Novel therapeutic radiopharmaceuticals and methods

Abstract

The present disclosure provides novel radiopharmaceutical conjugates for tumor targeting and delivery, and their therapeutic and / or theragnostic use.

Description

Attorney Docket No: 119289-861234NOVEL THERAPEUTIC RADIOPHARMACEUTICALS AND METHODSFIELD OF THE DISCLOSURE

[0001] The present disclosure relates to novel radiopharmaceuticals and methods of making and using such radiopharmaceuticals.BACKGROUND

[0002] Current radiotherapeutics are used for systemic treatment of multiple malignancies - however all treated patients will progress and die. Current radiotherapeutic agents are based on p-particle-emitting radionuclides that are effective at extending life and abating symptoms in many patients - although they are never curative, and they may have significant toxicities. To overcome these deficiencies, new radiopharmaceuticals are needed.SUMMARY OF THE DISCLOSUREFORMULA (I)

[0003] One aspect of the present disclosure provides a conjugate of Formula (I), also referred to herein as TRAIL, comprising a tumor-targeting moiety T; an Auger-electron-emitting radionuclide RA to bring radiation to cancer cells; a DNA-intercalating moiety DI to localize the conjugate to DNA, and a Linker linking the T, RA and DI components. In one aspect, the linker is non-cleavable. In another aspect, the Linker is cleavable, which allows the release of T, RA and / or DI from the TRAIL conjugate upon exposure to a cleaving condition. In yet another aspect, the Linker is a cleavable linker selected from a biodegradable linker, a biocompatible linker, an enzymatic cleavable linker, a pH sensitive linker, a photo sensitive linker, a radiation sensitive linker, or any combination thereof. In one aspect, the bond between the Linker and T, the Linker and RA, and the Linker and DI each independently comprises an amide bond, an amine bond, an imide bond, an ester bond, an acrylate bond, a disulfide bond, a sulfhydryl bond, an alkylsulfide bond, a1106498195 1Attorney Docket No: 119289-861234 sulfonyl bond, a succinimide bond, a phosphate bond, a phosphoramidate bond, a peptide bond, an oligopeptide bond, a biocompatible polymer bond, a biodegradable bond, a maleimide bond, a click chemistry bond, a photo-cleavable bond, a radiation-cleavable bond, or any combination thereof.

[0004] In another aspect, the tumor-targeting moiety T is a moiety selected from Prostate Specific Membrane Antigen (PSMA), Fibroblast Activation protein, a carbonic anhydrase (for example, carbonic anhydrase IX), Herceptin, RGD, EGF, PDGF, VEGF, hyaluronan, AS-1411 , GBI-10, biotin, vitamin H, vitamin B-7, folates, Lectins, Lactoferrin, transferrin, integrin, Mannose derivates, bombesin, bradykinin, mesothelin, hepsin, mucin, estrogen receptor, milk fat globulin, telomerases, nuclear matrix proteins, prostatic acid phosphatase, SCCA, OCA, pancreas cancer associated antigen (PaA), or antibodies, antibody fragments or antigenic fragments of EGFR, HER2, HER3, HER4, BRCA1 CA19-9, CA59, CA-125, CD4, CD19, CD20, CD22, CD23, CD276, CD30, CD32, CD324, CD33, CD34, CD44, CD5, CD52, CD70, CD71 , CD79b, CDH1 , CDH6, CDH19, CDH17, CEA, CTLA4, DLL3, EphA2, FLT3, GRP78, insulin receptor, IGFR, VEGF, VEGFR1 , VEGFR2, VEGFR3, Ly1 , Lym1 , Lym2, MPL, c-MET, MUC1 , MUC2, MUC3, MUC16, MUC18, NRP1 , PD1 , PD-L1 , PTK7, TGFp, TNF.TROP2, TFM1 , LAMP1 , VLA4, MART1 / MelanA, gp100, tyrosinase, TRP1 , TRP2, NY-ESO-1 , CDK-4, p-catenin, MUM1 , Caspase-8, KIAA0205, HPVE7, SART1 , PRAME, p15, BAGE1 , DAGE1 , RAGE1 , NAG, TAG72, CA125, p21ras, p53, HPV16-E7, SSX , HOM-MEL55, NY-COL2, HOM-HD397, HOM-RCC-1 .14, HOM-HD21 , HOM-NSCLC11 , HOM-MEL-2.4, HOM-TES11 , RCC-3.1.3, NY-ESO-1 , MAGE1 , MAGE2, MAGE3, MAGE4 MAGE-11 , GAGE1 , GAGE6, Ha-ras, RAF, GD2, GD3, GM2, TF, sTn, gp75, EBV-LMP1 , EBV-LMP2, HPV-F4, HPV-F6, HPV-F7, PSA), AFP, CO17-1A, GA733, gp72, p-HCG, gp43, HSP-70, p17 mel, gp43, HMW, HOJ-1 , melanoma gangliosides, TAG-72, MZ2-E, PEM, LK26, Thomsen-Friedenreich (T), HCG, pancreatic oncofetal antigen, cancer antigens 15-3, 19-9, 549, 195, PROTACs, topoisomerase inhibitor, leukemia inhibitory factor or any combination thereof. In one aspect, the Auger-electron- emitting radionuclide RA comprises iodine-125, iodine-131 , gallium-68, fluoride 18, terbium-161 , or lutetium-177. In one aspect, the DNA-intercalating moiety DI comprises acridine orange, propidium iodide, DAPI (4',6-diamidino-2-phenylindole), berberine, ethidium bromide, proflavine, or thalidomide. In one aspect, the Linker2106498195 1Attorney Docket No: 119289-861234 comprises a moiety of Formula (I I A), wherein X is C, P, P-OH, or S; n2 is zero or an integer of 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; Ri is selected from the group consisting of hydroxyl, -NH2, -C(O)NH2, -NHC(O), -C(O)OH, -C(O)OMe, -C(O)R5, -C(O)NR5, -OC(O)Rs, -C(O)ORS-, -N3; -C1-C15 alkyl, -C2-Cis alkenyl, -C2-Cis alkynyl; R2is selected from the substituted or unsubstituted group consisting of -C1-C15 alkyl, -C2-Ci5 alkenyl, -C2-Cis alkynyl, and Rs is selected from the group consisting of hydrogen, C1-C6 alkyl, C2-Ce alkenyl, and C2-Ce alkynyl. In yet another aspect, the Linker comprises a moiety of Formula (I IB).Formula (HA) Formula (IIB)

[0005] In another aspect, the Linker has a structure of Formula (IIC), wherein X is C, P, P-OH, or S; n1 , n2, n3, n4 and n5 each is independently zero or an integer of 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; R1 is selected from the group consisting of hydroxyl, -NH2, -C(O)NH2, -NHC(O), -C(O)OH, -C(O)OMe, -C(O)Rs, -C(O)NRs, -OC(O)Rs, -C(O)ORs-, -N3; -C1-C15 alkyl, -C2-Cis alkenyl, -C2-Cis alkynyl; R2and R3 each is independently selected from the substituted or unsubstituted group consisting of -C1-C15 alkyl, -C2-Cis alkenyl, -C2-Cis alkynyl, and Rs is selected from the group consisting of hydrogen, Ci-Ce alkyl, C2-Ce alkenyl, and C2-Ce alkynyl.Formula (IIC)

[0006] In one aspect, the Linker is or comprises a moiety of HE4, PEG, or a moiety of the following structure Formula (HD):3106498195 1Attorney Docket No: 119289-861234Formula (HD)

[0007] In one aspect, the conjugate has a structure of Formula (III), wherein X is C, P, P-OH, or S; n1 , n2, n3, n4 and n5 each is independently zero or an integer of 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; Ri is selected from the group consisting of hydroxyl, -NH2, -C(O)NH2, -NHC(O), -C(O)OH, -C(O)OMe, -C(O)Rs, -C(O)NR5, -OC(O)R5, -C(O)OR5-, -N3; -C1-C15 alkyl, -C2-Ci5 alkenyl, -C2-Ci5 alkynyl; R2and R3 each is independently selected from the substituted or unsubstituted group consisting of -C1-C15 alkyl, -C2-Ci5 alkenyl, -C2-Cis alkynyl, and Rs is selected from the group consisting of hydrogen, Ci-Ce alkyl, C2-Cs alkenyl, and C2-Ce alkynyl.FORMULA (III)

[0008] In various aspects, the conjugate has one of the following structures:4106498195 1Attorney Docket No: 119289-861234

[0009] In various aspects, the present disclosure is directed to a building block compound selected from LP-1 ; LP-1-1 ; L-2; Simplified LP-2; and LP-3.

[0010] In various aspects, the present disclosure is directed to a conjugate selected from TRAIL-1 ; TRAIL-2; TRAIL-3; TRAIL-4; TRAIL-5; TRAIL-6; TRAIL-7; TRAI L-8; TRAI L-9; TRAI L-10; TRAI L-11 ; and TRAI L-12.

[0011] In various aspects, the present disclosure is directed to a conjugate selected from Click_1 ; Click_2; Click_3; Click_4; Click_5; Click_6; Click_7; and Click_8.

[0012] In yet another aspect, the present disclosure encompasses a composition comprising an amount of the conjugate, and a carrier. The amount of the conjugate may be a pharmaceutically effective amount, and the carrier may be a pharmaceutically acceptable carrier. In yet another aspect, the amount may be a diagnostically effective amount, and the carrier may be a diagnostically acceptable carrier. In yet another aspect, the amount may be a theragnostically effective amount, and the carrier may be a theragnostically acceptable carrier.5106498195 1Attorney Docket No: 119289-861234

[0013] In various aspects, a composition according to the present disclosure may be formulated for parenteral delivery or for enteral delivery, such as oral delivery. In another aspect, the composition may be formulated for injection. In various aspects, a composition formulated for injection may formulated for intravenous injection, intratumor injection, subcutaneous injection, intramuscular injection, or intrathecal injection. In yet another aspect, the composition is prepared in a unit dosage form.

[0014] In additional aspects, the present disclosure provides a method of inducing DNA scission, and / or a method for treating a tumor in a subject. Such methods comprise administering to a subject in need thereof a therapeutically effective amount of the conjugate or a composition thereof. In another aspect, the present disclosure provides a theragnostic method by administering a theragnostically effective amount of the conjugate or a composition thereof, to a subject in need thereof. In various aspects of any of the methods, the subject may have or may be suspected of having a cancer or a tumor. A cancer or tumor may be breast cancer, lung cancer, colorectal cancer, colon cancer, skin cancer, bladder cancer, kidney cancer, or prostate cancer.

[0015] In another aspect, the present disclosure encompasses a synthesis method for the conjugate, the method comprises the steps shown in Scheme 1 :6106498195 1Attorney Docket No: 119289-861234SCHEME 1REFERENCE TO COLOR FIGURES

[0016] The application file contains at least one photograph executed in color. Copies of this patent application publication with color photographs will be provided by the Office upon request and payment of the necessary fee.BRIEF DESCRIPTION OF THE FIGURES

[0017] FIGURE 1 depicts an conjugate and release mechanism of an exemplary TRAIL conjugate.

[0018] FIGURE 2 depicts exemplary control molecules against an exemplary TRAIL conjugate.

[0019] FIGURE 3 provides another exemplary TRAIL conjugate that incorporates radiometal of terbium-161 .

[0020] FIGURE 4 depicts an exemplary synthesis route of an exemplary TRAIL conjugate.7106498195 1Attorney Docket No: 119289-861234

[0021] FIGURES 5-17 depict further aspects and embodiments of the disclosure.

[0022] FIGURES 18-22 provide data regarding aspects and embodiments of the disclosure.DETAILED DESCRIPTION

[0023] The present disclosure is based in part on the design and synthesis of a novel class of radiopharmaceuticals, and their use in imaging, therapeutic, diagnostic, and theragnostic applications. In one aspect, a radiopharmaceutical as disclosed herein is a conjugate molecule comprising at least the following four components: (1) a tumor-targeting moiety T for targeting cancer cells, e.g., prostate cancer; (2) an Auger-electron-emitting radionuclide (i.e., with a certain type of radioactivity); (3) a DNA-intercalating moiety to localize the compound to DNA; and (4) a Linker linking the component (1) through (3) and optionally allowing release of the components from the conjugate. The novel radiopharmaceutical conjugate is also referred to herein as a “TRAIL” conjugate throughout, referring T as the tumortargeting moiety, RA for radionuclide; I for DNA intercalating moiety; and L for linker.I. Radiopharmaceutical TRAIL Conjugates

[0024] Various TRAIL conjugates are designed and synthesized. In one aspect, the TRAIL conjugate is of Formula (I), having a tumor-targeting moiety T to target the tumor; an Auger-electron-emitting radionuclide RA to bring the radiation to cancer cells; a DNA-intercalating moiety DI to localize the conjugate to DNA, and a Linker linking T, RA and DI. In one aspect, the linker is non-cleavable. In another aspect, the Linker is cleavable, which allows the release of T, RA and / or DI from the conjugate upon exposure to a cleaving condition. In yet another aspect, the Linker is a cleavable linker selected from a biodegradable linker, a biocompatible linker, an8106498195 1Attorney Docket No: 119289-861234 enzymatic cleavable linker, a pH sensitive linker, a photo sensitive linker, a radiation sensitive linker, or any combination thereof. In another aspect, the bond between Linker-T, Linker-RA, or Linker-DI each independently comprises an amide bond, an amine bond, an imide bond, an ester bond, an acrylate bond, a disulfide bond, a sulfhydryl bond, an alkylsulfide bond, a sulfonyl bond, a succinimide bond, a phosphate bond, a phosphoramidate bond, a peptide bond, an oligopeptide bond, a biocompatible polymer bond, a biodegradable bond, a maleimide bond, a click chemistry bond, a photo-cleavable bond, a radiation-cleavable bond, or any combination thereof. In another aspect, the tumor-targeting moiety T is a moiety selected from the group consisting of Prostate Specific Membrane Antigen (PSMA), Fibroblast Activation protein, a carbonic anhydrase (for example, carbonic anhydrase IX), Herceptin, RGD, EGF, PDGF, VEGF, hyaluronan, AS-1411 , GBI-10, biotin, vitamin H, vitamin B-7, folates, Lectins, Lactoferrin, transferrin, integrin, Mannose derivates, bombesin, bradykinin, mesothelin, hepsin, mucin, estrogen receptor, milk fat globulin, telomerases, nuclear matrix proteins, prostatic acid phosphatase , SCCA, OCA, pancreas cancer associated antigen (PaA), or antibodies, antibody fragments or antigenic fragments of EGFR, HER2, HER3, HER4, BRCA1 CA19-9, CA59, CA-125, CD4, CD19, CD20, CD22, CD23, CD276, CD30, CD32, CD324, CD33, CD34, CD44, CD5, CD52, CD70, CD71 , CD79b, CDH1 , CDH6, CDH19, CDH17, CEA, CTLA4, , DLL3, EphA2, FLT3, GRP78, insulin receptor, IGFR, VEGF, VEGFR1 , VEGFR2, VEGFR3, Ly1 , Lym1 , Lym2, MPL, c-MET, MUC1 , MUC2, MUC3, MUC16, MUC18, NRP1 , PD1 , PD-L1 , PTK7, TGFp, TNF.TROP2, TFM1 , LAMP1 , VLA4, MARTI / MelanA, gp1OO, tyrosinase, TRP1 , TRP2, NY-ESO-1 , CDK- 4, p-catenin, MUM1 , Caspase-8, KIAA0205, HPVE7, SART1 , PRAME, p15, BAGE1 , DAGE1 , RAGE1 , NAG, TAG72, CA125, p21 ras, p53, HPV16-E7, SSX , HOM- MEL55, NY-COL2, HOM-HD397, HOM-RCC-1.14, HOM-HD21 , HOM-NSCLC11 , HOM-MEL-2.4, HOM-TES11 , RCC-3.1.3, NY-ESO-1 , MAGE1 , MAGE2, MAGE3, MAGE4 MAGE-11 , GAGE1 , GAGE6, Ha-ras, RAF, GD2, GD3, GM2, TF, sTn, gp75, EBV-LMP1 , EBV-LMP2, HPV-F4, HPV-F6, HPV-F7, PSA), AFP, C017-1A, GA733, gp72, p-HCG, gp43, HSP-70, p17 mel, gp43, HMW, HOJ-1 , melanoma gangliosides, TAG-72, MZ2-E, PEM, LK26, Thomsen-Friedenreich (T), HCG, pancreatic oncofetal antigen, cancer antigens 15-3, 19-9, 549, 195, PROTACs, topoisomerase inhibitor, leukemia inhibitory factor or any combination thereof. In one aspect, the Auger- electron-emitting radionuclide RA comprises iodine-125, iodine-131 , gallium-68,9106498195 1Attorney Docket No: 119289-861234 fluoride 18, terbium-161 , or lutetium-177. In one aspect, the DNA-intercalating moiety DI comprises acridine orange, berberine, ethidium bromide, proflavine, or thalidomide. In one aspect, the Linker comprises a moiety of Formula (HA), wherein X is C, P, P-OH, or S; n2 is zero or an integer of 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; Ri is selected from the group consisting of -NH2, -C(O)NH2, -NHC(O), -C(O)OH, -C(O)OMe, -C(O)Rs, -C(O)NRs, -OC(O)Rs, -C(O)ORs-, -NS; -C1-C15 alkyl, -C2-C15 alkenyl, -C2-Cis alkynyl; R2is selected from the substituted or unsubstituted group consisting of -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl, and Rs is selected from the group consisting of hydrogen, Ci-Ce alkyl, C2-C6 alkenyl, and C2-C6 alkynyl. In yet another aspect, the Linker comprises a moiety of Formula (II B).Formula (HA) Formula (IIB)

[0025] In another aspect, the Linker has a structure of Formula (IIC), wherein X is C, P, P-OH, or S; n1 , n2, n3, n4 and n5 each is independently zero or an integer of 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; R1 is selected from the group consisting of hydroxyl, -NH2, -C(O)NH2, -NHC(O), -C(O)OH, -C(O)OMe, -C(O)Rs, -C(O)NRs, -OC(O)Rs, -C(O)ORs-, -Ns; -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl; R2and R3 each is independently selected from the substituted or unsubstituted group consisting of -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl. Rs is selected from the group consisting of hydrogen, Ci-Ce alkyl, C2-C6 alkenyl, and C2-C6 alkynyl.10106498195 1Attorney Docket No: 119289-861234Formula (HC)

[0026] In yet another aspect, the Linker is or comprises a moiety of HE4, PEG, or a moiety of the following structure Formula (HD):Formula (HD)

[0027] In an embodiment, the TRAIL conjugate comprises the following structures:11106498195 1Attorney Docket No: 119289-861234LP-2 lysosomal cleavage at pH 4.5,LP-3 non-cleavable

[0028] In another aspect, the TRAIL conjugate has a structure of Formula (III), wherein X is C, P, or S; n1 , n2, n3, n4 and n5 each is independently zero or an integer of 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; Ri is selected from the group consisting of hydroxyl, -NH2, -C(O)NH2, -NHC(O), -C(O)OH, -C(O)OMe, -C(O)Rs, -C(O)NR5, - OC(O)Rs, -C(O)ORS-, -NS; -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl; R2and Rs each is independently selected from the substituted or unsubstituted group consisting of -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl. The Rs is selected from the group consisting of hydrogen, Ci-Ce alkyl, C2-C6 alkenyl, and C2-C6 alkynyl.12106498195 1Attorney Docket No: 119289-861234Formula (III)

[0029] In yet another aspect, the TRAIL conjugate has the following compound:

[0030] In an aspect, the TRAIL conjugate has the following structure after Cu- catalyzed click reaction:13106498195 1Attorney Docket No: 119289-861234wherein, R =LP-2-p-hydroxy phenyl propionate containing 1-125 as Radionuclide (TRAIL- 1 ); R = Simplified LP-2-p-hydroxy phenyl propionate containing 1-125 Radionuclide (TRAIL-2); R = LP-3-p-hydroxy phenyl propionate containing I- 125 Radionuclide (TRAIL-3); R = LP-2-DOTA containing Lu-177 or Tb-161 as Radionuclide (TRAIL-4); R = Simplified LP-2-DOTA containing Lu-177 or Tb- 161 Radionuclide (TRAIL-5); or R = LP-3-DOTA containing Lu-177 or Tb-161 Radionuclide (TRAIL-6).

[0031] In an aspect, the TRAIL conjugate has the following structure after Cu- catalyzed click reaction:wherein R = LP-2-p-hydroxy phenyl propionate containing 1-125 as Radionuclide (TRAIL-7); R = Simplified LP-2-p-hydroxy phenyl propionate containing 1-125 Radionuclide (TRAIL-8); R = LP-3-p-hydroxy phenyl propionate containing 1-125 Radionuclide (TRAIL-9); R = LP-2-DOTA containing Lu-177 or Tb-161 as Radionuclide (TRAIL-10); R = Simplified LP-2-DOTA containing Lu-177 or Tb- 161 Radionuclide (TRAIL-1 1 ); R = LP-3-DOTA containing Lu-177 or Tb-161 Radionuclide (TRAIL-12).

[0032] In an aspect, the TRAIL conjugate has the following structure TRAIL-1 after Cu-catalyzed click reaction:14106498195 1Attorney Docket No: 119289-861234LP-2-p-hydroxy phenyl propionate containing 1-125 as a radionuclide

[0033] In another aspect, the TRAIL conjugate has the following structureTRAIL-2 after Cu-catalyzed click reaction:, wherein R =Simplified LP-2-p-hydroxy phenylpropionate containing 1-125 Radionuclide

[0034] In another aspect, the TRAIL conjugate has the following structureTRAIL-3 after Cu-catalyzed click reaction:15106498195 1Attorney Docket No: 119289-861234, wherein R= LP-3-p-hydroxy phenylpropionate containing 1-125 Radionuclide.

[0035] In another aspect, the TRAIL conjugate has the following structureTRAIL-4 after Cu-catalyzed click reaction:, wherein R = LP-2-DOTA containing Lu-177 or Tb-161 Radionuclide.

[0036] In another aspect, the TRAIL conjugate has the following structureTRAIL-5 after Cu-catalyzed click reaction:16106498195 1Attorney Docket No: 119289-861234Simplified LP-2-DOTAcontaining Lu-177 or Tb-161.

[0037] In another aspect, the TRAIL conjugate has the following structureTRAIL-6 after Cu-catalyzed click reaction:, wherein R = LP-3-DOTA containing Lu-167 or Tb-161.

[0038] In one aspect the TRAIL conjugate is TRAIL-1 :TRAIL-117106498195 1Attorney Docket No: 119289-861234

[0039] In another aspect the TRAIL conjugate is TRAIL-2:TRAIL-2

[0040] In another aspect the TRAIL conjugate is TRAIL-3:TRAIL-3

[0041] In another aspect the TRAIL conjugate is TRAIL-4:18106498195 1Attorney Docket No: 119289-861234TRAIL-4

[0042] In another aspect the TRAIL conjugate is TRAIL-5:TRAIL-5

[0043] In another aspect the TRAIL conjugate is TRAIL-6:19106498195 1Attorney Docket No: 119289-861234TRAIL-6

[0044] In one aspect, TRAIL-1 is likely to be cleaved in endosomes at pH 5.5. In another aspect, TRAIL-2 is likely to be cleaved in lysosomes at pH 4.5.

[0045] In another aspect, TRAIL-4 is likely to be cleaved in endosomes at pH 5.5. In another aspect, TRAIL-5 is likely to be cleaved in lysosomes at pH 4.5.

