Glypican-3 targeting compositions and uses thereof

Abstract

Provided herein are GPC-3 binding peptides and non-radionuclide conjugates thereof, and compositions comprising the same. The GPC-3 targeting peptides and non-radionuclide conjugates thereof have avidity to GPC3. The GPC-3 targeting peptides described herein can optionally further comprise a linker connecting the GPC-3 targeting peptide to a non-radionuclide moiety. The non-radionuclide conjugates described herein comprises the GPC-3 targeting peptide linked to a non-radionuclide moiety, either directly or through a linker. Further provided herein are methods of diagnosing and / or treating a disease or condition characterized by GPC-3 expression (e.g., overexpression), such as various cancers, by administering the described peptides and non-radionuclide conjugates thereof, or compositions thereof.

Description

Attorney Docket No. 137521-10320GLYPICAN-3 TARGETING COMPOSITIONS AND USES THEREOFCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Application No. 63 / 730,701, filed on December 11, 2024, which is incorporated herein by reference in its entirety for any purpose.JOINT RESEARCH AGREEMENT

[0002] Subject matter disclosed herein was developed, and the claimed invention was made by, or on behalf of, one or more parties to a Joint Research Agreement (JRA), within the meaning of 35 U.S.C. § 100(h) and 37 C.F.R. § 1.9(e), that was in effect on or before the effective filing date of the claimed invention. Said one or more parties to the JRA consist of PeptiDream, Inc. (Kanagawa, Japan) and RayzeBio, Inc. (San Diego, CA, U.S.A.). The claimed invention was made as a result of activities undertaken within the scope of said Joint Research Agreement.SEQUENCE LISTING

[0003] This application contains a Sequence Listing XML file, which has been submitted electronically in .xml format as part of the specification and is incorporated herein by reference in its entirety. Said XML file, created on October 22, 2025, is named "137521 -10320_SL.xml,” and is 1,767,112 bytes in size.TECHNICAL FIELD

[0004] The present disclosure relates to conjugates comprising peptides that bind to glypican-3 (GPC3) and to compositions comprising such conjugates and uses thereof. In an aspect, the present disclosure relates to the use of said conjugates in preventing, suppressing or treating a disease or disorder characterized by overexpression or decreased expression of GPC3 in diseased tissue, such as in a tumor where GPC3 is overexpressed.BACKGROUND

[0005] Glypican-3 (GPC3) is a heparan sulfate (HS) glycoprotein, belonging to the sulfate heparan proteoglycan family, and which is anchored on the cell membrane surface by phosphatidylinositol (GPI) anchor. The GPC3 core protein comprises 580 amino acids, with a molecular weight of about 70 kDa. It is cut by furin (Furin), generating a 40 kDa N-terminal subunit and a 30 kDa C-terminal subunit, connected to each other by a disulfide bond. GPC3's two HS side chain is combined at the position close to the C end (Takahiro Nishida, Hiroaki Kataoka. Glypican 3-Targeted Therapy in Hepatocellular Carcinoma, Cancer 2019; 11 (9): 1339).

[0006] GPC3 can play an important role in the cell proliferation of embryo layer tissue. Deletion of GPC3 gene can cause excessive growth syndrome, namely Simpson-Golabi-Behmel syndrome (SGBS). GPC3 can be clearly expressed throughout the entire fetal stage, and after birth to adult stage, except for placental, breast, mesodermal, ovarian, lung and kidney tissue with weak expression, other normal tissues have no obvious expression.

[0007] Abnormal GPC3 expression has been found in multiple tumour tissues of adult, such as hepatocellular carcinoma (HCC), lung squamous carcinoma, gastric cancer, ovarian cancer and so on. Especially highly expressed in HCC cells, GPC3 improves autocrine / paracrine canonical Wnt signal transmission, and promotes growth and invasion of HCC cells (Capurro Ml, Xiang Y-Y, Lobe C, Filmus J. Glypican-3 promotes the growth of hepatocellular carcinoma by-stimulating canonical Wnt signaling; Cancer Res 2005, 65(14): 6245-54.). Immunohistochemical staining detection shows that about 70% of HCC patient tumor tissue exhibits high GPC3 protein expression (Capurro M, Wanless IR, Sherman M, et al. Glypican-3: a novel serum and histochemical marker for hepatocellular carcinoma;1MEl\59069747.vlAttorney Docket No. 137521-10320Gastroenterology 2003, 125(1) :89-97). Thus, GPC3 is considered as a candidate target for tumor treatment.

[0008] Codrituzumab (also known as GC33 antibody) is a recombinant humanized monoclonal antibody developed in Japan by Chugai Pharmaceutical Co., which binds to the region of GPC3 protein proximal membrane end. GC33 antibody targets GPC3 positive HCC cells, and can generate antibody dependent cell toxicity (ADCC). In phase I clinical trials, Codrituzumab shows good immune tolerance. The HCC patient can generate an anti-tumour effect (Ikeda M, Ohkawa S, Okusaka T, et al. Japanese phase I study of GC33, a-antibody against glypican-3 for advanced hepatocellular carcinoma. Cancer Sci. 2014, 105, 455-462). However, in the phase II clinical trial with 185 late-stage liver cancer patients, the therapeutic efficacy of Codrituzumab was not impressive compared with the control group.

[0009] Therefore, novel GPC3-binding peptides and compositions comprising the GPC3-binding peptide are both useful and desired.SUMMARY

[0010] Provided herein is a peptide that has avidity for Glypican 3 (GPC3), or a pharmaceutically acceptable salt thereof, wherein the peptide comprises an amino acid sequence of Formula I:Formula I, wherein R is -CH2-CO-; and,(i)Xi is MeLys, MeLysAc, MeAla, MeGlu, MeGIn, or CrpG, wherein the N-terminal nitrogen of Xi is attached to the C(O) of R;X2 is Melle, MeGIn, MeLys, MeHseMe, MeLysCOpipzaa, or MeGlu;X3 is Asp, MeAsp, Hgn, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, Hva, or (S)Mor(2CO);X4 is MeGIn, MeLys, MeAspapCOpipzaa, MeGlu, or CrpG;X5is F4COO, MeF4COO, Y, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A;Xe is TMe, altMe, lie, or Leu;X7 is TMe, lie, Vai, Leu, Gcpe, or Eva;X8is Tyr, Cha, F3OH, F4aao, F4COO, F4aa, Yae, YaeCOpipzaa, F4F, F4U, Aph, 3Py6NH2, 4Py, F4(CONMeOH), F4CONHMe, F4CONMe2, or (PyrrCOO)A;X9is MeNal27N, MeF3Me4C, MeF34dOMe, or MeW7N;X10 is Gly, MeGly, D-Ala, D-Ser, or D-Pro;X11 is 3Py6Ph, 3Py6Pyrazol1 , F41 Me4Pyz, 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61ap4, F41 apPyz, F42Py, F44Py, or Bph4C; and,X12 is MeCys or Cys, wherein sulfur of the X12 side chain is covalently attached to R, and2MEl\59069747.vlAttorney Docket No. 137521-10320 carboxylic acid of X12 is replacedwherein Xi is MeGlu or CrpG; or X2 is MeLys; or X3 is MeAsp, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, (S)Mor(2CO), or Hva; or X4 is MeGlu, or CrpG; or X5 is MeF4C00, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A; or Xe is alTMe or Leu; or X? is TMe, Vai, Leu, Gcpe, or Eva; or Xs is Cha, F3OH, F4F, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A; or X9is MeF3Me4C, MeF34dOMe, or MeW7N; or X is MeGly, D-Ala, D-Ser, or D-Pro; or Xu is 3Py6Pyrazol1 , 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61 ap4, F41apPyz, F42Py, F44Py, or Bph4C; or X12 is Cys; or(ii) Xi to X12 have a sequence from Table 1 , 2, 3, 4, or 9A, or 20A (and, in some aspects, from Table 1 , 2, 3, 4, or 9A).

[0011] In certain embodiments, in the peptide or pharmaceutically acceptable salt thereof, Xi to X12 of the peptide of Formula (I) have a sequence from Table 1 , 2, 3, 4, or 9A, or 20A (and, in some aspects, from Table 1 , 2, 3, 4, or 9A).

[0012] Also provided herein is a peptide that has avidity for Glypican 3 (GPC3), or a pharmaceutically acceptable salt thereof, wherein the peptide comprises an amino acid sequence of Formula II:Formula II wherein:R* is -CH2-S-CH2-CO-; each Z is independently selected from H and Me; and,(i)A1 is the side chain of Lys, LysAc, Ala, Glu, Gin, or CrpG;3MEl\59069747.vlAttorney Docket No. 137521-10320A2 is the side chain of lie, Gin, Lys, HseMe, LysCOpipzaa, or Glu;A3 is the side chain of Asp, Hgn, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, Hva, or (S)Mor(2CO);A4 is the side chain of Gin, Lys, AspapCOpipzaa, Glu, or CrpG;A5is the side chain of F4COO, Y, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A;Ae is the side chain of TMe, altMe, lie, or Leu;Az is the side chain of Tme, lie, Vai, Leu, Gcpe, or Eva;As is the side chain of Tyr, Cha, F3OH, F4aao, F4COO, F4aa, Yae, YaeCOpipzaa, F4F, F4U, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A;A9 is the side chain of Nal27N, F3Me4C, F34dOMe, or W7N;Aw is the side chain of Gly, D-Ala, D-Ser, or D-Pro;An is the side chain of 3Py6Ph, 3Py6Pyrazol1 , F41 Me4Pyz, 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61 ap4, F41 apPyz, F42Py, F44Py, or Bph4C; and, wherein A1 is the side chain of Glu or CrpG; or A2 is the side chain of Lys; or A3 is the side chain of Asp, Ser, Thr, Ala, or Lys, Glu, Asn, diMeDap, (S)Mor(2CO), or Hva; or A4 is the side chain of Glu or CrpG; or A5 is the side chain of F4COO, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A; or As is the side chain of alTMe or Leu; or A7 is the side chain of Tme, Vai, Leu, Gcpe, or Eva; or As is the side chain of Cha, F3OH, F4F, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A; or A9is the side chain of F3Me4C, MeF34dOMe, or MeW7N; or Aw is the side chain of Gly, D-Ala, D-Ser, or D-Pro; or An is the side chain of 3Py6Pyrazol1 , 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61 ap4, F41 apPyz, F42Py, F44Py, or Bph4C; or(ii) A1 to A11 correspond to the side chains of Xi to Xi 1 of a sequence from T able 1 , 2, 3, 4, or 9A, or 20A (and, in some aspects, from Table 1 , 2, 3, 4, or 9A).

[0013] In certain embodiments, in the peptide or pharmaceutically acceptable salt thereof, Xi to Xw of the peptide of Formula (I) have a sequence from Table 1 , 2, 3, 4, 9A, or 20A. In certain embodiments, in the peptide or pharmaceutically acceptable salt thereof, Xi to Xw of the peptide of Formula (I) have a sequence from Table 1 , 2, 3, 4, or 9A. In certain embodiments, in the peptide or pharmaceutically acceptable salt thereof, Xi to X12 of the peptide of Formula (I) have the sequence of SEQ ID NO: 226, 520, or 521. In certain embodiments, in the peptide or pharmaceutically acceptable salt thereof, Xi to X12 of the peptide of Formula (I) have the sequence of SEQ ID NO: 226. In certain embodiments, in the peptide or pharmaceutically acceptable salt thereof, Xi to X12 of the peptide of Formula (I) have the sequence of SEQ ID NO: 520. In certain embodiments, in the peptide or pharmaceutically acceptable salt thereof, Xi to X12 of the peptide of Formula (I) have the sequence of SEQ ID NO: 521 .

[0014] In certain embodiments, the peptide is not any of SEQ ID Nos: 1-72. In certain embodiments, the peptide is not a peptide from Table A.

[0015] Further provided herein is a conjugate comprising the peptide or pharmaceutically acceptable salt thereof of the invention disclosed herein, covalently linked to a non-radionuclide moiety.

[0016] In certain embodiments, in the conjugate or pharmaceutically acceptable salt thereof, the non-radionuclide moiety is a substance or a payload molecule selected from the group consisting of: a nucleotide, a small molecule4MEl\59069747.vlAttorney Docket No. 137521-10320(e.g., with a M.W. of less than about 1000 Da), a medium sized molecule (e.g., with a M.W. of about 1 ,000-2,500 Da), a large sized molecule (e.g., with a M.W. of >2,500 Da), a polymer compound, a protein, a peptide, a tag, a biological fragment, a carrier including pharmaceutical compound, or a combination thereof.

[0017] In certain embodiments, the peptide of the conjugate has a sequence from Table 1 , 2, 3, 4, 9A, or 20A. In certain embodiments, the peptide of the conjugate has a sequence from T able 1 , 2, 3, 4, or 9A. In certain embodiments, the peptide of the conjugate comprises the sequence of any one of SEQ ID NOs: 226, 520, and 521. In certain embodiments, the peptide of the conjugate comprises the seqeunce of any one of SEQ ID NOs: 226. In certain embodiments, the peptide of the conjugate comprises the seqeunce of any one of SEQ ID NOs: 520. In certain embodiments, the peptide of the conjugate comprises the seqeunce of any one of SEQ ID NOs: 521. In certain embodiments, the conjugate is a conjugate from Table 5, 6, 7, 8, 9B, or 20B. In certain embodiments, the conjugate is a conjugate from Table 5, 6, 7, 8, or 9B.

[0018] In certain embodiments, the non-radionuclide moiety is covalently attached to the side chain of the first amino acid of the peptide (Xi, Ai), the side chain of the second amino acid of the peptide (X2, A2), the side chain of the third amino acid of the peptide (X3, A3), the side chain of the fourth amino acid of the peptide (X4, A4), the side chain of the eighth amino acid of the peptide (Xs, As), the C-terminus of the C-terminal amino acid of the peptide (X12, Q, when Q is -NH2); or the side chain of the thirteenth amino acid (X13, Q, when Q is D-Lys) when present.

[0019] In certain embodiments, the conjugate or pharmaceutically acceptable salt thereof further comprises a linker that covalently connects the peptide to the non-radionuclide moiety.

[0020] In certain embodiments, the linker comprises 3 to 10 intervening non-hydrogen, organic atoms between the non-radionuclide moiety and the peptide.

[0021] In certain embodiments, the peptide is directly attached to the non-radionuclide moiety.

[0022] In certain embodiments, the conjugate has the following structure (SEQ ID NOs: 641-643, respectively):5MEl\59069747.vlAttorney Docket No. 137521-10320

[0023] Also provided herein is a conjugate or pharmaceutically acceptable salt thereof having the followingwherein Xis a non-radionuclide moiety as used herein.

[0024] Also provided herein is a conjugate or pharmaceutically acceptable salt thereof having the following structure (SEQ ID NO: 614):6MEl\59069747.vlAttorney Docket No. 137521-10320wherein Xis a non-radionuclide moiety as used herein.

[0025] Also provided herein is a conjugate or pharmaceutically acceptable salt thereof having the following structure (SEQ ID NO: 615):7MEl\59069747.vlAttorney Docket No. 137521-10320 wherein X is a non-radionuclide moiety as used herein.

[0026] In certain embodiments, the conjugate is not a conjugate of Table B, or wherein the conjugate is a conjugate from Table 5, 6, 7, 8, 9B, or 20B.

[0027] Further provided herein is a pharmaceutical composition comprising the peptide or pharmaceutically acceptable salt thereof of the invention described herein, and a pharmaceutically acceptable excipient or carrier.

[0028] Further provided herein is a pharmaceutical composition comprising the conjugate or pharmaceutically acceptable salt thereof of the invention as described herein, and a pharmaceutically acceptable excipient or carrier.

[0029] Further provided herein is a method of treating a disease or disorder characterized by overexpression of glypican 3 (GPC3) in a subject in need of treatment, the method comprising administering to the subject a therapeutically effective amount of the peptide or pharmaceutically acceptable salt thereof of the invention described herein, the conjugate or pharmaceutically acceptable salt thereof of the invention described herein, or the pharmaceutical composition of the invention described herein.

[0030] Further provided herein is a method of diagnosing a disease or disorder characterized by overexpression of GPC3 in a subject in need thereof, comprising administering to the subject, or contacting a sample from the subject in vitro, the peptide or pharmaceutically acceptable salt thereof of the invention described herein, the conjugate or pharmaceutically acceptable salt thereof of the invention described herein, or the pharmaceutical composition of the invention described herein; and detecting binding to GPC3 by the peptide or the conjugate, thereby determining expression level of GPC3 in the subject.

[0031] In certain embodiments, the disease or disorder is cancer, such as a solid tumor or a hematological cancer.

[0032] In certain embodiments, the cancer is hepatocellular carcinoma, squamous cell carcinoma of the lung, lung adenocarcinoma, germ cell tumors, hepatoblastoma, wilms tumor, malignant rhabdoid tumors, rhabdomyosarcoma, liposarcoma, thyroid cancers, pancreatic cancer, small bowel cancer, small cell neuroendocrine carcinoma (SONG), hormonally treated, castration resistant prostatic adenocarcinoma, ovarian cancer, gastric cancer, esophageal carcinoma, or malignant melanoma.

[0033] Further provided herein is a kit, tester, or composition for determining expression level of GPC3 in a sample, wherein the kit, tester, or composition comprises the peptide or pharmaceutically acceptable salt thereof of the invention described herein, the conjugate or pharmaceutically acceptable salt thereof of the invention described herein, or the pharmaceutical composition of the invention described herein.

[0034] In certain embodiments, the kit, tester, or composition is adapted for use in a method of diagnosing a disease or disorder characterized by an overexpression or a decreased expression of GPC3.

[0035] In certain embodiments, the sample is from a subject suspected of having a disease or disorder characterized by an overexpression or a decreased expression of GPC3.

[0036] Further provided herein is a use of the peptide, conjugate, or pharmaceutically acceptable salt thereof of the invention described herein, in the manufacture of a medicament for diagnosing and / or treating a disease or disorder characterized by an overexpression or a decreased expression of GPC3.

[0037] Further provided herein is a peptide, conjugate, or pharmaceutically acceptable salt thereof the invention described herein, for use in diagnosing and / or treating a disease or disorder characterized by an overexpression or a decreased expression of GPC3.8MEl\59069747.vlAttorney Docket No. 137521-10320

[0038] In all aspects of this disclosure the substance or payload molecule excludes any radioactive materials. Examples of the substance that are excluded are: radioisotope, radionuclide or radiopharmaceutical (as defined herein below), or any compound having radioactive component. In all aspects of this disclosure, the substance further excludes any chelators for radioisotope or radionuclide conjugation, regardless of whether the chelator is connected to the peptide directly or via a linker. Accordingly, a complex, conjugate or Peptide-Drug Conjugate (PDC) described herein does not encompass any compound containing a chelator for radioisotope conjugation, and does not encompass a radioisotope.

[0039] Since the peptide of the present technology has the ability to bind to the GPC3, it is possible for the peptide to target and transport compounds having pharmacological actions to the GPC3, such as low molecular weight compounds, middle molecular weight compounds, high molecular weight compounds, peptides, proteins, antibodies, and nucleic acids.

[0040] Other aspects and features of the present disclosure will become apparent to those ordinarily skilled in the art upon review of the following description of specific embodiments in conjunction with the accompanying drawings.INCORPORATION BY REFERENCE

[0041] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference for the specific purposes identified herein.

[0042] All features of embodiments which are described in this disclosure are not mutually exclusive and can be combined with one another. For example, elements of one embodiment can be utilized in the other embodiments without further mention. A detailed description of specific embodiments is provided herein below with reference to the accompanying drawings in which:DETAILED DESCRIPTION

[0043] It should be understood that both the general descriptions and the detailed description below are merely illustrative and descriptive and do not limit the present technology of the present application. Those of skill in the art will recognize that there are numerous variations and modifications of this present disclosure, which are encompassed within its scope.

[0044] Although various features of the present disclosure may be described in the context of a single embodiment, the features may also be provided separately or in any suitable combination. Conversely, although the present disclosure may be described herein in the context of separate embodiments for clarity, the present disclosure may also be implemented in a single embodiment.

[0045] The headings used in the present specification are for structural purposes only and must not be construed as limiting the subject matter described.

[0046] In the present specification, the use of the singular form includes the plural form unless otherwise specified. In the present specification, the use of "or (or)” means "and / or (and / or)” unless otherwise stated. Furthermore, terms such as "element” or "component” encompass both an element and a component including one unit and an element and a component including two or more subunits unless when otherwise specified.

[0047] The recitation herein of numerical ranges by endpoints is intended to include all numbers subsumed within that range (e.g., a recitation of 1 to 5 includes 1 , 1.5, 2, 2.75, 3, 3.80, 4, 4.32, and 5).

[0048] All terms are intended to be understood as they would be understood by a person skilled in the art. Unless9MEl\59069747.vlAttorney Docket No. 137521-10320 defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the disclosure pertains.

