２ ) ４ NH ２ , -(CH ２ ) ３ NHC (=NH)NH ２ , and -CH ２ imidazolyl, wherein the alkyl is selected from 1 to 3 R ａ 6. The compound according to any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof, wherein the indolyl, phenyl, and imidazolyl are optionally substituted with 1 to 3 OH groups.

7. R ７ , R ８ , and R ９ Two of the are hydrogen and the other is CH ３ , C.H. ２ CH (CH ３ ) ２ , CH(CH ３ ) CH ２ CH ３ , CH(CH ３ ) CH ２ CH ３ CH (CH ３ ) ２ , C.H. ２ OH, CH ２ COOH, CH(OH)CH ３ , (CH ２ ) ２ C(O)NH ２ , -(CH ２ ) ３ -NHC(=NH)NH ２ , -(CH ２ ) ４ -NH ２ , and -(CH ２ ) ２ 7. The compound of claim 6, or a pharmaceutically acceptable salt thereof, wherein the compound is selected from the group consisting of:

8. R ８ and R ９ together with the atoms to which they are bonded, １－６ The compound according to any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof, which forms a nitrogen-containing 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 groups selected from alkyl and halogen.

9. R ８ and R ９ and R 1 and R 2 are independently substituted or unsubstituted, or a pharmaceutically acceptable salt thereof.

10. R １０ is optionally substituted C １－６ Alkyl and CH ２ 10. The compound according to any one of claims 1 to 9, or a pharmaceutically acceptable salt thereof, selected from indolyl.

11. R １１ and R １２ But both are C １－６ alkyl, or together with the atom to which they are attached, C １－６ The compound according to any one of claims 1 to 10, which forms a 3- to 10-membered heterocyclyl group optionally substituted with 1 to 3 groups selected from alkyl, phenyl, and halogen, or a pharmaceutically acceptable salt thereof.

12. R １１ and R １２ are taken together to form a group selected from pyrrolidinyl, piperidinyl, and tetrahydroisoquinolinyl, and said pyrrolidinyl, piperidinyl, and tetrahydroisoquinolinyl are selected from C １－６ 12. The compound according to claim 11, or a pharmaceutically acceptable salt thereof, optionally substituted with 1 to 3 groups selected from alkyl, phenyl, and halogen.

13. R １１ and R １２ together form a pyrrolidinyl, and said pyrrolidinyl is C １－６ 12. The compound according to claim 11, or a pharmaceutically acceptable salt thereof, optionally substituted with 1 to 3 groups selected from alkyl, phenyl, and halogen.

14. R １３ But NH ２ 14. The compound according to any one of claims 1 to 13, wherein:

15. R １３ but, 【Chemistry 9】 or 【Chemistry 10】 14. The compound according to any one of claims 1 to 13, wherein:

16. R １４ is hydrogen and R0aC(O)- (wherein R0a is C １－３ 16. The compound of claim 15, or a pharmaceutically acceptable salt thereof, wherein the alkyl group is selected from the group consisting of PEG, PEG 1-24, PEG 2-25, PEG 3-26, PEG 4-27, PEG 5-28, PEG 6-29, PEG 7-29, PEG 8-29, PEG 9-29, PEG 10-29, PEG 11-29, PEG

17. R １４ is represented by Formula II and Formula III: 【Chemistry 11】 wherein n is an integer selected from 1 to 24, each m is an integer independently selected from 1 to 12, and R １５ is hydrogen, C(O)(CH ２ ) ２ NHC(O)OC(CH ３ ) ３ C(O)CH ２ NH(NOTA-1), and R １４ (ii), (iii), (iv) or (v) 16. The compound of claim 15, or a pharmaceutically acceptable salt thereof, which is a polyethylene glycol polymer selected from:

18. R １４ However, deferoxamine, 1,4,7,10-tetraacetic acid, diethylenetriaminepentaacetic acid, ethylenediaminetetraacetic acid, (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetra(methylenephosphonic) acid, (1R,4R,7R,10R)-α'α"α'''-tetramethyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid, 1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid, H ４ Oktapa, H. ６ Hospa, H2 Dedopa, H ５ Decapa, H2Azapa, HOPO, DO2A, 1,4,7,10-tetrakis(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane, 1,4,7-triazacyclononane-N,N',N"-triacetic acid, 1,4,8,11-tetraazabicyclo[6.6.2]hexadecane-4,11-dicetic acid (dicetic 16. The compound of claim 15, or a pharmaceutically acceptable salt thereof, wherein the chelating moiety is selected from: 1,4,7,10-tetraazacyclododecane, 1,4,8,11-tetraazacyclotetradecane, an octadentate chelating agent, a hexadentate chelating agent, a phosphonate chelating agent, a macrocyclic chelating agent, a chelating agent comprising a macrocyclic terephthalamide ligand, a bifunctional chelating agent, fusarinine C and fusarinine C derivative chelating agents, triacetylfusarinine C, ferrioxamine E, ferrioxamine B, and ferrichrome A, wherein the chelating moiety optionally comprises a positron-emitting isotope as an imaging agent.

19. R １４ but, (ii) 【Chemistry 12】 is selected from 16. The compound of claim 15, or a pharmaceutically acceptable salt thereof, wherein (ii) and (iv) are optionally chelated to a positron-emitting isotope.

20. The positron-emitting isotope is ６８ Ga, １８ F. ６４ Cu, and １８ 20. The compound of claim 19, selected from F-Al.

21. The compound of any one of claims 1 to 17, comprising a chelating moiety.

22. the chelating moiety ６８ Ga, １８ F, and ６４ 22. The compound of claim 21, comprising a positron-emitting isotope selected from Cu.

23. Structural formula Ia: 【Chemistry 13】 (In the formula, R ｘ and R ｙ is hydrogen, hydroxyl, C １－６ independently selected from alkyl and halogen 2. The compound of claim 1, wherein said Formula Ia is represented by: wherein said Formula Ia optionally comprises one or more positron-emitting imaging agents, or a pharmaceutically acceptable salt thereof.

24. A compound or a pharmaceutically acceptable salt thereof selected from: Table 1

25. 2. The macrocyclic peptide of claim 1, or a pharmaceutically acceptable salt thereof, selected from the following: Table 2 where * represents the amino acid attachment point between the amino acid leucine (L), phenylalanine (F), or tyrosine (Y) and the sulfur on the amino acid cysteine ​​(C).

26. 26. The compound of any one of claims 1 to 25, or a pharmaceutically acceptable salt thereof, which is a binding agent for Granzyme B.

27. 26. The compound according to any one of claims 1 to 25, or a pharmaceutically acceptable salt thereof, which is an inhibitor of Granzyme B.

28. A method for imaging granzyme B in a cell or tissue, comprising contacting the cell or tissue with a compound of any one of claims 1 to 25 or a pharmaceutically acceptable salt thereof, and imaging the cell or tissue using a suitable imaging technique, thereby imaging granzyme B in the cell or tissue.

29. A method for treating a disease in which granzyme B is involved, comprising administering to a subject in need thereof a compound according to any one of claims 1 to 25 or a pharmaceutically acceptable salt thereof.

30. 26. The use according to any one of claims 1 to 25, or a pharmaceutically acceptable salt thereof, for treating a disease in which granzyme B is involved.

31. 26. The use according to any one of claims 1 to 25, or a pharmaceutically acceptable salt thereof, for imaging granzyme B in said cells or tissues.

32. A pharmaceutical composition comprising a compound according to any one of claims 1 to 25 and a pharmaceutically acceptable carrier or excipient.

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