[0046] In one aspect, LP3 clicked derivatives have the following structures:20106498195 1Attorney Docket No: 119289-861234

[0047] In one aspect, present disclosure encompasses the synthesis of the TRAIL conjugate. One exemplary synthesis is given in Scheme 1.SCHEME 121106498195 1Attorney Docket No: 119289-861234II. Tumor-Targeting Moiety: T

[0048] The present disclosure encompasses a tumor-targeting moiety T. In one aspect, the tumor-targeting moiety T comprises a moiety of Prostate Specific Membrane Antigen (PSMA), Fibroblast Activation protein, Herceptin, RGD, EGF, PDGF, VEGF, hyaluronan, AS-1411 , GBI-10, biotin, vitamin H, vitamin B-7, folates, Lectins, Lactoferrin, transferrin, integrin, Mannose derivates, bombesin, bradykinin, mesothelin, hepsin, mucin, estrogen receptor, milk fat globulin, telomerases, nuclear matrix proteins, prostatic acid phosphatase , SCCA, OCA, pancreas cancer associated antigen (PaA), or antibodies, antibody fragments or antigenic fragments of EGFR, HER2, HER3, HER4, BRCA1 CA19-9, CA59, CA-125, CD4, CD19, CD20, CD22, CD23, CD276, CD30, CD32, CD324, CD33, CD34, CD44, CD5, CD52, CD70, CD71 , CD79b, CDH1 , CDH6, CDH19, CDH17, CEA, CTLA4, , DLL3, EphA2, FLT3, GRP78, insulin receptor, IGFR, VEGF, VEGFR1 , VEGFR2, VEGFR3, Ly1 , Lym1 , Lym2, MPL, c-MET, MUC1 , MUC2, MUC3, MUC16, MUC18, NRP1 , PD1 , PD-L1 , PTK7, TGFp, TNF.TROP2, TFM1 , LAMP1 , VLA4, MART1 / MelanA, gp100, tyrosinase, TRP1 , TRP2, NY-ESO-1 , CDK-4, p-catenin, MUM1 , Caspase-8, KIAA0205, HPVE7, SART1 , FRAME, p15, BAGE1 , DAGE1 , RAGE1 , NAG, TAG72, CA125, p21ras, p53, HPV16-E7, SSX , HOM-MEL55, NY-COL2, HOM-HD397, HOM-RCC-1 .14, HOM-HD21 , HOM-NSCLC11 , HOM-MEL-2.4, HOM-TES11 , RCC- 3.1.3, NY-ESO-1 , MAGE1 , MAGE2, MAGE3, MAGE4 MAGE-11 , GAGE1 , GAGE6, Ha-ras, RAF, GD2, GD3, GM2, TF, sTn, gp75, EBV-LMP1 , EBV-LMP2, HPV-F4, HPV-F6, HPV-F7, PSA), AFP, CO17-1A, GA733, gp72, P-HCG, gp43, HSP-70, p17 mel, gp43, HMW, HOJ-1 , melanoma gangliosides, TAG-72, MZ2-E, PEM, LK26, Thomsen-Friedenreich (T), HCG, pancreatic oncofetal antigen, cancer antigens 15- 3, 19-9, 549, 195, PROTACs, topoisomerase inhibitor, leukemia inhibitory factor or any combination thereof. In one aspect, the tumor-targeting moiety T comprises a moiety having the structure:22106498195 1Attorney Docket No: 119289-861234III. Radionuclide: RA

[0049] The present disclosure encompasses an Auger-electron-emitting radionuclide RA. In one aspect, the radionuclide RA comprises iodine-125, iodine- 131 , gallium-68, fluoride 18, terbium-161 , or lutetium-177. In one aspect, the Augerelectron-emitting radionuclide RA comprises a moiety having a moiety of:23106498195 1Attorney Docket No: 119289-861234IV. DNA Intercalating Moiety: I or DI

[0050] The present disclosure encompasses a DNA-intercalating moiety DI. A DNA-intercalating agent is a compound capable of inserting itself into the DNA structure of a cell and binds to the DNA, which results in DNA damage. In cancer treatment, DNA intercalating agents may kill cancer cells by either damaging their DNA and / or stopping them from dividing. In one aspect, the DNA-intercalating moiety DI comprises acridine orange, propidium iodide, berberine, ethidium bromide, proflavine, or thalidomide. In one aspect, the DNA-intercalating moiety DI comprises a moiety ofV. Linker

[0051] The present disclosure encompasses a Linker linking T, RA and / or DI.In one aspect, the linker is non-cleavable. In another aspect, the Linker is cleavable, which allows the release of T, RA and / or DI from the TRAIL conjugate upon exposure to a cleaving condition. In yet another aspect, the Linker is a cleavable linker selected from a biodegradable linker, a biocompatible linker, an enzymatic cleavable linker, a pH sensitive linker, a photo sensitive linker, a radiation sensitive linker, or any combination thereof. Enzymatic cleavage is a process wherein enzymes, such as peptidases or proteases, break peptide or peptidomimetic bonds. A pH sensitive linker may be acid- or base-liable, allowing the cleavage upon stimulation of pH change. In another aspect, subjecting the TRAIL conjugate to light (photo) or radiation may also trigger the cleavage of components T, RA, and / or DI from the TRAIL conjugate when the Linker is photo or radiation sensitive. In one aspect, the Linker comprises a moiety of Formula (I I A), wherein X is C, P, P-OH, or S; n2 is zero or an integer of 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; Ri is selected from the group consisting of hydroxyl, -NH2, -C(O)NH2, -NHC(O), -C(O)OH, -C(O)OMe, - C(O)R5, -C(O)NR5, -OC(O)R5, -C(O)OR5-, -N3; -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl; R2 is selected from the substituted or unsubstituted group consisting of -C1-24106498195 1Attorney Docket No: 119289-861234C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl, and Rs is selected from the group consisting of hydrogen, Ci-Ce alkyl, C2-C6 alkenyl, and C2-C6 alkynyl. In yet another aspect, the Linker comprises a moiety of Formula (I I B).Formula (HA) Formula (IIB)

[0052] In one aspect, such linking comprises a conjugate bond. In one aspect, the conjugate bond with T, RA, and / or DI each independently comprises an amide bond, an amine bond, an imide bond, an ester bond, an acrylate bond, a disulfide bond, a sulfhydryl bond, an alkylsulfide bond, a sulfonyl bond, a succinimide bond, a phosphate bond, a phosphoramidate bond, a peptide bond, an oligopeptide bond, a biocompatible polymer bond, a biodegradable bond, a maleimide bond, a click chemistry bond, a photo-cleavable bond, or any combination thereof. In one aspect, such Linker is cleavable linker and capable of releasing T, RA and / or DI simultaneously, sequentially, or at different time points. In one aspect, the linker L is of Formula (IIC):Formula (IIC)

[0053] In Formula (IIC), X is C, P, P-OH, or S; n1 , n2, n3, n4 and n5 each is independently zero or an integer of 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10; R1 is selected from the group consisting of hydroxyl, -NH2, -C(O)NH2, -NHC(O), -C(O)OH, -C(O)OMe, - C(O)R5, -C(O)NR5, -OC(O)R5, -C(O)OR5-, -N3; -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl; R2 and R3 each is independently selected from the substituted or unsubstituted group consisting of -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl. The25106498195 1Attorney Docket No: 119289-861234Rs is selected from the group consisting of hydrogen, Ci-Ce alkyl, C2-C6 alkenyl, and C2-C6 alkynyl. In another aspect, the Linker comprises a moiety of HE4 or PEG (such as PEG12), or a moiety shown below as Formula (I I D):Formula (HD)VI. METHODS OF USE

[0054] The present disclosure encompasses methods of use of the novel TRAIL conjugates. The conjugates may be used in various medical fields, such as in disease therapy, in disease diagnosis, and / or as theragnostics. One such use is to induce DNA scission and / or to treat a tumor or cancer in a subject. In one aspect, such method comprises administering the TRAIL conjugate in an amount sufficient to induce DNA scission, and / or to kill or reduce tumor or cancer cells. The subject comprises, but not limited to, mammals and non-mammals. The subject may be a human being. The subject may have or is suspect of having a cancer and / or a tumor. In one aspect, the subject is having or suspected of having breast cancer, lung cancer, colorectal cancer, colon cancer, skin cancer, bladder cancer, kidney cancer, or prostate cancer.

[0055] The present disclosure encompasses TRAIL conjugates that are capable of inducing tumor DNA strand scissions, such as upon oxidation at the DNA backbones. Reactive oxygen species are widely generated in biological systems. Intracellular production of active oxygen species, such as radical -OH, 02“ and H2O2, is associated with, among other things, the arrest of cell proliferation. Appropriate oxygen activity modulation may promote chemical reactions to trigger DNA scission in tumor cells, to degrade tumor polypeptides and polynucleotides by converting26106498195 1Attorney Docket No: 119289-861234 tumor proteins into peroxides that cleave the DNA backbone. Similarly, generation of oxidative stress in response to various external stimuli has been implicated in the activation of transcription factors and to the triggering of apoptosis. The present disclosure encompasses use of the TRAIL conjugates to initiate apoptosis signaling leading eventually to tumor cell death, and / or to the activation of several protooncogenes, and / or the activation of tumor suppressor genes.

[0056] The present disclosure provides methods of use by administering to a subject in need thereof an effective amount of the TRAIL conjugates and / or compositions thereof. Such use can be a therapeutic use, a diagnostic use, an imaging use, a theragnostic use, or any combination thereof. Such effective amount the TRAIL conjugate, or the compositions thereof, may be a pharmaceutical effective amount, a diagnostic effective amount, an imaging effective amount, a theragnostic effective amount. The subject, such as a human being or other mammals, may be having or suspected of having a cancer or a tumor. In one aspect, the subject is having or suspected of having breast cancer, lung cancer, colorectal cancer, colon cancer, skin cancer, bladder cancer, kidney cancer, or prostate cancer.

[0057] The present disclosure also provides a theragnostic method by administering the TRAIL conjugates and / or their compositions thereof to a subject. Theragnostic method is a treatment using diagnostic imaging to identify if target receptors are present on cancer cells, followed by precision radiation treatment that target these receptors. One advantage of the TRAIL conjugate in the present disclosure is that they are both diagnostic and therapeutic due to the isotope radiation and intercalating effect.VII. Diseases and Disorders of Tumor or Cancer

[0058] As it will be recognized by individuals skilled in the art, cancer as used throughout the instant disclosure may be one or more neoplasm or cancer, may be a solid tumor cancer and / or a soft tissue cancer. The solid tumor cancer comprises, but not limited to colorectal cancer, pancreatic cancer, primary liver cancers, kidney cancer, ovarian cancer, uterine cancer, lung cancer, breast cancer, prostate cancer, sarcomas, adipose tissue cancer, a subtype or any combination thereof. A soft tissue cancer or soft tissue sarcomas is understood as malignant neoplasms in the27106498195 1Attorney Docket No: 119289-861234 muscles, tendons, fat, blood vessels, lymphatic vessels, nerves and tissues around the joints. Normally, soft tissue sarcomas in adults can form in almost any part of the body, but they are more common in the head, neck, arms, legs, trunk and abdomen. Examples of soft tissue cancer are, but are not limited to, Solitary Fibrous Tumor (TFS), Hemangiopericytoma (HPC), Ewing’s sarcoma, synovial sarcoma, rhabdomyosarcoma, and myxofibrosarcoma. The neoplasm may be malignant or benign, the cancer may be primary or metastatic; the neoplasm or cancer may be early stage or late stage. Non-limiting examples of neoplasms or cancers that may be treated include acute lymphoblastic leukemia, acute myeloid leukemia, adrenocortical carcinoma, AIDS-related cancers, AIDS-related lymphoma, anal cancer, appendix cancer, astrocytomas (childhood cerebellar or cerebral), basal cell carcinoma, bile duct cancer, bladder cancer, bone cancer, brainstem glioma, brain tumors (cerebellar astrocytoma, cerebral astrocytoma / malignant glioma, ependymoma, medulloblastoma, supratentorial primitive neuroectodermal tumors, visual pathway and hypothalamic gliomas), breast cancer, bronchial adenomas / carcinoids, Burkitt lymphoma, carcinoid tumors (childhood, gastrointestinal), carcinoma of unknown primary, central nervous system lymphoma (primary), cerebellar astrocytoma, cerebral astrocytoma / malignant glioma, cervical cancer, childhood cancers, chronic lymphocytic leukemia, chronic myelogenous leukemia, chronic myeloproliferative disorders, colon cancer, cutaneous T-cell lymphoma, desmoplastic small round cell tumor, endometrial cancer, ependymoma, esophageal cancer, Ewing’s sarcoma in the Ewing family of tumors, extracranial germ cell tumor (childhood), extragonadal germ cell tumor, extrahepatic bile duct cancer, eye cancers (intraocular melanoma, retinoblastoma), gallbladder cancer, gastric (stomach) cancer, gastrointestinal carcinoid tumor, gastrointestinal stromal tumor, germ cell tumors (childhood extracranial, extragonadal, ovarian), gestational trophoblastic tumor, gliomas (adult, childhood brain stem, childhood cerebral astrocytoma, childhood visual pathway and hypothalamic), gastric carcinoid, hairy cell leukemia, head and neck cancer, hepatocellular (liver) cancer, Hodgkin lymphoma, hypopharyngeal cancer, hypothalamic and visual pathway glioma (childhood), intraocular melanoma, islet cell carcinoma, Kaposi sarcoma, kidney cancer (renal cell cancer), laryngeal cancer, leukemias (acute lymphoblastic, acute myeloid, chronic lymphocytic, chronic myelogenous, hairy cell), lip and oral cavity cancer, liver cancer (primary), lung cancers (non-small cell, small cell), lymphomas28106498195 1Attorney Docket No: 119289-861234(AIDS-related, Burkitt, cutaneous T-cell, Hodgkin, non-Hodgkin, primary central nervous system), macroglobulinemia (Waldenstrom), malignant fibrous histiocytoma of bone / osteosarcoma, medulloblastoma (childhood), melanoma, intraocular melanoma, Merkel cell carcinoma, mesotheliomas (adult malignant, childhood), metastatic squamous neck cancer with occult primary, mouth cancer, multiple endocrine neoplasia syndrome (childhood), multiple myeloma / plasma cell neoplasm, mycosis fungoides, myelodysplastic syndromes, myelodysplastic / myeloproliferative diseases, myelogenous leukemia (chronic), myeloid leukemias (adult acute, childhood acute), multiple myeloma, myeloproliferative disorders (chronic), nasal cavity and paranasal sinus cancer, nasopharyngeal carcinoma, neuroblastoma, nonHodgkin lymphoma, non-small cell lung cancer, oral cancer, oropharyngeal cancer, osteosarcoma / malignant fibrous histiocytoma of bone, ovarian cancer, ovarian epithelial cancer (surface epithelial-stromal tumor), ovarian germ cell tumor, ovarian low malignant potential tumor, pancreatic cancer, pancreatic cancer (islet cell), paranasal sinus and nasal cavity cancer, parathyroid cancer, penile cancer, pharyngeal cancer, pheochromocytoma, pineal astrocytoma, pineal germinoma, pineoblastoma and supratentorial primitive neuroectodermal tumors (childhood), pituitary adenoma, plasma cell neoplasia, pleuropulmonary blastoma, primary central nervous system lymphoma, prostate cancer, rectal cancer, renal cell carcinoma (kidney cancer), renal pelvis and ureter transitional cell cancer, retinoblastoma, rhabdomyosarcoma (childhood), salivary gland cancer, sarcoma (Ewing family of tumors, Kaposi, soft tissue, uterine), Sezary syndrome, skin cancers (nonmelanoma, melanoma), skin carcinoma (Merkel cell), small cell lung cancer, small intestine cancer, soft tissue sarcoma, squamous cell carcinoma, squamous neck cancer with occult primary (metastatic), stomach cancer, supratentorial primitive neuroectodermal tumor (childhood), T-Cell lymphoma (cutaneous), testicular cancer, throat cancer, thymoma (childhood), thymoma and thymic carcinoma, thyroid cancer, thyroid cancer (childhood), transitional cell cancer of the renal pelvis and ureter, trophoblastic tumor (gestational), unknown primary site (adult, childhood), ureter and renal pelvis transitional cell cancer, urethral cancer, uterine cancer (endometrial), uterine sarcoma, vaginal cancer, visual pathway and hypothalamic glioma (childhood), vulvar cancer, Waldenstrom macroglobulinemia, and Wilms tumor (childhood). The cancer stages include advanced-stage, unresectable, or metastatic solid tumor cancers, including in subjects that have failed, or become intolerant,29106498195 1Attorney Docket No: 119289-861234 resistant, or refractory to an existing cancer therapy. The current methods may apply to cancer of various stages and may apply to a cancer subject comorbidity with other diseases or disorders.

[0059] The method comprises administering a therapeutically effective, a theragnostically effective, and / or diagnostic effective amount of TRAIL conjugates or their compositions to a subject when the subject is determined to have, is having, or is suspecting of having anyone of the above disclosed conditions or is determined to an eligible candidate for the therapy.VIII. Compositions, Dosage Forms and Administration Regimen

[0060] One aspect of the disclosure encompasses a pharmaceutical composition or formulation for delivery of the TRAIL conjugate. A pharmaceutical composition or formulation comprises an effective amount of the active, and any pharmaceutically acceptable salt thereof.

[0061] Pharmaceutically acceptable salts of the TRAIL conjugate comprise, without limitation, acetate, aspartate, benzoate, bitartrate, citrate, formate, gluconate, glucuronate, glutamate, fumarate, hydrochloride, hydrobromide, hydroiodide, hypophosphite, isobutyrate, isocitrate, lactate, malate, maleate, meconate, methylbromide, methanesulfonate, monohydrate, mucate, nitrate, oxalate, phenylpropionate, phosphate, phthalate, propionate, pyruvate, salicylate, stearate, succinate, sulfate, tannate, tartrate, terephthalate, valerate, and the like.

[0062] The amount of TRAIL conjugate in the composition may be a therapeutically effective amount. As used herein “therapeutically effective amount” or “therapeutically effective dosage” refers to an amount that is effective to achieve a desired therapeutic result. In some embodiments, the desired therapeutic result is DNA scissions and / or tumor cell death. For example, the therapeutically effective amount may be an amount that induces DNA scission by at least 10%, preferably 20% or more, preferably 25% or more, preferably 30% or more, preferably 35% or more, preferably 40% or more, preferably 45% or more, preferably 50% or more, preferably 60% or more, preferably 70% or more, preferably 80% or more, preferably 90% or more.30106498195 1Attorney Docket No: 119289-861234

[0063] In some cases, the amount of the TRAIL conjugate in the composition is more than about 1 pg. For example, the TRAIL composition may comprise the TRAIL conjugate in about 2 pg or more, about 5 pg or more, about 10 pg or more, about 100 pg or more, about 500 pg or more, about 1000 pg or more, about 1500 pg or more, about 2000 pg or more, about 2500 pg or more, about 3000 pg or more, about 3500 pg or more, about 4000 pg or more, about 4500 pg or more, about 5000 pg or more, about 5500 pg or more, about 6000 pg or more, about 6500 pg or more, about 7000 pg or more, about 7500 pg or more, about 8000 pg or more, about 8500 pg or more, about 9000 pg or more, about 9500 pg or more, about 10 mg or more, about 20 mg or more, about 30 mg or more, about 40 mg or more, about 50 mg or more, about 60 mg or more, about 70 mg or more, about 80 mg or more, about 90 mg or more, about 100 mg or more, about 150 mg or more, about 200 mg or more, about 250 mg or more, about 300 mg or more, about 350 mg or more, about 400 mg or more, about 450 mg or more, about 500 mg or more, about 550 mg or more, about 600 mg or more, about 650 mg or more, about 700 mg or more, about 800 mg or more, about 900 mg or more, or about 1 g or more.

[0064] Additionally or alternatively, the amount of TRAIL conjugate in the TRAIL composition may be about 0.01 wt.% to about 95 wt.%, about 0.1 wt.% to about 95 wt.%, about 1 wt.% to about 95 wt.%, about 5 wt.% to about 95 wt.%, about 10 wt.% to about 95 wt.%, about 15 wt.% to about 95 wt.%, about 20 wt.% to about 95 wt.%, about 30 wt.% to about 95 wt.%, about 40 wt.% to about 95 wt.%, about 50 wt.% to about 95 wt.%, about 60 wt.% to about 95 wt.%, about 70 wt.% to about 95 wt.%, about 80 wt.% to about 95 wt.%; about 0.01 wt.% to about 85 wt.%, about 0.1 wt.% to about 85 wt.%, about 1 wt.% to about 85 wt.%, about 5 wt.% to about 85 wt.%, about 10 wt.% to about 85 wt.%, about 15 wt.% to about 85 wt.%, about 20 wt.% to about 85 wt.%, about 30 wt.% to about 85 wt.%, about 40 wt.% to about 85 wt.%, about 50 wt.% to about 85 wt.%, about 60 wt.% to about 85 wt.%, about 70 wt.% to about 85 wt.%; about 0.01 wt.% to about 75 wt.% about 0.1 wt.% to about 75 wt.%, about 1 wt.% to about 75 wt.%, about 5 wt.% to about 75 wt.%, about 10 wt.% to about 75 wt.%, about 15 wt.% to about 75 wt.%, about 20 wt.% to about 75 wt.%, about 30 wt.% to about 75 wt.%, about 40 wt.% to about 75 wt.%, about 50 wt.% to about 75 wt.%, about 60 wt.% to about 75 wt.%; about 0.01 wt.% to about 65 wt.%, about 0.1 wt.% to about 65 wt.%, about 1 wt.% to about 65 wt.%, about 5 wt.% to31106498195 1Attorney Docket No: 119289-861234 about 65 wt.%, about 10 wt.% to about 65 wt.%, about 15 wt.% to about 65 wt.%, about 20 wt.% to about 65 wt.%, about 30 wt.% to about 65 wt.%, about 40 wt.% to about 65 wt.%, about 50 wt.% to about 65 wt.%; about 0.01 wt.% to about 55 wt.%, about 0.1 wt.% to about 55 wt.%, about 1 wt.% to about 55 wt.%, about 5 wt.% to about 55 wt.%, about 10 wt.% to about 55 wt.%, about 15 wt.% to about 55 wt.%, about 20 wt.% to about 55 wt.%, about 30 wt.% to about 55 wt.%, about 40 wt.% to about 55 wt.%; about 0.01 wt.% to about 45 wt.%, about 0.1 wt.% to about 45 wt.%, about 1 wt.% to about 45 wt.%, about 5 wt.% to about 45 wt.%, about 10 wt.% to about 45 wt.%, about 15 wt.% to about 45 wt.%, about 20 wt.% to about 45 wt.%, about 30 wt.% to about 45 wt.%; about 0.01 wt.% to about 35 wt.%, about 0.1 wt.% to about 35 wt.%, about 1 wt.% to about 35 wt.%, about 5 wt.% to about 35 wt.%, about 10 wt.% to about 35 wt.%, about 15 wt.% to about 35 wt.%, about 20 wt.% to about 35 wt.%; about 0.01 wt.% to about 25 wt.%, about 0.1 wt.% to about 25 wt.%, about 1 wt.% to about 25 wt.%, about 5 wt.% to about 25 wt.%, about 10 wt.% to about 25 wt.%; about 0.1 wt.% to about 15 wt.%, about 1 wt.% to about 15 wt.%, about 5 wt.% to about 15 wt.%, about 10 wt.% to about 15 wt.%; about 0.01 wt.% to about 25 wt.%, about 0.1 wt.% to about 10 wt.%, about 1 wt.% to about 10 wt.%, or about 5 wt.% to about 10 wt.%, including ranges and subranges thereof, based on the total weight of the TRAIL composition.