[0049] The following definitions supplement those in the art and are directed to the current application and are not to be imputed to any related or unrelated case, e.g., to any commonly owned patent or application. Although any methods and materials similar or equivalent to those described herein can be used in the practice for testing of the present disclosure, the preferred materials and methods are described herein. Accordingly, the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting.I. Definitions

[0050] As used herein and in the appended claims, the singular forms "a,” "an,” and "the” include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "an agent” includes a plurality of such agents, and reference to "the cell” includes reference to one or more cells (or to a plurality of cells) and equivalents thereof known to those skilled in the art, and so forth. When ranges are used herein for physical properties, such as molecular weight, or chemical properties, such as chemical formulae, all combinations and subcombinations of ranges and specific embodiments therein are intended to be included.

[0051] The term "about” or "approximately” can mean within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, i.e., the limitations of the measurement system. For example, "about” can mean within 1 or more than 1 standard deviation, per the practice in the art. Alternatively, "about” can mean a range of up to 20%, up to 15%, up to 10%, up to 5%, or up to 1 % of a given value. Alternatively, particularly with respect to biological systems or processes, the term can mean within an order of magnitude, within 5-fold, or within 2-fold, of a value.

[0052] The term "moiety” {i.e. in Table 1.1) refers to a specific segment or functional group of a molecule. Chemical moieties are often recognized chemical entities embedded in or appended to a molecule. In some embodiments, the term "moiety” describes a conjugate or payload molecule added at the defined position or amino acid. Some examples of a "moiety” are, but are not limited to, a chemical compound, payload molecule, or a linker, wherein the linker is optionally connected to a chemical compound, wherein the chemical compound is, for example, a payload molecule, chemical label, capture agent, or fluorophore.

[0053] As used herein, the term "conjugate” refers to a peptide described herein that is bound to a moiety or nonradionuclide. For example, a peptide described herein may be bound to a chemical compound, payload molecule, or a linker, wherein the linker is optionally connected to a chemical compound, wherein the chemical compound is, for example, a payload molecule, chemical label, capture agent, or fluorophore.

[0054] As used herein, the term "radiopharmaceutical” refers to a conjugate comprising a radionuclide. In some embodiments, the radionuclide is covalently bound to the conjugate in the radiopharmaceutical. In some embodiments, the radionuclide is bound to the conjugate through a metal chelator.

[0055] The term "comprising” (and related terms such as "comprise” or "comprises” or "having” or "including”) are to be construed in an open, inclusive sense, that is, as "including, but not limited to.” The term "comprising” (and related terms such as "comprise” or "comprises” or "having” or "including”) is not intended to exclude that in other certain embodiments, for example, an embodiment of any composition of matter, composition, method, or process, or the like,10MEl\59069747.vlAttorney Docket No. 137521-10320 described herein, "consist of' or "consist essentially of' the described features.

[0056] "Amino" refers to the -NH2 radical.

[0057] "Cyano" refers to the -CN radical.

[0058] "Nitro" refers to the -NO2 radical.

[0059] "Oxo" refers to the =0 radical.

[0060] "Imino" refers to the =N-H radical.

[0061] "Oximo" refers to the =N-OH radical.

[0062] "Hydrazino" refers to the =N-NH2 radical.

[0063] "Hydroxy” or "hydroxyl” refers to the -OH radical.

[0064] "Hydroxyamino” refers to the -NH-OH radical.

[0065] "Acyl” refers to a substituted or unsubstituted alkylcarbonyl, substituted or unsubstituted alkenylcarbonyl, substituted or unsubstituted alkynylcarbonyl, substituted or unsubstituted cycloalkylcarbonyl, substituted or unsubstituted heterocycloalkylcarbonyl, substituted or unsubstituted arylcarbonyl, substituted or unsubstituted heteroarylcarbonyl, amide, or ester, wherein the carbonyl atom of the carbonyl group is the point of attachment. Unless stated otherwise specifically in the specification, an alkylcarbonyl group, alkenylcarbonyl group, alkynylcarbonyl group, cycloalkylcarbonyl group, amide group, or ester group is optionally substituted, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like.

[0066] "Alkyl” refers to an optionally substituted straight-chain, or optionally substituted branched-chain saturated hydrocarbon monoradical. An alkyl group can have from one to about twenty carbon atoms, from one to about ten carbon atoms, or from one to six carbon atoms. Examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, 2-methyl-1 -propyl, 2-methyl-2-propyl, 2-methyl-1 -butyl, 3-methyl-1 -butyl, 2-methyl-3-butyl, 2,2-dimethyl-1- propyl, 2-methyl-1 -pentyl, 3-methyl-1 -pentyl, 4-methyl-1 -pentyl, 2-methyl-2-pentyl, 3-methyl-2-pentyl, 4-methyl-2- pentyl, 2,2-dimethyl-1 -butyl, 3,3-dimethyl-1 -butyl, 2-ethyl-1 -butyl, n-butyl, isobutyl, sec-butyl, t-butyl, n-pentyl, isopentyl, neopentyl, tert-amyl, and hexyl, and longer alkyl groups, such as heptyl, octyl, and the like. Whenever it appears herein, a numerical range such as “Ci-Ce alkyl” means that the alkyl group consists of 1 carbon atom, 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms or 6 carbon atoms, although the present definition also covers the occurrence of the term "alkyl” where no numerical range is designated. In some embodiments, the alkyl is a C1-C10 alkyl, a C1-C9 alkyl, a Ci-Cs alkyl, a C1-C7 alkyl, a Ci-Ce alkyl, a C1-C5 alkyl, a C1-C4 alkyl, a C1-C3 alkyl, a C1-C2 alkyl, or a Ci alkyl. Unless stated otherwise specifically in the specification, an alkyl group is optionally substituted, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like. In some embodiments, the alkyl is optionally substituted with oxo, halogen, -CN, -CF3, -OH, - OMe, -NH2, -NO2, or -C=CH. In some embodiments, the alkyl is optionally substituted with oxo, halogen, -CN, - CF3, -OH, or -OMe. In some embodiments, the alkyl is optionally substituted with halogen.

[0067] "Alkylene” refers to a straight or branched divalent hydrocarbon chain. Unless stated otherwise specifically in the specification, an alkylene group may be optionally substituted, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like. In some embodiments, an alkylene is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, an alkylene is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, the alkylene is11MEl\59069747.vlAttorney Docket No. 137521-10320 optionally substituted with halogen. In some embodiments, the alkylene is -CH2-, -CH2CH2-, -CH2CH2CH2-, or - CH2CH(CH3)CH2-. In some embodiments, the alkylene is -CH2-. In some embodiments, the alkylene is -CH2CH2-. In some embodiments, the alkylene is -CH2CH2CH2-.

[0068] "Alkenyl” refers to an optionally substituted straight-chain, or optionally substituted branched-chain hydrocarbon monoradical having one or more carbon-carbon double-bonds. In some embodiments, an alkenyl group has from two to about ten carbon atoms, or two to about six carbon atoms. The group may be in either the cis or trans configuration about the double bond(s), and should be understood to include both isomers. Examples include, but are not limited to, ethenyl (-CHOH2), 1-propenyl (-CH2CHOH2), isopropenyl [-C(CH3)=CH2], butenyl, 1 ,3-butadienyl, and the like. Whenever it appears herein, a numerical range such as "C2-C6 alkenyl” means that the alkenyl group may consist of 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, or 6 carbon atoms, although the present definition also covers the occurrence of the term "alkenyl” where no numerical range is designated. In some embodiments, the alkenyl is a C2-C10 alkenyl, a C2-C9 alkenyl, a C2-C8 alkenyl, a C2-C7 alkenyl, a C2-C6 alkenyl, a C2- C5 alkenyl, a C2-C4 alkenyl, a C2-C3 alkenyl, or a C2 alkenyl. Unless stated otherwise specifically in the specification, an alkenyl group is optionally substituted, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like. In some embodiments, an alkenyl is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, an alkenyl is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, the alkenyl is optionally substituted with halogen.

[0069] The term "alkenylene” or "alkenylene chain” refers to an optionally substituted straight or branched divalent hydrocarbon chain in which at least one carbon-carbon double bond is present linking the rest of the molecule to a radical group. In some embodiments, the alkenylene is -CH=CH-, -CH2CH=CH-, or -CHOHCH2-. In some embodiments, the alkenylene is -CH=CH-. In some embodiments, the alkenylene is -CH2CH=CH-. In some embodiments, the alkenylene is -CH=CHCH2-.

[0070] “Alkynyl” refers to an optionally substituted straight-chain or optionally substituted branched-chain hydrocarbon monoradical having one or more carbon-carbon triple-bonds. In some embodiments, an alkynyl group has from two to about ten carbon atoms, more preferably from two to about six carbon atoms. Examples include, but are not limited to, ethynyl, 2-propynyl, 2-butynyl, 1 ,3-butadiynyl, and the like. Whenever it appears herein, a numerical range such as "C2-C6 alkynyl” means that the alkynyl group may consist of 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, or 6 carbon atoms, although the present definition also covers the occurrence of the term "alkynyl” where no numerical range is designated. In some embodiments, the alkynyl is a C2-C10 alkynyl, a C2-C9 alkynyl, a C2-C8 alkynyl, a C2-C7 alkynyl, a C2-C6 alkynyl, a C2-C5 alkynyl, a C2-C4 alkynyl, a C2-C3 alkynyl, or a C2 alkynyl. Unless stated otherwise specifically in the specification, an alkynyl group is optionally substituted, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like. In some embodiments, an alkynyl is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or - NO2. In some embodiments, an alkynyl is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, the alkynyl is optionally substituted with halogen. The term "alkynylene” refers to an optionally substituted straight-chain or optionally substituted branched-chain divalent hydrocarbon having one or more carboncarbon triple-bonds.12MEl\59069747.vlAttorney Docket No. 137521-10320

[0071] “Alkylamino” refers to a radical of the formula -N(Ra)2 where Rais an alkyl radical as defined, or two Ra, taken together with the nitrogen atom, can form a substituted or unsubstituted C2-C7 heterocy loal ky I ring. Unless stated otherwise specifically in the specification, an alkylamino group may be optionally substituted, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like. In some embodiments, an alkylamino is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, an alkylamino is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, the alkylamino is optionally substituted with halogen.

[0072] "Alkoxy” refers to a radical of the formula -ORawhere Rais an alkyl radical as defined. Unless stated otherwise specifically in the specification, an alkoxy group may be optionally substituted, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like. In some embodiments, an alkoxy is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, an alkoxy is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, the alkoxy is optionally substituted with halogen.

[0073] “Aminoalkyl” refers to an alkyl radical, as defined above, that is substituted by one or more amines. In some embodiments, the alkyl is substituted with one amine. In some embodiments, the alkyl is substituted with one, two, or three amines. Hydroxyalkyl include, for example, aminomethyl, aminoethyl, aminopropyl, aminobutyl, or aminopentyl. In some embodiments, the hydroxyalkyl is aminomethyl.

[0074] The term "aryl” refers to a radical comprising at least one aromatic ring wherein each of the atoms forming the ring is a carbon atom. Aryl groups can be optionally substituted. Examples of aryl groups include, but are not limited to phenyl, and naphthyl. In some embodiments, the aryl is phenyl. Depending on the structure, an aryl group can be a monoradical or a diradical ( / .e., an arylene group). Unless stated otherwise specifically in the specification, the term "aryl” or the prefix "ar-”(such as in "aralkyl”) is meant to include aryl radicals that are optionally substituted. In some embodiments, an aryl group comprises a partially reduced cycloalkyl group defined herein (e.g., 1 ,2- dihydronaphthalene). In some embodiments, an aryl group comprises a fully reduced cycloalkyl group defined herein (e.g., 1 ,2,3,4-tetrahydronaphthalene). When aryl comprises a cycloalkyl group, the aryl is bonded to the rest of the molecule through an aromatic ring carbon atom. An aryl radical can be a monocyclic or polycyclic (e.g., bicyclic, tricyclic, or tetracyclic) ring system, which may include fused, spiro or bridged ring systems. Unless stated otherwise specifically in the specification, an aryl may be optionally substituted, for example, with halogen, amino, alkylamino, aminoalkyl, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, heteroalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, -S(O)2NH-Ci-C6alkyl, and the like. In some embodiments, an aryl is optionally substituted with halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, -NO2, -S(O)2NH2, -S(O)2NHCH3, -S(O)2NHCH2CH3, -S(O)2NHCH(CH3)2, -S(O)2N(CH3)2, or -S(O)2NHC(CH3)3. In some embodiments, an aryl is optionally substituted with halogen, methyl, ethyl, -CN, - CF3, -OH, or -OMe. In some embodiments, the aryl is optionally substituted with halogen. In some embodiments, the aryl is substituted with alkyl, alkenyl, alkynyl, haloalkyl, or heteroalkyl, wherein each alkyl, alkenyl, alkynyl, haloalkyl, heteroalkyl is independently unsubstituted, or substituted with halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2

[0075] The term “cycloalkyl” refers to a monocyclic or polycyclic non-aromatic radical, wherein each of the atoms forming the ring ( / .e. skeletal atoms) is a carbon atom. In some embodiments, cycloalkyls are saturated or partially13MEl\59069747.vlAttorney Docket No. 137521-10320 unsaturated. In some embodiments, cycloalkyls are spirocyclic or bridged compounds. In some embodiments, cycloalkyls are fused with an aromatic ring (in which case the cycloalkyl is bonded through a non-aromatic ring carbon atom). Cycloalkyl groups include groups having from 3 to 10 ring atoms. Representative cycloalkyls include, but are not limited to, cycloalkyls having from three to ten carbon atoms, from three to eight carbon atoms, from three to six carbon atoms, or from three to five carbon atoms. Monocyclic cycloalkyl radicals include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. In some embodiments, the monocyclic cycloalkyl is cyclopentyl. In some embodiments, the monocyclic cycloalkyl is cyclopentenyl or cyclohexenyl. In some embodiments, the monocyclic cycloalkyl is cyclopentenyl. Polycyclic radicals include, for example, adamantyl, 1 ,2- dihydronaphthalenyl, 1 ,4-dihydronaphthalenyl, tetrainyl, decalinyl, 3,4-dihydronaphthalenyl-1 (2H)-one, spiro[2.2]pentyl, norbornyl and bicycle[1.1.1]pentyl. Unless otherwise stated specifically in the specification, a cycloalkyl group may be optionally substituted. Representative cycloalkyls include, but are not limited to, cycloalkyls having from three to fifteen carbon atoms (e.g., C3-C15 fully saturated cycloalkyl or C3-C15 cycloalkenyl), from three to ten carbon atoms (e.g., C3-C10 fully saturated cycloalkyl or C3-C10 cycloalkenyl), from three to eight carbon atoms (e.g., C3-C8 fully saturated cycloalkyl or C3-C8 cycloalkenyl), from three to six carbon atoms (e.g., C3-C6 fully saturated cycloalkyl or C3-C6 cycloalkenyl), from three to five carbon atoms (e.g., C3-C5 fully saturated cycloalkyl or C3-C5 cycloalkenyl), or three to four carbon atoms (e.g., C3-C4 fully saturated cycloalkyl or C3-C4 cycloalkenyl). In some embodiments, the cycloalkyl is a 3- to 6-membered cycloalkyl. In some embodiments, the cycloalkyl is a 5- to 6- membered cycloalkyl. Monocyclic cycloalkyls include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. Polycyclic cycloalkyls or carbocycles include, for example, adamantyl, norbornyl, decalinyl, bicyclo[3.3.0]octane, bicyclo[4.3.0]nonane, cis-decalin, trans-decalin, bicyclo[2.1.1]hexane, bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, bicyclo[3.2.2]nonane, and bicyclo[3.3.2]decane, and 7,7-dimethyl-bicyclo[2.2.1]heptanyl. Partially saturated cycloalkyls include, for example, cyclopentenyl, cyclohexenyl, cycloheptenyl, and cyclooctenyl. Unless stated otherwise specifically in the specification, a cycloalkyl is optionally substituted, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like. In some embodiments, a cycloalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, a cycloalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, the cycloalkyl is optionally substituted with halogen.

[0076] "Halo” or "halogen” refers to bromo, chloro, fluoro, or iodo. In some embodiments, halogen is fluoro or chloro. In some embodiments, halogen is fluoro.

[0077] "Haloalkyl” refers to an alkyl radical, as defined above, that is substituted by one or more halogens. In some embodiments, the alkyl is substituted with one, two, or three halogens. In some embodiments, the alkyl is substituted with one, two, three, four, five, or six halogens. Haloalkyl can include, for example, iodoalkyl, bromoalkyl, chloroalkyl, and fluoroalkyl. For example, "fluoroalkyl" refers to an alkyl radical, as defined above, that is substituted by one or more fluoro radicals, as defined above, for example, trifluoromethyl, difluoromethyl, fluoromethyl, 2,2,2-trifluoroethyl, 1 -fluoromethyl-2-fluoroethyl, and the like. In some embodiments, the alkyl part of the fluoroalkyl radical is optionally substituted as defined above for an alkyl group.

[0078] “Heteroal ky I” refers to an alkyl group in which one or more skeletal atoms of the alkyl are selected from an atom other than carbon, e.g., oxygen, nitrogen {e.g., -NH-, -N(alkyl)-), sulfur, or combinations thereof. A heteroalkyl is14MEl\59069747.vlAttorney Docket No. 137521-10320 attached to the rest of the molecule at a carbon atom of the heteroalkyl. In one aspect, a heteroalkyl is a Ci-Ce heteroalkyl wherein the heteroalkyl is comprised of 1 to 6 carbon atoms and one or more atoms other than carbon, e.g., oxygen, nitrogen (e.g. -NH-, -N(alkyl)-), sulfur, or combinations thereof wherein the heteroalkyl is attached to the rest of the molecule at a carbon atom of the heteroalkyl. Examples of such heteroalkyl are, for example, -CH2-O-CH2-, — CH2-N(alkyl)-CH2-, -CH2-N(aryl)-CH2-, -OCH2CH2O-, -OCH2CH2OCH2CH2O-, or -OCH2CH2OCH2CH2OCH2CH2O-. Unless stated otherwise specifically in the specification, a heteroalkyl is optionally substituted for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like. In some embodiments, a heteroalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, a heteroalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, the heteroalkyl is optionally substituted with halogen.

[0079] As used herein, a "heteroalkylene” refers to divalent heteroalkyl group. Examples of such heteroalkylene are, for example, -CH2-O-CH2-, -CH2-N(alkyl)-CH2-, -CH2-N(aryl)-CH2-, -OCH2CH2O-, -OCH2CH2OCH2CH2O-, or - OCH2CH2OCH2CH2OCH2CH2O-.

[0080] The term “heterocycloalkyl” refers to a cycloalkyl group that includes at least one hetero ring atom, e.g., a heteroatom selected from nitrogen, oxygen, and sulfur. Unless stated otherwise specifically in the specification, the heterocycloalkyl radical may be a monocyclic, or bicyclic ring system, which may include fused (when fused with an aryl or a heteroaryl ring, the heterocycloalkyl is bonded through a non-aromatic ring atom) or bridged ring systems. The nitrogen, carbon or sulfur atoms in the heterocyclyl radical may be optionally oxidized. The nitrogen atom may be optionally quaternized. The heterocycloalkyl radical is partially or fully saturated. Examples of heterocycloalkyl radicals include, but are not limited to, dioxolanyl, thienyl[1 ,3]dithianyl, tetrahydroquinolyl, tetrahydroisoquinolyl, decahydroquinolyl, decahydroisoquinolyl, imidazolinyl, imidazolidinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl, piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl, thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl, thiomorpholinyl, thiamorpholinyl, 1-oxo-thiomorpholinyl, 1,1-dioxo-thiomorpholinyl. Representative heterocycloalkyls include, but are not limited to, heterocycloalkyls having from two to fifteen carbon atoms (e.g., C2- C15 fully saturated heterocycloalkyl or C2-C15 heterocycloalkenyl), from two to ten carbon atoms (e.g., C2-C10 fully saturated heterocycloalkyl or C2-C10 heterocycloalkenyl), from two to eight carbon atoms (e.g., C2-C8 fully saturated heterocycloalkyl or C2-C8 heterocycloalkenyl), from two to seven carbon atoms (e.g., C2-C7 fully saturated heterocycloalkyl or C2-C7 heterocycloalkenyl), from two to six carbon atoms (e.g., C2-C6 fully saturated heterocycloalkyl or C2-C7 heterocycloalkenyl), from two to five carbon atoms (e.g., C2-C5 fully saturated heterocycloalkyl or C2-C5 heterocycloalkenyl), or two to four carbon atoms (e.g., C2-C4 fully saturated heterocycloalkyl or C2-C4 heterocycloalkenyl). The term heterocycloalkyl also includes all ring forms of carbohydrates, including but not limited to monosaccharides, disaccharides and oligosaccharides. Unless otherwise noted, heterocycloalkyls have from 2 to 12 carbons in the ring. In some embodiments, heterocycloalkyls have from 2 to 10 carbons in the ring. In some embodiments, heterocycloalkyls have from 2 to 10 carbons in the ring and 1 or 2 N atoms. In some embodiments, heterocycloalkyls have from 2 to 10 carbons in the ring and 3 or 4 N atoms. In some embodiments, heterocycloalkyls have from 2 to 12 carbons, 0-2 N atoms, 0-2 O atoms, 0-2 P atoms, and 0-1 S atoms in the ring. In some embodiments, heterocycloalkyls have from 2 to 12 carbons, 1-3 N atoms, 0-1 O atoms, and 0-1 S atoms in the ring. It is understood15MEl\59069747.vlAttorney Docket No. 137521-10320 that when referring to the number of carbon atoms in a heterocycloalkyl, the number of carbon atoms in the heterocycloalkyl is not the same as the total number of atoms (including the heteroatoms) that make up the heterocycloalkyl ( / .e. skeletal atoms of the heterocycloalkyl ring). Unless stated otherwise specifically in the specification, a heterocycloalkyl is optionally substituted, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like. In some embodiments, a heterocycloalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, a heterocycloalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, - CF3, -OH, or -OMe. In some embodiments, the heterocycloalkyl is optionally substituted with halogen.

[0081] "Heteroaryl” refers to a ring system radical comprising carbon atom(s) and one or more ring heteroatoms selected from the group consisting of nitrogen, oxygen, phosphorous, and sulfur, and at least one aromatic ring. In some embodiments, heteroaryl is monocyclic, bicyclic or polycyclic. Illustrative examples of monocyclic heteroaryls include pyridinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, pyridazinyl, triazinyl, oxadiazolyl, thiadiazolyl, furazanyl, indolizine, indole, benzofuran, benzothiophene, indazole, benzimidazole, purine, quinolizine, quinoline, isoquinoline, cinnoline, phthalazine, quinazoline, quinoxaline, 1 ,8-naphthyridine, and pteridine. Illustrative examples of monocyclic heteroaryls include pyridinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, pyridazinyl, triazinyl, oxadiazolyl, thiadiazolyl, and furazanyl. Illustrative examples of bicyclic heteroaryls include indolizine, indole, benzofuran, benzothiophene, indazole, benzimidazole, purine, quinolizine, quinoline, isoquinoline, cinnoline, phthalazine, quinazoline, quinoxaline, 1 ,8-naphthyridine, and pteridine. In some embodiments, heteroaryl is pyridinyl, pyrazinyl, pyrimidinyl, thiazolyl, thienyl, thiadiazolyl or furyl. In some embodiments, a heteroaryl contains 0-6 N atoms in the ring. In some embodiments, a heteroaryl contains 1-4 N atoms in the ring. In some embodiments, a heteroaryl contains 4-6 N atoms in the ring. In some embodiments, a heteroaryl contains 0-4 N atoms, 0-1 O atoms, 0-1 P atoms, and 0-1 S atoms in the ring. In some embodiments, a heteroaryl contains 1-4 N atoms, 0-1 O atoms, and 0-1 S atoms in the ring. In some embodiments, heteroaryl is a C1-C9 heteroaryl. In some embodiments, monocyclic heteroaryl is a C1-C5 heteroaryl. In some embodiments, monocyclic heteroaryl is a 5-membered or 6-membered heteroaryl. In some embodiments, a bicyclic heteroaryl is a C6-C9 heteroaryl. In some embodiments, a heteroaryl group comprises a partially reduced cycloalkyl or heterocycloalkyl group defined herein (e.g., 7,8-dihydroquinoline). In some embodiments, a heteroaryl group comprises a fully reduced cycloalkyl or heterocycloalkyl group defined herein (e.g., 5,6,7,8-tetrahydroquinoline). When heteroaryl comprises a cycloalkyl or heterocycloalkyl group, the heteroaryl is bonded to the rest of the molecule through a heteroaromatic ring carbon or hetero atom. A heteroaryl radical can be a monocyclic or polycyclic (e.g., bicyclic, tricyclic, or tetracyclic) ring system, which may include fused, spiro or bridged ring systems. Unless stated otherwise specifically in the specification, a heteroaryl is optionally substituted, for example, with halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like. In some embodiments, a heteroaryl is optionally substituted with halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, a heteroaryl is optionally substituted with halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, the heteroaryl is optionally substituted with halogen.

[0082] The term "moiety” refers to a specific segment or functional group of a molecule. Chemical moieties are16MEl\59069747.vlAttorney Docket No. 137521-10320 often recognized chemical entities embedded in or appended to a molecule.

[0083] The terms "treat,” "prevent,” "ameliorate,” and "inhibit,” as well as words stemming therefrom, as used herein, do not necessarily imply 100% or complete treatment, prevention, amelioration, or inhibition. Rather, there are varying degrees of treatment, prevention, amelioration, and inhibition of which one of ordinary skill in the art recognizes as having a potential benefit or therapeutic effect. In this respect, the disclosed methods can provide any amount of any level of treatment, prevention, amelioration, or inhibition of the disorder in a mammal. For example, a disorder, including symptoms or conditions thereof, may be reduced by, for example, about 100%, about 90%, about 80%, about 70%, about 60%, about 50%, about 40%, about 30%, about 20%, or about 10%. Furthermore, the treatment, prevention, amelioration, or inhibition provided by the methods disclosed herein can include treatment, prevention, amelioration, or inhibition of one or more conditions or symptoms of the disorder, e.g., cancer or an inflammatory disease.

[0084] In certain embodiments, the term "treatment” or grammatical variations thereof refers to therapeutic treatment, and excludes prevention or prophylactic intervention.

[0085] In certain embodiments, "treating” includes the concepts of "alleviating,” which refers to lessening the frequency of occurrence or recurrence, or the severity, of any symptoms or other ill effects related to a disorder and / or the associated side effects. In certain embodiments, the term "treating” also encompasses the concept of "managing” which refers to reducing the severity of a particular disease or disorder in a patient or delaying its recurrence, e.g., lengthening the period of remission in a patient who had suffered from the disease.

[0086] In certain embodiments, the term "prevent” or "preventing” as related to a disease opdisorder can refer to a compound that in a statistical sample, reduces the occurrences of the disorder or condition in the treated sample relative to an untreated control sample, or delays the onset or reduces the severity of one or more symptoms of the disorder or condition relative to the untreated control sample.

[0087] The term "therapeutically effective amount" as used herein refers to an amount effective at the dosage and duration necessary to achieve the desired therapeutic result. A therapeutically effective amount of the composition may vary depending on factors such as the individual's condition, age, sex, and weight, and the ability of the protein to elicit the desired response of the individual. A therapeutically effective amount can also be an amount that exceeds any toxic or deleterious effect of the composition that would have a beneficial effect on the treatment.

[0088] The term "optional” or "optionally” means that the subsequently described event or circumstance may or may not occur, and that the description includes instances where said event or circumstance occurs and instances in which it does not. For example, "optionally substituted alkyl” means either "alkyl” or "substituted alkyl” as defined above. Further, an optionally substituted group may be un-substituted {e.g., -CH2CH3), fully substituted {e.g., -CF2CF3), monosubstituted {e.g., -CH2CH2F) or substituted at a level anywhere in-between fully substituted and mono-substituted {e.g., -CH2CHF2, -CH2CF3, -CF2CH3, -CFHCHF2, etc.).

[0089] As used herein, the term "substituent" means positional variables on the atoms of a core molecule that are substituted at a designated atom position, replacing one or more hydrogens on the designated atom, provided that the designated atom's normal valency is not exceeded, and that the substitution results in a stable compound. Combinations of substituents and / or variables are permissible only if such combinations result in stable compounds. A person of ordinary skill in the art should note that any carbon as well as heteroatom with valences that appear to be17MEl\59069747.vlAttorney Docket No. 137521-10320 unsatisfied as described or shown herein is assumed to have a sufficient number of hydrogen atom(s) to satisfy the valences described or shown. In certain instances one or more substituents having a double bond (e.g., "oxo" or "=O") as the point of attachment may be described, shown or listed herein within a substituent group, wherein the structure may only show a single bond as the point of attachment to the core structure. A person of ordinary skill in the art would understand that, while only a single bond is shown, a double bond is intended for those substituents.

[0090] For the purpose of the disclosure, one event of "substitution” of an amino acid or an amino sequence is not considered two separate events of one deletion plus one addition. Thus, for the avoidance of doubt, as an example, a sequence change of "up to two deletion, substitution and / or addition” includes one deletion and one substitution, one deletion and one addition (at a different position), one substitution and one addition, one deletion only, one substitution only, one addition only, two deletions, two substitutions, two additions, etc. The deletion, addition, or substitution position may be at one or both ends of the peptide, or in the middle of the peptide.

[0091] The term "optionally substituted” or "substituted” means that the referenced group is optionally substituted with one or more additional group(s). For example, "optionally substituted” or "substituted” can mean that the referenced group is optionally substituted with one or more substituents individually and independently selected from D, halogen, -CN, -NH2, -NH(alkyl), -N(alkyl)2, -OH, -CO2H, -CO2alkyl, -C(=O)NH2, -C(=O)NH(alkyl), -C(=O)N(alkyl)2, - S(=O)2NH2, -S(=O)2NH(alkyl), -S(=O)2N(alkyl)2, alkyl, cycloalkyl, fluoroalkyl, heteroalkyl, alkoxy, fluoroalkoxy, heterocycloalkyl, aryl, heteroaryl, aryloxy, alkylthio, arylthio, alkylsulfoxide, arylsulfoxide, alkylsulfone, and arylsulfone. In some other embodiments, optional substituents are independently selected from D, halogen, -CN, -NH2, -NH(CH3), -N(CH3)2, -OH, -CO2H, -CO2(Ci-C4alkyl), -C(=O)NH2, -C(=O)NH(Ci-C4alkyl), -C(=O)N(Ci-C4alkyl)2, -S(=O)2NH2, - S(=O)2NH(Ci-C4alkyl), -S(=O)2N(Ci-C4alkyl)2, Ci-C4alkyl, Cs-Cecycloalkyl, Ci-C4fluoroalkyl, Ci-C4heteroalkyl, Ci- C4alkoxy, Ci-C4fluoroalkoxy, -SCi-C4alkyl, -S(=O)Ci-C4alkyl, and -S(=O)2Ci-C4alkyl. In some embodiments, an "optionally substituted” group is independently substitued with 1-6 substituents selected from halogen, -CN, oxo, -OH, -SF5, -SH, -S(=O)Ci-C3alkyl, -S(=O)2Ci-C3alkyl, -S(=O)2NH2, -S(=O)2NHCi-C3alkyl, -S(=O)2N(Ci-C3alkyl)2, - S(=O)(=NCi-C3alkyl)(Ci-C3alkyl), -NH2, -NHCi-C3alkyl, -N(Ci-C3alkyl)2, -N=S(=O)(Ci-C3alkyl)2, -C(=O)Ci-C3alkyl, - C(=O)OH, -C(=O)OCi-C3alkyl, -C(=O)NH2, -C(=O)NHCi-C3alkyl, -C(=O)N(Ci-C3alkyl)2, -P(=O)(Ci-C3alkyl)2, Ci- Cealkyl, Ci-Cealkoxy, Ci-Cehaloalkyl, Ci-Cehaloalkoxy, Ci-Cehydroxyalkyl, Ci-Ceaminoalkyl, Ci-Ceheteroalkyl, aryl, heteroaryl, heterocycloalkyl and cycloalkyl. In some embodiments, an "optionally substituted” group is independently substitued with 1-6 substituents selected from halogen, -CN, oxo, -OH, -SF5, -SH, -S(=O)Ci-C3alkyl, -S(=O)2Ci-C3alkyl, -S(=O)2NH2, -S(=O)2NHCi-C3alkyl, -S(=O)2N(Ci-C3alkyl)2, -S(=O)(=NCi-C3alkyl)(Ci-C3alkyl), -NH2, -NHCi-C3alkyl, - N(Ci-C3alkyl)2, -N=S(=O)(Ci-C3alkyl)2, -C(=O)Ci-C3alkyl, -C(=O)OH, -C(=O)OCi-C3alkyl, -C(=O)NH2, -C(=O)NHCi- C3alkyl, -C(=O)N(Ci-C3alkyl)2, -P(=O)(Ci-C3alkyl)2, Ci-C3alkyl, Ci-C3alkoxy, Ci-C3haloalkyl, Ci-C3haloalkoxy, C1- C3hydroxyalkyl, Ci-C3aminoalkyl, Ci-C3heteroalkyl, 5-6 membered heterocycloalkyl and Cs-Cecycloalkyl. In some embodiments, an "optionally substituted” group is independently substitued with 1-6 substituents selected from halogen, oxo, -OH, -NH2, -NHCi-C3alkyl, -N(Ci-C3alkyl)2, -C(=O)OH, -C(=O)NH2, Ci-C3alkyl, Ci-C3alkoxy, C1- C3haloalkyl, Ci-C3haloalkoxy, Ci-C3hydroxyalkyl, Ci-C3aminoalkyl, Ci-C3heteroalkyl, and Cs-Cecycloalkyl. In some embodiments, optional substituents are independently selected from D, halogen, -CN, -NH2, -OH, -NH(CH3), -N(CH3)2, -NH(cyclopropyl), -CH3, -CH2CH3, -CF3, -OCH3, and -OCF3. In some embodiments, substituted groups are substituted with one or two of the preceding groups. In some embodiments, an optional substituent on an aliphatic carbon atom18MEl\59069747.vlAttorney Docket No. 137521-10320(acyclic or cyclic) includes oxo (=0). When indicating the number of substituents, the term "one or more” means from one substituent to the highest possible number of substitutions, i.e. replacement of one hydrogen up to replacement of all hydrogens by substituents. In some embodiments, an "optionally substituted” group is unsubstituted. In some embodiments, an "optionally substituted” group is independently substitued with 1-6 substituents. In some embodiments, an "optionally substituted” group is independently substitued with 1-3 substituents. In some embodiments, an "optionally substituted” group is independently substitued with 1-2 substituents.

[0092] The term "unsubstituted” means that the specified group bears no substituents.

[0093] Certain compounds described herein may exist in tautomeric forms, and all such tautomeric forms of the compounds being within the scope of the disclosure.

[0094] Unless otherwise stated, structures depicted herein are also meant to include all stereochemical forms of the structure; i.e., the R and S configurations for each asymmetric center. Therefore, single stereochemical isomers as well as enantiomeric and diastereomeric mixtures of the present compounds are within the scope of the disclosure.

[0095] The term "peptide” as used herein refers to a compound that includes two or more amino acids. A peptide described herein can comprise one or more unnatural amino acids. The term "peptide” also encompasses peptide mimetics. In the present disclosure, the term "amino acid” is used in its broadest meaning and it embraces not only natural amino acids but also derivatives thereof and artificial amino acids. For example, the term "amino acid” encompasses unnatural amino acids.

[0096] The term “peptoid” as used herein refers to an N-substituted glycine. A peptoid can be optionally substituted. A peptoid can optionally comprise additional substitutions at the alpha-carbon.

[0097] As used herein, the term "unnatural amino acid” refers to an amino acid other than the 20 canonical amino acids. The 20 canonical amino acids refer to alanine (ala or A), arginine (arg or R), asparagine (asn or N), aspartic acid (asp or D), cysteine (cys or C), glutamine (gin or Q), glutamic acid (glu or E), glycine (gly or G), histidine (his or H), isoleucine (lie or I), leucine (leu or L), lysine (lys or K), methionine (met or M), phenylalanine (phe or F), proline (pro or P), serine (ser or S), threonine (thr or T), tryptophan (trp or W), tyrosine (tyr or Y), and valine (val or V).

[0098] The term "protein” as used herein refers to a polypeptide {i.e., a string of at least 3 amino acids linked to one another by peptide bonds). Proteins can include moieties other than amino acids {e.g., may be glycoproteins, proteoglycans, etc.) and / or can be otherwise processed or modified. A protein can be a complete polypeptide as produced by and / or active in a cell (with or without a signal sequence). In some embodiments, a protein is or comprises a characteristic portion such as a polypeptide as produced by and / or active in a cell. A protein can include more than one polypeptide chain. For example, polypeptide chains can be linked by one or more disulfide bonds or associated by other means.

[0099] The term "peptide mimetic” or "mimetic” refers to biologically active compounds that mimic the biological activity of a peptide or a protein but are no longer entirely peptidic in chemical nature, e.g.„ they can contain nonpeptide bonds (that are, bonds other than amide bonds between amino acids). As used herein, the term peptide mimetic is used in a broader sense to include molecules that are no longer completely peptidic in nature, such as pseudo-peptides, semi-peptides and peptoids. Whether completely or partially non-peptide, peptide mimetics described herein can provide a spatial arrangement of reactive chemical moieties that closely resemble the three- dimensional arrangement of active groups in the subject amino acid sequence or subject molecule on which the19MEl\59069747.vlAttorney Docket No. 137521-10320 peptide mimetic is based. As a result of this similar active-site geometry, the peptide mimetic can have effects on biological systems that are similar to the biological activity of the subject entity.

[0100] In some embodiments, the peptide mimetics are substantially similar in both three-dimensional shape and biological activity to the subject amino acid sequence or subject molecule on which the peptide mimetic is based. An example is described in the paper "Tritiated D-ala1 -Peptide T Binding”, Smith C. S. et al., Drug Development Res., 15, pp. 371-379 (1988). A second method is altering cyclic structure for stability, such as N to C interchain imides and lactams (Ede etal. in Smith and Rivier (Eds.) "Peptides: Chemistry and Biology”, Escom, Leiden (1991), pp. 268-270). An example of this is provided in conformationally restricted thymopentin-like compounds, such as those disclosed in US4457489. A third method is to substitute peptide bonds in the subject entity by pseudopeptide bonds that confer resistance to proteolysis.

[0101] The term "organic atoms” refers to atoms which would be found in organic compounds, such as carbon, hydrogen, nitrogen, oxygen, surfur, phosphorus, fluorine, chlorine, bromine, or iodine

[0102] Ranges provided herein are understood to be shorthand for all of the values within the range. For example, a range of 1 to 50 is understood to include any number, combination of numbers, or sub-range from the group consisting of 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49, or 50, as well as all intervening decimal values between the aforementioned integers such as, for example, 1.1 , 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, and 1.9. With respect to sub-ranges, "nested sub-ranges” that extend from either end point of the range are specifically contemplated. For example, a nested sub-range of an exemplary range of 1 to 50 may comprise 1 to 10, 1 to 20, 1 to 30, and 1 to 40 in one direction, or 50 to 40, 50 to 30, 50 to 20, and 50 to 10 in the other direction.

[0103] As used herein, Ci-Cx(or Ci-X) includes C1-C2, C1-C3... Ci-Cx. By way of example only, a group designated as “C1-C4” indicates that there are one to four carbon atoms in the moiety, i.e. groups containing 1 carbon atom, 2 carbon atoms, 3 carbon atoms or 4 carbon atoms. Thus, by way of example only, “C1-C4 alkyl” indicates that there are one to four carbon atoms in the alkyl group, i.e., the alkyl group is selected from among methyl, ethyl, propyl, isopropyl, n-butyl, iso-butyl, sec-butyl, and t-butyl. Also, by way of example, C0-C2 alkylene includes a direct bond, -CH2-, and -CH2CH2- linkages.

[0104] The term "cyclized” or "cyclization” as used herein means that two amino acids apart from each other by at least one amino acid bind directly or bind indirectly to each other in one peptide to form a cyclic structure in the molecule. In some cases, the two amino acids bind via a linker or the like. It is not necessary for all the atoms in the cycle to be amino acid backbone atoms. For example, the sidechain of an amino acid (such as the thiol group of a Cys) can be linked to the amino group of another amino acid to form the cycle. In some embodiment, cyclization is not formed between two Cys residue sidechains (e.g., cyclization is not due to the formation of a disulfide bond between two amino acids).

[0105] The term "subject” or "patient” encompasses mammals. Examples of mammals include, but are not limited to, any member of the Mammalian class: humans, non-human primates such as chimpanzees, and other apes and monkey species; farm animals such as cattle, horses, sheep, goats, swine; domestic animals such as rabbits, dogs, and cats; laboratory animals including rodents, such as rats, mice and guinea pigs, and the like. In one aspect, the mammal is a companion animal such as a dog or a cat. In one aspect, the mammal is a human.20MEl\59069747.vlAttorney Docket No. 137521-10320

[0106] Percent sequence identity can be calculated using computer programs or direct sequence comparison. Preferred computer program methods to determine identity between two sequences include, but are not limited to, the GCG program package, FAST A, BLASTP, and TBLASTN (see, e.g., D. W. Mount, 2001 , Bioinformatics: Sequence and Genome Analysis, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.). The BLASTP and TBLASTN programs are publicly available from NCBI and other sources. The Smith Waterman algorithm can also be used to determine percent identity. Exemplary parameters for amino acid sequence comparison include the following: 1) algorithm from Needleman and Wunsch (J. Mol. Biol., 48:443-453 (1970)); 2) BLOSSUM62 comparison matrix from Hentikoff and Hentikoff (Proc. Nat. Acad. Sci. USA., 89: 10915-10919 (1992)) 3) gap penalty=12; and 4) gap length penalty =4. A program useful with these parameters can be publicly available as the "gap” program (Genetics Computer Group, Madison, Wis.). The aforementioned parameters are the default parameters for polypeptide comparisons (with no penalty for end gaps). Alternatively, polypeptide sequence identity can be calculated using the following equation: % identity = (the number of identical residues) / (alignment length in amino acid residues)*100. For this calculation, alignment length may include internal gaps and mismatches but does not include terminal unaligned residues. Alternatively, polypeptide sequence identity can be calculated using the following equation: % identity = (the number of identical residues) / (the number of total amino acid residues in the shorter of the two sequences being compared)*100.