[0065] Alternatively or additionally, the TRAIL composition may include other anti-tumor or anti-cancer actives, or any combination thereof in an amount of about 0.01 wt.% to about 95 wt.%, about 0.1 wt.% to about 95 wt.%, about 1 wt.% to about 95 wt.%, about 5 wt.% to about 95 wt.%, about 10 wt.% to about 95 wt.%, about 15 wt.% to about 95 wt.%, about 20 wt.% to about 95 wt.%, about 30 wt.% to about 95 wt.%, about 40 wt.% to about 95 wt.%, about 50 wt.% to about 95 wt.%, about 60 wt.% to about 95 wt.%, about 70 wt.% to about 95 wt.%, about 80 wt.% to about 95 wt.%; about 0.01 wt.% to about 85 wt.%, about 0.1 wt.% to about 85 wt.%, about 1 wt.% to about 85 wt.%, about 5 wt.% to about 85 wt.%, about 10 wt.% to about 85 wt.%, about 15 wt.% to about 85 wt.%, about 20 wt.% to about 85 wt.%, about 30 wt.% to about 85 wt.%, about 40 wt.% to about 85 wt.%, about 50 wt.% to about 85 wt.%, about 60 wt.% to about 85 wt.%, about 70 wt.% to about 85 wt.%; about 0.01 wt.% to about 75 wt.%, about 0.1 wt.% to about 75 wt.%, about 1 wt.% to about 75 wt.%, about 5 wt.% to about 75 wt.%, about 10 wt.% to about 75 wt.%, about 1532106498195 1Attorney Docket No: 119289-861234 wt.% to about 75 wt.%, about 20 wt.% to about 75 wt.%, about 30 wt.% to about 75 wt.%, about 40 wt.% to about 75 wt.%, about 50 wt.% to about 75 wt.%, about 60 wt.% to about 75 wt.%; about 0.01 wt.% to about 65 wt.% about 0.1 wt.% to about 65 wt.%, about 1 wt.% to about 65 wt.%, about 5 wt.% to about 65 wt.%, about 10 wt.% to about 65 wt.%, about 15 wt.% to about 65 wt.%, about 20 wt.% to about 65 wt.%, about 30 wt.% to about 65 wt.%, about 40 wt.% to about 65 wt.%, about 50 wt.% to about 65 wt.%; about 0.01 wt.% to about 55 wt.%, about 0.1 wt.% to about 55 wt.%, about 1 wt.% to about 55 wt.%, about 5 wt.% to about 55 wt.%, about 10 wt.% to about 55 wt.%, about 15 wt.% to about 55 wt.%, about 20 wt.% to about 55 wt.%, about 30 wt.% to about 55 wt.%, about 40 wt.% to about 55 wt.%; about 0.01 wt.% to about 45 wt.%, about 0.1 wt.% to about 45 wt.%, about 1 wt.% to about 45 wt.%, about 5 wt.% to about 45 wt.%, about 10 wt.% to about 45 wt.%, about 15 wt.% to about 45 wt.%, about 20 wt.% to about 45 wt.%, about 30 wt.% to about 45 wt.%; about 0.01 wt.% to about 35 wt.%, about 0.1 wt.% to about 35 wt.%, about 1 wt.% to about 35 wt.%, about 5 wt.% to about 35 wt.%, about 10 wt.% to about 35 wt.%, about 15 wt.% to about 35 wt.%, about 20 wt.% to about 35 wt.%; about 0.01 wt.% to about 25 wt.%, about 0.1 wt.% to about 25 wt.%, about 1 wt.% to about 25 wt.%, about 5 wt.% to about 25 wt.%, about 10 wt.% to about 25 wt.%; about 0.01 wt.% to about 15 wt.% about 0.1 wt.% to about 15 wt.%, about 1 wt.% to about 15 wt.%, about 5 wt.% to about 15 wt.%, about 10 wt.% to about 15 wt.%; about 0.01 wt.% to about 10 wt.% about 0.1 wt.% to about 10 wt.%, about 1 wt.% to about 10 wt.%, or about 5 wt.% to about 10 wt.%, including ranges and subranges thereof, based on the total weight of the TRAIL composition.

[0066] Additionally or alternatively, the TRAIL composition may comprise more than one active including at least one TRAIL conjugate. In one aspect, the composition comprises one TRAIL conjugate and one known anti-cancer agent. The different actives in the formulation may have a weight ratio ranges from 1 :100 to 100:1 , 1 :50 to 50:1 , 1 :20 to 20:1 1 :10 to 10:1 , 1 :9 to 10:1 , 1 :8 to 10:1 , 1 :7 to 10:1 , 1 :6 to 10:1 , 1 :5 to 10:1 , 1 :4 to 10:1 , 1 :3 to 10:1 , 1 :2 to 10:1 , 1 :1 to 10:1 , 1 :10 to 9:1 , 1 :10 to 8:1 , 1:10 to 7:1 , 1 :10 to 6:1 , 1 :10 to 5:1 , 1:10 to 4:1, 1 :10 to 3:1, 1 :10 to 2:1 , or 1 :10 to 1 :1 , including ranges and subranges thereof. In one instance, the weight ratio of the one TRAIL conjugate to the one known anti-cancer agent is about 1 :10 to 10:1 , about 1 :9 to 10:1 , about 1 :8 to 10:1 , about 1 :7 to 10:1 , about 1 :6 to 10:1 , about33106498195 1Attorney Docket No: 119289-8612341 :5 to 10:1 , about 1 :4 to 10:1 , about 1 :3 to 10:1 , about 1 :2 to 10:1 , about 1 :1 to 10:1 , about 1 :10 to 9:1 , about 1 :10 to 8:1 , about 1 :10 to 7:1 , about 1 :10 to 6:1 , about 1 :10 to 5:1 , about 1 :10 to 4:1 , about 1 :10 to 3:1 , about 1 :10 to 2:1 , or about 1 :10 to 1 :1 , including ranges and subranges thereof.

[0067] The amount of TRAIL conjugate in the composition, may be a diagnostically effective amount. As used herein “diagnostically effective amount” refers to an amount that is effective to achieve a detecting limit in a living body. In some embodiments, the desired diagnostically effective amount may be an amount that sufficient to be detected in an imaging tool, including but not limited to PET, MRI, or CT. The amount of TRAIL conjugate in the composition may be more than about 1 pg. For example, the amount is at or about 2 pg or more, about 5 pg or more, about 10 pg or more, about 100 pg or more, about 500 pg or more, about 1000 pg or more, about 1500 pg or more, about 2000 pg or more, about 2500 pg or more, about 3000 pg or more, about 3500 pg or more, about 4000 pg or more, about 4500 pg or more, about 5000 pg or more, about 5500 pg or more, about 6000 pg or more, about 6500 pg or more, about 7000 pg or more, about 7500 pg or more, about 8000 pg or more, about 8500 pg or more, about 9000 pg or more, about 9500 pg or more, about 10 mg or more, about 20 mg or more.

[0068] The amount of TRAIL conjugate in the composition may be a theragonistically effective amount. As used herein “theragnostically effective amount” refers to an amount that is sufficient to identify if target receptors are present on cancer cells, followed by precision radiation treatment that target these receptors. In some embodiments, the desired theragnostically effective amount may be an amount that sufficient to be coupled with in an imaging tool, including but not limited to PET, MRI, or CT. The amount of compound of Formula (I) through Formula (VI), such as metal chelates in an isotope form, may be more than about 1 pg. For example, the amount is at or about 2 pg or more, about 5 pg or more, about 10 pg or more, about 100 pg or more, about 500 pg or more, about 1000 pg or more, about 1500 pg or more, about 2000 pg or more, about 2500 pg or more, about 3000 pg or more, about 3500 pg or more, about 4000 pg or more, about 4500 pg or more, about 5000 pg or more, about 5500 pg or more, about 6000 pg or more, about 6500 pg or more, about 7000 pg or more, about 7500 pg or more, about 8000 pg or more, about34106498195 1Attorney Docket No: 119289-8612348500 pg or more, about 9000 g or more, about 9500 pg or more, about 10 mg or more, about 20 mg or more.

[0069] The TRAIL composition can be formulated and administered to a subject by several different routes and means. For instance, the composition can generally be administered parenterally, intraperitoneally, intravascularly, topically, transdermally, subcutaneously, or intrapulmonarily in dosage unit forms containing conventional nontoxic pharmaceutically or diagnositically acceptable adjuvants, carriers, excipients, and vehicles as desired. The term parenteral as used herein includes subcutaneous, intravenous, intramuscular, intrathecal, or intrasternal, or infusion techniques. In one aspect, the therapeutically effective amount of TRAIL conjugate can range from about 0.5 mcg to about 200 mg, about 1 mcg to about 180 mg, about 5 mcg to about 150 mg, or about 10 mcg to about 120 mg. The therapeutically effective amount of TRAIL conjugate can range from about 0.5 mcg / day to about 100 mg / day, from about 1 mcg to about 60 mg / day, from about 20 mcg to about 50 mg / day, from about 20 mcg to about 30 mg / day, or from about 15 mcg to about 25 mg / day. Administering the TRAIL composition may provide the TRAIL conjugate at blood levels of at or about 1 ng / ml, at or about 2 ng / ml, at or about 4 ng / ml, at or about 5 ng / ml, at or about 6 ng / ml, at or about 8 ng / ml, at or about 10 ng / ml, at or about 12 ng / ml, at or about 15 ng / ml, at or about 20 ng / ml, at or about 22 ng / ml, at or about 24 ng / ml, at or about 26 ng / ml, at or about 28 ng / ml, at or about 30 ng / ml, at or about 32 ng / ml, at or about 34 ng / ml, at or about 36 ng / ml, at or about 38 ng / ml, at or about 40 ng / ml, at or about 42 ng / ml, at or about 44 ng / ml, 46 ng / ml, at or about 48 ng / ml, at or about 50 ng / ml.

[0070] A TRAIL composition can be administered to the subject daily or more than once daily. For example, the composition can be administered to the subject once, twice or three times per day. The duration of each administration can vary from few seconds to about several hours, such as in infusion administration. Further, the TRAIL composition can be administered every 2, 3, 4, 5, 6, 7, 14, or every 30 days. Such treatment regimen is subject to alter and adjust based on subject’s individual needs and response sensitivity. The TRAIL composition can be administered over a period ranging from about 1 day to about 1 year, from about 1 day to about 1 week, from about 3 days to about 1 month, from about 2 weeks to about 6 months, or from about 2 months to about 4 months. The TRAIL composition can also be administered35106498195 1Attorney Docket No: 119289-861234 over a period of about 1 day, about 7 days, about 30 days, about 60 days, about 120 days, or about 180 days or more. In some aspect, TRAIL composition is administered over a period of about 57 weeks, about 148 weeks, about 208 weeks, indefinitely, or until resolution of the condition being treated.

[0071] A pharmaceutical formulation comprises one or more pharmaceutically acceptable excipients. Non-limiting examples of excipients include chemical enhancers, humectants, pressure sensitive adhesives, antioxidants, solubilizers, thickening agents, plasticizers, adjuvants, carriers, excipients, vehicles, coatings, and any combinations thereof. One or more excipients can be selected for oral, transdermal, parenteral, intraperitoneal, intravascular, subcutaneous, by inhalation spray, rectal, or intrapulmonary administration.

[0072] A TRAIL composition can in general be formulated for improving patient compliance, preventing a subject from removing the drug-delivery device. For instance, the TRAIL composition could be formulated for improved patient compliance and preventing removal of a drug-delivery device by providing formulations for extended delivery. Extended delivery can range for periods ranging from more than one day, to months. This may be especially relevant for patients with compromised cognitive and / or motor-control abilities. Extended delivery for periods can range from about 1 day to about 1 year, from about 1 day to about 1 week, from about 3 days to about 1 month, from about 2 weeks to about 6 months, or from about 2 months to about 4 months.

[0073] Extended-release formulations could be used for substantially continuous delivery of drug at a preselected rate. For example, the TRAIL composition can be delivered at a rate of from about 1 mg to about 100 mg / day, from about 40 to about 60 mg / day, or from about 10 to about 30 gm / day. Appropriate amounts of TRAIL composition can be readily determined by the ordinarily skilled artisan based upon, for example, the intended duration of administration of the active by the extended-release formulation, the delivery mechanism, the specific formulation, and the relative potency of the drug among other factors. / . Binders

[0074] Non-limiting examples of binders suitable for the formulations of various aspects include starches, pregelatinized starches, gelatin,36106498195 1Attorney Docket No: 119289-861234 polyvinylpyrrolidone, cellulose, methylcellulose, sodium carboxymethylcellulose, ethylcellulose, polyacrylamides, polyvinyloxoazolidone, polyvinylalcohols, C12-C18 fatty acid alcohols, polyethylene glycol, polyols, saccharides, oligosaccharides, polypeptides, oligopeptides, and combinations thereof. The polypeptide may be any arrangement of amino acids ranging from about 100 to about 300,000 Daltons.

[0075] The binder can be introduced into the mixture by granulating into a solid form, including but not limited to a crystal, a particle, a powder, or any other finely divided solid form known in the art. Alternatively, the binder can be dissolved or suspended in a solvent and sprayed onto the mixture in a granulation device as a binder fluid during granulation. / / . Diluents

[0076] Non-limiting examples of diluents (also referred to as “fillers” or “thinners”) include carbohydrates, inorganic compounds, and biocompatible polymers, such as polyvinylpyrrolidone (PVP). Other non-limiting examples of diluents include dibasic calcium sulfate, tribasic calcium sulfate, starch, calcium carbonate, magnesium carbonate, microcrystalline cellulose, dibasic calcium phosphate, tribasic calcium phosphate, magnesium carbonate, magnesium oxide, calcium silicate, talc, modified starches, saccharides such as sucrose, dextrose, lactose, microcrystalline cellulose, fructose, xylitol, and sorbitol, polyhydric alcohols; starches; pre-manufactured direct compression diluents; and mixtures of any of the foregoing.Hi. Disintegrants

[0077] Disintegrants can be effervescent or non-effervescent. Non-limiting examples of non-effervescent disintegrants include starches such as corn starch, potato starch, pregelatinized and modified starches thereof, sweeteners, clays, such as bentonite, micro-crystalline cellulose, alginates, sodium starch glycolate, gums such as agar, guar, locust bean, karaya, pectin, and tragacanth. Suitable effervescent disintegrants include but are not limited to sodium bicarbonate in combination with citric acid, and sodium bicarbonate in combination with tartaric acid. iv. Preservatives37106498195 1Attorney Docket No: 119289-861234

[0078] Non-limiting examples of preservatives include, but are not limited to, ascorbic acid and its salts, ascorbyl palmitate, ascorbyl stearate, anoxomer, N- acetylcysteine, benzyl isothiocyanate, m-aminobenzoic acid, o-aminobenzoic acid, p- aminobenzoic acid (PABA), butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), caffeic acid, canthaxantin, alpha-carotene, beta-carotene, beta-caraotene, beta-apo-carotenoic acid, camosol, carvacrol, catechins, cetyl gallate, chlorogenic acid, citric acid and its salts, clove extract, coffee bean extract, p-coumaric acid, 3,4-dihydroxybenzoic acid, N,N’-diphenyl-p-phenylenediamine (DPPD), dilauryl thiodipropionate, distearyl thiodipropionate, 2,6-di-tert-butylphenol, dodecyl gallate, edetic acid, ellagic acid, erythorbic acid, sodium erythorbate, esculetin, esculin, 6-ethoxy-1 ,2-dihydro-2,2,4-trimethylquinoline, ethyl gallate, ethyl maltol, ethylenediaminetetraacetic acid (EDTA), eucalyptus extract, eugenol, ferulic acid, flavonoids (e.g., catechin, epicatechin, epicatechin gallate, epigallocatechin (EGC), epigallocatechin gallate (EGCG), polyphenol epigallocatechin-3-gallate), flavones (e.g., apigenin, chrysin, luteolin), flavonols (e.g., datiscetin, myricetin, daemfero), flavanones, fraxetin, fumaric acid, gallic acid, gentian extract, gluconic acid, glycine, gum guaiacum, hesperetin, alpha-hydroxybenzyl phosphinic acid, hydroxycinammic acid, hydroxyglutaric acid, hydroquinone, N-hydroxysuccinic acid, hydroxytryrosol, hydroxyurea, rice bran extract, lactic acid and its salts, lecithin, lecithin citrate; R-alpha-lipoic acid, lutein, lycopene, malic acid, maltol, 5-methoxy tryptamine, methyl gallate, monoglyceride citrate; monoisopropyl citrate; morin, betanaphthoflavone, nordihydroguaiaretic acid (NDGA), octyl gallate, oxalic acid, palmityl citrate, phenothiazine, phosphatidylcholine, phosphoric acid, phosphates, phytic acid, phytylubichromel, pimento extract, propyl gallate, polyphosphates, quercetin, trans-resveratrol, rosemary extract, rosmarinic acid, sage extract, sesamol, silymarin, sinapic acid, succinic acid, stearyl citrate, syringic acid, tartaric acid, thymol, tocopherols (i.e., alpha-, beta-, gamma- and delta-tocopherol), tocotrienols (i.e., alpha-, beta-, gamma- and delta-tocotrienols), tyrosol, 38hosphor acid, 2,6-di- tert-butyl-4-hydroxymethylphenol (i.e., lonox 100), 2,4-(tris-3’,5’-bi-tert-butyl-4 - hydroxybenzyl)-mesitylene (i.e., lonox 330), 2,4,5-trihydroxybutyrophenone, ubiquinone, tertiary butyl hydroquinone (TBHQ), thiodipropionic acid, trihydroxy butyrophenone, tryptamine, tyramine, uric acid, vitamin K and derivates, vitamin Q10, wheat germ oil, zeaxanthin, or combinations thereof.38106498195 1Attorney Docket No: 119289-861234 Flavor-modifying agents

[0079] Suitable flavor-modifying agents include flavorants, taste-masking agents, sweeteners, and the like. Flavorants include, but are not limited to, synthetic flavor oils and flavoring aromatics and / or natural oils, extracts from plants, leaves, flowers, fruits, and combinations thereof. Other non-limiting examples of flavors include cinnamon oils, oil of Wintergreen, peppermint oils, clover oil, hay oil, anise oil, eucalyptus, vanilla, citrus oils such as lemon oil, orange oil, grape and grapefruit oil, fruit essences including apple, peach, pear, strawberry, raspberry, cherry, plum, pineapple, and apricot.

[0080] Taste-masking agents include but are not limited to cellulose hydroxypropyl ethers (HPC) such as Klucel®, Nisswo HPC and PrimaFlo HP22; low- substituted hydroxypropyl ethers (L-HPC); cellulose hydroxypropyl methyl ethers (HPMC) such as Seppifilm-LC, Pharmacoat®, Metolose SR, Opadry YS, PrimaFlo, MP3295A, Benecel MP824, and Benecel MP843; methylcellulose polymers such as Methocel® and Metolose®; Ethylcelluloses (EC) and mixtures thereof such as E461 , Ethocel®, Aqualon®-EC, Surelease; Polyvinyl alcohol (PVA) such as Opadry AMB; hydroxyethylcelluloses such as Natrosol®; carboxymethylcelluloses and salts of carboxymethylcelluloses (CMC) such as Aualon®-CMC; polyvinyl alcohol and polyethylene glycol co-polymers such as Kollicoat IR®; monoglycerides (Myverol), triglycerides (KLX), polyethylene glycols, modified food starch, acrylic polymers and mixtures of acrylic polymers with cellulose ethers such as Eudragit® EPO, Eudragit® RD100, and Eudragit® E100; cellulose acetate phthalate; sepifilms such as mixtures of HPMC and stearic acid, cyclodextrins, and mixtures of these materials. In other aspects, additional taste-masking agents contemplated are those described in U.S. Pat. Nos. 4,851 ,226; 5,075,114; and 5,876,759, each of which is hereby incorporated by reference in its entirety.

[0081] Non-limiting examples of sweeteners include glucose (corn syrup), dextrose, invert sugar, fructose, and mixtures thereof (when not used as a carrier); saccharin and its various salts such as the sodium salt; dipeptide sweeteners such as aspartame; dihydrochalcone compounds, glycyrrhizin; Stevia rebaudiana (Stevioside); chloro derivatives of sucrose such as sucralose; sugar alcohols such as sorbitol, mannitol, sylitol, hydrogenated starch hydrolysates and the synthetic39106498195 1Attorney Docket No: 119289-861234 sweetener 3,6-dihydro-6-methyl-1 ,2,3-oxathiazin-4-one-2,2-dioxide, particularly the potassium salt (acesulfame-K), and sodium and calcium salts thereof. vi. Lubricants and c / lidants

[0082] The lubricant compositions may be utilized to lubricate ingredients that form a pharmaceutical composition. As a glidant, the lubricant facilitates removal of solid dosage forms during the manufacturing process. Non-limiting examples of lubricants and glidants include magnesium stearate, calcium stearate, zinc stearate, hydrogenated vegetable oils, sterotex, polyoxyethylene monostearate, talc, polyethylene glycol, sodium benzoate, sodium lauryl sulfate, magnesium lauryl sulfate, and light mineral oil. The pharmaceutical composition will generally comprise from about 0.01 % to about 10% by weight of a lubricant. In some aspects, the pharmaceutical composition will comprise from about 0.1 % to about 5% by weight of a lubricant. In a further aspect, the pharmaceutical composition will comprise from about 0.5% to about 2% by weight of a lubricant. vii. Dispersants

[0083] Dispersants may include but are not limited to starch, alginic acid, polyvinylpyrrolidones, guar gum, kaolin, bentonite, purified wood cellulose, sodium starch glycolate, isoamorphous silicate, and microcrystalline cellulose as high hydrophilic-lipophilic balance (HLB) emulsifier surfactants. viii. Colorants

[0084] Depending upon the aspect of the disclosure, it may be desirable to include a coloring agent. Suitable color additives include but are not limited to food, drug and cosmetic colors (FD&C), drug and cosmetic colors (D&C), or external drug and cosmetic colors (Ext. D&C). These colors or dyes, along with their corresponding lakes, and certain natural and derived colorants, may be suitable for use in various aspects of the disclosure. ix. pH modifiers

[0085] Non-limiting examples of pH modifiers include citric acid, acetic acid, tartaric acid, malic acid, fumaric acid, lactic acid, phosphoric acid, sorbic acid, benzoic acid, sodium carbonate and sodium bicarbonate.40106498195 1Attorney Docket No: 119289-861234 x. Chelating agents

[0086] A chelating agent may be included as an excipient to immobilize oxidative groups, including but not limited to metal ions, to inhibit the oxidative degradation of the morphinan by these oxidative groups. Non-limiting examples of chelating agents include lysine, methionine, glycine, gluconate, polysaccharides, glutamate, aspartate, and disodium ethylenediaminetetraacetate (Na2EDTA). xi. Antimicrobial agents

[0087] An antimicrobial agent may be included as an excipient to minimize the degradation of the compound according to this disclosure by microbial agents, including but not limited to bacteria and fungi. Non-limiting examples of antimicrobials include parabens, chlorobutanol, phenol, calcium propionate, sodium nitrate, sodium nitrite, Na2EDTA, and sulfites including but not limited to sulfur dioxide, sodium bisulfite, and potassium hydrogen sulfite. xii. Release-controlling polymers

[0088] Release-controlling polymers may be included in the various aspects of the solid dosage pharmaceutical compositions incorporating compounds according to this disclosure. In one aspect, the release-controlling polymers may be used as a tablet coating. In other aspects, including but not limited to bilayer tablets, a releasecontrolling polymer may be mixed with the granules and other excipients prior to the formation of a tablet by a known process including but not limited to compression in a tablet mold. Suitable release-controlling polymers include but are not limited to hydrophilic polymers and hydrophobic polymers.