[0107] It is appreciated that certain features of the disclosure, which are, for clarity, described in the context of separate embodiments, can also be provided in combination in a single embodiment. Conversely, various features of the disclosure, which are, for brevity, described in the context of a single embodiment, can also be provided separately or in any suitable subcombination. For example, a conjugate of this disclosure can comprise any peptide ligand described herein {e.g., a peptide of Formula (I) or (II), or Table 1 , 2, 3, 4, or 9A), any moiety described herein, and optionally a linker described herein (e.g., a linker of Formula (11-1), (11-1 a), or (11-1 b)).

[0108] Unless special definitions are given, the terminology used in relation to analytical chemistry, synthetic organic chemistry, and medical chemistry and pharmaceutical chemistry described in the present specification, as well as their procedures and techniques, are well known and commonly used in the field of the present art. Standard techniques may be used for chemical synthesis and chemical analysis. Those defined from among such techniques and procedures can be found in, for example, "K.J. Jensen, P.T. Shelton, S.L. Pedersen, Peptide Synthesis and Applications, 2nd Edition, Springer, 2013” and the like, and these are incorporated into the present specification by reference for all purposes. All patents, applications, published applications, and other publications, and other data referred to throughout the entire disclosure, when permitted, are incorporated into the present specification by reference.II. Conjugates

[0109] Provided herein are conjugates that have avidity for Glypican 3 (GPC3), or a pharmaceutically acceptable salt thereof, and pharmaceutical compositions comprising the conjugates. The conjugates described herein excludes conjugates with radionuclides. The conjugates and compositions can be useful for treating cancer. The conjugates and compositions can also be useful in imaging and disease diagnosis.

[0110] In one aspect, described herein is a conjugate that comprises a peptide that has avidity for Glypican 321MEl\59069747.vlAttorney Docket No. 137521-10320(GPC3). In some embodiments, the GPC3 is a human GPC3. The peptide can be cyclic or acyclic, and it can be monocyclic, bicyclic or polycyclic. In one aspect, described herein is a conjugate that comprises a cyclic peptide. In some embodiments, the peptide (such as cyclic peptide) is configured to bind to a target. A conjugate described herein can further comprise a linker that covalently connects the peptide to a moiety that excludes metal chelator for conjugating radionuclides.

[0111] In another aspect, described herein is a conjugate that comprises a peptide that has avidity for Glypican 3 (GPC3). In some embodiments, the GPC3 is a human GPC3. The peptide can be cyclic or acyclic, and it can be monocyclic, bicyclic or polycyclic. In one aspect, described herein is a conjugate that comprises a cyclic peptide and a moiety that excludes a metal chelator that is configured to bind with a radionuclide. In some embodiments, the peptide (such as cyclic peptide) is configured to bind to a target. A conjugate described herein can further comprise a linker that links a moiety excluding a covalent radionuclide to the peptide.

[0112] Peptides, linkers, and radionuclides suitable for use in the conjugates provided herein are as described herein.

[0113] In some embodiments, the conjugate comprises a moiety that excludes a metal chelator configured to bind with a radionuclide, wherein the moiety is bound to the peptide. In some embodiments, the moiety is covalently linked to the peptide. In some embodiments, the conjugate further comprises a linker that connects the peptide with the moiety. In some embodiments, the linker covalently connects the peptide with the moiety. In some embodiments, the linker covalently attaches the moiety to the N-terminus of the peptide. In some embodiments, the linker covalently attaches the moiety to the C-terminus of the peptide. In some embodiments, the linker is attached to the peptide via a non-terminal amino acid residue of the peptide. In some embodiments, the linker is attached to amino acid Xi. In some embodiments, the linker is attached to amino acid X2. In some embodiments, the linker is attached to amino acid X3. In some embodiments, the linker is attached to amino acid X4. In some embodiments, the linker is attached to amino acid X5. In some embodiments, the linker is attached to amino acid XB. In some embodiments, the linker is attached to amino acid X7. In some embodiments, the linker is attached to amino acid Xs. In some embodiments, the linker is attached to amino acid X9. In some embodiments, the linker is attached to amino acid X10. In some embodiments, the linker is attached to amino acid Xu. In some embodiments, the linker is attached to amino acid X12. In some embodiments, the linker is attached to amino acid Xi, X2, X3, X4, Xs, or X12. In some embodiments, the linker is attached to a lysine of the peptide. In some embodiments, the linker comprises one or more amino acid residues. In some embodiments, the linker comprises a lysine residue, an alanine residue, a glycine residue, a D- phenylalanine residue, a histidine residue, a dAtb residue, or a D-glutamate residue. In some embodiments, the linker comprises 2 amino acids selected from the group consisting of lysine, alanine, glycine, D-phenylalanine, histidine, dAtb, and D-glutamate. In some embodiments, the linker is a C1-6 heteroalkylene. In some embodiments, the peptide is directly attached to the moiety that excludes metal chelator.

[0114] In some embodiments, the moiety is covalently attached to the side chain of the first amino acid of the peptide (Xi, A1), the side chain of the second amino acid of the peptide (X2, A2), the side chain of the third amino acid of the peptide (X3, A3), the side chain of the fourth amino acid of the peptide (X4, A4), the side chain of the eighth amino acid of the peptide (Xs, As), or to the C-terminus of the C-terminal amino acid of the peptide (X12, Q, when Q is -NH2); or wherein the peptide comprises a thirteenth amino acid and the moiety is covalently attached to the side22MEl\59069747.vlAttorney Docket No. 137521-10320 chain of the thirteenth amino acid (X13, Q, when Q is D-Lys).

[0115] In some embodiments, the conjugate comprises a covalently bound moiety other than a radionuclide. In some embodiments, the conjugate further comprises a linker that connects the peptide with the covalently bound moiety. In some embodiments, the linker covalently connects the peptide with the covalently bound moiety. In some embodiments, the linker covalently attaches the covalently bound moietyto the N-terminus of the peptide. In some embodiments, the linker covalently attaches the covalently bound moietyto the C-terminus of the peptide. In some embodiments, the linker is attached to the peptide via a non-terminal amino acid residue of the peptide. In some embodiments, the linker is attached to amino acid Xi. In some embodiments, the linker is attached to amino acid X2. In some embodiments, the linker is attached to amino acid X3. In some embodiments, the linker is attached to amino acid X4. In some embodiments, the linker is attached to amino acid X5. In some embodiments, the linker is attached to amino acid Xe. In some embodiments, the linker is attached to amino acid X7. In some embodiments, the linker is attached to amino acid Xs. In some embodiments, the linker is attached to amino acid X9. In some embodiments, the linker is attached to amino acid X10. In some embodiments, the linker is attached to amino acid Xu. In some embodiments, the linker is attached to amino acid X12. In some embodiments, the covalently bound moiety other than radionuclide is directly covalently bound to an amino acid comprising an aromatic ring. In some embodiments, the covalent moiety other than radionuclide is directly covalently bound to X4, X5, Xs, X9, or Xu. In some embodiments, the covalent moiety other than radionuclide is directly covalently bound to Xi, X2, X3, X4, Xs, or X12.

[0116] In some embodiments, described herein is a conjugate comprising: (a) a targeting moiety that comprises a monocyclic peptide that has avidity for Glypican 3 (GPC3), or a pharmaceutically acceptable salt thereof, and (b) a moiety other than a radionuclide, wherein the moiety is bound to the peptide. In some embodiments, described herein is a conjugate comprising: a monocyclic peptide that is configured to bind with GPC3 and a moiety other than a radionuclide. In some embodiments, the monocyclic peptide is cyclized by a non-disulfide bond. In some embodiments, the monocyclic peptide does not comprise a disulfide bond. In some embodiments, the monocyclic peptide comprises 5 to 20 amino acid residues. In some embodiments, the monocyclic peptide comprises 7 to 14 amino acid residues. A conjugate described herein can further comprises a linker that covalently connects the cyclic peptide to the moiety other than a radionuclide.

[0117] In some embodiments, a herein-described conjugate is in a salt form (e.g., pharmaceutically acceptable salt form). In some embodiments, a herein-described conjugate is in a free-base form.

[0118] In some embodiments, a conjugate of the present disclosure is a conjugate from Table 5, 6, 7, 8, or 9B, wherein the moiety is a non-radionuclide moiety. In some embodiments, a conjugate of the present disclosure is a conjugate from Table 5, 6, 7, 8, 9B, or 20B, wherein the moiety is a non-radionuclide moiety. In some embodiments, a conjugate of the present disclosure comprises a peptide of Table 1, 2, 3, 4, or 9A and a moiety (a non-radionuclide moiety), as shown in Table 6. In some embodiments, a conjugate of the present disclosure comprises a peptide of Table 20A and a moiety (a non-radionuclide moiety), as shown in Table 20B.Conjugate Having GPC3 Binding and / or Antagonizing Avidity

[0119] Glypican-3 (GPC3) is a protein that in humans is encoded by the GPC3 gene. GPC3 may be upregulated in multiple cancers, often correlating with disease progression, metastasis and poor prognosis e.g., in solid tumors23MEl\59069747.vlAttorney Docket No. 137521-10320 such as hepatocellular, lung, gastric, and ovarian.

[0120] GPC3 belongs to the sulfate heparin proteoglycan family, and is anchored on the cell membrane surface by phosphatidylinositol (GPI) anchor.

[0121] GPC3 can play an important role in the cell proliferation of embryo layer tissue. Deletion of GPC3 gene can cause excessive growth syndrome, namely Simpson-Golabi-Behmel syndrome (SGBS). GPC3 can be expressed throughout the entire fetal stage, and after birth to adult stage, except for placental, breast, mesodermal, ovarian, lung and kidney tissue with weak expression, other normal tissues have no obvious expression.

[0122] In some embodiments, the conjugate of the present disclosure binds to GPC3. In some embodiments, the conjugate has GPC3 antagonistic activity. In some embodiments, the conjugate binds to human GPC3 (hGPC3) and has hGPC3 antagonistic activity, such as inhibiting the ability of GPC3 to promote or stabilize Wnt I Frizzled interaction and / or downstream signaling.

[0123] As used herein, the term "GPC3” refers to any form of GPC3 and a variant thereof for retaining at least a part of the activity of GPC3. The GPC3 includes all the native sequences of GPC3 in mammals such as, for example, humans, dogs, cats, horses, and cows, unless otherwise specifically described as human GPC3 (hGPC3). One exemplification of GPC3 is hGPC3 (Gene ID:2719), which is human GPC3 and is a protein having an amino acid sequence (Isoform 1, P51654-1): MAGTVRTACLWAMLLSLDFPGQAQPPPPPPDATCHQVRSFFQRLQPGLKWVPETPVPGSDLQVCLPKGPTCCSR KMEEKYQLTARLNMEQLLQSASMELKFLIIQNAAVFQEAFEIWRHAKNYTNAMFKNNYPSLTPQAFEFVGEFFTDVS LYILGSDINVDDMVNELFDSLFPVIYTQLMNPGLPDSALDINECLRGARRDLKVFGNFPKLIMTQVSKSLQVTRIFLQAL NLGIEVINTTDHLKFSKDCGRMLTRMWYCSYCQGLMMVKPCGGYCNWMQGCMAGWEIDKYWREYILSLEELVN GMYRIYDMENVLLGLFSTIHDSIQYVQKNAGKLTTTIGKLCAHSQQRQYRSAYYPEDLFIDKKVLKVAHVEHEETLSSR RRELIQKLKSFISFYSALPGYICSHSPVAENDTLCWNGQELVERYSQKAARNGMKNQFNLHELKMKGPEPWSQIIDK LKHINQLLRTMSMPKGRVLDKNLDEEGFESGDCGDDEDECIGGSGDGMIKVKNQLRFLAELAYDLDVDDAPGNSQQ ATPKDNEISTFHNLGNVHSPLKLLTSMAISWCFFFLVH (SEQ ID NO: 500).

[0124] As used herein, the expression "has avidity for GPC3” or "binds to GPC3” indicates the activity of binding to GPC3 over (e.g., substantially or significantly over) background or non-specific binding. Binding site of the peptide of the present disclosure on the GPC3 is not limited, the peptide can bind to anywhere on the GPC3 protein. Binding to GPC3 may be measured by any method for measuring known intermolecular binding. In a non-limiting manner, for example, this may be determined by competitive binding assays such as surface plasmon resonance (SPR) assays, scatter analysis and / or radioimmunoassays (RIA), enzyme immunoassays (EIA), and sandwich and competitive assays, and in any suitable manner which is known, including different variants of the examples given that are known in the technical field.

[0125] In one aspect, the binding affinity of the peptide or a conjugate thereof of the present disclosure is at most 100 nM as determined by Kd in surface plasmon resonance (SPR) analysis. In some implementations, the Kd of the peptides of the conjugates disclosed herein is 100 nM or less, 50 nM or less, 30 nM or less, 20 nM or less, 10 nM or less, 5 nM or less, 4 nM or less, 3 nM or less, 2 nM or less, 1 nM or less, 0.9 nM or less, 0.5 nM or less, 0.4 nM or less, 0.3 nM or less, 0.2 nM or less, 0.1 nM or less, 0.09 nM or less, 0.08 nM or less, 0.07 nM or less, 0.06 nM or less, 0.05 nM or less, 0.04 nM or less, 0.03 nM or less, 0.02 nM or less, 0.01 nM or less.24MEl\59069747.vlAttorney Docket No. 137521-10320

[0126] In some embodiments, a peptide or conjugate thereof described herein has a binding affinity to a human GPC3 of at most 1 , 5, 10, 50, 100, 200, 500, 1000, 5000 or 10,000 nM as determined by Kd in surface plasmon resonance (SPR) analysis. In some embodiments, a peptide or conjugate thereof described herein has a binding affinity to a human GPC3 of at most 100nM as determined by Kd in surface plasmon resonance (SPR) analysis. In some embodiments, a peptide or conjugate thereof described herein has a binding affinity to a human GPC3 of at most 1 nM as determined by Kd in surface plasmon resonance (SPR) analysis. In some embodiments, a peptide or conjugate thereof described herein has a binding affinity to a human GPC3 of at most 2 nM as determined by Kd in surface plasmon resonance (SPR) analysis. In some embodiments, a peptide or conjugate thereof described herein has a binding affinity to a human GPC3 of at most 5 nM as determined by Kd in surface plasmon resonance (SPR) analysis. In some embodiments, a peptide or conjugate thereof described herein has a binding affinity to a human GPC3 of at most 10 nM as determined by Kd in surface plasmon resonance (SPR) analysis.

[0127] In one aspect, the binding affinity of the peptide or conjugate thereof of the present disclosure is at most 100 nM as determined by Kd in surface plasmon resonance (SPR) analysis. In some embodiments, the Kd of the peptide or conjugate thereof of the present disclosure is 100 nM or less, 50 nM or less, 30 nM or less, 20 nM or less, 10 nM or less, 5 nM or less, 4 nM or less, 3 nM or less, 2 nM or less, 1 nM or less, 0.9 nM or less, 0.5 nM or less, 0.4 nM or less, 0.3 nM or less, 0.2 nM or less, 0.1 nM or less, 0.09 nM or less, 0.08 nM or less, 0.07 nM or less, 0.06 nM or less, 0.05 nM or less, 0.04 nM or less, 0.03 nM or less, 0.02 nM or less, 0.01 nM or less.GPC3 Peptide Binding Ligand

[0128] In one aspect, the disclosure relates to a peptide or a conjugate comprising the peptide (e.g., a binding peptide) that has avidity for Glypican-3 (GPC3). The GPC3 can be a mammalian GPC3. The GPC3 can be a human GPC3. The GPC3 can be a wild-type or mutated GPC3. In some embodiments, the peptide I conjugate of the disclosure comprises two or more peptides, which can be the same or different. The peptide can be linear or cyclic. In some embodiments, the peptide is monocyclic. The peptide can comprise any suitable number of amino acid residues. In some embodiments, the peptide comprises from 5 to 50, 6 to 40, 7 to 30, 8 to 25, 12 to 25, or 9 to 20 amino acid residues. In some embodiments, the peptide comprises from 5 to 14 amino acid residues. In some embodiments, the peptide comprises from 7 to 12 amino acid residues. In some embodiments, the peptide comprises from 8 to 12 amino acid residues. In some embodiments, the peptide comprises from 8 to 10 amino acid residues. In some embodiments, the peptide comprises from 7 to 13 amino acid residues. In some embodiments, the peptide comprises from 12 to 15 amino acid residues. In some embodiments, the peptide comprises from 13 to 14 amino acid residues. In some embodiments, the peptide comprises 6 amino acid residues. In some embodiments, the peptide comprises 7 amino acid residues. In some embodiments, the peptide comprises 8 amino acid residues. In some embodiments, the peptide comprises 9 amino acid residues. In some embodiments, the peptide comprises 10 amino acid residues. In some embodiments, the peptide comprises 11 amino acid residues. In some embodiments, the peptide comprises 12 amino acid residues. In some embodiments, the peptide comprises 13 amino acid residues. In some embodiments, the peptide comprises 14 amino acid residues. In some embodiments, the peptide comprises 15 amino acid residues. In some embodiments, the peptide comprises 16 amino acid residues. In some embodiments, the peptide consists of 6 amino acid residues. In some embodiments, the peptide consists of 7 amino acid residues. In some embodiments, the25MEl\59069747.vlAttorney Docket No. 137521-10320 peptide consists of 8 amino acid residues. In some embodiments, the peptide consists of 9 amino acid residues. In some embodiments, the peptide consists of 10 amino acid residues. In some embodiments, the peptide consists of 11 amino acid residues. In some embodiments, the peptide consists of 12 amino acid residues. In some embodiments, the peptide consists of 13 amino acid residues. In some embodiments, the peptide consists of 14 amino acid residues. In some embodiments, the peptide consists of 15 amino acid residues. In some embodiments, the peptide consists of 16 amino acid residues. In some embodiments, the conjugate comprises a monocyclic peptide of 6, 7, 8, 9, 10, 11 , 12, 13, 14, or 15 amino acid residues. A peptide described herein can be a binding peptide that binds to GPC3. In some embodiments, the binding peptide consists of 6 to 20 amino acid residues. In some embodiments, the binding peptide consists of 7 to 12 amino acid residues. In some embodiments, the binding peptide consists of 10 to 12 amino acid residues. In some embodiments, the binding peptide consists of 8 to 12 amino acid residues. In some embodiments, the binding peptide is monocyclic. In some embodiments, the peptide of the present technology is an isolated peptide. In some embodiments, the peptide of the present technology is a purified peptide.

[0129] In one aspect, the peptide or a pharmaceutically acceptable salt thereof has avidity for Glypican 3 (GPC3), wherein the peptide comprises an amino acid sequence of Formula I:Formula I, wherein R is -CH2-CO-; and, either (I)Xi is MeLys, MeLysAc, MeAla, MeGlu, MeGIn, or CrpG, wherein the N-terminal nitrogen of Xi is attached to the C(O) of R;X2 is Melle, MeGIn, MeLys, MeHseMe, MeLysCOpipzaa, or MeGlu;X3 is Asp, MeAsp, Hgn, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, Hva, or (S)Mor(2CO);X4 is MeGIn, MeLys, MeAspapCOpipzaa, MeGlu, or CrpG;X5is F4COO, MeF4COO, Y, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A;Xe is TMe, altMe, lie, or Leu;X7 is TMe, lie, Vai, Leu, Gcpe, or Eva;X8is Tyr, Cha, F3OH, F4aao, F4COO, F4aa, Yae, YaeCOpipzaa, F4F, F4U, Aph, 3Py6NH2, 4Py, F4(CONMeOH), F4CONHMe, F4CONMe2, or (PyrrCOO)A;X9is MeNal27N, MeF3Me4C, MeF34dOMe, or MeW7N;X10 is Gly, MeGly, D-Ala, D-Ser, or D-Pro;X11 is 3Py6Ph, 3Py6Pyrazol1 , F41 Me4Pyz, 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61ap4, F41 apPyz, F42Py, F44Py, or Bph4C; and, X12 is MeCys or Cys, wherein sulfur of the X12 side chain is covalently attached to R, and26MEl\59069747.vlAttorney Docket No. 137521-10320 carboxylic acid of X12 is replacedwherein Xi is MeGlu or CrpG; or X2 is MeLys; or X3 is MeAsp, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, (S)Mor(2CO), or Hva; or X4 is MeGlu, or CrpG; or X5 is MeF4C00, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A; or Xe is alTMe or Leu; or X? is TMe, Vai, Leu, Gcpe, or Eva; or Xs is Cha, F3OH, F4F, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A; or X9is MeF3Me4C, MeF34dOMe, or MeW7N; or X is MeGly, D-Ala, D-Ser, or D-Pro; or Xu is 3Py6Pyrazol1 , 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61 ap4, F41apPyz, F42Py, F44Py, or Bph4C; or X12 is Cys; or (ii) Xi to X12 have a sequence from Table 1 , 2, 3, 4, 9A, or 20A (and, in some further aspects, from Table 1 , 2, 3, 4, or 9A).