[0089] Suitable hydrophilic release-controlling polymers include, but are not limited to, cellulose acetate, cellulose diacetate, cellulose triacetate, cellulose ethers, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methylcellulose, microcrystalline cellulose, nitrocellulose, crosslinked starch, agar, casein, chitin, collagen, gelatin, maltose, mannitol, maltodextrin, pectin, pullulan, sorbitol, xylitol, polysaccharides, ammonia alginate, sodium alginate, calcium alginate, potassium alginate, propylene glycol alginate, alginate sodium carmellose, calcium carmellose, carrageenan, fucoidan, furcellaran, carrageen’s gum, ghatti gum, guar gum, karaya gum, locust bean gum, okra gum, tragacanth gum, scleroglucan gum, xanthan gum, hypnea, laminaran, acrylic polymers, acrylate polymers, carboxyvinyl polymers,41106498195 1Attorney Docket No: 119289-861234 copolymers of maleic anhydride and styrene, copolymers of maleic anhydride and ethylene, copolymers of maleic anhydride propylene or copolymers of maleic anhydride isobutylene), crosslinked polyvinyl alcohol and poly N-vinyl-2-pyrrolidone, diesters of polyglucan, polyacrylamides, polyacrylic acid, polyamides, polyethylene glycols, polyethylene oxides, poly(hydroxyalkyl methacrylate), polyvinyl acetate, polyvinyl alcohol, polyvinyl chloride, polystyrenes, polyvinylpyrrolidone, anionic and cationic hydrogels, and combinations thereof. x / 77. Coatings

[0090] A solid dosage comprising a compound according to this disclosure may comprise a coating, wherein such a coating may control release of the compound, function as a moisture barrier, or buffer or modify pH. A “control releasing coat” or “controlled release coat” as used herein is defined to mean a functional coat which can for example comprise at least one pH independent polymer, pH dependent polymer (for example enteric or reverse enteric type polymers), soluble polymer, insoluble polymer, lipids, lipidic materials, or combinations thereof. The coating, when applied onto a dosage form, may slow (for example when applied to a normal release matrix dosage form), further slow (for example when applied to a controlled release matrix dosage form) or modify the rate of release of a compound according to this disclosure when applied to an uncoated dosage form. For example, the control releasing coat can be designed such that when the control releasing coat is applied to a dosage form, the dosage form in conjunction with the control releasing coat can exhibit the release of the compound according to this disclosure, such as a “modified-release”, “controlled-release”, “sustained-release”, “extended-release”, “delayed-release”, “prolonged-release,” or combinations thereof. The “control releasing coat” may optionally comprise additional materials that may alter the functionality of the control releasing coat.

[0091] The term “moisture barrier” as used herein is one which impedes or retards the absorption of moisture. Compounds according to this disclosure may be hygroscopic and, as such, may be susceptible to decomposition over time under highly humid conditions. The proportion of the components of the moisture barrier and the amount of the moisture barrier optionally applied onto the control-releasing coating or onto the core are typically such that the moisture barrier does not fall within the USP definition and requirement for an enteric coat. Suitably, the moisture42106498195 1Attorney Docket No: 119289-861234 barrier may comprise an enteric and / or acrylic polymer, suitably an acrylic polymer, optionally a plasticizer, and a permeation enhancer. The permeation enhancer is a hydrophilic substance, which allows water to enter without physical disruption of the coating. The moisture barrier may additionally comprise other conventional inert excipients, which may improve processing of an extended-release formulation.

[0092] Coating and matrix materials which may be used in accordance with the present disclosure are those known in the art for use in controlled-release formulations, such as synthetic polymers of the polyvinyl type, e.g., polyvinylchloride, polyvinylacetate and copolymers thereof, polyvinylalcohol, and polyvinylpyrrolidone; synthetic polymers of the polyethylene type, e.g., polyethylene and polystyrene; acrylic acid polymers; biopolymers or modified biopolymers, such as cellulosic polymers, shellac and gelatin; fats, oils, higher fatty acids and higher alcohols (i.e., acids and alcohols containing alkyl chains of at least 10 carbon atoms), for example aluminum monostearate, cetylalcohol, hydrogenated beef tallow, hydrogenated castor oil, 12-hydroxystearyl alcohol, glyceryl mono- or dipalmitate; glyceryl mono-, di- or tri-stearate; myristyl alcohol, stearic acid, stearyl alcohol, and polyethylene glycols; waxes; sugars and sugar alcohols.

[0093] The pH-buffering properties of a coating may be strengthened by introducing into the coating substances chosen from a group of compounds usually used in antacid formulations, for example magnesium oxide, hydroxide or carbonate, aluminum or calcium hydroxide, carbonate or silicate; composite aluminum / magnesium compounds, for example AI2O3 6MgO CC>2-12H2O, (Mg6Al2(OH)ieCO3-4H2O), MgO Al2O3-2SiO2.nH2O, aluminum bicarbonate coprecipitate or similar compounds; or other pharmaceutically acceptable pH- buffering compounds, for example the sodium, potassium, calcium, magnesium and aluminum salts of phosphoric, carbonic, citric or other suitable, weak, inorganic or organic acids; or suitable organic bases, including basic amino acids; and salts or combinations thereof.

[0094] A pH-dependent coating serves to release the drug in desired areas of the gastrointestinal (Gl) tract, e.g., the stomach or small intestine. When a pH- independent coating is desired, the coating is designed to achieve optimal release regardless of pH-changes in the environmental fluid, e.g., the Gl tract. When the coating is formulated to release a compound according to this disclosure in the43106498195 1Attorney Docket No: 119289-861234 intestines (especially the upper small intestines), the coating is often called an “enteric coating”. A pH-dependent coating may include, but is not limited to, acrylic acid polymers and copolymers, for example polymers formed from acrylic acid, methacrylic acid, methyl acrylate, ammonio methylacrylate, ethyl acrylate, methyl methacrylate and / or ethyl methacrylate (e.g., Eudragit™); cellulosic polymers such as hydroxypropyl cellulose, hydroxyethyl cellulose, hydroxypropyl methyl cellulose, methyl cellulose, ethyl cellulose, cellulose acetate, cellulose acetate phthalate (CAP), cellulose acetate trimellitate, hydroxypropylmethyl cellulose phthalate, hydroxypropylmethyl cellulose succinate and carboxymethylcellulose sodium; shellac (purified lac); vinyl polymers and copolymers such as polyvinyl pyrrolidone, polyvinyl acetate, polyvinylacetate phthalate (PVAP), vinylacetate crotonic acid copolymer, and ethylene-vinyl acetate copolymers; zein; and salts and combinations thereof. ix. Nanoparticles

[0095] The TRAIL compositions may comprise nanoparticles wherein each of the nanoparticles comprises a carrier entity and at least one TRAIL conjugate, wherein the active is arranged on an outside surface of the nanoparticles and wherein the nanoparticles are capable of binding to a predetermined epitope in vivo. In another aspect, provided herein are nanoparticle compositions comprising nanoparticles, wherein each of the nanoparticles comprises at least one TRAIL conjugate, wherein the TRAIL conjugate is arranged on a surface of the nanoparticles such that the binding with subject’s receptors or cells is directed outward from that surface and wherein the nanoparticles are capable of binding to a predetermined epitope in vivo. In other embodiments, the nanoparticles multimerize, e.g. dimerize. Multimerization may be observed as multiples of the weight or size of the unit molecule, e.g. 160 nm particles multimerize to about 320 nm, 480 nm, 640 nm, etc. In some embodiments, less than 20% of the population are multimers. In some embodiments, more than 80% of the population are multimers. x. Liposomes

[0096] In one alternative embodiment, a liposome delivery vehicle may be utilized. Liposomes, depending upon the embodiment, may be used for delivery of a TRAIL composition comprising compounds of Formula (I) or Formula (II), or their metal chelates thereof, in view of their structural and chemical properties. Generally,44106498195 1Attorney Docket No: 119289-861234 liposomes are spherical vesicles with a phospholipid bilayer membrane. The lipid bilayer of a liposome may fuse with other bilayers (e.g., the cell membrane), thus delivering the contents of the liposome to cells.

[0097] Liposomes may be comprised of a variety of diverse types of phospholipids having varying hydrocarbon chain lengths. Phospholipids generally comprise two fatty acids linked through glycerol phosphate to one of a variety of polar groups. Suitable phospholipids include phosphatidic acid (PA), phosphatidylserine (PS), phosphatidylinositol (PI), phosphatidylglycerol (PG), diphosphatidyl glycerol (DPG), phosphatidylcholine (PC), and phosphatidylethanolamine (PE). The fatty acid chains comprising the phospholipids may range from about 6 to about 26 carbon atoms in length, and the lipid chains may be saturated or unsaturated. Suitable fatty acid chains include (common name presented in parentheses) n-dodecanoate (laurate), n- tetradecanoate (myristate), n- hexadecanoate (palmitate), n-octadecanoate (stearate), n-eicosanoate (arachidate), n-docosanoate (behenate), n-tetracosanoate (lignocerate), cis-9-hexadecenoate (palmitoleate), cis-9-octadecanoate (oleate), cis,cis-9, 12- octadecandienoate (linoleate), all cis-9, 12, 15-octadecatrienoate (linolenate), and all cis-5,8,11 ,14- eicosatetraenoate (arachidonate). The two fatty acid chains of a phospholipid may be identical or different. Acceptable phospholipids include dioleoyl PS, dioleoyl PC, distearoyl PS, distearoyl PC, dimyristoyl PS, dimyristoyl PC, dipalmitoyl PG, stearoyl, oleoyl PS, palmitoyl, linolenoyl PS, and the like.

[0098] The phospholipids may come from any natural source, and, as such, may comprise a mixture of phospholipids. For example, egg yolk is rich in PC, PG, and PE, soybeans contain PC, PE, PI, and PA, and animal brain or spinal cord is enriched in PS. Phospholipids may come from synthetic sources too. Mixtures of phospholipids having a varied ratio of individual phospholipids may be used. Mixtures of different phospholipids may result in liposome compositions having advantageous activity or stability of activity properties. The above mentioned phospholipids may be mixed, in optimal ratios with cationic lipids, such as N-(1-(2,3- dioleolyoxy)propyl)-N,N,N- trimethyl ammonium chloride, 1 , 1 ’-dioctadecyl-3, 3, 3’, 3’- tetramethylindocarbocyanine perchloarate, 3,3’-deheptyloxacarbocyanine iodide, 1 ,T-dedodecyl-3, 3, 3’, 3’- tetramethylindocarbocyanine perchloarate, 1 ,T-dioleyl- 3,3,3’,3’-tetramethylindo carbocyanine methanesulfonate, N-4-45106498195 1Attorney Docket No: 119289-861234(delinoleylaminostyryl)-N-methylpyridinium iodide, or 1 ,1 ,-dilinoleyl-3,3,3’,3’- tetramethylindocarbocyanine perchloarate.

[0099] Liposomes may optionally comprise sphingolipids, in which spingosine is the structural counterpart of glycerol and one of the one fatty acids of a phosphoglyceride, or cholesterol, a major component of animal cell membranes. Liposomes may optionally contain pegylated lipids, which are lipids covalently linked to polymers of polyethylene glycol (PEG). PEGs may range in size from about 500 to about 10,000 Daltons.

[0100] Liposomes may further comprise a suitable solvent. The solvent may be an organic solvent or an inorganic solvent. Suitable solvents include, but are not limited to, dimethylsulfoxide (DMSO), methylpyrrolidone, N-methylpyrrolidone, acetro nitrile, alcohols, dimethylformamide, tetrahydrofuran, or combinations thereof. xi. Emulsions

[0101] The TRAIL composition may be formulated as part of a microemulsion. Microemulsions are generally clear, thermodynamically stable solutions comprising an aqueous solution, a surfactant, and “oil.” The “oil” in this case, is the supercritical fluid phase. The surfactant rests at the oil-water interface. Any of a variety of surfactants are suitable for use in microemulsion formulations including those described herein or otherwise known in the art. The aqueous microdomains suitable for use according to the present disclosure generally will have characteristic structural dimensions from about 5 nm to about 100 nm. Aggregates of this size are poor scatterers of visible light and hence, these solutions are optically clear. As will be appreciated by a skilled artisan, microemulsions can and will have a multitude of different microscopic structures including sphere, rod, or disc shaped aggregates. In one embodiment, the structure may be micelles, which are the simplest microemulsion structures that are generally spherical or cylindrical objects. Micelles are like drops of oil in water, and reverse micelles are like drops of water in oil. In an alternative embodiment, the microemulsion structure is the lamellae. It comprises consecutive layers of water and oil separated by layers of surfactant. The “oil” of microemulsions optimally comprises phospholipids.

[0102] Any of the phospholipids detailed above for liposomes are suitable for embodiments directed to microemulsions. A composition comprising at least one46106498195 1Attorney Docket No: 119289-861234 anti-tumor therapeutic derivative may be encapsulated in a microemulsion by any method generally known in the art.

[0103] In yet another embodiment, the TRAIL conjugate may be delivered in a dendritic macromolecule, or a dendrimer. Generally, a dendrimer is a branched treelike molecule, in which each branch is an interlinked chain of molecules that divides into two new branches (molecules) after a certain length. This branching continues until the branches (molecules) become so densely packed that the canopy forms a globe. Generally, the properties of dendrimers are determined by the functional groups at their surface. For example, hydrophilic end groups, such as carboxyl groups, would typically make a water-soluble dendrimer. Alternatively, phospholipids may be incorporated in the surface of a dendrimer to facilitate absorption across the skin. Any of the phospholipids detailed for use in liposome embodiments are suitable for use in dendrimer embodiments. Any method generally known in the art may be utilized to make dendrimers and to encapsulate compositions as disclosed herein. For example, dendrimers may be produced by an iterative sequence of reaction steps, in which each additional iteration leads to a higher order dendrimer. Consequently, they have a regular, highly branched 3D structure, with nearly uniform size and shape. Furthermore, the final size of a dendrimer is typically controlled by the number of iterative steps used during synthesis. A variety of dendrimer sizes are suitable for use according to the present disclosure. Generally, the size of dendrimers may range from about 1 nm to about 100 nm.IX. Dosage Forms

[0104] One aspect of the disclosure encompasses dosage forms made from the TRAIL conjugate. The dosage form may comprise TRAIL conjugate in an amount from about 1 microgram to about 50 g, from about 1 mcg to about 5,000 mg, from about 1 mcg to about 1 ,000 mg, from about 1 mcg to about 500 mg, from about 50 mcg to about 400 mg, from about 75 mcg to about 150 mg, from about 150 mcg to about 200 mg, from about 40 mcg to about 150 mg, from about 80 mcg to about 120 mg, or from about 180 mcg to about 100 mg. For instance, the dosage form can comprise 1 , 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 120, 140, 160, 180, 200, 220, 240, 260, 280, or 300 or more mcg of the TRAIL conjugate. In some aspect, dosage47106498195 1Attorney Docket No: 119289-861234 forms can comprise from about 1 mcg to about 500 mg, about 1 mcg to about 400 mg, about 1 mcg to about 300 mg, about 1 mcg to about 250 mg, about 1 mcg to about 200 mg, about 1 mcg to about 150 mg, or about 1 mcg to about 100 mg of the TRAIL conjugate.

[0105] Dosage forms include those formulated for extended or slow release, and those formulated for immediate release. For example, an immediate release dosage form may include a crystalline form of TRAIL conjugate as the free base or as a TRAIL conjugate salt as disclosed herein. For example, a fast-dissolve oral dosage form may include for example a TRAIL conjugate salt, such as the HCI salt. Alternatively, a dosage form may include a crystalline form of the TRAIL conjugate or crystalline form of a salt of the TRAIL conjugate.

[0106] Dosage forms also include those formulated for topical administration. For instance, a dosage form can be formulated as one or more of a gel, ointment, emulsion, microemulsion, lipids, liposomes, nanosomes, solution, suspension, paste, gel, foam, spray, lotion, or cream. In one aspect, a topical administration dosage form is a transdermal patch. When the dosage form is formulated as a transdermal patch, the transdermal patch can contain from at or about 40 mcg to at or about 60 mg, from at or about 80 mcg to at or about 50 mg, or from about 180 mcg to at or about 40 mg of TRAIL conjugate in crystalline form.

[0107] Dosage forms can alternatively be formulated for oral administration. Dosage forms formulated for oral administration can be tablets to swallow, chew, or dissolve in water or under the tongue, capsules and chewable capsules, powders, granules, teas, drops, or liquid medications or syrups. In some aspect, the dosage form is an enteric coated oral formulation.

[0108] When the dosage form is an enteric coated oral formulation, the formulation can comprise from about 0.1 mcg to about 60 mg TRAIL conjugate, preferably from about 1 mcg to about 50 mg TRAIL conjugate.

[0109] Dosage forms also encompass those formulated for injection administrations, such as intravenous injection, intratumor injection, intramuscular injection, intraperitoneal injection, intraocular injection, subcutaneous injection, intradermal injection, or intrathecal injection. For example, an intravenous dosage48106498195 1Attorney Docket No: 119289-861234 form may comprise TRAIL conjugate dissolved in an aqueous or oil matrix suitable for intravenous injection.X. Kits

[0110] A further aspect of the present disclosure provides kits comprising one or more compositions for use according to the present disclosure. The kits may comprise a container to hold the TRAIL composition and an instruction of use. The kits generally include instructions for carrying out the methods detailed in the present disclosure. Instructions included in the kits may be affixed to packaging material or may be included as a package insert. While the instructions are typically written or printed materials, they are not limited to such. Any medium capable of storing such instructions and communicating them to an end user is contemplated by this disclosure. Such media include, but are not limited to, electronic storage media (e.g., magnetic discs, tapes, cartridges, chips), optical media (e.g., CD ROM), and the like. As used herein, the term “instructions” may include the address of an internet site that provides such instructions.DEFINITIONS

[0111] Unless defined otherwise, all technical and scientific terms used herein have the meaning commonly understood by a person skilled in the art to which this invention belongs. The following references provide one of skill with a general definition of many of the terms used in this invention: Singleton et al., Dictionary of Microbiology and Molecular Biology (2nded. 1994); The Cambridge Dictionary of Science and Technology (Walker ed., 1988); The Glossary of Genetics, 5thEd., R. Rieger et al. (eds.), Springer Verlag (1991); and Hale & Marham, The Harper Collins Dictionary of Biology (1991 ). As used herein, the following terms have the meanings ascribed to them unless specified otherwise.

[0112] When introducing elements of the present disclosure or the preferred aspects(s) thereof, the articles “a”, “an”, “the” and “said” are intended to mean that there are one or more of the elements. The terms “comprising”, “including” and “having” are intended to be inclusive and mean that there may be additional elements other than the listed elements.

[0113] The term “comprising” means “including, but not necessarily limited to”; it specifically indicates open-ended inclusion or membership in a so-described49106498195 1Attorney Docket No: 119289-861234 combination, group, series and the like. The terms “comprising” and “including” as used herein are inclusive and / or open-ended and do not exclude additional, unrecited elements or method processes. The term “consisting essentially of” is more limiting than “comprising” but not as restrictive as “consisting of.” Specifically, the term “consisting essentially of” limits membership to the specified materials or steps and those that do not materially affect the essential characteristics of the claimed invention.

[0114] The term “alkyl,” as used herein, describes groups that are preferably lower alkyl containing from one to eight carbon atoms in the principal chain and up to 20 carbon atoms. They may be straight or branched chain or cyclic and include methyl, ethyl, propyl, isopropyl, butyl, hexyl and the like.

[0115] The term “alkenyl,” as used herein, describes groups that are preferably lower alkenyl containing from two to eight carbon atoms in the principal chain and up to 20 carbon atoms. They may be straight or branched chain or cyclic and include ethenyl, propenyl, isopropenyl, butenyl, isobutenyl, hexenyl, and the like.

[0116] The term “alkynyl,” as used herein, describes groups that are preferably lower alkynyl containing from two to eight carbon atoms in the principal chain and up to 20 carbon atoms. They may be straight or branched chain and include ethynyl, propynyl, butynyl, isobutynyl, hexynyl, and the like.

[0117] The term “aromatic,” as used herein, alone or as part of another group denotes optionally substituted homo- or heterocyclic planar ring or ring system comprising delocalized electrons. These aromatic groups are preferably monocyclic (e.g., furan or benzene), bicyclic, or tricyclic groups containing from 5 to 14 atoms in the ring portion. The term “aromatic” encompasses “aryl” groups defined below.

[0118] The terms “aryl” or “Ar” as used herein, alone or as part of another group denote optionally substituted homocyclic aromatic groups, preferably monocyclic or bicyclic groups containing from 6 to 10 carbons in the ring portion, such as phenyl, biphenyl, naphthyl, substituted phenyl, substituted biphenyl, or substituted naphthyl.

[0119] The terms “carbocyclo” or “carbocyclic,” as used herein, alone or as part of another group denote optionally substituted, aromatic or non-aromatic, homocyclic ring or ring system in which all of the atoms in the ring are carbon, with50106498195 1Attorney Docket No: 119289-861234 preferably 5 or 6 carbon atoms in each ring. Exemplary substituents include one or more of the following groups: hydrocarbyl, substituted hydrocarbyl, alkyl, alkoxy, acyl, acyloxy, alkenyl, alkenoxy, aryl, aryloxy, amino, amido, acetal, carbamyl, carbocyclo, cyano, ester, ether, halogen, heterocyclo, hydroxy, keto, ketal, 51 hosphor, nitro, and thio.

[0120] The terms “halogen” or “halo,” as used herein, alone or as part of another group refer to chlorine, bromine, fluorine, and iodine.

[0121] The term “heteroatom,” as used herein, refers to atoms otherthan carbon and hydrogen. The term “heteroaromatic,” as used herein, alone or as part of another group denotes optionally substituted aromatic groups having at least one heteroatom in at least one ring, and preferably 5 or 6 atoms in each ring. The heteroaromatic group may have 1 to 4 oxygen, nitrogen, silicone, and / or sulfater atoms (e.g. 1 or 2 oxygen atoms and / or 1 to 4 nitrogen atoms) in the ring and is bonded to the remainder of the molecule through a carbon. Exemplary groups include fury I, benzofuryl, oxazolyl, isoxazolyl, oxadiazolyl, benzoxazolyl, benzoxadiazolyl, pyrrolyl, pyrazolyl, imidazolyl, triazolyl, tetrazolyl, pyridyl, pyrimidyl, pyrazinyl, pyridazinyl, indolyl, isoindolyl, indolizinyl, benzimidazolyl, indazolyl, benzotriazolyl, tetrazolopyridazinyl, carbazolyl, purinyl, quinolinyl, isoquinolinyl, imidazopyridyl, and the like. Exemplary substituents include one or more of the following groups: hydrocarbyl, substituted hydrocarbyl, alkyl, alkoxy, acyl, acyloxy, alkenyl, alkenoxy, aryl, aryloxy, amino, amido, acetal, carbamyl, carbocyclo, cyano, ester, ether, halogen, heterocyclo, hydroxy, keto, ketal, 51 hosphor, nitro, and thio.