[0130] In another aspects, the peptide or a pharmaceutically acceptable salt thereof has avidity for Glypican 3 (GPC3), wherein the peptide comprises an amino acid sequence of Formula II:Formula II wherein:R* is -CH2-S-CH2-CO-; each Z is independently selected from H and Me; and,either (I)A1 is the side chain of Lys, LysAc, Ala, Glu, Gin, or CrpG;A2 is the side chain of lie, Gin, Lys, HseMe, LysCOpipzaa, or Glu;A3 is the side chain of Asp, Hgn, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, Hva, or (S)Mor(2CO);A4 is the side chain of Gin, Lys, AspapCOpipzaa, Glu, or CrpG;27MEl\59069747.vlAttorney Docket No. 137521-10320A5is the side chain of F4C00, Y, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A;Ae is the side chain of TMe, altMe, lie, or Leu;Az is the side chain of Tme, lie, Vai, Leu, Gcpe, or Eva;As is the side chain of Tyr, Cha, F3OH, F4aao, F4COO, F4aa, Yae, YaeCOpipzaa, F4F, F4U, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A;A9 is the side chain of Nal27N, F3Me4C, F34dOMe, or W7N;Aw is the side chain of Gly, D-Ala, D-Ser, or D-Pro;An is the side chain of 3Py6Ph, 3Py6Pyrazol1 , F41 Me4Pyz, 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61 ap4, F41 apPyz, F42Py, F44Py, or Bph4C; and, wherein A1 is the side chain of Glu or CrpG; or A2 is the side chain of Lys; or A3 is the side chain of Asp, Ser, Thr, Ala, or Lys, Glu, Asn, diMeDap, (S)Mor(2CO), or Hva; or A4 is the side chain of Glu or CrpG; or A5 is the side chain of F4COO, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A; or As is the side chain of alTMe or Leu; or Az is the side chain of Tme, Vai, Leu, Gcpe, or Eva; or As is the side chain of Cha, F3OH, F4F, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A; or A9is the side chain of F3Me4C, MeF34dOMe, or MeW7N; or Aw is the side chain of Gly, D-Ala, D-Ser, or D-Pro; or An is the side chain of 3Py6Pyrazol1 , 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61 ap4, F41 apPyz, F42Py, F44Py, or Bph4C; or (ii) A1 to An correspond to the side chains of Xi to Xu of a sequence from Table 1 , 2, 3, 4, 9A, or 20A (and, in some further aspects, from Table 1 , 2, 3, 4, or 9A).

[0131] In certain embodiments, Xi to Xw of the peptide of Formula (I) have a sequence from Table 1 , 2, 3, 4, 9A, or 20A. In certain embodiments, Xi to X12 of the peptide of Formula (I) have a sequence from Table 1 , 2, 3, 4, or 9A. In certain embodiments, Xi to X12 of the peptide of Formula (I) have the sequence of SEQ ID NO: 226, 520, or 521 .

[0132] In certain embodiments, the peptide does not have a sequence from Table A. In certain embodiments, the peptide is not any of SEQ ID NOS: 1-72.

[0133] A list of natural amino acids is found in Table AA. It is understood that "Me” as a prefix for any of the amino acids of Table AA represents the N-methylated version of that amino acid (e.g. MeAla or MeA is N- methylalanine, etc.). It is also understood that “D-”, “D”, “L-“ or “L” as prefix for any of the amino acids of Table AA represents the D-stereoisomer or L-stereoisomer of that amino acid (e.g.: D-Ala or D-A is D-alanine, L-Ala or L-A is L-alanine, etc.).Table AA: List of Natural Amino Acids28MEl\59069747.vlAttorney Docket No. 137521-10320

[0134] Unless otherwise stated in the present specification, the following abbreviations for non-natural amino acids (abbreviations indicated in bolded text in the table below) are used according to the following meanings:29MEl\59069747.vlAttorney Docket No. 137521-103230MEl\59069747.vlAttorney Docket No. 137521-103231MEl\59069747.vlAttorney Docket No. 137521-103232MEl\59069747.vlAttorney Docket No. 137521-103233MEl\59069747.vlAttorney Docket No. 137521-103234MEl\59069747.vlAttorney Docket No. 137521-103235MEl\59069747.vlAttorney Docket No. 137521-103236MEl\59069747.vlAttorney Docket No. 137521-103237MEl\59069747.vlAttorney Docket No. 137521-103238MEl\59069747.vlAttorney Docket No. 137521-103239MEl\59069747.vlAttorney Docket No. 137521-103240MEl\59069747.vlAttorney Docket No. 137521-103241MEl\59069747.vlAttorney Docket No. 137521-10320Table A. Example peptide sequences with avidity to GPC3 ("Term” refers to the functional group at the C-terminus)42MEl\59069747.vlAttorney Docket No. 137521-1032043MEl\59069747.vlAttorney Docket No. 137521-1032044MEl\59069747.vlAttorney Docket No. 137521-10320Table B. Example conjugates of the present disclosure containing moiety (12mer cyclic peptides). "Term” refers to the functional group at the C-terminus.45MEl\59069747.vlAttorney Docket No. 137521-1032046MEl\59069747.vlAttorney Docket No. 137521-10320Table 1. Example peptide sequences with avidity to GPC3, from N-terminal amino acid position 1 to amino acid position 12 (Xi to X12 in Formula I, respectively)47MEl\59069747.vlAttorney Docket No. 137521-1032048MEl\59069747.vlAttorney Docket No. 137521-10320Table 2. Example peptide sequences with avidity to GPC3, from N-terminal amino acid position 1 to amino acid position 12 (Xi to X12 in Formula I, respectively), and amino acid position 13 if present (X13 in Formula I)49MEl\59069747.vlAttorney Docket No. 137521-1032050MEl\59069747.vlAttorney Docket No. 137521-1032051MEl\59069747.vlAttorney Docket No. 137521-1032052MEl\59069747.vlAttorney Docket No. 137521-10320Table 3. Example peptide sequences with avidity to GPC3, from N-terminal amino acid position 1 to amino acid position 12 (Xi to X12 in Formula I, respectively)Table 4. Example peptide sequences with avidity to GPC3, from N-terminal amino acid position 1 to amino acid position 12 (Xi to X12 in Formula I, respectively)53MEl\59069747.vlAttorney Docket No. 137521-1032054MEl\59069747.vlAttorney Docket No. 137521-10320Table 5. Example conjugates of the present disclosure comprise a moiety and sequence of amino acids from N-terminal position 1 to position 12 (Xi to X12 in Formula I, respectively), and position 13 (X13 in Formula I) if present, as shown in the table below. Each cysteine side chain of position 12 is covalently linked to the N-terminus of amino acid 1 via -CH2-CO-, thereby forming a cyclic peptide.55MEl\59069747.vlAttorney Docket No. 137521-1032056MEl\59069747.vlAttorney Docket No. 137521-10320Table 6. Example conjugates of the present disclosure comprise a moiety and sequence of amino acids from N-terminal position 1 to position 12 (Xi to X12 in Formula I, respectively), and position 13 (X13 in Formula I) if present, as shown in the table below. Each cysteine side chain of position 12 is covalently linked to the N-terminus of amino acid 1 via -CH2-CO-, thereby forming a cyclic peptide.57MEl\59069747.vlAttorney Docket No. 137521-10320Table 7. Example conjugates of the present disclosure comprise a moiety and sequence of amino acids from N-terminal position 1 to position 12 (Xi to X12 in Formula I, respectively), as shown in the table below. Each cysteine side chain of position 12 is covalently linked to the N-terminus of amino acid 1 via -CH2-CO-, thereby forming a cyclic peptide.Table 8. Example conjugates of the present disclosure comprise a moiety and sequence of amino acids from N-terminal position 1 to position 12 (Xi to X12 in Formula I, respectively), as shown in the table below. Each cysteine side chain of position 12 is covalently linked to the N-terminus of amino acid 1 via -CH2-CO-, thereby forming a cyclic peptide.58MEl\59069747.vlAttorney Docket No. 137521-10320Table 9A. Example peptide sequences with avidity to GPC3, from N-terminal amino acid position 1 to amino acid position 12 (Xi to X12 in Formula I, respectively)59MEl\59069747.vlAttorney Docket No. 137521-1032060MEl\59069747.vlAttorney Docket No. 137521-1032061MEl\59069747.vlAttorney Docket No. 137521-10320Table 9B. Example conjugates of the present disclosure comprise a moiety and sequence of amino acids from N-terminal position 1 to position 12 (Xi to X12 in Formula I, respectively), as shown in the table below. Each cysteine side chain of position 12 is covalently linked to the N-terminus of amino acid 1 via -CH2-CO-, thereby forming a cyclic peptide.62MEl\59069747.vlAttorney Docket No. 137521-1032063MEl\59069747.vlAttorney Docket No. 137521-1032064MEl\59069747.vlAttorney Docket No. 137521-10320Table 20A. Example peptide sequences with avidity to GPC3, from N-terminal amino acid position 1 to amino acid position 12 (Xi to X12 in Formula I, respectively)Table 20B. Example conjugates of the present disclosure comprise a non-radionuclide moiety and sequence of amino acids from N-terminal position 1 to position 12 (Xi to X12 in Formula I, respectively), as shown in the table below. Each cysteine side chain of position 12 is covalently linked to the N-terminus of amino acid 1 via - CH2-CO-, thereby forming a cyclic peptide.65MEl\59069747.vlAttorney Docket No. 137521-10301

[0135] A peptide described herein can be cyclized {e.g., macrocyclized). Cyclization can be achieved less ideally via a single disulfide bond, or more ideally via a peptide bond, alkyl bond, alkenyl bond, ester bond, thioester bond, ether bond, thioether bond, phosphate ether bond, azo bond, C— S— C bond, C— N— C bond, C=N— C bond, C=N— 0 bond, amide bond, lactam bridge, carbamoyl bond, urea bond, thiourea bond, amine bond, thioamide bond, or the like, but not limited to them. In some embodiments, the peptide is a cyclic peptide that is cyclized by a peptide bond, alkyl bond, alkenyl bond, ester bond, thioester bond, ether bond, thioether bond, phosphate ether bond, azo bond, C— N— C bond, C=N— C bond, C=N— 0 bond, amide bond, lactam bridge, carbamoyl bond, urea bond, thiourea bond, amine bond, or thioamide bond. In some embodiments, the cyclic peptide is cyclized by a thioether bond. In some embodiments, the cyclic peptide is cyclized via an oxime cyclization reaction. Cyclization of a peptide sometimes stabilizes the peptide structure and thereby enhance its affinity for a target, compared to that of the linear version of the same peptide. The cyclization can occur between the N- and C-terminus of a linear peptide, or it can occur between a terminal amino acid and a non-terminal amino acid of a linear peptide. In some embodiments, the cyclization occurs between two non-terminal amino acids of a linear peptide. In some embodiments, the peptide is cyclized via oxime cyclization. In some embodiments, the peptide is cyclized between cysteine and haloacyl. In some embodiments, the peptide comprises a haloacetyl group {e.g., chloroacetyl or bromoacetyl) at the N-terminus of a linear peptide. In some embodiments, the peptide comprises a haloacetyl group {e.g., chloroacetyl or bromoacetyl) at the C-terminus of a linear peptide. In some embodiments, the peptide comprises a Cys at the C-terminus of a linear peptide. In some embodiments, the peptide comprises a Cys at the N-terminus of a linear peptide. In some embodiments, the cyclization occurs via a thioether bond between Cys and a haloacetyl group. In some embodiments, the cyclization occurs between the N-terminus and the C-terminus of the linear peptide.

[0136] As amino acids for macrocyclization, for example, an amino acid having the following functional group A and an amino acid having a corresponding functional group B can be used (see Table 4A). Either the functional group A or the functional group B may be placed on the N-terminal side of a linear peptide. The amino acid having the functional group A and the amino acid having the functional group B can each be an N-terminal amino acid or C-terminal amino acid or a non-terminal amino acid. In some embodiments, an amino acid having the functional group A is placed at the N-terminus. In some embodiments, an amino acid having the functional group A is placed at the C-terminus. In some embodiments, an amino acid having the functional group A is placed at a non-terminal amino acid. In some embodiments, an amino acid having the functional group B is placed at the N-terminus. In some embodiments, an amino acid having the functional group B is placed at the C- terminus. In some embodiments, an amino acid having the functional group B is placed at a non-terminal amino acid.Table 4A. Functional groups for cyclization66MEl\59069747.vlAttorney Docket No. 137521-10301

[0137] In some embodiments, the amino acid (l-A) in Table 4A can be, for example, a chloroacetylated amino67MEl\59069747.vlAttorney Docket No. 137521-10301 acid. Exemplary chloroacetylated amino acids include N-chloroacetyl-L-alanine, N-chloroacetyl-L-phenylalanine, N-chloroacetyl-L-tyrosine, N-chloroacetyl-L-tryptophan, N-3-(2-chloroacetamido)benzoyl-L-phenylalanine, N-3-(2- chloroacetamido)benzoyl-L-tyrosine, N-3-(2-chloroacetamido)benzoyl-L-tryptophan, p-N-chloroacetyl-L- diaminopropanoic acid, y-N-chloroacetyl-L-diaminobutyric acid, cr-N-chloroacetyl-L-ornithine, s-N-chloroacetyl-L- lysine, N-3-chloromethylbenzoyl-L-tyrosine, and N-3-chloromethylbenzoyl-L-tryptophane and D-amino acid derivatives corresponding thereto (for example, N-Chloroacetyl-D-alanine, N-Chloroacetyl-D-phenylalanine, N- Chloroacetyl-D-tyrosine, and N-Chloroacetyl-D-tryptophan).

[0138] Examples of the amino acid (l-B) include, but are not limited to, cysteine, homocysteine, mercaptonorvaline, mercaptonorleucine, 2-amino-7-mercaptoheptanoic acid, 2-amino-8-mercaptooctanoic acid, and amino acids obtained by protecting the SH group of these amino acids and then eliminating the protecting group, and D-amino acid derivatives corresponding thereto.

[0139] The cyclization method can be carried out, for example, according to the method described in Kawakami, T. et al., Nature Chemical Biology 5, 888-890 (2009); Yamagishi, Y. et al., ChemBioChem 10, 1469- 1472 (2009); Sako, Y. et al., Journal of American Chemical Society 130, 7932-7934 (2008); or WC2008 / 117833.

[0140] In some embodiments, for example, the amino acid (ll-A) is selected from propargylglycine, homopropargylglycine, 2-amino-6-heptynoic acid, 2-amino-7-octynoic acid, and 2-amino-8-nonynoic acid can be used. In addition, 4-pentynoylated or 5-hexynoylated amino acids can also be used. Examples of the 4- pentynoylated amino acids include N-(4-pentenoyl)-L-alanine, N-(4-pentenoyl)-L-phenylalanine, N-(4-pentenoyl)- L-tyrosine, N-(4-pentenoyl)-L-tryptophan, N-3-(4-pentynoylamido)benzoyl-L-phenylalanine, N-3-(4- pentynoylamido)benzoyl-L-tyrosine, N-3-(4-pentynoylamido)benzoyl-L-tryptophan, |3-N-(4-pentenoyl)-L- diaminopropanoic acid, y-N-(4-pentenoy l)-L-diaminobuty ric acid, cr-N-(4-pentenoyl)-L-ornithine, and E-N-(4- pentenoyl)-L-lysine, and D-amino acid derivatives corresponding thereto.

[0141] In some embodiments, for example, the amino acid (ll-B) is selected from azidoalanine, 2-amino-4- azidobutanoic acid, azidoptonorvaline, azidonorleucine, 2-amino-7-azidoheptanoic acid, and 2-amino-8- azidooctanoic acid can be used. In addition, azidoacetylated or 3-azidopentanoylated amino acids can also be used. Examples of the azidoacetylated amino acids include N-azidoacety l-L-al ani ne, N-azidoacetyl-L- phenylalanine, N-azidoacetyl-L-tyrosine, N-azidoacetyl-L-tryptophan, N-3-(4-pentynoylamido)benzoyl-L- phenylalanine, N-3-(4-pentynoylamido)benzoyl-L-tyrosine, N-3-(4-pentynoylamido)benzoyl-L-tryptophan, |3-N- azidoacetyl-L-diaminopropanoic acid, y-N-azidoacetyl-L-diaminobutyric acid, o-N-azidoacetyl-L-ornithine, and E- N-azidoacetyl-L-lysine, and D-amino acid derivatives corresponding thereto.

[0142] The cyclization method can be performed, for example, according to the method described in Sako, Y. et al., Journal of American Chemical Society 130, 7932-7934 (2008) or WC2008 / 117833.

[0143] Examples of amino acid (lll-A) include, but are not limited to, N-(4-aminomethyl-benzoyl)- phenylalanine (AMBF) and 4-3-aminomethyltyrosine.

[0144] Examples of the amino acid (lll-B) include, but are not limited to, 5-hydroxytryptophan (WoH). The cyclization method can be performed, for example, according to the method described in Yamagishi, Y. et al., ChemBioChem 10, 1469-1472 (2009) or WC2008 / 117833.68MEl\59069747.vlAttorney Docket No. 137521-10301

[0145] Examples of the amino acid (IV-A) include, but are not limited to, 2-amino-6-chloro-hexynoic acid, 2- amino-7-chloro-heptynoic acid, and 2-amino-8-chloro-octynoic acid.

[0146] Examples of the amino acid (IV-B) include, but are not limited to, cysteine, homocysteine, mercaptonorvaline, mercaptonorleucine, 2-amino-7-mercaptoheptanoic acid, and 2-amino-8-mercaptooctanoic acid, amino acids obtained by protecting the SH group of these amino acids and then eliminating the protecting group, and D-amino acid derivatives corresponding thereto. The cyclization method can be performed, for example, according to the method described in WO2012 / 074129.

[0147] Examples of the amino acid (V-A) include, but are not limited to, N-3-chloromethylbenzoyl-L- phenylalanine, N-3-chloromethylbenzoyl-L-tyrosine, and N-3-chloromethylbenzoyl-L-tryptophane.

[0148] Examples of the amino acid (V-B) include, but are not limited to, cysteine, homocysteine, mercaptonorvaline, mercaptonorleucine, 2-amino-7-mercaptoheptanoic acid, and 2-amino-8-mercaptooctanoic acid, and amino acids obtained by protecting the SH group of these amino acids and then eliminating the protecting group, and D-amino acid derivatives corresponding thereto.

[0149] The amino acids l-A to V-A and l-B to V-B can be introduced into the peptide in a known manner by chemical synthesis or translation and synthesis described herein. In some embodiments, the cyclization reaction comprises forming a thioether bond using an amino acid comprising a sulfanyl group, e.g., cysteine, homocysteine, mercaptonorvaline, mercaptovaline, mercaptonorleucine, 2-amino-7-mercaptoheptanoic acid, and 2-amino-8-mercaptooctanoic acid.

[0150] A peptide described herein can comprise one or more negatively charged amino acids and / or one or more positively charged amino acids. Positively charged amino acids include, for example, lysine, arginine, histidine, and amino acids that contain additional amine groups. Positively charged amino acids can comprise a heteroaryl substitution such as pyridine, imidazole, pyrazole, or triazole that has one or more ring nitrogen atoms. Negatively charged amino acids include, for example, amino acids that contain an additional carboxylic acid group such as glutamic acid or the like.

[0151] In some embodiments, a cyclic peptide has a net charge of -3 to +1. In some embodiments, the cyclic peptide has a net charge of -3. In some embodiments, the cyclic peptide has a net charge of -2. In some embodiments, the cyclic peptide has a net charge of -1 . In some embodiments, the cyclic peptide has a net charge of 0. In some embodiments, the cyclic peptide has a net charge of +1 . In some embodiments, a cyclic peptide has a net charge of at most -4. In some embodiments, the cyclic peptide has a net charge of -4. In some embodiments, a cyclic peptide has a net charge of at least +2. In some embodiments, the cyclic peptide has a net charge of +2. In some embodiments, the cyclic peptide has a net charge of +3. The net charge can be determined by aggregating the charge of each of the Xi to X12 amino acids (or each of the amino acid in the peptide). For example, aspartic acid (D) and glutamic acid (E) each has a charge of -1, lysine (K), arginine (R) and histidine (H) each has a charge of +1, and the rest of the canonical amino acids each has a charge of 0.