[0122] The terms “heterocyclo” or “heterocyclic,” as used herein, alone or as part of another group denote optionally substituted, fully saturated or unsaturated, monocyclic or bicyclic, aromatic or non-aromatic groups having at least one heteroatom in at least one ring, and preferably 5 or 6 atoms in each ring. The heterocyclo group may have 1 or 2 oxygen atoms and / or 1 to 4 nitrogen atoms in the ring and is bonded to the remainder of the molecule through a carbon or heteroatom. Exemplary heterocyclo groups include heteroaromatics as described above. Exemplary substituents include one or more of the following groups: hydrocarbyl, substituted hydrocarbyl, alkyl, alkoxy, acyl, acyloxy, alkenyl, alkenoxy, aryl, aryloxy, amino, amido, acetal, carbamyl, carbocyclo, cyano, ester, ether, halogen, heterocyclo, hydroxy, keto, ketal, 51 hosphor, nitro, and thio.51106498195 1Attorney Docket No: 119289-861234

[0123] The terms “hydrocarbon” and “hydrocarbyl,” as used herein, refer to organic compounds or radicals consisting exclusively of the elements carbon and hydrogen. These moieties include alkyl, alkenyl, alkynyl, and aryl moieties. These moieties also include alkyl, alkenyl, alkynyl, and aryl moieties substituted with other aliphatic or cyclic hydrocarbon groups, such as alkaryl, alkenaryl and alkynaryl. Unless otherwise indicated, these moieties preferably comprise 1 to 20 carbon atoms.

[0124] The compounds described herein may have asymmetric centers. Compounds of the present disclosure containing an asymmetrically substituted atom may be isolated in optically active or racemic form. All chiral, diastereomeric, racemic forms and all geometric isomeric forms of a structure are intended, unless the specific stereochemistry or isomeric form is specifically indicated.

[0125] As used herein, the terms “disease”, “disorder” or “medical condition” are used interchangeably. Each includes, but is not limited to, any condition or disease manifesting as one or more physical and / or psychological symptoms for which treatment is desirable and includes previously and newly identified diseases and other disorders.

[0126] As used herein, the term “anticancer”, “anti-tumor” and “antineoplastic” may be used interchangeably, for an activity in combating cancer or tumor. Classes of anticancer agents include, but are not limited to, chemotherapeutic agents, cytotoxins, antimetabolites, alkylating agents, protein kinase inhibitors, anthracyclines, antibiotics, antimitotic agents (e.g. antitubulin agents), corticosteroids, radiopharmaceuticals, and proteins (e.g. cytokines, enzymes, or interferons). Specific examples include, but are not limited to docetaxel, gemcitabine, imatinib (Gleevec®), 5-fluorouracil, 9-aminocamptothecin, amine-modified geldanamycin, doxorubicin, paclitaxel (Taxol®), procarbazine, hydroxyurea, meso e- chlorin, cisplatin, Gd(+3) compounds, asparaginase, and radionuclides (e.g 1-131 , Y- 90, ln-111 , and Tc-99m). There are many anticancer agents known in the art and many continue to be developed.

[0127] The terms “treat,” “treating,” and “treatment” are meant to include alleviating or abrogating a condition, or one or more of the symptoms associated with the condition; or alleviating or eradicating the cause(s) of the condition itself.52106498195 1Attorney Docket No: 119289-861234

[0128] The terms “manage,” “managing,” and “management” encompass preventing the recurrence of the specified disease, disorder, or condition in a patient who has already suffered from the disease, disorder, or condition, and / or lengthening the time that a patient who has suffered from the disease, disorder, or condition remains in remission. The terms encompass modulating the threshold, development and / or duration of the disease, disorder, or condition, or changing the way that a patient responds to the disease, disorder, or condition.

[0129] The terms “prevent,” “preventing,” and “prevention” are meant to include a method of delaying and / or precluding the onset of a disorder, disease, or condition, and / or its attendant symptoms; barring a subject from acquiring a disorder, disease, or condition; or reducing a subject’s risk of acquiring a disorder, disease, or condition

[0130] As used herein, the term “effective amount” refers to an amount of an agent (such as a mixture of RNAs) that provides a desired biological, therapeutic, preventive, theragnostic, and / or prophylactic result. That result can be reduction, amelioration, palliation, lessening, delaying, prevention, and / or alleviation of one or more of the signs, symptoms, or causes of a disease (such as advanced stage solid tumor cancer). In some embodiments, an effective amount comprises an amount sufficient to cause a solid tumor / lesion to shrink. In some embodiments, an effective amount is an amount sufficient to decrease the growth rate of a solid tumor (such as to suppress tumor growth). In some embodiments, an effective amount is an amount sufficient to delay tumor development. In some embodiments, an effective amount is an amount sufficient to prevent or delay tumor recurrence.

[0131] In some aspect, an effective amount is an amount sufficient to increase a subject’s immune response to a tumor or a cancer, such that tumor growth and / or size and / or metastasis is reduced, delayed, ameliorated, and / or prevented. An effective amount can be administered in one or more administrations. In some embodiments, administration of an effective amount (e.g., of a composition comprising mRNAs) may: (i) reduce the number of cancer cells; (ii) reduce tumor size; (iii) inhibit, retard, slow to some extent and may stop cancer cell infiltration into peripheral organs; (iv) inhibit (e.g., slow to some extent and / or block or prevent) metastasis; (v) inhibit tumor growth; (vi) prevent or delay occurrence and / or53106498195 1Attorney Docket No: 119289-861234 recurrence of tumor; and / or (vii) relieve to some extent one or more of the symptoms associated with the cancer.

[0132] The term “co-administered” or “co-administration” or the like as used herein refers to administration of two or more agents concurrently, simultaneously, or essentially at the same time, either as part of a single formulation or as multiple formulations that are administered by the same or different routes. “Essentially at the same time” as used herein means within about 1 minute, 5 minutes, 10 minutes, 15 minutes, 30 minutes, 1 hour, 2 hours, or 6 hours period of each other.

[0133] As used herein, the term “subject” means that preferably the subject is a mammal, such as a human, but can also be an animal, e.g., domestic animals (e.g., dogs, cats and the like), farm animals (e.g., cows, sheep, pigs, horses and the like) and laboratory animals (e.g., cynomolgus monkey, rats, mice, guinea pigs and the like).

[0134] As used herein, the administration of an agent or drug to a subject or patient includes self-administration and the administration by another. It is also to be appreciated that the various modes of treatment or prevention of medical conditions as described are intended to mean “substantial”, which includes total but also less than total treatment or prevention, and wherein some biologically or medically relevant result is achieved.EXAMPLES

[0135] The publications discussed above are provided solely for their disclosure before the filing date of the present application. Nothing herein is to be construed as an admission that the invention is not entitled to antedate such disclosure by virtue of prior invention.

[0136] The following examples are included to demonstrate the disclosure. It should be appreciated by those of skill in the art that the techniques disclosed in the following examples represent techniques discovered by the inventors to function well in the practice of the disclosure. Those of skill in the art should, however, in light of the present disclosure, appreciate that many changes could be made in the disclosure and still obtain a like or similar result without departing from the spirit and54106498195 1Attorney Docket No: 119289-861234 scope of the disclosure, therefore all matter set forth is to be interpreted as illustrative and not in a limiting sense.Example 1. Study OverviewThe present disclosure relates to a new class of low-molecular-weight (LMW) radiotherapeutics, comprising an Auger-electron-emitting radionuclide (i.e. a very energetic but short pathlength type of radioactivity), a cell-targeting moiety to bring the therapy to cancer cells, a DNA-intercalating moiety to localize the compound to DNA, and a cleavable linker to ensure release of the other components of the agent from the lysosomal environment. Molecular components capable of associating in a modular fashion are assembled to prepare this unique class of molecules, referred to as “TRAIL” in recognition of the Targeting moiety (such as urea-based scaffold that binds prostate-specific membrane antigen), the RAdionuclide (such as the pure Auger-emitter iodine- 125), an Intercalating moiety (such as acridine orange), and a cleavable Linker (such as a pH-dependent moiety). The non-limiting examples provided herein leverage established models of prostate cancer and various proof- of-concept experiments.Example 2. Study Design

[0137] The present study is to develop a class of modular, low-molecular- weight (LMW) radiotherapeutic agents for the targeted delivery of lethal doses of radiation to the DNA of cancer cells. This study represents the first of its kind and enables creation of a generalizable platform for synthesis of a range of agents with potential efficacy against a wide number of cancers. Current clinical approaches to radiotherapeutic agents are based on p-particle-emitting radionuclides that are effective at extending life and abating symptoms in many patients - but they are never curative and they can have significant toxicities. To address those deficiencies, a new class of agents TRAIL conjugates are designed, allowing the specificity of a Targeting moiety for a cell-surface biomarker that will release a therapeutic RAdionuclide moiety - attached to a DNA Intercalator- once the molecule is within the lysosome and following activation of a cleavable Linker (TRAIL). TRAIL conjugates are a modular concept, with any of the four components55106498195 1Attorney Docket No: 119289-861234 being modifiable and optimizable. The initial study utilizes prostate-specific membrane antigen (PSMA)-expressing prostate cancer (PCa) as a model system. PSMA is a type II transmembrane glycoprotein that is commonly over-expressed on the tumors of men with PCa. There are multiple reasons for utilizing this enzymatic target, e.g. (1) numerous model systems for PSMA-expressing and PSMA-non- expressing tumors have been extensively characterized, (2) PSMA is an established clinical biomarker and theranostic target that is constitutively internalized after binding, making it an ideal target for imaging and therapy, (3) LMW radiotherapeutic approaches are well established in routine clinical uses, and (4) although current radiotherapeutic compounds are effective, eventually all patients will progress on them. Certain radionuclides may have specific advantages that could be accentuated by using TRAIL conjugates for DNA localization. Notably, Auger-electron-emitting radionuclides have exceedingly high linear energy transfer and short path-lengths to cause DNA damage relative to their commonly used p-particle-emitting analogues.

[0138] The long-term goal of the present study is to develop molecules with improved efficacy and tolerability for the radiotherapeutic management of men with PCa - and subsequently apply those principles to therapies of other cancers. The pathway for the development of LMW radiopharmaceuticals has been established through the United States Food and Drug Administration (FDA) approval of multiple such therapies for prostate cancer, mid-gut neuroendocrine tumors, and pheochromocytoma / paraganglioma. The present study is to improve the targeting and efficacy of LMW radiopharmaceuticals, while potentially decreasing their off- target effects and mitigating side-effects. The present study includes the following specific aims:Aim 1. Synthetic optimization of TRAIL lead compounds and key control molecules.

[0139] The present study synthesizes a PCa-targeted TRAIL agent utilizing a urea-based PSMA-binding moiety, an appended group for the Auger-emitting radionuclide, a DNA intercalator, and an intracellular-pH-responsive cleavable linker. A series of control molecules are synthesized that are each missing one of the key components of the complete scaffold of TRAIL.56106498195 1Attorney Docket No: 119289-861234Aim 2. Radiolabeling and evaluation of physicochemical and in vitro biological properties.

[0140] The TRAIL conjugates and key control molecules are radiolabeled with iodine- 125, a pure Auger-electron emitter. It is hypothesized that delivering Augerelectron-emitting radionuclides to the tumor cells and promoting localization of the internalized complex to DNA could facilitate therapeutic efficacy of the radiopharmaceuticals. The present study also includes examining the binding affinity, internalization, DNA intercalation, and radiation induced double-strand DNA damage caused by the TRAIL conjugates and control molecules from Aim 1 to predict their potential therapeutic efficacy.Aim 3. Initial in vivo evaluation of the optimized compounds from Aims 1 and 2.

[0141] The TRAIL conjugates and negative controls are investigated via quantitative biodistribution in normal organs and LNCaP and PC3 tumor xenografts.Impacts

[0142] It cannot be underestimated the impact that LMW therapeutic radiopharmaceuticals have had on clinical practice. However, ultimately, many patients are primary refractory to currently available agents and all patients who live long enough would eventually become refractory. Improving the efficacy of therapeutic radiopharmaceuticals, while also decreasing off-target effects, is a distinct need that may be addressed by the TRAIL modular approach. Additionally, there is an established pathway to clinical translation and commercialization of PSMA targeted TRAIL conjugates. Industry leaders from the Center for Probe Development and Commercialization (CPDC) are to provide translational guidance, support for regulatory filings and GMP manufacturing of the probe. The CPDC has expertise needed to move the proposed radiopharmaceutical from the lab, into clinical trials and to ultimately attract venture investments to advance the technology to market. To assess the potential or efficacy of TRAIL conjugate and their compositions as DNA cleaving or scission agents, various physiochemical testing and biological assays are performed.Example 3: Introduction and Terminologies57106498195 1Attorney Docket No: 119289-861234

[0143] Throughout the disclosure, the abbreviations listed in TABLE 1 were used. In recent years, low-molecular-weight (LMW) radiotherapeutic agents have revolutionized care of patients with metastatic or unresectable cancers, with United States Food and Drug Administration approvals of compounds for paraganglioma / pheochromocytoma, midgut neuroendocrine tumors, and metastatic, castration-resistant prostate cancer (PCa). To date, all the approved, tumor-targeted LMW radiotherapeutic agents utilize b-particle emission as the means of tumoricidal radiation. In many ways, LMW radiotherapeutics have joined chemotherapy and immunotherapy / cell-based therapies among the broad categories of widely applicable systemic treatments for cancer. However, with currently available agents, when patients are followed-up long-term after treatment, eventually all individuals who live long enough will subsequently progress and eventually die of the effects of their metastatic disease. This emphasizes the need for innovative approaches to LMW radiotherapeutics which might include different classes of radionuclides (e.g. a- or Auger-electron-emitters) or medicinal chemistry refinement of the targeting scaffolds to afford improved specificity, improved target localization, and / or improved pharmacokinetics / pharmacodynamics.TABLE 1 : Abbreviations Used58106498195 1Attorney Docket No: 119289-861234

[0144] To address some of the key shortcomings of existing approaches, a novel route with a widely applicable platform of LMW radiotherapeutics is designed. The designed novel class of therapeutic molecules utilizes a Targeting moiety for a cell-surface protein that will release a therapeutic RAdionuclide moiety - attached to a DNA Intercalator - once the molecule is within the lysosome and following activation of a cleavable Linker. Given the concatenation of structural features, this class of agents is dubbed as TRAIL conjugates. Some exemplary TRAIL conjugates include:

[0145] Such compounds capitalize on the intersection of three complementary (and possibly synergistic) fundamental alterations to existing agents. The first is the incorporation of iodine-125, an Auger-electron-emitting radionuclide with an exceedingly high linear-energy-transfer (LET) that has previously been shown to be an effective radionuclide for therapeutic applications in micrometastatic cancer models. Iodine-125 is readily available and capable of being leveraged with existing radiohalogenation chemistry. Iodine-125 has a long half-life, which may be an underrecognized aspect to optimizing radiotherapeutic efficacy. The second major59106498195 1Attorney Docket No: 119289-861234 alteration involves the incorporation of a DNA-chelating moiety, which localizes the entire structure to DNA and maximizes the efficacy of the high-LET, short pathlength Auger electrons. The third major alteration is the inclusion of a cleavable linker between the cancer-targeting moiety and the “warhead” of the molecule (i.e. the intercalation and radionuclide-containing portions of the agent). That cleavable linker ensures that the warhead is free to reach the nucleus and localize to DNA after the agent is endocytosed and trafficked to the lysosome. The initial target for validating TRAIL conjugates is metastatic, castration resistant PCa. That is an appealing target because (1) of the approval of a LMW radiotherapeutic agent in that space, suggesting a regulatory pathway to eventual approval, (2) the urea-based scaffold being widely utilized for imaging and therapy of PCa, and the well-characterized models of PSMA expression across a variety of different models.

[0146] Based on the above overall approach, details are provided to show (1) the cold synthesis of a series of TRAIL analogs and optimization of the overall structure through in vitro experiments, (2) radiosynthesis of the optimized compound, and (3) in vivo biodistribution and preliminary cellular toxicity data. Future work is to extend these ideas into patient-derived xenografts (PDXs) as a step towards eventual clinical translation for men with metastatic, castration-resistant PCa. In the longer term, the platform and modular approach of TRAIL conjugates permits their application to a wide range of cancers via the utilization of appropriate targeting moieties. The present disclosure involves both the initial concept and the eventual clinical applicability.Example 4: Synthesis and Optimization of TRAIL Molecules

[0147] An exemplary synthesis utilizes LMW radiotherapeutic agents having key aspects of the proposed overall modular approach. Briefly, the DNA intercalator acridine orange was functionalized to 10-(4-amino-butyl)-3,6-bis-dimethylamino- acridinium with a yield that was a notable improvement on the current literature, largely attributed in an initial step by using acetonitrile as solvent over p-xylene. The primary benefit of acetonitrile lied on its optimal effect for nucleophilic reactions as a polar aprotic solvent. The second step in the reaction sequence produced the key intermediate proceeded by the same mechanism described for the Gabriel60106498195 1Attorney Docket No: 119289-861234Phthalimide synthesis. This reaction conditions are further optimized to achieve higher yields. The key intermediate obtained from LMW-acridine orange chemistry was subsequently used in the solid phase synthetic route for an agent containing the acridine orange tethered to a glutamate-urea-lysine PSMA-binding moiety and a DOTA chelator for potential labeling with radiometals - followed by purification and characterization.

[0148] The construct obtained above is similar to the proposed TRAIL conjugates, lacking just the pH-dependent cleavable linker and an Auger-electron- emitting radionuclide, which has no previously reported synthesis. However, 10-(4- Amino-butyl)-3,6-bis-dimethylamino-acridinium, was successfully synthesized in acceptable yield and high purity, superior synthetic strategies were employed that allowed for higher overall yield as well as significantly simpler purification methods. Further, solid-phase methodology facilitated the synthesis of a construct that resembled the proposed modular TRAIL conjugates.Aim 1. Synthetic optimization of TRAIL lead conjugates and key control molecules.

[0149] A PCa-targeted TRAIL conjugate is synthesized utilizing a urea-based PSMA-binding moiety, an appended group for the Auger-emitting radionuclide, a DNA intercalator, and an intracellular-pH-induced cleavable linker. A series of control molecules are also synthesized that are each missing one of the key components of the overall scaffold, some of the exemplary structures and synthesis routes are provided in FIGURE 1 and FIGURE 2. Both TRAIL conjugates and control compounds are evaluated for uptake, intracellular localization, and DNA binding with well-established dyes with DNA intercalating properties such as acridine orange, Proflavine (3,6-diaminoacridine) and methylene blue. These molecules are to accommodate a bifunctional labeling agent for bioconjugation with either a stable isotope or radiolabeling with an iodine-125 labeled prosthetic group. An alternative approach is the incorporation of well-established cyclic chelators such as DOTA and NOTA derivatives that form stable complexes with Auger-emitting radiometals such as terbium-161. These compounds can also be labeled with PET imaging radiometals such as gallium-68 and [18F]AIF to serve as theragnostic probes that could image and treat cancer based on the choice of radionuclide.61106498195 1Attorney Docket No: 119289-861234Aim 2. Radiolabeling and evaluation of physicochemical and in vitro biological properties.

[0150] The TRAIL lead conjugate and key control molecules are radiolabeled with iodine-125, a pure Auger-electron emitter. It is hypothesized that delivering auger-electron-emitting radionuclides to the tumor cells and promoting localization of the internalized complex to DNA facilitate therapeutic efficacy of the radiopharmaceuticals. Characterizations of binding affinity, internalization, and DNA intercalation of AO-constructs from Aim 1 are conducted to predict their potential therapeutic efficacy.Evaluation of physicochemical properties.

[0151] Before advancing to in vivo preclinical studies, each candidate is evaluated in vitro by measuring the following physicochemical properties: (a). Hydrophilicity. The hydrophilicity of each compound is assessed by measuring the octanol / water partition coefficient (logP) at pH 7.4 using analytical high-performance liquid chromatography (HPLC); and b). Hydrolytic stability. The hydrolysis of each candidate at physiological conditions (pH 7.4, 37 °C) is monitored by HPLC at 7 time points over a period of up to 24 h.Evaluation of radiolabeling efficiency.

[0152] The radiolabeling properties of synthesized candidates are assessed using low amounts of iodine-125 activity (<100 MBq). Radio-thin-layer chromatography (radio-TLC) is used to measure the 125 l-iodine incorporation or radiochemical conversion (RCC). Radio-HPLC is used for measuring the purity of radiolabeled candidates.Evaluation of in vitro biological properties.

[0153] Besides assessing the physicochemical and radiolabeling properties, each candidate is evaluated by the following biological properties. First is PSMA binding affinity. Binding affinity is measured in LNCaP cells that stably express PSMA, by displacing [125I]PSMA in a dose-dependent manner. The binding affinity is reported as the half maximal inhibitory concentration (IC50). Second is Human serum albumin (HSA) binding. Binding to plasma proteins such as HSA influences62106498195 1Attorney Docket No: 119289-861234 the blood-circulation times of radiopharmaceuticals and impacts their ability to bind to the biological target. HSA binding properties of candidates are assessed with an HPLC method using a Chiralpak HSA column. The results are reported as % HSA binding. Third is Human plasma stability. Human plasma stability is assessed by incubating peptide conjugates at 37°C for 72 hours and the fraction of the parent compound will be monitored by HPLC at several time points. Fourth / s DNA intercalation and radiation induced double-strand breaks. Alongside the cascade of extremely low-energy electrons, lodine-125 also produces hydroxyl radicals along the path of Auger electrons in the vicinity of the decay site. These decay processes single and double strand DNA breaks in supercoiled DNA resulting from direct energy transfer of Auger electrons as well as hydroxyl radical-mediated indirect effects. Gel electrophoresis of supercoiled plasmid issued to measure DNA damage caused by radio-iodinated TRAIL conjugates and control compounds, with and without the DNA intercalating and cleavable linkers. Gel mobility is affected by DNA conformation and length. Migration is faster for intact supercoiled conformation, whereas relaxed circular conformation DNA and linear form, resulting from single strand breaks and double strand breaks respectively are slower moving on the same gel assay. Imaging of bands provides quantification by defining areas of interest for each band and calculating the percentage distribution. Quantification of DNA bands containing undecayed 125-lodine bound to DNA, is performed by cutting regions of interest and measuring in the gamma counter.Aim 3. Initial in vivo evaluation of the optimized compounds from Aims 1 and 2.

[0154] The TRAIL conjugates and negative control compounds are investigated via quantitative biodistribution in normal organs and LNCaP and PC3 tumor xenografts.Biodistribution studies with the 125 l-labeled candidates in tumor-bearing mice (McMaster).

[0155] It is expected 2 candidates satisfying the above-mentioned criteria are advanced for further studies. They are produced via scaled up radiosynthesis starting with -100 MBq.63106498195 1Attorney Docket No: 119289-861234Evaluation of clinical grade radiolabeling efficiency.

[0156] The candidates are 125 Iodine-labeled using the optimized procedure, purified by solid phase extraction technique and reformulated in an aqueous buffer at pH 7.4-8.0 for safe injection in mice. The isolated radiochemical yield (RCY) is calculated by dividing the activity of the purified peptide in a final formulation by the starting activity of iodine-125. The purity and molar activity (Am) of the final product are determined by using radio-HPLC and the identity will be confirmed by coinjection with the non-radioactive reference.In vivo Biodistribution studies.

[0157] For the preclinical evaluation of each 125 l-iodine candidate, 24 male NCr nude mice are implanted with 0.5-4 million LNCaP cells. About three weeks post-implantation, the visible tumors are in 50 mg to 400 mg range. Four mice are injected in the tail-vein with 0.5-1 MBq of the 125 l-labeled candidate in a baseline experiment. In the other four mice, the 125 l-labeled tracer are co-injected with 100 pg PSMA inhibitor in a blocking experiment. Four mice are sacrificed at each of the 4h, 48h, and 168h time-points, and organs are harvested to assess the ex vivo biodistribution. While rate of metabolism in vivo is not always easy to predict, in vitro hydrolytic and plasma stability assays described in Aim 2 provide a useful tool for predicting in vivo metabolic stability. Therefore, most of the candidates advanced to preclinical studies are sufficiently stable, but the predictive capabilities are improved through screening additional candidates, incorporating various DNA intercalators and / or Auger-emitting radiometals, and evaluating their performance in vivo.

[0158] High uptake of radioligands in liver and gall bladder generally correlates with high lipophilicity (i.e. logP > -1.5). To address this challenge, a range of more hydrophilic conjugates are prepared that incorporate PEG units. Retention in the kidneys is more difficult to predict but must be avoided, especially for therapeutic radiopharmaceuticals. While there is no “silver bullet” to prevent kidney uptake, the repeats of the PEG unit in the linker have been reported to accelerate the wash out of radioligands from the kidneys. Accumulation of RPs in the targeted tumor is the most important measure of success, and it generally correlates with the affinity of the RH for the targeted receptor. Most successful RPs, including PSMA-targeting peptides, have Ki values in a single digit nanomolar range. Therefore, the desirable64106498195 1Attorney Docket No: 119289-861234 range is approached, and subtle changes to the linker domain provide the necessary increase in potency. A longer linker (e.g. HE4 or PEG12) that creates additional space between the binding domain and the cleavable linker / intercalator motifs improves potency. The synthesis of these more potent candidates is underway, and they are expected to exhibit higher tumor uptake and retention, while also exhibiting the enhanced PK properties described above.Thresholds.

[0159] Tumor uptake must be >5 % ID / g at the 4h time-point, >2 % injected dose / gram (ID / g) at the 48h time-point, and >0.3 % ID / g at the 168h time-point. At the 4h time-point, the tumor-to-blood ratio must be >1 .0, the tumor-to-liver ratio must be >1.0, and the tumor-to-muscle ratio must be >5.0. At the 48h and 168h time-point, the tumor-to-blood ratios must be >2.0, the tumor-to-liver ratio must be >2.0, and the tumor-to-muscle ratio must be >5.0. In all mice, the activity should wash out from all non-targeted organs. Minimum requirements at all time-points are <125 % ID / g in the kidneys, <50 % ID / g in the liver and gallbladder, and <25 % ID / g in all other organs, including blood. A lower % ID / g in non-targeted organs is preferred.Example 5: Peptide synthesis and in vitro biological assaysSynthesis of the PSMA-targetinq molecules incorporating cleavable linkers and AO motifs.

[0160] Peptide conjugates are synthesized using solution and solid phase peptide synthesis (SPPS) methods according to previously established procedures.Hydrolytic stability.

[0161] Solution of conjugate (100 pM) in phosphate buffer (pH 7.4) is kept in a thermostated autosampler of the HPLC system at 37 °C and the aliquots are injected at several time-points over 24 hours.Metabolic stability.

[0162] Solution of conjugate (100 pM) in human plasma is incubated in human plasma at 37 °C for 72 hours. Aliquots are taken at several time-points, deproteinated by addition of ice-cold acetonitrile and analyzed by HPLC. logD measurement.65106498195 1Attorney Docket No: 119289-861234

[0163] To a solution of conjugate (100 pM) in a phosphate buffer (pH 7.4), equivoluminal amount of 1 -octanol is added and the mixture is vigorously stirred and then centrifuged to separate the layers. Aliquots from each layer are taken and injected on HPLC in triplicates to calculate relative concentrations and the LogD is calculated as the log(Coctanol / Cwater).DNA Intercalation.

[0164] The interaction of the ligands is studies using fluorescence spectroscopy. Titrations are performed with duplex DNA to obtain apparent dissociation constants (KD) for the different ligands.DNA damage assay.

[0165] Supercoiled plasmid DNA in Tris-HCI at pH 7.0 are incubated with 0.01-0.1 MBq / ml of all control compounds containing iodine-125 for up to 25 h to determine the effect of plasmid concentration, activity, and radionuclide on induction of DNA damage. Plasmid samples are mixed with loading dye and the mixture run on a 0.8% agarose gel at 200 V for 30 min. Agarose gels are imaged after electrophoresis using and analysis performed using Imaged to quantify the intact DNA, single strand breaks, and linear band resulting from double strand breaks.Human serum albumin (HSA) binding.

[0166] Solution of conjugate (100 pM) in a phosphate buffer (pH 7.4) is injected on HPLC system equipped with a Chiralpak HSA column in triplicates. The calibration curve of retention time (tp) vs HSA is made using 9 reference compounds with known HSA values prior to each series of measurement. HSA binding of each peptide is calculated as a function of tR using the calibration curve.PSMA binding affinity.

[0167] LNCaP cells expressing PSMA are seeded at 0.25-0.3 million cells / well in 24 well plates for 30-36 hours before the assay. The growth medium is then replaced by the buffer (400 pL) and the cells are incubated for 1 hour at 37 °C. Mixtures of [ 125 l]PSMA inhibitor and tested compounds at different concentrations (10-5 M to 10-12 M) are added, the cells are incubated for 1 hour at 37 °C, washed with ice cold PBS and detached. The radioactivity is then measured using the gamma counter to calculate the Ki values.66106498195 1Attorney Docket No: 119289-861234Biodistribution.

[0168] 24 male NCr nude mice are implanted with 2 million LNCaP cells. The tumors are allowed to develop over 21 days and then the mice are injected with 0.5- 1 MBq of the candidate tracer. The mice are then euthanized, dissected and the radioactivity in critical organs are measured using the gamma counter and expressed as percent injected dose per gram (%ID / g).Example 6. Alternative Approaches and Directions.Biodistribution and tumor-treatment studies with 161 Tb-labeled candidates in tumorbearing mice.

[0169] A potential alternative to demonstrate the advantage of the TRAIL radioligand therapies is to replace iodine-125 with terbium-161 . Terbium-161 has been validated as a potential alternative to lutetium-177, the most clinically used medium energy beta emitter therapeutic radionuclide. 161 Tb has similar beta energy emission, half-life and chemical properties to 177 Lu. However, in addition to beta-radiation, 161 Tb decay also results in Auger electrons, which may make it a promising TRAIL therapeutic radionuclide. While previous reports have demonstrated comparable therapeutic efficacy resulting from exchange of lutetium- 177 for terbium-161 , it is anticipated that the beta energy and the additional release of conversion and Auger electron in close proximity to the DNA, facilitated by the cleavable linker and intercalation of the radioligand labeled with 161 Tb, expand the therapeutic window by significantly increasing the delivered dose to the site of disease. FIGURE 3 provides an alternative TRAIL architecture that could incorporate radiometals such as terbium-161.Tumor mice therapy monitoring study with the lead 125 l-labeled candidate.

[0170] Following successful completion of the proposed synthesis and biodistribution studies, the optimal dose of 125 l-labeled TRAIL conjugate is evaluated in a combined double dose therapy monitoring and ex vivo biodistribution study. For each experiment, 12 male LNCaP mice models are developed as described above and divided in three cohort of 4 animals. Cohort 1 is injected via the tail-vein with 37 MBq of the 125 l-labeled candidate, cohort 2 is injected with 74 MBq of the same RP and cohort 3 is injected with saline. Tumors are monitored twice per week until the passing of the animal or until 4 weeks p.i. and the ex vivo67106498195 1Attorney Docket No: 119289-861234 biodistribution study are performed at the endpoint. Similar biodistribution and therapy studies are conducted with the 177 Lu- and 161 Tb labeled lead TRAIL conjugates.Translation into Human Subjects.

[0171] It will be understood that 125 l-and / or 161 Tb-labeled TRAIL conjugates can be used in human subjects.Example 7. Synthesis of LP2.Summary of LP2

[0172] The synthetic route is based on a procedure68106498195 1Attorney Docket No: 119289-86123469106498195 1Attorney Docket No: 119289-861234

[0174] Experimental for largest scale run:

[0175] General procedure for preparation of compound 2

[0176] To a solution of compound 1 (40.0 g, 335 mmol, 1 .00 eq.) in H2O (400 mL) was added Cbz-CI (57.2 g, 335 mmol, 47.9 mL, 1.00 eq.) and NaHCOs (141 g, 1.68 mol, 65.3 mL, 5.00 eq.). The mixture was stirred at 25 °C for 16 hrs. LC-MS 1 (MD00900-31-p1a1 ) showed one main peak (Rt = 0.303 min) with desired mass. The resultant was filtered and concentrated under freezing dry to give compound 2 (80.0 g, crude) as colorless oil and it confirmed by LC-MS (MD00900-31-p1 az, Rt = 0.318 min).

[0177] LC-MS 1 : MD00900-31-p1a1 , Rt = 0.303 min, MS cal.: 253.2, MS observed: [M+Na]+= 275.9

[0178] LC-MS: MD00900-31-p1az, Rt= 0.318 min, MS cal.: 253.2, MS observed: [M+Na]+= 276.0

[0179] General procedure for preparation of compound 3

[0180] To a solution of compound 2 (80.0 g, 315 mmol, 10.0 eq.) in DCM (800 mL) was added TBS-CI (47.6 g, 315 mmol, 38.8 mL, 1.00 eq.) and imidazole (21.5 g, 315 mmol, 1.00 eq.). The mixture was stirred at 25 °C for 12 hrs. LC-MS 1 (MD00900-32-P1A1 ) showed the main peak (Rt = 0.624 min) with desired mass. The reaction mixture was removed under reduced pressure. The residue was purified by column chromatography (SiCk, DCM / MeOH = 100 / 1 to 40 / 1 ). Compound 3 (46.0 g, 92.1 mmol, 29.1 % yield over two steps, 73.6% purity) was obtained as colorless oil and it confirmed by LC-MS (MD00900-32-P1 B1 , Rt = 0.622 min).70106498195 1Attorney Docket No: 119289-861234

[0181] LC-MS1 : MD00900-32-P1A1 , Rt = 0.624 min, MS cal.:367.5, MS observed: [M+H]+= 368.2

[0182] LC-MS: MD00900-32-P1 B1 , Rt = 0.622 min, MS cal.:367.5, MS observed: [M+H]+= 368.1

[0183] General procedure for preparation of compound 4

[0184] DCC (27.790 g, 134.690 mmol, 27.245 mL, 1.1 eq.) and DMAP (14.959 g, 122.445 mmol, 1 eq.) were addeed to a solution of compound 3 (45 g, 122.445 mmol, 1 eq.) and compound 4-1 (28.958 g, 244.891 mmol, 2 eq.) in DCM (450 mL). The mixture was stirred at 25 °C for 1 hr. LC-MS 1 (MD00842-83-P1A1 ) showed compound 3 was consumed completely, and identified the desired mass at Rt = 0.803 min. The reaction solution was filtered. The filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (SiC>2, Petroleum ether / Ethyl acetate = 1 / 0 to 0 / 1). Compound 4 (20.0 g, 39.6 mmol, 32.3% yield, 92.7% purity) was obtained as colorless oil and it confirmed by LC-MS (MD00842-83-P1 H1 , Rt = 2.329 min).

[0185] LC-MS 1 : MD00842-83-P1A1 , Rt = 0.803 min, MS cal.: 467.7, MS observed: [M+Na]+= 490.2

[0186] LC-MS: MD00842-83-P1 H1 , Rt = 2.329 min, MS cal.: 467.7, MS observed: [M+Na]+= 490.3

[0187] General procedure for preparation of compound 5

[0188] To a solution of Pd / C (1.5 g, 1.410 mmol, 10% purity, 4.741 e-2 eq.) inTHF (10.0 mL) was added compound 4 (15 g, 29.728 mmol, 1 eq.) in THF (140 mL)71106498195 1Attorney Docket No: 119289-861234 under H2. The mixture was stirred at 25 °C for 12 hrs. LC-MS 1 (MD00842-84- P1A33) showed compound 4 was consumed, and detected the desired mass (Rt =1 .449 min). The reaction mixture was filtered carefully under N2 atmosphere. The resultant was concentrated under reduced pressure to give a crude compound 5 (10.0 g, crude).

[0189] LC-MS 1 : MD00842-84-P1A33, Rt = 1.449 min, MS cal.: 333.6, MS observed: [M+H]+= 334.4

[0190] General procedure for preparation of compound 6-1

[0191] To a solution of compound 6-1A (1 g, 7.745 mmol, 1 eq.) and HOSu (1.159 g, 10.068 mmol, 1.3 eq.) in DCM (10 mL) was added EDCI (1.930 g, 10.068 mmol, 1 .3 eq.). The mixture was stirred at 25 °C for 2 hrs. TLC indicated compound 6-1A was consumed completely, and detected the new spot. The reaction mixture was diluted with DCM (10 mL) and washed with H2O (20 mL). The combined organics were washed with brine (50 mL). Dried over Na2SO4, filtered and concentrated under reduced pressure. Compound 6-1 (1.7 g, 7.516 mmol, crude) was obtained as light yellow oil and used for the next step directly.

[0192] General procedure for preparation of compound 6

[0193] To a solution of compound 5 (7 g, 20.982 mmol, 1 eq.) and compound 6-1 (5.221 g, 23.081 mmol, 1.1 eq.) in DMF (70.0 mL) was added DIEA (5.424 g, 41 .965 mmol, 6.953 mL, 2 eq.). The mixture was stirred at 25 °C for 1 hr. LC-MS 1 (MD00842-86-P1A1 1 ) showed compound 5 was consumed completely, and detected the desired mass (Rt = 0.766 min). The reaction mixture was partitioned72106498195 1Attorney Docket No: 119289-861234 between 100 mL of ethyl acetate and 300 mL H2O. The organic phase was washed with 50.0 mL of 10.0% aqueous NaCI and 60.0 mL of brine. Further it dried over anhydrous Na2SO4. The solvent was removed under reduced pressure. Compound 6 (9.50 g, crude) was obtained as colorless oil and it was confirmed by LC-MS (MD00842-86-P1 EA, Rt = 0.765 min).

[0194] LC-MS 1 : MD00842-86-P1A11 , Rt = 0.766 min, MS cal.: 444.7, MS observed: [M+H]+= 445.2

[0195] LC-MS: MD00842-86-P1 EA, Rt = 0.765 min, MS cal.: 444.7, MS observed: [M+H]+= 445.3

[0196] General procedure for preparation of compound 76 7

[0197] To a solution of compound 6 (2.5 g, 5.622 mmol, 1 eq.) in MeOH (25.0 mL) was added PPTS (423.814 mg, 1 .686 mmol, 0.3 eq.) under N2. The mixture was stirred at 25 °C for 120 hrs. LC-MS 1 (MD00842-92-P1 A4) showed compound 6 was remained (Rt = 0.762 min), and detected the desired mass (Rt = 0.498 min). The resultant was removed under reduced pressure. The residue was purified by column chromatography (SiO2, DCM / MeOH = 1 / 0 to 0 / 1 ). Compound 7 (6.00 g, 17.648 mmol, 70.0% yield over three steps, 97.2% purity) was obtained as yellow oil and it was confirmed by LC-MS (MD00842-92-P1 G1 , Rt = 1.403 min).

[0198] Note: combined purification of multiple batches of materials

[0199] LC-MS 1 : MD00842-92-P1A4, Rt = 0.498 min, MS cal.: 330.4, MS observed: [M+Na]+= 353.2

[0200] LC-MS: MD00842-92-P1G1 , Rt = 1.403 min, MS cal.: 330.4, MS observed: [M+Na]+= 353.0

[0201] General procedure for preparation of compound 873106498195 1Attorney Docket No: 119289-861234

[0202] Compound 7 (6 g, 17.648 mmol, 1 eq.) in DCM (60.0 mL) and Py (31 .069 g, 392.777 mmol, 31 .703 mL, 22.256 eq.) in DCM (60.0 mL) were added to a solution of (PhO)2P(O)H (24.385 g, 104.126 mmol, 20.037 mL, 5.9 eq.) in DCM (60.0 mL). The mixture was stirred at 25 °C for 4 hrs followed by the addition of 2- trimethylsilylethanol (18.365 g, 155.306 mmol, 22.261 mL, 8.8 eq.) in DCM (60.0 mL). The mixture was stirred at 25 °C for another 12 hrs. LC-MS 1 (MD00842-94- P1A2) showed compound 7 was consumed, and detected the desired mass (Rt = 1 .891 min). Volatiles were removed by rotary evaporation. The leftover oil was redissolve in EA and washed sequentially with 1 N HCI (aq.) (50.0 mL, 3x), sat. NaHCOs (aq.) (50.0 mL, 2x), sat. CuSO4 (aq) (30.0 mL, 2x) and Brine (30.0 mL, 1x). The organic layer was dried over Na2SO4, and the organics were removed under reduced pressure. Compound 8 (15.0 g, crude) was obtained as colorless liquid and it was confirmed by LC-MS (MD00842-94-P1 B2, Rt = 2.001 min).

[0203] LC-MS 1 : MD00842-94-P1A2, Rt = 1.891 min, MS cal.: 494.6, MS observed: [M+Na]+= 517.2

[0204] LC-MS: MD00842-94-P1 B2, Rt = 2.001 min, MS cal.: 494.6, MS observed: [M+Na]+= 517.2

[0205] General procedure for preparation of compound 9-1 B

[0206] (BOC)2O (15.129 g, 69.321 mmol, 15.925 mL, 1 eq.) was added to a solution of compound 9-1A (10 g, 69.321 mmol, 12.392 mL, 1 eq.) in EtOH (100 mL). The mixture was stirred at 25 °C for 1 hr. LC-MS 1 (MD00842-59-P1A111 ) showed compound 9-1 A was remained (Rt = 0.252 min) along with the desired mass of the74106498195 1Attorney Docket No: 119289-861234 product (Rt = 0.927 min). The resultant was purified by prep-HPLC (HOI condition). Compound 9-1 B (5.5 g, 21 .291 mmol, 30.714% yield, 94.6% purity) was obtained as colorless oil and confirmed by LCMS (MD00842-59-P1 F2, Rt = 0.365 min).

[0207] LC-MS 1 : MD00842-59-P1A111 , Rt = 0.927 min, MS cal.: 244.3, MS observed: [M+H]+= 245.4

[0208] LC-MS: MD00842-59-P1 F2, Rt = 0.365 min, MS cal.: 244.3, MS observed: [M+H]+= 245.2

[0209] General procedure for preparation of compound 9-1 CNFmoc9-1B 9-1 C

[0210] NaHCOs (3.577 g, 42.583 mmol, 1.657 mL, 2 eq.) and Fmoc-OSu (7.182 g, 21 .291 mmol, 1 eq.) were addede to a solution of compound 9-1 B (5.5 g, 21.291 mmol, 1 eq.) in THF (27.5 mL) and H2O (27.5 mL). The mixture was stirred at 25 °C for 1 hr. LC-MS 1 (MD00842-73-P1A1 ) showed compound 9-1 B was consumed and detected the desired mass (Rt = 0.734 min). The resultant was washed sequentially with DCM (100 mL, x2), H2O (50 mL, x2), and brine (100 mL, x1 ). The organics was dried over Na2SO4, and removed under reduced pressure. Compound 9-1 C (10 g, crude) was obtained as colorless oil and it was confirmed by LCMS (MD00842-73-P1 EA, Rt = 0.737 min).

[0211] LC-MS 1 : MD00842-73-P1A1 , Rt = 0.734 min, MS cal.: 466.6, MS observed: [M+H]+= 467.4

[0212] LC-MS: MD00842-73-P1 EA, Rt = 0.737 min, MS cal.: 466.6, MS observed: [M+H]+= 467.3

[0213] General procedure for preparation of compound 9-1moc NFmocI 25 °C, 1 hr I9-1 C 9-175106498195 1Attorney Docket No: 119289-861234

[0214] Compound 9-1C (10 g, 21.431 mmol, 1 eq.) was added in HCI / dioxane (100 mL) and stirred at 25 °C for 1 hr. LC-MS 1 (MD00842-74-P1A2) showed compound 9-1 C was consumed, and detected the desired mass (Rt =1.316 min). The reaction mixture was removed under reduced pressure, and the residue was purified by prep-HPLC (HCI condition). Compound 9-1 (6.5 g, 16.973 mmol, 79.198% yield over two steps, 95.7% purity) was obtained as a yellow solid and it was confirmed by LCMS (MD00842-74-P1 DD1 , Rt = 0.441 min).

[0215] LC-MS 1 : MD00842-74-P1A2, Rt = 1.316 min, MS cal.: 366.4, MS observed: [M+H]+= 367.2

[0216] LC-MS: MD00842-74-P1 DD1 , Rt = 0.441 min, MS cal.: 366.4, MS observed: [M+H]+= 367.3

[0217] General procedure for preparation of compound 9

[0218] Compound 9-1 (5.50 g, 14.362 mmol, 5.074e-1 eq.), BrCCI3(28.058 g, 141 .509 mmol, 13.945 mL, 5 eq.) and TEA (8.591 g, 84.905 mmol, 11 .769 mL, 3 eq.) were added to a solution of compound 8 (14 g, 28.302 mmol, 1 eq.) in ACN (140 mL) at -15 °C. The mixture was stirred at 25 °C for 50 mins. LC-MS 1 (MD00842-97- P1A1 ) showed compound 8 was consumed, and detected the desired mass (Rt = 2.615 min). The residue was purified by prep-HPLC (TFA condition). Compound 9 (2.20 g, 2.43 mmol, 8.58% yield over two steps, 94.9% purity) was obtained as colorless oil and it was confirmed by LC-MS (MD00842-97-P1 D3, Rt = 2.615 min).

[0219] LC-MS 1 : MD00842-97-P1A1 , Rt = 2.615 min, MS cal.: 859.1 , MS observed: [M+H]+= 859.5

[0220] LC-MS: MD00842-97-P1 D3, Rt = 2.615 min, MS cal.: 859.1 , MS observed: [M+H]+= 859.4

[0221] General procedure for preparation of compound 1076106498195 1Attorney Docket No: 119289-861234

[0222] TEA (4.818 g, 47.614 mmol, 6.6 mL, 19.593 eq.) was added to a solution of compound 9 (2.2 g, 2.430 mmol, 1 eq.) in DMF (15.4 mL). The mixture was stirred at 25 °C for 7 hrs. LC-MS 1 (MD00842-99-P1 A3) showed compound 9 was consumed, and detected the desired mass (Rt = 0.561 min). The resulatnt was washed sequentially with EA (20.0 mL, x2), H2O (60.0 mL, x2), and Brine (20.0 mL, x1). The organics was dried over Na2SO4, and removed under reduced pressure. Compound 10 (1 .60 g, crude) was obtained as brown oil and it was confirmed by LC- MS (MD00842-99-P1 EA, Rt = 1.523 min).