[0152] In some embodiments, a cyclic peptide has a net charge of -3 to +1 . In some embodiments, the cyclic peptide has a net charge of -3. In some embodiments, the cyclic peptide has a net charge of -2. In some embodiments, the cyclic peptide has a net charge of -1 . In some embodiments, the cyclic peptide has a net69MEl\59069747.vlAttorney Docket No. 137521-10301 charge of 0. In some embodiments, the cyclic peptide has a net charge of +1 . The net charge can be determined by aggregating the charge of each of the amino acids of the cyclic peptide.

[0153] In some embodiments, a cyclic peptide does not contain any S-S bond.

[0154] In some embodiments, a peptide of the present disclosure can be cyclized by forming a group as illustrated in Table 4B.Table 4B. Ring Closing Groups (m and n are independently 0 or an integer from 1 to 6.)

[0155] In some embodiments, m is 0 and n is 0. In some embodiments, m is 0. In some embodiments, m is 1. In some embodiments, m is 2. In some embodiments, m is 3. In some embodiments, m is 4. In some embodiments, m is 5. In some embodiments, m is 6. In some embodiments, n is 0. In some embodiments, n is 1. In some embodiments, n is 2. In some embodiments, n is 3. In some embodiments, n is 4. In some embodiments, n is 5. In some embodiments, n is 6.

[0156] In some embodiments, a peptide of the present disclosure can be cyclized by reacting a first functional group with a second functional group, see Table 4C. In some embodiments, the first functional group is located at the N-terminus. In some embodiments, the first functional group is located at a non-terminal amino acid. In some embodiments, the second functional group is located at the C-terminus. In some embodiments, the second functional group is located at a non-terminal amino acid.70MEl\59069747.vlAttorney Docket No. 137521-10301Table 4C. Formation of Ring Closing Groups00157] In some embodiments, a conjugate comprising any one of peptide of Table 1 may further comprise amino acid residues at the N and / or C terminus of the peptide, which is not part of the cyclic structure. In some embodiments, the conjugate further comprises a linker.A peptide described herein can be a peptide mimetic. For example, the peptide can comprise non-peptide bonds and it can comprise one or more unnatural amino acids. Unless stated otherwise, each of the amino acid in a peptide described herein (except the natural amino acid glycine) can independently be in its D or L form. Both D and L forms are encompassed by the present disclosure.

[0158] In the present disclosure, the term amino acid embraces derivatives of amino acids. The derivatives include, for example, amino acids obtained by modifying a natural amino acid constituting a protein produced by cellular DNA-encoded biological matter. Examples of such non-natural amino acids include hydroxyproline and hydroxylysine, which are amino acids having a hydroxyl group introduced therein, and diaminopropionic acid, which is an amino acid having an amino group introduced therein.

[0159] A peptide described herein can comprise an N-substituted amino acid. In some embodiments, the N- substituted amino acid is a derivative of tryptophan, phenylalanine, tyrosine, arginine, histidine, isoleucine, leucine, lysine, or valine. In some embodiments, the N-substitution is an N-alkyl, such as N-methyl and N-ethyl. In some embodiments, the N-substitution is N-methyl. In some embodiments, the N-substitution is an N-aryl, such as N-phenyl or N-biphenyl. In some embodiments, the N-substitution is an N-heteroaryl such as N-pyridyl. In some embodiments, the N-substituted amino acid is at the N-terminus of the peptide. In some embodiments, the N-substituted amino acid is a non-terminal amino acid.

[0160] In some embodiments, peptides described herein comprise one or more amino acids in Tables 5A to 5G.71MEl\59069747.vlAttorney Docket No. 137521-10301Table 5A. Exemplary Amino Acids at N or C-terminusTable 5B. Exemplary Amino Acids That Crosslink With A PeptideTable 5C. D-amino AcidsTable 5D. Exemplary N-alkylamino Acids

[0161] Exemplary alkyl groups for Table 5D include methyl, ethyl, and propyl groups.Table 5E. Exemplary Peptoid Blocks72MEl\59069747.vlAttorney Docket No. 137521-10301Table 5F. Exemplary Unnatural Amino Acids

[0162] Amino acids used in the disclosed peptides can be substituted with similar amino acids. In some embodiments, an amino acid can be substituted with another amino acid with similar hydrophobicity. In some embodiments, an amino acid can be substituted with another amino acid with similar hydrophilicity. In some embodiments, an amino acid can be substituted with another amino acid with similar size. In some embodiments, an amino acid can be substituted with another amino acid with similar charge. In some embodiment, an amino acid can be substituted with another amino acid with a similar functional group. In some embodiments, an amino acid can be substituted with another amino acid with the same functional group.

[0163] In some embodiments, an amino acid described herein can be replaced with a variant thereof. Examples of an amino acid substitution or variant include derivatives having an amine, amide, ester, or carboxyl group as the C-terminus and / or N-terminus thereof. Additional examples of amino acid / peptide variants include those obtained by modification such as phosphorylation, alkylation (e.g., methylation), acetylation, adenylylation, ADP-ribosylation, or glycosylation and fused protein obtained by fusion with another peptide or protein. These variants can be prepared by those skilled in the art in a known manner or a method based thereon. An amino acid variant further encompasses the amino acids that have the same functional groups but with different lengths73MEl\59069747.vlAttorney Docket No. 137521-10301 of the side chain (e.g., LysAc vs. OrnAc and cysteine vs. homocysteine). An amino acid variant further encompasses amino acids with a different aromatic moiety compared to the canonical amino acid (e.g., the indole in tryptophan vs the 7-azaindole in 7-AzaTrp; the phenyl in phenylalanine vs the pyridine in 4Py). An amino acid variant further encompasses amino acids with optional substituents, i.e., optionally substituted amino acid. In some embodiments, the optionally substituted amino acid is optionally substituted with one or more substituents independently selected from halogen, hydroxyl, cyano, amino, amide, nitro, ureido, Ci-Ce alkyl, Ci- Ce alkoxy, Ce-C aryl, C3-C6 cycloalkyl, 6-10 membered heterocycloalkyl, and 6-10 membered heteroaryl. In some embodiments, the optionally substituted amino acid is optionally substituted with one or more substituents independently selected from halogen, -CN, -NH2, -NH(alkyl), -N(alkyl)2, oxo, -OH, -CO2H, -CO2alkyl, -C(=O)NH2, -C(=O)NH(alkyl), -C(=O)N(alkyl)2, -S(=O)2NH2, -S(=O)2NH(alkyl), -S(=O)2N(alkyl)2, alkyl, cycloalkyl, fluoroalkyl, heteroalkyl, alkoxy, fluoroalkoxy, heterocycloalkyl, aryl, heteroaryl, aryloxy, alkylthio, arylthio, alkylsulfoxide, arylsulfoxide, alkylsulfone, and arylsulfone. In some embodiments, substituents may include any substituents described herein, for example: halogen, hydroxy, oxo (=0), thioxo (=S), cyano (-CN), nitro (-NO2), imino (=N-H), oximo (=N-OH), hydrazino (=N-NH2), SF5, -Rb-ORa, -Rb-OC(O)-Ra, -Rb-OC(O)-ORa, -Rb-OC(O)-N(Ra)2, -Rb-N(Ra)2, -Rb-C(O)Ra, -Rb-C(O)ORa, -Rb-C(O)N(Ra)2, -Rb-O-Rc-C(O)N(Ra)2, -Rb-N(Ra)C(O)ORa, -Rb-N(Ra)C(O)Ra, - Rb-N(Ra)S(O)tRa(where t is 1 or 2), -Rb-S(O)tRa(where t is 1 or 2), -Rb-S(O)tORa(where t is 1 or 2), and -Rb-S(O)tN(Ra)2 (where t is 1 or 2); and alkyl, alkenyl, alkynyl, aryl, aralkyl, aralkenyl, aralkynyl, cycloalkyl, cycloalkylalkyl, and heterocycle, any of which may be optionally substituted by alkyl, alkenyl, alkynyl, halogen, haloalkyl, haloalkenyl, haloalkynyl, oxo (=0), thioxo (=S), cyano (-CN), nitro (-NO2), imino (=N-H), oximo (=N- OH), hydrazine (=N-NH2), -Rb-ORa, -Rb-OC(O)-Ra, -Rb-OC(O)-ORa, -Rb-OC(O)-N(Ra)2, -Rb-N(Ra)2, -Rb-C(O)Ra, -Rb-C(O)ORa, -Rb-C(O)N(Ra)2, -Rb-O-Rc-C(O)N(Ra)2, -Rb-N(Ra)C(O)ORa, -Rb-N(Ra)C(O)Ra, -Rb-N(Ra) S(O)tRa(where t is 1 or 2), -Rb-S(O)tRa(where t is 1 or 2), -Rb-S(O)tORa(where t is 1 or 2) and -Rb-S(O)tN(Ra)2 (where t is 1 or 2); wherein each Rais independently selected from hydrogen, alkyl, cycloalkyl, cycloalkylalkyl, aryl, aralkyl, and heterocycle, wherein each Ra, valence permitting, may be optionally substituted with alkyl, alkenyl, alkynyl, halogen, haloalkyl, haloalkenyl, haloalkynyl, oxo (=0), thioxo (=S), cyano (-CN), nitro (-NO2), imino (=N-H), oximo (=N-OH), hydrazine (=N-NH2), -Rb-0Ra, -Rb-0C(0)-Ra, -Rb-0C(0)-0Ra, -Rb-OC(O)-N(Ra)2, -Rb-N(Ra)2, -Rb-C(0)Ra, -Rb-C(0)0Ra, -Rb-C(O)N(Ra)2, -Rb-O-Rc-C(O)N(Ra)2, -Rb-N(Ra)C(O)ORa, -Rb-N(Ra)C(O)Ra, -Rb-N(Ra)S(O)tRa(where t is 1 or 2), -Rb-S(O)tRa(where t is 1 or 2), -Rb-S(O)tORa(where t is 1 or 2) and -Rb-S(O)tN(Ra)2 (where t is 1 or 2); and wherein each Rbis independently selected from a direct bond or a straight or branched alkylene, alkenylene, or alkynylene chain, and each Rcis a straight or branched alkylene, alkenylene or alkynylene chain.

[0164] In some embodiments, a variant of an amino acid is selected from amino acids having one, two or three substituents based on the amino acid, and wherein the substituents are independently selected from halogen, -CN, -NH2, -NH(Ci-C3alkyl), -N(Ci-C3alkyl)2, oxo, -OH, -CO2H, -CO2-C1-C3alkyl, -C(=O)NH2, - C(=O)NH(Ci-C3alkyl), -C(=O)N(Ci-C3alkyl)2, -S(=O)2NH2, -S(=O)2NH(Ci-C3alkyl), -S(=O)2N(Ci-C3alkyl)2, Ci-C6alkyl, Ci-Ce heteroalkyl, Ci-Ce alkoxy, Ce-Cw aryl, C3-Ce cycloalkyl, 6-10 membered heterocycloalkyl, and 6-10 membered heteroaryl. In some embodiments, the variant is selected from amino acids having one or two74MEl\59069747.vlAttorney Docket No. 137521-10301 substituents based on the amino acid, and wherein the substituents are independently selected from halogen, - CN, -NH2, -NH(Ci-C3alkyl), -N(Ci-C3alkyl)2, oxo, -OH, -CO2H, -CO2-Ci-C3alkyl, -C(=O)NH2, -C(=O)NH(Ci- C3alkyl), -C(=O)N(Ci-C3alkyl)2, and Ci-Ce alkyl. In some embodiments, the variant is selected from amino acids having one or two substituents based on the amino acid, and wherein the substituents are independently selected from halogen, -CN, -NH2, -NH(Ci-C3alkyl), -N(Ci-C3alkyl)2, and Ci-Ce alkyl. In some embodiments, the variant is selected from amino acids having one or two substituents based on the amino acid, and wherein the substituents are independently selected from Ci-Ce alkyl.

[0165] In some embodiments, a variant of an amino acid is selected from amino acids that have the similar hydrophilicity or hydrophobicity compared to the amino acid. Thus, in some embodiments, a positively charged amino acid can be a variant of another positively charged amino acid. In some embodiments, a negatively charged amino acid can be a variant of another negatively charged amino acid. In some embodiments, a zwitterionic amino acid can be a variant of another zwitterionic amino acid.

[0166] In some embodiments, a hydrophilic amino acid has an electrically charged side chain. In some embodiments, a hydrophilic amino acid has a positive charge. In some embodiments, a hydrophilic amino acid has a negative charge. In some embodiments, a hydrophilic amino acid is zwitterionic (e.g., KCOpipzaa). In some embodiments, a hydrophilic amino acid comprises a -OH, COOH, -NH- or NH2moiety. In embodiments, a hydrophilic amino acid comprises -OH, -C(O)OH, -NHC(=NH)NH2, -NHC(O)NH2, -C(O)NH2, or -NHC(O)CH3In some embodiments, a hydrophilic amino acid comprises a side chain of Ci-Cehydroxyalkyl, Ci-Ceaminoalkyl, -Co- 6 alkylene-NH-C(=NH)-NH2, -Co-6 alkylene-CO-NH2, -Co-6 alkylene-COOH, or -NH-CO-C1-6 alkyl.

[0167] In some embodiments, a hydrophobic amino acid is not charged. In some embodiments, a hydrophobic amino acid contains at least 2 contiguous carbon atoms. In some embodiments, a hydrophobic amino acid comprises at least 3 contiguous carbon atoms, either linear or branched. In some embodiments, a hydrophobic amino acid comprises at least 4 contiguous carbon atoms, either linear or branched. In some embodiments, a hydrophobic amino acid comprises at least 5 contiguous carbon atoms, either linear or branched. In some embodiments, a hydrophobic amino acid comprises an ethylene moiety in the side chain. In some embodiments, a hydrophobic amino acid comprises a propylene moiety in the side chain. In some embodiments, a hydrophobic amino acid comprises a butylene moiety in the side chain. In some embodiments, a hydrophobic amino acid comprises phenyl moiety. In some embodiments, a hydrophobic amino acid comprises a heteroaryl moiety. In some embodiments, a hydrophobic amino acid is Trp, Tyr, Phe, or derivatives thereof.

[0168] In some embodiments, a variant of an amino acid is selected from amino acids that have the same functional group as the amino acid, and wherein the variant has a different length of a side chain compared to the amino acid. In some embodiments, a variant of an amino acid is selected from amino acids that have the same charge compared to the amino acid. In some embodiments, a variant of an amino acid is selected from amino acids that have the same polarity compared to the amino acid. In some embodiments, an amino acid comprising an aromatic group can be a variant of another amino acid having an aromatic group. In some embodiments, an amino acid comprising a phenyl can be a variant of another amino acid having a phenyl. In some embodiments, an amino acid comprising a heteroaryl can be a variant of another amino acid having a75MEl\59069747.vlAttorney Docket No. 137521-10301 heteroaryl. Amino acids having an aromatic group include, but are not limited to, F, W, F4G, 4Py ,3Py, F4COO, F3G, 3Py6COO, 4Py2NH2, 3Py5COO, F3COO, 3Py6NHAc, F4C, F40Me, Nal2, F3aao, F4aa, F4aao, 3Py6NHaa, F3CON, F4F, F4OEt, F4Me, F4CON, F4CONPEG4Me, F30Me, YaeCOpipzaa, F4aaopipzaa, 4Pdo, 3Py6CON, F4amC0pipzaa, F4u, F4CONdMe, F4ms, F34dOMe, F3C, F3CONdMe, 5lnda, F3aa, 3Py6NHae, 3Py6OMe, MeNal2, MeNal27N, MeF34diox, MeF34dOMe, MeF4T, MeW7N, MeF3C4Me, MeF3Me4C, Bph, 3Py6Ph, F41 Me4Pyz, F43Pyz, F44Pyz, F41Pyz, F41 Me3Pyz, F41 Et4Pyz, F41 MeOe4Pyz, F41 MeOp4Pyz, F44thp, F4Ac4Pip, PhNva, PhNIe, Yph, Ybn, F4tb, F4OPr, Yae, Me3Py, W5N and MeWIMe.

[0169] In some embodiments, an amino acid comprising a cycloalkyl group can be a variant of another amino acid having a cycloalkyl group. In some embodiments, an amino acid comprising a heterocycloalkyl group can be a variant of another amino acid having a heterocycloalkyl group.

[0170] In some embodiments, a variant of an amino acid is selected from amino acids that have similar polarity and / or charge with the amino acid. For example, in some embodiments, a polar, uncharged amino acid can be a variant of another polar, uncharged amino acid (e.g., Hgn, Q, S, T, Qglucamine),

[0171] In some embodiments, a variant of an amino acid has the same number of hydrogen donor as the amino acid. In some embodiments, a variant of an amino acid has the same number of hydrogen acceptor as the amino acid.

[0172] In some embodiments, the variant has a molecular weight that does not vary for more than 14, 28, 30, 45 or 60 g / mol compared to the amino acid. In some embodiments, the variant has a molecular weight that does not vary for more than 14 g / mol compared to the amino acid. In some embodiments, the variant has a molecular weight that does not vary for more than 50 g / mol compared to the amino acid. In some embodiments, the variant has a molecular weight that does not vary for more than 28 g / mol compared to the amino acid.

[0173] An amino acid variant further encompasses amino acids wherein a functional group is substituted with another functional group having similar properties, e.g., a cysteine can be substituted with a homocysteine. In some embodiments, an aryl functional group can be substituted with an aryl or heteroaryl group. In some embodiments, a heteroaryl functional group can be substituted with an aryl or heteroaryl group. In some embodiments, an amino functional group can be substituted with an NH(alkyl) group.

[0174] As used herein, the expression "conservative amino acid substitution” refers to a substitution of functionally equivalent or similar amino acids. A conservative amino acid substitution in a peptide brings about a static change to the amino acid sequence of the peptide. For example, one or two or more amino acids having similar polarity act functionally equivalent to each other and bring about a static change in the amino acid sequence of the peptide. In general, a substitution within a certain group may be considered conservative regarding structure and function. However, as is clear to a person having ordinary skill in the art, the role played by a defined amino acid residue may be determined by its implication in the three-dimensional structure of the molecule containing the amino acid. For example, a cysteine residue in an oxidized-type (disulfide) form may have a lower polarity than that of a reduced-type (thiol) form. The long aliphatic part of the arginine side chain may constitute structurally and functionally important features. Furthermore, the side chain (tryptophan, tyrosine, phenylalanine) including an aromatic ring may contribute to ion-aromatic interaction or cation-pi interaction. In76MEl\59069747.vlAttorney Docket No. 137521-10301 such a case, even if the amino acids having these side chains are substituted for amino acids belonging to the acidic or non-polar groups, they may be structurally and functionally conservative. There is a possibility that residues such as proline, glycine, cysteine (disulfide foam) have a direct effect on the three-dimensional structure of the main chain and often may not be substituted without structural distortion.

[0175] Conservative amino acid substitution, as shown below, includes specific substitution based on the similarity of side chains (for example, substitutions are described in Lehninger, Biochemistry, Revised 2nd Edition, published in 1975, pp. 73 to 75: L. Lehninger, Biochemistry, 2nd edition, pp. 73 to 75, Worth Publisher, New York (1975)), incorporated herein by reference, and typical substitution.

[0176] Hydrophobic amino acids include amino acids that exhibit hydrophobicity, including alanine (also referred to as "Ala” or simply "A”), glycine (also referred to as “Gly” or simply “G”), valine (also referred to as "Vai” or simply "V”), leucine (also referred to as "Leu” or simply “L”), isoleucine (also referred to as "lie” or simply "I”), proline (also referred to as "Pro” or simply “P”), phenylalanine (also referred to as "Phe” or simply “F”), tryptophan (also referred to as Trp” or simply “W”), tyrosine (also referred to as "Tyr” or simply “Y”), and methionine (also referred to as "Met” or simply “M”).

[0177] Exemplary hydrophobic amino acids may be further divided into the following groups:• Aliphatic amino acids: Amino acids having a fatty acid or hydrogen in the side chain, including e.g., Ala, Gly, Vai, lie, and Leu.• Aliphatic / branched-chain amino acids: Amino acids having a branched fatty acid in the side chain, including e.g., Vai, lie, and Leu.• Aromatic amino acids: Amino acids having an aromatic ring in the side chain, including e.g., Trp, Tyr, and Phe.

[0178] In some embodiments, a hydrophobic amino acid has a C1-C8 alkyl, cycloalkyl, or heterocycloalkyl, wherein the alkyl, cycloalkyl, and heterocycloalkyl are each independently, optionally substituted. In some embodiments, a hydrophobic amino acid has a C1-C8 alkyl, cycloalkyl, or heterocycloalkyl, wherein the alkyl, cycloalkyl, and heterocycloalkyl are each unsubstituted. In some embodiments, a hydrophobic amino acid has a C3-C6 unsubstituted alkyl. In certain embodiments, a hydrophobic amino acid is I, Eva, all, TMe, SMe, Gcpr, Gcpe, Gthp, dMeS, TdMe, or Cbg.