[0223] LC-MS 1 : MD00842-99-P1A3, Rt = 0.561 min, MS cal.: 636.9, MS observed: [M+H]+= 637.4

[0224] LC-MS: MD00842-99-P1 EA, Rt = 1.523 min, MS cal.: 636.9, MS observed: [M+H]+= 637.8

[0225] General procedure for preparation of compound 11-1C11-iA 11-ic

[0226] Compound 11-1A (4.5 g, 16.959 mmol, 1 eq.) and compound 11-1B (11.961 g, 42.396 mmol, 2.5 eq.) were dissolved in p-xylene (50 mL). The mixture was stirred at 130 °C for 16 hrs. LC-MS 1 (MD00574-23-P1A1JPC) showed compound 11-1 A was consumed completely and desired mass (Rt = 1.711 min) was detected. The reaction mixture was filtered and the solid obtained was washed with77106498195 1Attorney Docket No: 119289-861234 acetone, and dried under reduced pressure. The residue was purified by column chromatography (SiC>2, DCM / MeOH = 1 / 0 to 0 / 1 ). Compound 11-1C (7 g, 12.755 mmol, 75.213% yield, 85.2% purity) was obtained as a red solid and confirmed it by LC-MS (MD00574-23-P1A1 , Rt = 1.721 min).

[0227] LC-MS 1 : MD00574-23-P1A1JPC, Rt = 1.711 min, MS cal.: 467.5, MS observed: [M+H]+= 468.3

[0228] LC-MS: MD00574-23-P1A1 , Rt = 1.721 min, MS cal.: 467.5, MS observed: [M+H]+= 468.3

[0229] General procedure for preparation of compound 11-1 D11-1C 11-1D

[0230] N2H4.H2O (6.745 g, 134.736 mmol, 9 eq.) was added to a suspension of compound 11-1 C (7 g, 14.971 mmol, 1 eq.) in a mixture of MeOH and EtOH (3 / 1 , 70 mL). The mixture was stirred at 30 °C for 16 hrs. LC-MS 1 (MD00574-29- P1A1JPC) showed compound 11-1C was consumed completely and the main peak (Rt = 1.054 min) with desired mass was detected. After addition of 100 mL of concentrated HCI (1 M), white solid of phthalic acid hydrazide was precipitated. After filtration the pH of the solution was adjusted to about 9 with 3M NaOH. The mixture was freeze-dried and the crude product was purified by prep-HPLC (TFA condition). Compound 11-1 D (5 g, 13.542 mmol, 90.454% yield, 91.4% purity) was obtained as a red solid and confirmed it by LC-MS (MD00574-29-P1 A1 , Rt = 0.303 min) and HPLC (MD00574-29-P1 B1 , Rt = 1.591 min).

[0231] LC-MS 1 : MD00574-29-P1A1JPC, Rt = 1.054 min, MS cal.: 337.4, MS observed: [M+H]+= 338.3

[0232] LC-MS: MD00574-29-P1A1 , Rt = 0.303 min, MS cal.: 337.4, MS observed: [M+H]+= 637.8

[0233] HPLC: MD00574-29-P1 B1 , Rt = 1.591 min, purity: 91.4%78106498195 1Attorney Docket No: 119289-861234

[0234] General procedure for preparation of compound 11-1 F11-1 D 11-1 F

[0235] DIEA (2.068 g, 16.001 mmol, 2.644 mL, 2 eq.), HOBt (2.162 g, 16.001 mmol, 2 eq.) and EDCI (3.067 g, 16.001 mmol, 2 eq.) were added to a solution of compound 11-1 D (2.7 g, 8.000 mmol, 1 eq.) and compound 11-1E (3.868 g, 8.801 mmol, 1.1 eq.) in DMF (30 mL). The mixture was stirred at 20 °C for 2 hrs. LC-MS 1 (MD00574-32-P1A01JPC) showed compound 11-1D was consumed completely, and the main peak (Rt = 2.328 min) with desired mass was detected. The reaction mixture was diluted with H2O (100 mL) and extracted with DCM (50 mL x3). The combined organics were washed with brine (150 mL). Dried over Na2SO4, filtered and concentrated under reduced pressure. The residue was purified by column chromatography (SiO2, DCM / MeOH = 1 / 0 to 0 / 1). Compound 11-1F (3.6 g, 3.705 mmol, 46.304% yield, 78.1 % purity) was obtained as a red solid and confirmed it by LC-MS (MD00574-32-P1A2, Rt = 0.518 min) and HPLC (MD00574-32-P1 B2, Rt = 3.708 min).

[0236] LC-MS 1 : MD00574-32-P1A01JPC, Rt = 2.328 min, MS cal.: 758.9, MS observed: [M+H]+= 759.7

[0237] LC-MS: MD00574-32-P1A2, Rt = 0.518 min, MS cal.: 758.9, MS observed: [M+H]+= 759.5

[0238] HPLC: MD00574-32-P1 B2, Rt = 3.708 min, purity: 78.1 %

[0239] General procedure for preparation of compound 11-111-1F 11-179106498195 1Attorney Docket No: 119289-861234

[0240] FA (43.920 g, 954.258 mmol, 36.000 mL, 201.180 eq.) was added to compound 11-1 F (3.6 g, 4.743 mmol, 1 eq.) and stirred at 20 °C for 2 hrs. LC-MS 1 (MD00574-35-P1A4JPC) showed compound 11-1F was consumed completely, and main peak (Rt = 0.463 min) with desired mass was detected. The resultant was purified by prep-HPLC (TFA condition). Compound 11-1 (2.5 g, 3.333 mmol, 70.264% yield, 93.7% purity) was obtained as a red solid and confirmed it by LC-MS (MD00574-35-P1 A1 , Rt = 1 .953 min) and HPLC (MD00574-35-P1 B1 , Rt = 3.035 min).

[0241] LC-MS 1 : MD00574-35-P1A4JPC, Rt = 0.463 min, MS cal.: 702.8, MS observed: [M+H]+=703.5

[0242] LC-MS: MD00574-35-P1A1 , Rt = 1.953 min, MS cal.: 702.8, MS observed: [M+H]+= 703.6

[0243] HPLC: MD00574-35-P1 B1 , Rt = 3.035 min, purity: 93.7%

[0244] General procedure for preparation of compound 11

[0245] DIEA (974.021 mg, 7.536 mmol, 1.249 mL, 3 eq.), HATU (1.910 g, 5.024 mmol, 2 eq.) were added to a solution of compound 10 (1 .6 g, 2.512 mmol, 1 eq.) and compound 11-1 (1.894 g, 2.512 mmol, 1 eq.) in DMF (16.0 mL). The mixture was stirred at 25 °C for 1 hr. LC-MS 1 (MD00842-100-P1A1 ) showed compound 10 was consumed, and the desired mass (Rt = 2.140 min) was detected. The resultant was purified by prep-HPLC (TFA condition). Compound 11 (800 mg, 552 pmol, 21 .9% yield over two steps, 91 .3% purity) was obtained as a red solid and confirmed it by LC-MS (MD00842-100-P1 F1X, Rt = 2.138 min).

[0246] LC-MS 1 : MD00842-100-P1A1 , Rt = 2.140 min, MS cal.: 1321.7, MS observed: [M+H]+= 1322.6

[0247] LC-MS: MD00842-100-P1 F1X, Rt = 2.138 min, MS cal.: 1321.7, MS observed: [M+H]+= 1322.780106498195 1Attorney Docket No: 119289-861234

[0248] General procedure for preparation of compound 12

[0249] TEA (1.4 ml_) was added to a solution of compound 11 (710 mg, 490.432 pmol, 1 eq.) in DMF (5.6 m L). The mixture was stirred at 25 °C for 5 hrs. LC-MS 1 (MD00842-110-P1A2) showed compound 11 was consumed, and detected the desired mass (Rt = 1.583 min). The resultant was purified by prep-HPLC (TFA condition). Compound 12 (400 mg, 311.981 pmol, 63.614% yield, 85.757% purity) was obtained as red oil and its was confirmed by LCMS (MD00842-110-P1 F1 , Rt = 1.776 min) and HPLC (MD00842-110-P1 F3, Rt = 3.897 min).

[0250] LC-MS 1 : MD00842-110-P1 A2, Rt = 1.583 min, MS cal.: 1099.5, MS observed: [M+H]+= 1099.5

[0251] LC-MS: MD00842-110-P1 F1 , Rt = 1.776 min, MS cal.: 1099.5, MS observed: [M+H]+= 1100.7

[0252] HPLC: MD00842-1 10-P1 F3, Rt = 3.897 min, purity: 85.757%

[0253] General procedure for preparation of LP2

[0254] 18-crown-6 (197.907 mg, 748.755 pmol, 2.4 eq.) and CsF (56.869 mg, 374.377 pmol, 13.820 pL, 1.2 eq.) were added to a solution of compound 12 (400 mg, 311 .981 pmol, 1 eq.) in DMF (4 mL). The mixture was stirred at 25 °C for 2.5 hrs. LC-MS 1 (MD00842-111-P1 A2) showed compound 2 was consumed, and desired mass (Rt = 0.427 min) was detected. The residue was purified by prep-HPLC (neutral condition). The main fraction was freeze-dried together with potassium bicarbonate and subjected to column desalination operation. LP2 (150 mg, 159.669 pmol, 51.179% yield, 95.7% purity) was obtained as red oil and it was confirmed by81106498195 1Attorney Docket No: 119289-861234LCMS (MD00842-111-P1Z1 , Rt = 0.427 min) and HPLC (MD00842-111-P1Z3, Rt = 2.228 min).

[0255] LC-MS 1 : MD00842-111-P1 A2, Rt = 0.427 min, MS cal.: 899.0, MS observed: [M+H]+= 900.5

[0256] LC-MS: MD00842-111-P1Z1 , Rt = 0.427 min, MS cal.: 899.0, MS observed: [M+H]+= 900.5

[0257] HPLC: MD00842-1 11-P1Z3, Rt = 2.228 min, purity: 95.7%

[0258] General procedure for preparation of LP 2_p-hydroxyphenyl propionic acid NHS ester

[0259] p-hydroxyphenyl propionic acid NHS ester (21 .016 mg, 79.834 pmol, 5 eq.) in DMF (2 mL) was added to a solution of LP 2 (15 mg, 15.967 pmol, 1 eq.) in H2O (600 pL, 100 mM KHCO3). The mixture was stirred at 25 °C for 1 hr. LC-MS 1 (MD00842-119-P1 A11 ) showed LP 2 was consumed and detected the desired mass (Rt = 0.901 min). The residue was purified by prep-HPLC (neutral condition) LP 2_p-hydroxyphenyl propionic acid NHS ester (27 mg, contained 27 mg KHCO3, 25.396 pmol, N / A yield, 98.5% purity) was obtained as a yellow solid which was confirmed by LCMS (MD00842-119-P1 A, Rt = 0.440 min) and HPLC (MD00842-119-P1 B, Rt = 2.453 min).

[0260] LC-MS 1 : MD00842-119-P1A11 , Rt = 0.901 min, MS cal.: 1047.2, MS observed: [M+H]+= 1047.6

[0261] LC-MS: MD00842-119-P1 A, Rt = 0.440 min, MS cal.: 1047.2, MS observed: [M+H]+= 1048.2

[0262] HPLC: MD00842-119-P1 B, Rt = 2.453 min, purity: 98.5%

[0263] General procedure for preparation of compound A82106498195 1Attorney Docket No: 119289-861234

[0264] CuSO4(2.552 mg, 15.990 pmol, 2.454 pL, 1 eq.) and VcNa (6.335 mg, 31 .980 pmol, 2 eq.) and BTTAA (6.660 mg, 15.990 pmol, 1 eq.) were added successively to a solution of LP 2 _p-hydroxyphenyl propionic acid NHS ester (17 mg, 15.990 pmol, 1 eq.) and LP 1 (9.416 mg, 15.990 pmol, 1 eq.) in THF (68 pL) and MeOH (68 pL) and H2O (34 pL, 100 mM KHCO3). The mixture was stirred at 25 °C for 1 hr. LC-MS 1 (MD00842-122-P1A1 ) showed LP 2_p-hydroxyphenyl propionic acid NHS ester was consumed, and detected the desired mass (Rt = 0.698 min). The residue was purified by prep-HPLC (neutral condition). Compound A (10 mg, contained 10 mg KHCO3, 6.220 pmol, N / A yield, N / A purity) was obtained as a yellow solid confirmed by LCMS (MD00842-122-P1 A_1 , Rt = 0.698 min).

[0265] LC-MS 1 : MD00842-122-P1 A1 , Rt = 0.698 min, MS cal.: 1607.8, MS observed: [M+2H]2+= 804.5

[0266] LC-MS: MD00842-122-P1 A_1 , Rt = 0.698 min, MS cal.: 1607.8, MS observed: [M+2H]2+= 804.6Example 8. Synthesis of LP1.

[0267] Summary of LP 1 :

[0268] Project Summary

[0269] The synthetic route is based on a procedure:83106498195 1Attorney Docket No: 119289-861234

[0270] Experimental for largest scale run:

[0271] General procedure for preparation of compound 1-3NHFmoc

[0272] To a cooled (0 °C) solution of compound 1-1 (8 g, 17.354 mmol, 1.00 eq., HCI) and TEA (7.00 g, 69.4 mmol, 9.66 mL, 4.00 eq.) in DCM (160 mL) was added triphosgene (2.06 g, 6.94 mmol, 1 .08 mL, 0.400 eq.) portion-wise at 0 °C under N2 atmosphere. The resulted mixture was stirred for 0.5 h at 0 °C before dropwise addition of compound 1-2 (5.13 g, 17.3 mmol, 1.00 eq., HCI). The reaction mixture was warmed to 25 °C and stirred for further 0.5 h. LCMS-1 (MD00574-2- P1A01JPC2) showed compound 1-1 was consumed completely, and detected the desired mass (MS cal.: 709.3, MS observed: [M+H]+= 710.6). The reaction mixture was poured into icecold-water (160 mL) and stirred for 5 min. The organic layer was isolated and the aqueous layer was extracted with DCM (80 mL x 3). The combined organics were washed with brine (250 mL), and dried over Na2SO4. Finally, it was concentrated under reduced pressure to give compound 1-3 (crude, 18.0 g) and it was confirmed via LCMS (MD00574-2-P1A1_MX2).

[0273] LCMS-1 : MD00574-2-P1A01JPC2, Rt = 2.51 min, MS cal.: 709.3, MS observed: [M+H]+= 710.6.84106498195 1Attorney Docket No: 119289-861234

[0274] LCMS: MD00574-2-P1A1_MX2, Rt = 0.63 min, MS cal.: 709.3, MS observed: [M+H]+= 710.5.

[0275] General procedure for preparation of compound 1-4

[0276] 20% TEA / DMF (180 mL) was added to compound 1-3 (18.0 g, 25.3 mmol, 1 .00 eq.) and it was stirred at 25 °C for 3 hrs. LCMS-1 (MD00574-3- P1A1_MX4) showed compound 1-3 was consumed completely, and detected the desired mass (MS cal.: 487.3, MS observed: [M+H]+= 488.4). The reaction was purified by prep-HPLC (0.1 % TFA condition, ELSD). Compound 1-4 (3.80 g, 6.26 mmol, 24.7% yield for two steps, 99.2% purity (ELSD), CFsCOOH) was obtained as a light yellow solid and confirmed via LCMS (MD00574-3-P1 A3) and HNMR (MD00574-3-P1 H1).

[0277] LCMS-1 : MD00574-3-P1A1_MX4, Rt = 1.44 min, MS cal.: 487.3, MS observed: [M+H]+= 488.4.

[0278] LCMS: MD00574-3-P1A3, Rt = 1.42 min, MS cal.: 487.3, MS observed: [M+H]+= 488.4.

[0279] 1H NMR: MD00574-3-P1 H1 , 400 MHz, DMSO_d6

[0280] 5: 7.72 (br s, 3 H), 6.28 - 6.38 (m, 2 H), 4.04 (td, J = 8.38, 5.25 Hz, 1 H), 3.93 - 4.00 (m, 1 H), 2.68 - 2.82 (m, 2 H), 2.14 - 2.31 (m, 2 H), 1.81 - 1.93 (m, 1 H), 1 .58 - 1 .70 (m, 2 H), 1.53 (br d, J = 4.75 Hz, 2 H), 1 .35 - 1 .43 (m, 27 H), 1 .33 (br d, J = 7.50 Hz, 2 H).

[0281] General procedure for preparation of compound 1-685106498195 1Attorney Docket No: 119289-861234

[0282] DIEA (2.44 g, 18.9 mmol, 3.13 mL, 3.00 eq.), HOBt (1.28 g, 9.47 mmol, 1.50 eq.) and EDCI (2.42 g, 12.6 mmol, 2.00 eq.) were successively added to the mixture of compound 1-4 (3.80 g, 6.31 mmol, 1.00 eq., CFsCOOH) and compound 1- 5 (2.19 g, 9.47 mmol, 1.50 eq.) in DMF (40.0 mL). The mixture was stirred at 25 °C for 2 hrs. LCMS-1 (MD00574-5-P2A1JPC) showed compound 1-4 was consumed completely, and detected the desired mass (MS cal.: 700.4, MS observed: [M+H]+= 701 .5). The reaction mixture was added to sat. NaHCOs (200 mL), and extracted with ethyl acetate (200 mL * 3). The combined organic layer washed with brine (600 mL). Further dried it over anhydrous sodium sulfate, and concentrated it under reduced pressure. Compound 1-6 (crude, 5.3 g) was obtained as a light yellow oil which was confirmed via LCMS (MD00574-5-P2A2_CP2).

[0283] LCMS-1 : MD00574-5-P2A1JPC, Rt = 2.15 min, MS cal.: 700.4, MS observed: [M+H]+= 701.5.

[0284] LCMS: MD00574-5-P2A2_CP2, Rt = 2.15 min, MS cal.: 700.4, MS observed: [M+H]+= 701.5.

[0285] General procedure for preparation of compound 1-7

[0286] Formic acid (50.0 ml_) was added to compound 1-6 (5.3 g, 7.562 mmol, 1.00 eq.) and stirred at 40 °C for 16 hrs. LCMS-1 (MD00574-6-P1A1 _IPC) showed compound 1-6 was consumed completely, and detected the desired mass (MS cal.: 432.2, MS observed: [M+H]+= 433.3). The reaction mixture was concentrated under86106498195 1Attorney Docket No: 119289-861234 reduced pressure and lyophilized with ACN and H2O. Compound 1-7 (crude, 3.30 g) was obtained as a light yellow solid and confirmed via LCMS (MD00574-6- P2A4_CP) and HNMR (MD00574-6-P1 B1 ).

[0287] LCMS-1 : MD00574-6-P1 A1JPC, Rt = 0.37 min, MS cal.: 432.2, MS observed: [M+H]+= 433.3.

[0288] LCMS: MD00574-6-P2A4_CP, Rt = 0.39 min, MS cal.: 432.2, MS observed: [M+H]+= 433.3.

[0289] 1H NMR: MD00574-6-P1 B1 , 400 MHz, DMSO_d6

[0290] 6: 8.30 (s, 1 H), 6.32 (br dd, J = 18.51 , 7.25 Hz, 2 H), 3.95 (br d, J = 4.38 Hz, 2 H), 2.96 - 3.06 (m, 2 H), 2.71 - 2.79 (m, 4 H), 2.16 - 2.22 (m, 2 H), 2.04 (br t, J = 7.13 Hz, 2 H), 1.67 - 1.86 (m, 2 H), 1.45 - 1.55 (m, 6 H), 1.21 - 1.36 (m, 6 H).

[0291] General procedure for preparation of LP1

[0292] To a solution of compound 1-7 (2.1 g, 4.85 mmol, 1 .00 eq.) in DMF (20 mL) was added DIEA (1.25 g, 9.71 mmol, 1.60 mL, 2.00 eq.) and compound 1-8 (1 .063 g, 4.37 mmol, 0.90 eq.). The mixture was stirred at 20 °C for 2 hrs. LCMS-1 (MD00574-14-P1A02JPC) showed the desired mass (MS cal.: 560.2, MS observed: [M+H]+= 561.3) after the mentioned time. The resultant was purified by prep-HPLC (0.1 % TFA condition). LP1 (500 mg, 849 pmol, 17.4% yield for three steps, 95.2% purity) was obtained as an off-white solid and confirmed it via LCMS (MD00574-14- P1A2), HPLC (MD00574-14-P1 B4) and HNMR (MD00574-14-P1 H2).

[0293] LCMS-1 : MD00574-14-P1A02JPC, Rt= 1.19 min, MS cal.: 560.2, MS observed: [M+H]+= 561.3.

[0294] LCMS: MD00574-14-P1 A2, Rt = 0.33 min, MS cal.: 560.2, MS observed: [M+H]+= 561.3.

[0295] HPLC: MD00574-14-P1 B4, Rt = 1.95 min, purity: 95.2%.87106498195 1Attorney Docket No: 119289-861234

[0296] 1H NMR: MD00574-14-P1 H2, 400 MHz, DMSO_d6

[0297] 5: 12.13- 12.86 (m, 3 H), 8.52 (brt, J = 5.00 Hz, 1 H), 7.83 (brd, J = 7.63 Hz, 2 H), 7.73 (br s, 1 H), 7.55 (br d, J = 7.50 Hz, 2 H), 6.30 (br dd, J = 13.95, 8.07 Hz, 2 H), 4.36 (s, 1 H), 4.00-4.13 (m, 2 H), 3.19-3.25 (m, 2 H), 2.99 (brd, J =5.38 Hz, 2 H), 2.24 (br d, J = 7.00 Hz, 2 H), 2.03 (br t, J = 7.25 Hz, 2 H), 1.91 (br d, J = 7.38 Hz, 1 H), 1.62 - 1.73 (m, 2 H), 1.50 (brs, 5 H), 1.35 (brd, J = 7.13 Hz, 2 H), 1.26 (brd, J= 6.13 Hz, 4 H).Example 9. Synthesis of LP32 5

[0298] To a solution of 1 (365 mg, 1.08 mmol) in CH3CN (10 mL) was added DIPEA (750 pL, 4.43 mmol) and HATU (500 mg, 1.31 mmol), respectively. To this mixture, a solution of compound 2 (383 mg, 0.87 mmol) in 5 mL CH3CN was added dropwise and the solution stirred at room temperature for 2 h or sonicate to 1 h. The solution was concentrated under reduced pressure and the product isolated by column chromatography (CH2Ch-MeOH, 19:1), affording a red solid, (615 mg, 75%). Rf: 0.38 (5% MeOH in DCM).1H-NMR (600 MHz, CD2CI2): 58.28 (s, 1 H), 7.74 - 7.67 (m, 4H), 7.54 - 7.49 (m, 1 H), 7.46 (dd, J = 7.5, 1.0 Hz, 1 H), 7.39 - 7.33 (m, 2H), 7.28 - 7.22 (m, 2H), 7.04 (d, J = 9.0 Hz, 2H), 6.55 - 6.46 (m, 3H), 5.58 - 5.49 (m, 1 H), 4.64 - 4.50 (m, 2H), 4.22-4.17 (m, 1H), 4.16-4.12 (m, 1H), 4.07-3.98 (m, 2H), 3.50-3.40 (m, 2H), 3.27 (s, 12H), 2.27-2.22 (m, 2H), 2.05-2.00 (m, 1H) 1.99- 1.95 (m, 1H), 1.90 -1.82 (m, 3H), 1.66-1.60 (m, 3H), 1.42 (s, 9H). LCMS m / z 759.4360 (M+, 100%).