[0179] Hydrophilic amino acids include amino acids that exhibit hydrophilicity, including e.g., serine (also referred to as "Ser” or simply “S”), threonine (also referred to as "Thr” or simply “T”), cysteine (also referred to as "Cys” or simply “C”), asparagine (also referred to as "Asn” or simply “N”), glutamine (also referred to as "Gin” or simply “Q”), aspartic acid (also referred to as "Asp” or simply “D”), glutamic acid (also referred to as “Glu” or simply “E”), Elysine (also referred to as "Lys” or simply “K”), arginine (also referred to as "Arg” or simply “R”), and histidine (also referred to as "His” or “H”).

[0180] Exemplary hydrophilic amino acids may be further divided into the following groups:• Acidic amino acids: Amino acids whose side chains exhibit acidity, including Asp and Glu.• Basic amino acids: Amino acids whose side chains exhibit basicity, including Lys, Arg, and His.77MEl\59069747.vlAttorney Docket No. 137521-10301• Neutral amino acids: Amino acids whose side chains exhibit neutrality, including Ser, Thr, Asn, Gin, and Cys.

[0181] Exemplary hydrophilic amino acids include, for example, D, Q, A, E, S, N, T, C, H, or a variant thereof (including D-amino acid such as da and variations such as KCOpipzetOH, Cit, COmeglumine, KCOpipzaa, A4paa, Hgl, F4COO, KCOpip4COO, KAc, Hgn, Har, DapCOpipzaa, Acpr, Cba, NleCOO, NleOH, Atb, Nva, Nle, DapAc, Abu, Nmm, Ndm, Ncit, Cit or the like). In some aspect, hydrophilic amino acid is optionally N-methylated such as MeD.

[0182] In some embodiments, a peptide described herein comprises an amino acid that affects the direction of the main chain, e.g., Gly and Pro. In some embodiments, a peptide described herein comprises a sulfur- containing amino acid, e.g., Cys and Met. In some embodiments, a peptide described herein comprises an amino acid that comprises an aromatic ring, which can be optionally substituted. Amino acids comprising an aromatic ring include, e.g., F (Phe; phenylalanine), Y (Tyr: tyrosine), W (Trp; tryptophan).

[0183] In some embodiments, W or a variant thereof can be W, an amino acid having a heteroatom in the indole ring of W in the side chain, an amino acid in which the hydrogen of NH in the indole ring of W is substituted, or an amino acids having a substituent in the benzene ring of W, or the like.

[0184] In some embodiments, F or a variant thereof can be F (phenylalanine), an amino acid comprising a phenyl, 6 membered heteroaryl (such as pyridine, pyrimidine, pyrazine), indazolyl, or naphthalyl in its side chain, wherein said phenyl, heteroaryl, indazolyl or naphthalyl is optionally substituted. In some embodiments, F or a variant thereof can be F (phenylalanine), an amino acid comprising a phenyl, naphthalyl, 6 membered heteroaryl, or bicyclic heteroaryl in its side chain, wherein said phenyl, heteroaryl, or naphthalyl is optionally substituted. In some embodiments, F or a variant thereof can be F (phenylalanine), an amino acid comprising a phenyl, pyridinyl, indazolyl, or naphthalyl in its side chain, wherein said phenyl, pyridinyl, indazolyl or naphthalyl is optionally substituted with one or more substituents each independently selected from halogen, -C1 -3alkyl, -OH, -NH2, -ON, -C(=O)OH, -C(=O)NH2, -NHC(=O)CH3, -C1-3alkylene-C(=O)OH, -C1-3alkylene-C(=O)NH2, -O-C1- 3alkylene-C(=O)OH, -O-C1-3alkylene-C(=O)NH2, -C1-3alkylene-C(=O)-5- to 6-membered heterocycloalkylene- C1-3alkylene-C(=O)OH, -O-C1-3alkylene-C(=O)- 5- to 6-membered heterocycloalkylene-C1-3alkylene-C(=O)OH, -C1-3alkylene-NHC(=O)- 5- to 6-membered heterocycloalkylene-C1-3alkylene-C(=O)OH, -O-C1-3alkylene- NHC(=O)- 5- to 6-membered heterocycloalkylene-C1-3alkylene-C(=O)OH, and -NH-C1-3alkylene-C(=O)OH, - NHC(=O)NH2, -Ci.3alkylene-C(NH2)-COOH, -NH-CO-CH3, -NH-Ci.3alkylene-NH2, -C(=O)-N(CH2)2, -S(=O)2-CH3, - Ci-3alkylene-NH-C(=O)- 5- to 6-membered heterocycloalkylene-Ci-3alkylene-C(=O)OH, and -O-Ci.3alkylene-NH- C(=O)- 5- to 6-membered heterocycloalkylene-Ci-3alkylene-C(=O)OH, -O-phenyl, -O-Ci-salkylene-phenyl, pyridinyl, imidazolyl, pyrazolyl, N-Ci-salkylene pyrazolyl, N-Ci-3alkylene(-O-Ci-3alkyl) pyrazolyl, pyranyl, tetrahydropyranyl, piperidinyl, N-Ci-sal ky lene-C(=O)-piperidiny I ; or the like. In some aspect, F or a variant thereof is optionally N-methylated.

[0185] In some embodiments, F or a variant thereof can be F (phenylalanine), an amino acid wherein (i) the phenyl ring of F is substituted with 1 or 2 substituents each independently selected from -OH, -ON, -C1-3 alkyl, such as -CH3; (ii) a 6-membered heteroaryl ring optionally substituted by 1 or 2 substituents each independently78MEl\59069747.vlAttorney Docket No. 137521-10301 selected from -OH, -ON, - C1-3 alkyl, such as -CH3; or (iii-1) having a heteroatom in the phenyl ring of F in the side chain; (iii-2) a derivative amino acid of F in which a 6-membered heteroaryl ring in the side chain is substituted; or the like. In some aspect, F or a variant thereof is optionally N-methylated.

[0186] In some embodiments, Y or a variant thereof can be Y, an amino acid having a hydroxyphenyl ring, wherein the hydrogen atom in hydroxyphenyl of Y or of the variant is optionally substituted. In some embodiments, Y or a variant thereof can be Y, an amino acid having a hydroxyphenyl ring, wherein the hydrogen atom in hydroxyphenyl of Y or of the variant is optionally substituted with one or more substituents selected from -C1-3alkyl, -C1-3alkylene-C(=O)OH, halogen, -OH, -C(=O)OH, -O-CH3, -Ci-3alkylene-C(=O)- 5- to 6-membered heterocycloalkylene-Ci-3alkylene-C(=O)OH, and -Ci-3alkylene-NHC(=O)- 5- to 6-membered heterocycloalkylene- Ci-3alkylene-C(=O)OH. In some embodiments, a variant of W or a variant thereof can be W, W1 Me, or W7N or the like. In some embodiments, W or a variant thereof is optionally N-methylated (e.g., MeW1 Me or MeW7N).

[0187] In certain embodiments, F or Y or a variants thereof is Y, F3G, 3Py6COO, 4Py2NH2, 3Py5COO, F3COO, 3Py6NHAc, F, F4C, F4OMe, F4COO, Nal2, F3aao, F4aa, F4aao, 3Py6NHaa, 5Pdo, F3CON, F4F, F40Et, F4Me, F4CON, F4CONPEG4Me, F3OMe, YaeCOpipzaa, F4aaopipzaa, 4Pdo, 3Py6CON, Atp, Cha4cH, Cha4tH, Cha4cOMe, A1mor, F4amCOpipzaa, F4OMe, F4u, F4CONdMe, F4ms, F34dOMe, F3C, F3CONdMe, 5lnda, F3aa, 3Py6Nhae, 3Py6OMe, F4amCOpipzaa, Bph, 3Py6Ph, F41Me4Pyz, F43Pyz, F44Pyz, F41 Pyz, F41 Me3Pyz, F41 Et4Pyz, F41MeOe4Pyz, F41 MeOp4Pyz, F44thp, F4Ac4pip, PhNva, PhNIe, Yph, Ybn, F4tb, or F4oPr.

[0188] In some embodiments, a monocyclic aromatic amino acid can be an amino acid having a phenyl or monocyclic heteroaryl, each of which optionally substituted. In some embodiments, a monocyclic aromatic amino acid can be an amino acid having a phenyl or pyridinyl optionally substituted with one or more substituents each independently selected from halogen, -C1 -3alkyl, and trifluoromethyl. In some embodiment, a bicyclic aromatic amino acid can be an amino acid having a naphthalyl or bicyclic heteroaryl, each of which optionally substituted. In some embodiment, a bicyclic aromatic amino acid can be an amino acid having a naphthalyl, quinolyl, or indazolyl optionally substituted with one or more substituents each independently selected from H or C1 -3alkyl. In some embodiments, bicyclic aromatic acid can be W (tryptophan), or a variant thereof, or N-methylated thereof. In certain embodiments, a monocyclic aromatic amino acid is MeF3C3Me, MeF34dOMe, MeF4T,or M3FMe4C. In certain embodiments, a bicyclic aromatic amino acid is MeNal2, MeNal27N, MeF34diox, MeWI Me, or MeW7N.

[0189] In some embodiments, an amino acid described herein is N-alkylated.

[0190] In some embodiments, an amino acid described herein is not N-alkylated {e.g., an amino acid with -H on the alpha-amino group).

[0191] Examples of the amino acids include natural protein L-amino acids, unnatural amino acids, and chemically synthesized compounds having properties known in the art as characteristics of an amino acid. Examples of the unnatural amino acids include, but not limited to, o,o-disubsti tuted amino acids (such as o- methylalanine), N-alkyl-o-amino acids, D-amino acids, p-amino acids, and o-hydroxy acids, each having a backbone structure different from that of natural amino acids; amino acids (such as norleucine and79MEl\59069747.vlAttorney Docket No. 137521-10301 homohistidine) having a side-chain structure different from that of natural amino acids; amino acids (such as "homo” amino acids, homophenylalanine, and homohistidine) having extra methylene in the side chain thereof; and amino acids (such as cysteic acid) obtained by substituting a carboxylic acid functional amino group in the side chain thereof by a sulfonic acid group.

[0192] The peptides described herein can comprise one or more unnatural amino acids. Unnatural amino acids include, but are not limited to, (1) amino acids corresponding to an amino acid residue on a polypeptide subjected to modification after expression (ex. phosphorylated tyrosine, acetylated lysine, or farnesylated cysteine), (2) amino acids that cannot be used in expression on a ribosome but occur naturally, and (3) artificial amino acids that do not occur naturally (unnatural amino acids). Non-limiting examples of unnatural amino acids include: p-acetyl-L-phenylalanine, p-iodo-L-phenylalanine, p-methoxyphenylalanine, O-methyl-L-tyrosine, p- propargyloxyphenylalanine, p-propargyl-phenylalanine, L-3-(2-naphthyl)alanine, 3-methyl-phenylalanine, 0-4- allyl-L-tyrosine, 4-propyl-L-tyrosine, tri-O-acetyl-GIcNAcp-serine, L-Dopa, fluorinated phenylalanine, isopropyl-L- phenylalanine, p-azido-L-phenylalanine, p-acyl-L-phenylalanine, p-benzoyl-L-phenylalanine, Boronophenylalanine, O-propargyltyrosine, L-phosphoserine, phosphonoserine, phosphonotyrosine, p- bromophenylalanine, selenocysteine, p-amino-L- phenylalanine, isopropyl-L-phenylalanine, and azido-lysine (AzK). In some embodiments, the unnatural amino acid is an unnatural analogue of a tyrosine amino acid; an unnatural analogue of a glutamine amino acid; an unnatural analogue of a phenylalanine amino acid; an unnatural analogue of an alanine amino acid; an unnatural analogue of a serine amino acid; an unnatural analogue of a threonine amino acid; an alkyl, aryl, acyl, azido, cyano, halo, hydrazine, hydrazide, hydroxyl, alkenyl, alkynl, ether, thiol, sulfonyl, seleno, ester, thioacid, borate, boronate, phospho, phosphono, phosphine, heterocyclic, enone, imine, aldehyde, hydroxylamine, keto, or amino substituted amino acid; or a combination thereof. In some embodiments, the unnatural amino acid is an amino acid with a photoactivatable cross-linker; a spin-labeled amino acid; a fluorescent amino acid; a metal binding amino acid; a metal-containing amino acid; a photocaged and / or photoisomerizable amino acid; a biotin or biotin-analogue containing amino acid; a keto containing amino acid; an amino acid comprising polyethylene glycol or polyether; a heavy atom substituted amino acid; a chemically cleavable or photocleavable amino acid; an amino acid with an elongated side chain; an amino acid containing a toxic group; a sugar substituted amino acid; a carbon-linked sugar-containing amino acid; a redox-active amino acid; an a-hydroxy containing acid; an amino thio acid; an a, a-disubstituted amino acid; a p-amino acid; a cyclic amino acid other than proline or histidine, or an aromatic amino acid other than phenylalanine, tyrosine or tryptophan.

[0193] Unnatural amino acids include, for example, N-alkyl amino acids in which a natural amino acid described above is N-alkylated, e.g., those modified with lower alkyl groups (for example, of C1 to C5, C1 to C3, and C1) in which the nitrogen forming a peptide bond is branched or not branched. Exemplary N-alkyl amino acids include, e.g., N-ethyl amino acid, N-butyl amino acid, and N-methyl amino acid. Also included are amino acids to which a functional group is further added to the side chain of a natural amino acid or substituted for another functional group (for example, an amino acid having a substitution or an addition in a part such as an arylene group, an alkylene group, or the like of the side chain; an amino acid wherein the arylene group or the80MEl\59069747.vlAttorney Docket No. 137521-10301 alkyl group of the side chain has an increased C-number; an amino acid having a substitution in the aromatic ring of the side chain; a heterocyclic or condensed cyclic amino acid; or the like). Exemplary N-alkyl amino acids further include, e.g., N-alkyllysine and N-methyllysine.

[0194] In a non-limiting manner, unnatural amino acids include, but are not limited to N-methyl amino acids, F4G, 4Py, 3Py, Cit, KCOpipzaa, Eva, Ahp, F4COO, KCOpip4COO, Aib, Hpr, Sbu, MeDapCOpipzaa, Scbm, Scpe, HseBu, A4paa, Spent, Hgl, Hsecpe, Hgn, DapCOpipzaa, MeD, CmG, Medd, HseEt, HseiPr, CrmG, CeG, CrpG, MeHgl, MeCit, F3G, 3Py6COO, 4Py2NH2, 3Py5COO, F3COO, 3Py6NHAc, F4C, F4OMe, Nal2, F3aao, F4aa, F4aao, 3Py6NHaa, 5Pdo, F3CON, F4F, F40Et, F4Me, F4CON, F4CONPEG4Me, F3OMe, YaeCOpipzaa, F4aaopipzaa, 4Pdo, 3Py6CON, Atp, Cha4cH, Cha4tH, Cha4cOMe, A1mor, F4amCOpipzaa, Chg, Tbg, Gcpr, Gcpe, Acpr, Cba, Gthp, NleCOO, NleOH, Atb, Nva, Nle, DapAc, Abu, Ncit, dMeS, TdMe, Cbg, NvaOMe, NleOMe, AhpOMe, F4u, F4CONdMe, F4ms, F34dOMe, F3C, F3CONdMe, 5lnda, F3aa, 3Py6NHae, 3Py6OMe, MeNal2, MeNal27N, MeF34diox, MeF34dOMe, MeF4T, MeW7N, MeF3C4Me, MeF3Me4C, Bph, 3Py6Ph, F41 Me4Pyz, F43Pyz, F44Pyz, F41Pyz, F41 Me3Pyz, F41 Et4Pyz, F41MeOe4Pyz, F41 MeOp4Pyz, F44thp, F4Ac4Pip, PhNva, PhNIe, Yph, Ybn, F4tb, F4OPr, S3REt, Yae, datb, Me3Py, W5N, MeA4paa, Mel, MeA, MeG, MeV, MeT, all, TMe, MeQ, MeTMe, MeK, MeKAc, Har, KAc, dd, EtG, K(biotin), MeeG, CmpG, Nmm, Ndm, SMe, HseMe, SiPr, SPr, MeWI Me, MeQdMe, MeSMe, MeHseMe, MeKCOpipzaa, KCOpipzetOH, KCOmeglumine, MeK(de), MeK(H), MeK(df), MeK(datb), and the like. Note that D-amino acids such as da may be classified as D- amino acids, but they may also be classified according to the properties of their side chains, and N-methyl amino acids may be classified as N-alkyl amino acids and may also be classified according to the property of the side chain.

[0195] In some embodiments, the unnatural amino acids incorporated into the peptides include one or more of: 1) a ketone functional group (as found in para or meta acetyl-phenylalanine) that can be specifically reacted with hydrazines, hydroxylamines and their derivatives (Addition of the keto functional group to the genetic code of Escherichia coli. Wang L, Zhang Z, Brock A, Schultz P G. Proc Natl Acad Sci USA. 2003 Jan. 7; 100(1 ):56-61 ; Bioorg Med Chem Lett. 2006 Oct. 15; 16 (20): 5356-9. Genetic introduction of a diketone-containing amino acid into proteins. Zeng H, Xie J, Schultz P G), 2) azides (as found in p-azido-phenyl alanine) that can be reacted with alkynes via copper catalyzed "click chemistry” or strain promoted (3+2) cycloadditions to form the corresponding triazoles (Addition of p-azido-L-phenylalanine to the genetic code of Escherichia coli. Chin J W, Santoro S W, Martin A B, King D S, Wang L, Schultz P G. J Am Chem Soc. 2002 Aug. 7; 124(31 ):9026-7; Adding amino acids with novel reactivity to the genetic code of Saccharomyces cerevisiae. Deiters A, Cropp T A, Mukherji M, Chin J W, Anderson J C, Schultz P G. J Am Chem Soc. 2003 Oct. 1; 125(39): 11782-3), or azides that can be reacted with aryl phosphines, via a Staudinger ligation (Selective Staudinger modification of proteins containing p- azidophenylalanine. Tsao M L, Tian F, Schultz P G. Chembiochem. 2005 December; 6(12):2147-9), to form the corresponding amides, 3) alkynes that can be reacted with azides to form the corresponding triazole ( / n vivo incorporation of an alkyne into proteins in Escherichia coli. Deiters A, Schultz P G. Bioorg Med Chem Lett. 2005 Mar. 1; 15(5): 1521 -4), and 4) boronic acids (boronates) than can be specifically reacted with compounds containing more than one appropriately spaced hydroxyl group or undergo palladium mediated coupling with81MEl\59069747.vlAttorney Docket No. 137521-10301 halogenated compounds (Angew Chem Int Ed Engl. 2008; 47(43):8220-3. A genetically encoded boronate- containing amino acid., Brustad E, Bushey M L, Lee J W, Groff D, Liu W, Schultz P G).

[0196] The peptide of the present disclosure embraces various derivatives thereof. Examples of the derivatives include derivatives having an amide, ester, or carboxyl group as the C-terminus and / or N-terminus thereof. Additional examples of the derivatives of the peptide include those obtained by modification such as phosphorylation, methylation, acetylation, adenylylation, ADP-ribosylation, or glycosylation and fused protein obtained by fusion with another peptide or protein. These derivatives can be prepared by those skilled in the art in a known manner or a method based thereon.

[0197] In some embodiments, the peptide described herein comprises a basic amino acid. Examples of the basic amino acid include arginine, lysine, citrulline, ornithine, creatine, histidine, diaminobutanoic acid, and diaminopropionic acid.

[0198] In some embodiments, provided herein is a peptide having 90% or more sequence identity to any of sequences disclosed herein. In some embodiments, the sequence identity is at least 95% or 99%.

[0199] The peptides and conjugates of the present disclosure embrace salts thereof. Physiologically acceptable base or acid are used to create salt forms of the peptides or conjugates. Examples include addition salts with an inorganic acid (such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, or phosphoric acid), addition salts with an organic acid (such as p-toluenesulfonic acid, methanesulfonic acid, oxalic acid, p-bromophenylsulfonic acid, carboxylic acid, succinic acid, citric acid, benzoic acid, or acetic acid), inorganic bases (such as ammonium hydroxide, alkali or alkaline earth metal hydroxide, carbonate, or bicarbonate), and an amino acid.

[0200] In certain embodiments, the peptides and conjugates described herein comprises one or more unnatural amino acids that are not any one of the 20 canonical amino acids found in proteins. Representative unnatural amino acids that can be incorporated into the peptides and conjugates described herein are described herein.Linker

[0201] A peptide or conjugate described herein can comprise or be linked to one or more linkers. In some embodiments, the linker covalently attaches the peptide with the non-radionuclide moiety linked to the peptide. In some embodiments, the peptide attaches directly to the moiety without a linker. In some embodiments, the present disclosure describes linkers that function as a spacer.