[0299] Synthesis of compound 4881064981951Attorney Docket No: 119289-8612343 4

[0300] To a solution of 3 (108 mg, 0.13 mmol) in CH2CI2 (1 mL) was added TFA (1 mL) and the solution stirred at room temperature for 1 h. The solution was completely evaporated under vacuum, and the red solid residue was dried under high vacuum to yield 4 as a red solid (100 mg, >99%).1H-NMR (600 MHz, CD2CI2): 6 8.20 (s, 1 H), 8.03 (s, 1 H), 7.66 (d, J = 7.9 Hz, 2H), 7.63 (d, J = 9.3 Hz, 2H), 7.53 (d, J = 7.6 Hz, 1 H), 7.44 (d, J = 7.5 Hz, 1 H), 7.31 (q, J = 7.7 Hz, 3H), 7.25 - 7.18 (m, 3H), 6.99 (d, J = 9.3 Hz, 2H), 6.56 - 6.50 (m, 3H), 4.59 (q, J = 8.6 Hz, 2H), 4.23 - 4.18 (m, 1 H), 4.18 - 4.15 (m, 1 H), 4.05 - 3.99 (m, 1 H), 3.94 (t, J = 7.4 Hz, 1 H), 3.46 - 3.40 (m, 3H), 3.25 (s, 12H), 2.47 - 2.40 (m, 1 H), 2.40 - 2.36 (m, 1 H), 2.06 - 1.99 (m, 1 H), 1 .99 - 1 .95 (m, 1 H), 1 .90 - 1 .83 (m, 3H), 1 .75 - 1 .68 (m, 3H). LCMS m / z 702.3687 (M+, 100%).

[0301] Synthesis of compound 6A S 8

[0302] To a solution of 4 (514 mg, 0.73 mmol) in CH3CN (10 mL) was added DIPEA (520 pL, 2.97 mmol) and HATU (333.7 mg, 0.88 mmol), respectively. To this mixture, a solution of compound 5 (83.2 pL, 0.59 mmol) in 5 mL CH3CN was added dropwise and the solution stirred at room temperature for 2 h or sonicate to 1 h. The solution was concentrated under reduced pressure and the product isolated by column chromatography (CH2Cl2-MeOH, 19 : 1 ), affording a red solid, (449.5 mg, 93%). Rf: 0.5 (10% MeOH in DCM).1HNMR (600 MHz, CD2CI2): 5 8.27 (s, 1 H), 7.7389106498195 1Attorney Docket No: 119289-861234- 7.69 (m, 2H), 7.67 (d, J = 9.2 Hz, 2H), 7.54 (d, J = 7.5 Hz, 1 H), 7.46 (d, J = 7.5 Hz, 1 H), 7.35 (t, J= 7.5 Hz, 2H), 7.26-7.21 (m, 3H), 7.02 (d, J = 9.1 Hz, 2H), 6.55 (s, 2H), 6.10 (s, 1H), 6.02 (d, J= 7.2 Hz, 1H), 4.68-4.52 (m, 2H), 4.21 -4.14 (m, 1H), 4.14-4.10(m, 1H), 4.09-4.07 (m, 1H), 3.99 (t, J=7.5Hz, 1H), 3.49-3.39 (m, 2H), 3.27 (s, 12H), 3.22 (t, J= 7.0 Hz, 2H), 3.20-3.13 (m, 3H), 2.27-2.20 (m, 2H), 1.94-1.87 (m, 2H), 1.84 - 1.78 (m, 1 H), 1.72 - 1.62 (m, 6H), 1.51 -1.43 (m, 3H), 1.37-1.29 (m,6H). LCMS m / z 826.4776 (M+, 100%).

[0303] Synthesis of LP3 (7)€ I S, ?

[0304] 6 (100 mg, 0.12 mmol) was added to a flask containing 20 mL of 20% Piperidine in DMF (v / v) and stirred for 3 h. The orange mixture was then dried by rotary evaporation and was then dissolved in minimal DCM. This solution was passed through silica and washed with more DCM to remove reaction byproducts. DCM:MeOH (8:2) was then used to elute remaining material from the silica and dried to yield 120 mg of crude mixture. LP3 (compound 7, 20 mg, 27%) was purified from this mixture by HPLC as a sticky orange oil. Retention time (tr= 8.0 min).1H-NMR (600 MHz, D2O): 08.62 (s, 1H), 7.90 (d, J= 9.30 Hz, 2H), 7.73 (s, 1H), 7.25 (dd, J= 9.30, 2.10 Hz, 2H), 6.75 (s, 1H), 6.67 (d, J= 1.56, 2H), 4.70 (t, J= 7.92 Hz, 2H), 4.47 (s, 2H), 3.87 (t, J= 7.02 Hz, 1H), 3.38 (s, J= 6.3 Hz, 2H), 3.31 (s, 12H), 3.28 (t, J= 6.90 Hz, 2H), 3.08 (q, J= 6.06 Hz, 2H), 2.08 (p, J= 2.46 Hz, 2H), 1.85 (p, J= 2.40 Hz, 2H), 1.66 (p, J= 6.78 Hz, 2H), 1.56 (p, J= 7.08 Hz, 2H), 1.43 (p, J= 7.26 Hz, 2H), 1.31 (m, 8H). ESI+ m / z 604 (M+, 100%).

[0305] HPLC Method: Gradient of 30:70 of CH3CN:H2O / 0.1% TFA, hold for 10 min then, to 70:30 over 8 minutes and 70:30 to 30:70 over 1 minutes and hold for 1 min.

[0306] Click reaction of LP3 and LP1: Synthesis of compound 9901064981951Attorney Docket No: 119289-861234

[0307] To a solution of LP3 (7, 10.0 mg, 16.55 pmol) in 379 pL of DMF, (5.7 mg, 10.17 pmol) of LP1 (8) was added as solid. After stirring the solution for 1 min at RT, 110 pL of a mixture of BTTAA (50 mM) and CuSO4 (100 mM) in a molar ratio (5:1 ) was added to the stirred reaction solution. Then, sodium ascorbate (11 pL, 1 .0 M) was added to the reaction mixture and stirred at RT for 45 min. LC-MS showed complete consumption of LP1 (8) and the crude reaction mixture was purified on HPLC using semiprep column to give compound 9 as solid orange color (6.0 mg, 50%). Retention time (tr = 6.3 min).1H-NMR (600 MHz, CD3CN): 5 8.55 (s, 1 H), 8.20 (s, 1 H), 7.90 - 7.72 (m, 6H), 7.22 - 7.10 (m, 2H), 6.57 - 6.48 (m, 2H), 4.62 - 4.52 (m, 2H), 4.47 - 4.39 (m, 1 H), 4.39 - 4.32 (m, 2H), 4.23 - 4.16 (m, 1 H), 4.15 - 4.08 (m, 1 H), 3.83 - 3.77 (m, 1 H), 3.35 - 3.28 (m, 3H), 3.22 (s, 12H), 3.09 - 3.01 (m, 4H), 2.39 - 2.32 (m, 2H), 2.16 - 2.01 (m, 6H), 1.88 - 1.65 (m, 9H), 1.60 - 1.50 (m, 7H), 1.45 - 1.34 (m, 4H), 1 .34 - 1 .22 (m, 9H). LCMS m / z 1164.6600 (M+, 100%).

[0308] HPLC Method: Gradient, hold on 30:70 of CH3CN:H2O / 0.1 % TFA for 9 minutes, then to 90:10 over one minute, hold for 5 minutes and gradient to 30:70 over 1 minute.

[0309] Synthesis of compound 10:91106498195 1Attorney Docket No: 119289-861234

[0310] To a solution of 9 (5.0 mg, 4.29 nmol) in 500 pL of dry DMF and DIPEA (299.4 nmol), a solution of DOTA-NHS ester in dry DMF (10 mg, 19.96 nmol) of DOTA-NHS ester was added and the reaction mixture was stirred at RT for 24 h. The crude reaction mixture was purified on HPLC using semiprep column to give compound 10 as solid orange color (4.0 mg, 48%). Retention time (tr= 9.9 min).1H- NMR (600 MHz, D2O): 0 7.94 (s, 1 H), 7.87 (s, 1 H), 7.31 (t, J = 8.8 Hz, 4H), 7.22 (d, J = 7.9 Hz, 2H), 6.73 (d, J = 9.1 Hz, 2H), 5.73 (s, 2H), 4.20 (dd, J = 8.8, 5.2 Hz, 2H), 4.13 - 4.07 (m, 3H), 4.03 (dd, J = 8.6, 5.3 Hz, 2H), 3.88 (s, 4H), 3.50 - 3.00 (m, 30H), 2.98 (s, 12H), 2.43 (t, J = 7.4 Hz, 2H), 2.20 - 2.15 (m, 4H), 2.11 - 2.08 (m,92106498195 1Attorney Docket No: 119289-8612341 H), 1 .92 - 1 .85 (m, 1 H), 1 .64 - 1 .60 (m, 6H), 1 .58 - 1 .53 (m, 4H), 1 .50 - 1 .47 (m, 2H), 1.44 - 1.41 (S. 2H), 1.35 - 1.31 (m, 4H), 1.27 - 1.21 (m, 4H), 1.14 - 1.09 (m, 4H). LCMS m / z 1550.8392 (M+, 100%).

[0311] HPLC Method: Gradient of 5:95 to 70:30 of CH3CN:H2O / 0.1 % TFA over 14 minutes, hold for 1 .0 min at 70:30 to 5:95 over 1 minute.

[0312] Radiosynthesis of compound 10 with [161Tb]TbCh

[0313] To a solution of 10 (100 pg, 64.48 nmol) in acetate buffer (100 pL, 0.1 M, pH ~5.0), 20 pL (2.2 mCi) of [161Tb]TbCl3 was added, and the pH was adjusted to ~ 5.0 by 0.1 M sodium acetate to a total volume of 300 pL. The reaction mixture was incubated at 85 °C for 30 min then cooled to room temperature. 1 pL of the crude reaction mixture was spotted in a TLC at EOS, 1 h, and 3 h. Another 5 pL was taken and diluted with 100 pL formulated buffer for an HPLC injection. 150 pL (~1 .0 mCi) Of the crude reaction mixture was transferred into another low bind Eppendorf tube. To this solution, 150 pL of mouse plasma was added and incubated at 37 °C for 7 day. 1 pL of the [161Tb]compound 10 / plasma mixture was spotted in a TLC at 1 h, 24 h, 48 h, and 7 days.

[0314] Isolated Radiochemical Yield (n.d.c.) 2.2 mCi (100%), purity >98% and SA = 0.034 Ci / pmol.

[0315] Radio-TLCSG condition: 0.5 M of citrate buffer pH ~ 5.0

[0316] HPLC Method: Gradient of 5:95 to 70:30 of CH3CN:H2O / 0.1 % TFA over 14 minutes, hold for 1 .0 min at 70:30 to 5:95 over 1 minute.

[0317] Formulated Buffer: 0.1 M acetate buffer pH ~ 5.0

[0318] Stickiness Test: %

[0319] Stability Study: Stable at RT in formulation buffer up to 3 h at RT and 7 days in mouse plasma.93106498195 1Attorney Docket No: 119289-8612341H-NMR of compound 3 in CD2CI294106498195 1Attorney Docket No: 119289-8612341H-NMR of compound 4 in CD2CI295106498195 1Attorney Docket No: 119289-8612341H-NMR of compound 6 in CD2CI296106498195 1Attorney Docket No: 119289-8612341H-NMR of LP3 in CDsCN97106498195 1Attorney Docket No: 119289-8612341H-NMR of compound 9 in CD3CN98106498195 1Attorney Docket No: 119289-8612341H-NMR of compound 10 in D2O:LCMS of compound 1099106498195 1Attorney Docket No: 119289-861234AnalysesRadio-TLC Data1. Radio-TLC of free [161Tb]TbCl32. Radio-TLC of crude [lslTb]Tb compoundlO in the formulated buffer at EOS100106498195 1Attorney Docket No: 119289-8612343. Radio-TLC of crude [161Tb]Tb compoundlO in the formulated buffer at lh4. Radio-TLC of crude [lslTb]Tb compoundlO in the formulated buffer after 6 h at RT101106498195 1Attorney Docket No: 119289-861234HPLC-Data1. HPLC chromatogram of crude [161Tb]Tb compoundlO in formulated buffer at EOS.Minutes2. HPLC chromatogram of crude [161Tb]Tb compoundlO in formulated buffer after 1 h at RT102106498195 1Attorney Docket No: 119289-8612343. HPLC chromatogram of crude [161Tb]Tb compoundlO in formulated buffer after 3 h at RTStability of [161Tb]Tb compoundlO in Mouse Plasma1. Radio-TLC of crude [lslTb]Tb compoundlO in the mouse plasma after 1 h at 37 °C103106498195 1Attorney Docket No: 119289-8612342. Radio-TLC of crude [161Tb]Tb compoundlO in the mouse plasma after 24 h at 37 °C3. Radio-TLC of crude [161Tb]Tb compoundlO in the mouse plasma after 7 days at 37 °C104106498195 1

Claims

Attorney Docket No: 119289-861234We claim:1 . A conjugate of Formula I:T ALinkerDiFormulawherein:T comprises a tumor-targeting moiety;RA comprises a moiety of an Auger-electron-emitting radionuclide;DI comprises a DNA-intercalating moiety; andLinker comprises a cleavable or non-cleavable linker, linking T, RA and / or DI.

2. The conjugate of claim 1 , wherein the tumor-targeting moiety T is selected from a group consisting of a moiety that binds to Prostate Specific Membrane Antigen (PSMA), Fibroblast Activation protein, a carbonic anhydrase (for example, carbonic anhydrase IX), Herceptin, RGD, EGF, PDGF, VEGF, hyaluronan, AS- 1411 , GBI-10, biotin, vitamin H, vitamin B-7, folates, Lectins, Lactoferrin, transferrin, integrin, Mannose derivates, bombesin, bradykinin, mesothelin, hepsin, mucin, estrogen receptor, milk fat globulin, telomerases, nuclear matrix proteins, prostatic acid phosphatase , SCCA, OCA, pancreas cancer associated antigen (PaA), antibody fragments or antigenic fragments of EGFR, HER2, HER3, HER4, BRCA1 CA19-9, CA59, CA-125, CD4, CD19, CD20, CD22, CD23, CD276, CD30, CD32, CD324, CD33, CD34, CD44, CD5, CD52, CD70, CD71 , CD79b, CDH1 , CDH6, CDH19, CDH17, CEA, CTLA4, , DLL3, EphA2, FLT3, GRP78, insulin receptor, IGFR, VEGF, VEGFR1 , VEGFR2, VEGFR3, Ly1 , Lym1 , Lym2, MPL, c-MET, MUC1 , MUC2, MUC3, MUC16, MUC18, NRP1 , PD1 , PD- L1 , PTK7, TGFp, TNF,TROP2, TFM1 , LAMP1 , VLA4, MART1 / MelanA, gp1OO, tyrosinase, TRP1 , TRP2, NY-ESO-1 , CDK-4, p-catenin, MUM1 , Caspase-8, KIAA0205, HPVE7, SART1 , FRAME, p15, BAGE1 , DAGE1 , RAGE1 , NAG, TAG72, CA125, p21 ras, p53, HPV16-E7, SSX , HOM-MEL55, NY-COL2, HOM-105106498195 1Attorney Docket No: 119289-861234HD397, HOM-RCC-1.14, H0M-HD21 , H0M-NSCLC11 , HOM-MEL-2.4, HOM- TES11 , RCC-3.1.3, NY-ESO-1 , MAGE1 , MAGE2, MAGE3, MAGE4 MAGE-11 , GAGE1 , GAGE6, Ha-ras, RAF, GD2, GD3, GM2, TF, sTn, gp75, EBV-LMP1 , EBV-LMP2, HPV-F4, HPV-F6, HPV-F7, PSA), AFP, CO17-1A, GA733, gp72, p- HCG, gp43, HSP-70, p17 mel, gp43, HMW, HOJ-1 , melanoma gangliosides, TAG-72, MZ2-E, PEM, LK26, Thomsen-Friedenreich (T), HOG, pancreatic oncofetal antigen, cancer antigens 15-3, 19-9, 549, 195, PROTACs, topoisomerase inhibitor, leukemia inhibitory factor, and any combination thereof.

3. The conjugate of claim 1 or claim 2, wherein the Auger-electron-emitting radionuclide RA comprises a radioisotope selected from a group consisting of iodine-125, iodine-131 , fluorine-18, gallium-68, terbium-161 , and lutetium-177.

4. The conjugate of any one of claims 1-3, wherein the DNA-intercalating moiety DI is selected from a group consisting of acridine orange, propidium iodide, DAPI (4',6-diamidino-2-phenylindole), berberine, ethidium bromide, proflavine, and thalidomide.

5. The conjugate of any one of claims 1-4, wherein the Linker is a cleavable linker selected from a biodegradable linker, a biocompatible linker, an enzymatic cleavable linker, a pH sensitive linker, a photo sensitive linker, a radiation sensitive linker, or any combination thereof.

6. The conjugate of any one of claims 1-5, wherein the Linker comprises a moiety of Formula (HA):Formula (IIA) whereinX is C, P, P-OH, or S; n2 is zero or an integer of 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10;106106498195 1Attorney Docket No: 119289-861234Ri is selected from the group consisting of hydroxyl, -NH2, -C(O)NH2, -NHC(O), -C(O)OH, -C(O)OMe, -C(O)R5, -C(O)NR5, -OC(O)Rs, -C(O)ORs-, -N3; -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl; andR2 is selected from the substituted or unsubstituted group consisting of -C1- C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl.

7. The conjugate of any one of claims 1-6, wherein the linker comprises a moiety of Formula (II B):

8. The conjugate of any one of claims 1-7, wherein the Linker comprises a moiety of Formula (IIC) :Formula (IIC) whereinX is C, P, P-OH, or S; n1 , n2, n3, n4 and n5 each is independently zero or an integer of 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10;R1 is selected from the group consisting of hydroxyl, -NH2, -C(O)NH2, - NHC(O), -C(O)OH, -C(O)OMe, -C(O)Rs, -C(O)NR5, -OC(O)Rs, -C(O)ORs-, -N3; -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl;R2 and R3 each is independently selected from the substituted or unsubstituted group consisting of -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl; and107106498195 1Attorney Docket No: 119289-861234Rs is selected from the group consisting of hydrogen, Ci-Ce alkyl, C2-C6 alkenyl, and C2-C6 alkynyl.

9. The conjugate of any one of claims 1-8, wherein the Linker comprises a moiety of HE4, PEG, or a moiety of Formula (HD):Formula (HD)10. The conjugate of any one of claims 1-9, wherein Tumor-Targeting moiety T comprises a moiety of:108106498195 1Attorney Docket No: 119289-86123411 . The conjugate of any one of claims 1-10, wherein the DNA-intercalator DI comprises a moiety of:

12. The conjugate of any one of claims 1-11 , wherein the moiety of the Auger- electron-emitting radionuclide RA comprises a moiety of:

13. The conjugate of any one of claims 1-12, wherein the bond between Linker-T, Linker-RA, or Linker-DI each independently comprises an amide bond, an amine bond, an imide bond, an ester bond, an acrylate bond, a disulfide bond, a sulfhydryl bond, an alkylsulfide bond, a sulfonyl bond, a succinimide bond, a phosphate bond, a phosphoramidate bond, a peptide bond, an oligopeptide bond, a biocompatible polymer bond, a biodegradable bond, a maleimide bond, a click chemistry bond, a photo-cleavable bond, a radiation cleavable bond, or any combination thereof.

14. The conjugate of any one of claims 1-13, having a structure of Formula (III):109106498195 1Attorney Docket No: 119289-861234Formula (III) whereinX is C, P, P-OH, or S; n1 , n2, n3, n4 and n5 each is independently zero or an integer of 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10;Ri is selected from the group consisting of hydroxyl, -NH2, -C(O)NH2, - NHC(O), -C(O)OH, -C(O)OMe, -C(O)Rs, -C(O)NR5, -OC(O)Rs, -C(O)ORs-, - N3; -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl;R2 and R3 each is independently selected from the substituted or unsubstituted group consisting of -C1-C15 alkyl, -C2-C15 alkenyl, -C2-C15 alkynyl; andRs is selected from the group consisting of hydrogen, Ci-Cs alkyl, C2-C6 alkenyl, and C2-C6 alkynyl.

15. The conjugate of any one of claims 1-14, having the following structure:110106498195 1Attorney Docket No: 119289-86123416. A composition comprising an amount of the conjugate of any one of claims 1- 15, 31 , and 32, and a carrier.

17. The composition of claim 16, wherein the amount is a therapeutically effective amount, and the carrier is a pharmaceutically acceptable carrier.

18. The composition of claims 16-17, wherein the amount is a theragnostic effective amount, and the carrier is a theragnostically acceptable carrier.

19. The composition of any one of claims 16-18, wherein the composition comprises an oral composition or a parenteral composition.

20. The composition of claim 19, wherein the parenteral composition is an injectable composition.21 . The composition of any one of claims 16-20, wherein the composition is a unit dosage form.

22. A method of inducing DNA scission or cleavage in a subject in need thereof by administering an effective amount of the conjugate of any one of claims 1- 15, 31 , and 32 or the composition of any one of claims 16-21.

23. A method of treating a tumor in a subject in need thereof by administering an effective amount of the conjugate of any one of claims 1-15, 31 , and 32 or the composition of any one of claims 16-21 .111106498195 1Attorney Docket No: 119289-86123424. The method of claims 22-23, wherein the subject is having or suspected of having breast cancer, lung cancer, colorectal cancer, colon cancer, skin cancer, bladder cancer, kidney cancer, or prostate cancer.

25. The method of any one of claims 22-24, wherein the administration is selected from the group consisting of oral administration, intravenous administration, intratumor administration, intramuscular administration, intraperitoneal administration, intraocular administration, subcutaneous administration, intradermal administration, intrathecal administration, intranasal administration, sublingual administration, buccal administration, rectal administration, and vaginal administration.

26. Use of the conjugate of any one of claims 1-15, 31 , and 32 or the composition of any one of claims 16-21 , for inducing DNA scission or cleavage, and / or for treating a tumor or cancer in a subject in need thereof.

27. The use of claim 26, wherein the subject is having or suspected of having breast cancer, lung cancer, colorectal cancer, colon cancer, skin cancer, bladder cancer, kidney cancer, or prostate cancer.

28. A theragnostic method by administering the conjugate of any one of claims 1- 15, 31 , and 32 or the composition of any one of claims 16-21 to a subject in need thereof, wherein the subject is having or suspected of having breast cancer, lung cancer, colorectal cancer, colon cancer, skin cancer, bladder cancer, kidney cancer, or prostate cancer.

29. The theragnostic method of claim 28, wherein the administration is selected from the group consisting of oral administration, intravenous administration, intratumor administration, intramuscular administration, intraperitoneal administration, intraocular administration, subcutaneous administration, intradermal administration, intrathecal administration, intranasal administration, sublingual administration, buccal administration, rectal administration, and vaginal administration.112106498195 1Attorney Docket No: 119289-86123430. A building block selected from LP-1 ; LP-1-1 ; L-2; Simplified LP-2; and LP-3.

31. A conjugate selected from TRAIL-1 ; TRAIL-2; TRAIL-3; TRAIL-4; TRAIL-5; TRAIL-6; TRAIL-7; TRAIL-8; TRAIL-9; TRAIL-10; TRAIL-11 ; and TRAIL-12.

32. A conjugate selected from Click_1 ; Click_2; Click_3; Click_4; Click_5; Click_6; Click_7; and Click_8.113106498195 1

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