[0202] A peptide or conjugate described herein can comprise or be linked to a linker. The linker can comprise a number of intervening atoms (on a linear chain, excluding pendant groups or substituents) between the nonradionuclide moiety and the binding peptide described herein, thereby creating a distance between the nonradionuclide moiety and the binding peptide. In some embodiments, a linker comprises 10-100 intervening atoms between the moiety and the binding peptide. In some embodiments, a linker comprises 2-60 intervening atoms between the moiety and the binding peptide. In some embodiments, a linker comprises 2 to 20, 2 to 50, 5 to 15,5 to 25, 10 to 40, 30 to 60, or 10 to 20 intervening atoms between the moiety and the binding peptide. In some embodiments, a linker comprises 3 to 30 intervening atoms between the moiety and the binding peptide. In some82MEl\59069747.vlAttorney Docket No. 137521-10301 embodiments, a linker comprises 5 to 25 intervening atoms between the moiety and the binding peptide. In some embodiments, a linker comprises 6 to 18 intervening atoms between the moiety and the binding peptide. In some embodiments, a linker comprises 10 to 20 intervening atoms between the moiety and the binding peptide. In some embodiments, the linker comprises 3 to 10 intervening non-hydrogen, organic atoms between the moiety and the peptide.

[0203] The intervening atoms can comprise 1 or more carbons, and optionally one or more heteroatoms such as O and N. In some embodiments, the intervening atoms comprise 2 to 20, 2 to 50, 5 to 15, 5 to 25, 10 to 40, 30 to 60, or 10 to 20 carbons. In some embodiments, the intervening atoms comprise 0, 1, 2, 3, 4, 5, or 6 nitrogen. In some embodiments, the intervening atoms comprise 0, 1, 2, 3, 4, 5, 6, 7 or 8 oxygen. In some embodiments, the intervening atoms comprise 1 to 6 nitrogen and 0 to 4 oxygen.

[0204] A linker can comprise one or more amino acid residues. In some embodiments, the linker comprises 1 to 3, 1 to 5, 1 to 10, 5 to 10, or 5 to 20 amino acid residues. In some embodiments, the linker comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid residues. In some embodiments, the linker comprises 1 to 5 amino acid residues. For example, the linker can comprise one or more lysine (K) residues such as K, KK, or KKK sequences. In some embodiments, the linker comprises a lysine or a derivative thereof. In some embodiments, the linker comprises a lysine. In some embodiments, one or more amino acids of the linker are unnatural amino acids. In some embodiments, the linker comprises a lysine residue, an alanine residue, or both. In certain embodiments, the linker comprises one or more amino acids chosen from a lysine residue, an alanine residue, or a phenylalanine residue. In some embodiments, the linker comprises a lysine residue. In some embodiments, the linker comprises an alanine residue. In some embodiments, the linker comprises a lysine residue, an alanine residue, a glycine residue, a D-phenylalanine residue, a histidine residue, a dAtb residue, or a D-glutamate residue. In some embodiments, the linker comprises 2 amino acids selected from the group consisting of lysine, alanine, glycine, D-phenylalanine, histidine, dAtb, and D-glutamate.

[0205] A herein-described linker can attach to the N-terminus of the peptide, the C-terminus of the peptide, or a non-terminal amino acid of the peptide, or it can attach to the peptide through a combination of the above. In some embodiments, the linker is attached to the peptide via its N-terminus. In some embodiments, the linker is attached to the peptide via a cysteine residue at the C-terminus. In some embodiments, the linker is attached to the peptide via a cysteine residue at the N-terminus. In some embodiments, the linker is attached to the peptide via its C-terminus. In some embodiments, the linker is attached to the peptide via a non-terminal amino acid. The linker can be bonded to the peptide, the payload molecule, or both, for example, through a chemically reactive group. Exemplary chemically reactive groups include, but are not limited to, a free amino, imino, hydroxyl, thiol or carboxyl group (e.g., to the N- or C-terminus, to the epsilon amino group of one or more lysine residues, the free carboxylic acid group of one or more glutamic acid or aspartic acid residues, or to the sulfhydryl group of one or more cysteinyl residues). The site to which the linker is bound to the peptide can be a natural or unnatural amino acid of the peptide and / or it can be introduced into the peptide, e.g., by DNA recombinant technology (e.g., by introducing a cysteine or protease cleavage site in the amino acid sequence) or by protein biochemistry (e.g., reduction, pH adjustment or proteolysis). Exemplary methods for attaching the linker includes carbodiimide83MEl\59069747.vlAttorney Docket No. 137521-10301 reaction, reactions using bifunctional agents such as dialdehydes or imidoesters, Schiff base reaction, Suzuki- Miyaura cross-coupling reactions, Isothiocyanates as coupling agents, and click chemistry.

[0206] The linker can have a prescribed length thereby linking the moiety and the peptide while allowing an appropriate distance therebetween. In some embodiments, the linker has 1 to 100 atoms, 1 to 60 atoms, 1 to 30 atoms, 1 to 15 atoms, 1 to 10 atoms, 1 to 5, or 2 to 20 atoms in length. In some embodiments, the linker has 1 to 10 atoms in length. In some embodiments, the linker comprises 3 to 10 intervening non-hydrogen, organic atoms between the moiety and the peptide.

[0207] The linker can comprise flexible and / or rigid regions. Exemplary flexible linker regions include those comprising Gly and Ser residues (“GS” linker), glycine residues, alkylene chain, PEG chain, etc. Exemplary rigid linker regions include those comprising alpha helix-forming sequences (e.g., EAAAK (SEQ ID NO: 400)), prolinerich sequences, and regions rich in double and / or triple bonds.

[0208] In some embodiments, a linker may be further added to the cyclic peptide. Examples of the linker include an amino acid linker (peptide linker), a chemical linker, a fatty acid linker, a nucleic acid linker, a sugar chain linker, or the like, or it may be a complex, for example, a chemical linker, a peptide linker, or the like. Examples of the chemical linker include a PEG (polyethylene glycol) linker. For example, the PEG linker may comprise between 1 to 24 ethylene glycol units. Furthermore, the linker may be a fatty acid linker containing a divalent chemical moiety derived from a fatty acid. The linker includes at least one amino acid, and, for example, a glycine-rich peptide such as a peptide having a sequence [Gly-Gly-Gly-Gly-Ser] n (in the formula, n is 1, 2, 3, 4, 5, or 6) (SEQ ID NO: 401) such as that according to US Patent No. 7,271,149, incorporated by reference herein, or a serine-rich peptide linker according to US Patent No. 5,525,491, incorporated by reference herein, may be used. In a non-limiting manner, there are some cases where a physical property (for example, solubility) of the peptide may be changed by the addition of a linker. In one aspect, the amino acid linker includes an amino acid sequence according to any one of the peptides or conjugates in Tables 1-4 and 9A.

[0209] The linker may be added at any position. In some embodiments, the linker may be bound to Cys positioned on the C-terminal side or may be bound to an amino acid comprised in the cyclic peptide. In some embodiments, the linker is bound to Cys or variant thereof positioned on the C-terminal side. In some embodiments, the linker is added to the -COCH on the Cys or variant residue. It is possible to add one to several amino acids to the C-terminus of such Cys or cysteine variant, such as MeC, and then the liker is added to its terminus; for example, Gly is added to the C-terminus of Cys or cysteine variant, such as MeC, within the cyclic structure peptide, then the -COCH of the Gly is bound to linker, such as a PEG linker or an amino acid linker. In some embodiments, the linker is added to the side chain of an amino acid within the cyclic peptide. In some embodiments, the linker is added to the side chain of a lysine within the cyclic peptide. In some embodiments, the linker is added to the side chain of Xi, X2, X3, X4, X5, Xs, or X12 of Formula I.

[0210] The linker can be cleavable, e.g., under physiological conditions, e.g., under intracellular conditions, such that cleavage of the linker releases the moiety (sometimes referred to herein as "payload molecule”) in the intracellular environment. The linker can be, e.g., a peptidyl linker that is cleaved by an intracellular peptidase or protease enzyme, including, but not limited to, a lysosomal or endosomal protease. In some embodiments, the84MEl\59069747.vlAttorney Docket No. 137521-10301 peptidyl linker is at least two amino acids long or at least three amino acids long. Cleaving agents can include cathepsins B and D and plasmin. In other embodiments, the linker is not cleavable. In some embodiments, the linker is pH-sensitive, i.e., sensitive to hydrolysis at certain pH values. For example, the pH-sensitive linker can be hydrolyzable under acidic conditions. For example, a linker can be an acid-labile linker that is hydrolyzable in the lysosome (e.g., a hydrazone, semicarbazone, thiosemicarbazone, cis-aconitic amide, orthoester, acetal, ketal, or the like). Such linkers can be relatively stable under neutral pH conditions, such as those in the blood, but are unstable at below pH 5.5 or 5.0, the approximate pH of the lysosome. In some embodiments, the hydrolyzable linker is a thioether linker.

[0211] In some embodiments, the linker comprises an amino acid sequence, such as a combination of amino acid sequence and a flexible and / or rigid region.

[0212] In some embodiments, the linker comprises one or more of substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl. In some embodiments, the linker comprises substituted or unsubstituted C1-C30 alkylene. In some embodiments, the linker comprises substituted or unsubstituted C1-C30 heteroalkylene. In some embodiments, the linker comprises polyethylene glycol such as (-CH2-CH2-0-)I-IO.

[0213] In some embodiments, the linker has a structure ofFormula (11-1) wherein each L is independently -O-, -NRL-, -N(RL)2+-, -OP(=O)(ORL)O-, -S-, -S(=O)-, -S(=O)2-, =CH-, - C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, -OC(=O)NRL-, -NRLC(=O)O-, - NRLC(=O)NRL-, -NRLC(=S)NRL-, -CRL=N-, -N=CRL, -NRLS(=O)2-, -S(=O)2NRL-, -C(=O)NRLS(=O)2-, - S(=O)2NRLC(=O)-, substituted or unsubstituted C3-C15 cycloalkyl, substituted or unsubstituted C1-C12 heterocycloalkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted C1-C30 alkylene, substituted or unsubstituted C2-C30 alkenylene, substituted or unsubstituted C2-C30 alkynylene, substituted or unsubstituted C1-C30 heteroalkylene, -(C1-C30 alkylene)-O-, -0-(Ci-C3o alkylene)-, -(C1- C30 alkylene)-NRL- , -NRL-(CI-C3O alkylene)-, -(C1-C30 alkylene)-N(RL)2+-, -N(RL)2+-(Ci-C3o alkylene)-, or a click chemistry residue; and each RLis independently hydrogen, substituted or unsubstituted C1-C4 alkyl, substituted or unsubstituted C1-C4 heteroalkyl, substituted or unsubstituted C2-C6 alkenyl, substituted or unsubstituted C2-C5 alkynyl, substituted or unsubstituted C3-C8 cycloalkyl, substituted or unsubstituted C2-C7 heterocycloalkyl, substituted or unsubstituted aryl, or substituted or unsubstituted heteroaryl; and n is i to 20 (e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20).In some embodiments, the linker has a structure of, wherein each L is independently-O-, -NRL-, -N(RL)2+-, -OP(=O)(ORL)O-, -S-, -S(=O)-, -S(=O)2-, -CH=CH-, =CH-, -C C-, -C(=O)-, -C(=O)O-, -85MEl\59069747.vlAttorney Docket No. 137521-103010C(=0)-, -0C(=0)0-, -C(=O)NRL-, -NRLC(=O)-, -OC(=O)NRL-, -NRLC(=O)O-, -NRLC(=O)NRL-, -NRLS(=O)2-, - S(=O)2NRL-, -C(=O)NRLS(=O)2-, or -S(=O)2NRLC(=O)-.

[0214] In some embodiments, the linker of Formula (11-1) has a structure of Formula (11-1 a),Formula (11-1 a) wherein each of L1and L3is independently -O-, -NRL-, -N(RL)2-, -OP(=O)(ORL)O-, -S-, -S(=O)-, - S(=O)2-, -CH=CH-, =CH-, -C C-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, - OC(=O)NRL-, -NRLC(=O)O-, -NRLC(=O)NRL-, -NRLS(=O)2-, -S(=O)2NRL-, -C(=O)NRLS(=O)2-, or - S(=O)2NRLC(=O)-; andL2is absent, substituted or unsubstituted C1-C30 alkylene, or substituted or unsubstituted C1- C30 heteroalkylene.

[0215] In some embodiments, the linker comprises a structure of Formula (11-1 b),Formula (11-1 b) wherein each of L1and L5is independently -O-, -NRL-, -N(RL)2-, -OP(=O)(ORL)O-, -S-, -S(=O)-, - S(=O)2-, -CH=CH-, =CH-, -C C-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, - OC(=O)NRL-, -NRLC(=O)O-, -NRLC(=O)NRL-, -NRLS(=O)2-, -S(=O)2NRL-, -C(=O)NRLS(=O)2-, - S(=O)2NRLC(=O)-, substituted or unsubstituted 5-6 membered cycloalkyl, or substituted or unsubstituted 5-6 membered heterocycloalkyl; andL2, L3and L4are each independently absent, substituted or unsubstituted 5-6 membered cycloalkyl, substituted or unsubstituted 5-6 membered heterocycloalky, substituted or unsubstituted C1- C30 alkylene, or substituted or unsubstituted C1-C30 heteroalkylene.

[0216] In some embodiments, L1is -NH-.

[0217] In some embodiments, L2is absent. In some embodiments, L2is substituted or unsubstituted C1-C30 alkylene, or substituted or unsubstituted C1-C30 heteroalkylene. In some embodiments, L2is substituted or unsubstituted C1-C30 alkylene. In some embodiments, L2is substituted or unsubstituted C1-C30 heteroalkylene. In some embodiments, L2is substituted or unsubstituted C1-C18 alkylene, or substituted or unsubstituted C1-C18 heteroalkylene. In some embodiments, L2is optionally substituted. In some embodiments, L2is optionally substituted with one or more substituents selected from -OH, -SH, oxo, amino, Ci-Ce alkyl, Ci-Ce hydroxyalkyl, Ci-C6haloalkyl, Ci-C6aminoalkyl, -C(=O)ORL, -OC(=O)RL, -OC(=O)ORL, -C(=O)N(RL)2, -NRLC(=O)RL, - OC(=O)N(RL)2, and -NRLC(=O)ORL. In some embodiments, L2is C1-C30 heteroalkylene that is optionally substituted with one or more substituents selected from -OH, -SH, oxo, amino, Ci-Ce alkyl, Ci-Ce hydroxyalkyl,86MEl\59069747.vlAttorney Docket No. 137521-10301Ci-Ce haloalkyl, and Ci-Ce aminoalkyl. In some embodiments, L2 is optionally substituted with Ci-Ce alkyl which is further optionally substituted with one or more substituents chosen from -OH, -SH, oxo, amino, Ce-C aryl, 6- to 10- membered heteroaryl, -C(=O)ORL, -OC(=O)RL, -OC(=O)ORL, -C(=O)N(RL)2, -NRLC(=O)RL, - OC(=O)N(RL)2, and -NRLC(=O)ORL.

[0218] In some embodiments, L3is -NH-. In some embodiments, L3is absent.

[0219] In some embodiments, L4is absent. In some embodiments, L4is substituted or unsubstituted 5-6 membered cycloalkyl, substituted or unsubstituted 5-6 membered heterocycloalky, substituted or unsubstituted C1-C30 alkylene, or substituted or unsubstituted C1-C30 heteroalkylene.

[0220] In some embodiments, L5is -NH-. In some embodiments, L5is absent.

[0221] In some embodiments for Formula (11-1 b), L1is -O-, -N(methyl)-, -NH- or -C(=O)-; L5is -O-, - N(methyl)-, -NH- or -C(=O)-; L2, L3and L4are each independently absent, substituted or unsubstituted 5-6 membered cycloalkyl, substituted or unsubstituted 5-6 membered heterocycloalky, substituted or unsubstituted Ci-Ci2alkylene, or substituted or unsubstituted C1-C30 heteroalkylene, wherein L1is connected to the payload molecule and L5is connected to the GPC3 binding peptide.

[0222] In some embodiments for Formula (11-1 b), L2is unsubstituted Ci-Ci2alkylene, and L3and L4are absent.

[0223] In some embodiments, the linker comprises substituted or unsubstituted C1-C30 alkylene, Ci-Ci2alkylene, Ci-Cs alkylene, Ci-Ce alkylene, or C2-Ce alkylene. In some embodiments, the linker comprises C2-Ce alkylene. In some embodiments, the linker comprises C4-C6 alkylene.

[0224] In some embodiments, each of L1is independently -O-, -NRL-, -N(RL)2-, -OP(=O)(ORL)O-, -S-, - S(=O)-, -S(=O)2-, =CH-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, -OC(=O)NRL-, - NRLC(=O)O-, -NRLC(=O)NRL-, -NRLC(=S)NRL-, -CRL=N-, -N=CRL, -NRLS(=O)2-, -S(=O)2NRL-, - C(=O)NRLS(=O)2-, -S(=O)2NRLC(=O)-, substituted or unsubstituted C3-C15 cycloalkyl, substituted or unsubstituted Ci-Ci2heterocycloalkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted C1-C30 alkylene, substituted or unsubstituted C2-C3o alkenylene, substituted or unsubstituted C2-C3o alkynylene, or substituted or unsubstituted C1-C30 heteroalkylene, In some embodiments, L1is -O-, -NRL-, - OP(=O)(ORL)O-, -S-, -S(=O)-, -S(=O)2-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, - OC(=O)NRL-, -NRLC(=O)O-, -NRLC(=O)NRL-, -NRLC(=S)NRL-, -NRLS(=O)2-, -S(=O)2NRL-, -C(=O)NRLS(=O)2-, or -S(=O)2NRLC(=O)-. In some embodiments, L1is -O-, -NH-, -S(=O)-, -S(=O)2-, or -C(=O)-. In some embodiments, L1is -C(=O)NH- or -NHC(=O)-. In some embodiments, L1is substituted or unsubstituted C3-C15 cycloalkyl, or substituted or unsubstituted Ci-Ci2heterocycloalkyl. In some embodiments, L1is substituted or unsubstituted aryl or substituted or unsubstituted heteroaryl. In some embodiments, L1is substituted or unsubstituted C1-C30 alkylene. In some embodiments, L1is substituted or unsubstituted C2-C3o alkenylene. In some embodiments, L1is substituted or unsubstituted C1-C30 heteroalkylene. In some embodiments, L1is substituted or unsubstituted C5- C25 heteroalkylene. In some embodiments, L1is substituted or unsubstituted C5-Ci2heteroalkylene.

[0225] In some embodiments, each of L2is independently -O-, -NRL-, -N(RL)2-, -OP(=O)(ORL)O-, -S-, - S(=O)-, -S(=O)2-, =CH-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, -OC(=O)NRL-, -87MEl\59069747.vlAttorney Docket No. 137521-10301NRLC(=O)O-, -NRLC(=O)NRL-, -NRLC(=S)NRL-, -CRL=N-, -N=CRL, -NRLS(=O)2-, -S(=O)2NRL-, - C(=O)NRLS(=O)2-, -S(=O)2NRLC(=O)-, substituted or unsubstituted C3-C15 cycloalkyl, substituted or unsubstituted Ci-Ci2heterocycloalkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted C1-C30 alkylene, substituted or unsubstituted C2-C3o alkenylene, substituted or unsubstituted C2-C3o alkynylene, or substituted or unsubstituted C1-C30 heteroalkylene, In some embodiments, L2is -O-, -NRL-, - OP(=O)(ORL)O-, -S-, -S(=O)-, -S(=O)2-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, - OC(=O)NRL-, -NRLC(=O)O-, -NRLC(=O)NRL-, -NRLC(=S)NRL-, -NRLS(=O)2-, -S(=O)2NRL-, -C(=O)NRLS(=O)2-, or -S(=O)2NRLC(=O)-. In some embodiments, L2is -O-, -NH-, -S(=O)-, -S(=O)2-, or -C(=O)-. In some embodiments, L2is -C(=O)NH- or -NHC(=O)-. In some embodiments, L2is substituted or unsubstituted C3-C15 cycloalkyl, or substituted or unsubstituted Ci-Ci2heterocycloalkyl. In some embodiments, L2is substituted or unsubstituted aryl or substituted or unsubstituted heteroaryl. In some embodiments, L2is substituted or unsubstituted C1-C30 alkylene. In some embodiments, L2is substituted or unsubstituted C2-C3o alkenylene. In some embodiments, L2is substituted or unsubstituted C1-C30 heteroalkylene. In some embodiments, L2is substituted or unsubstituted C5- C25 heteroalkylene. In some embodiments, L2is substituted or unsubstituted C5-Ci2heteroalkylene.

[0226] In some embodiments, each of L3is independently -O-, -NRL-, -N(RL)2-, -OP(=O)(ORL)O-, -S-, - S(=O)-, -S(=O)2-, =CH-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, -OC(=O)NRL-, - NRLC(=O)O-, -NRLC(=O)NRL-, -NRLC(=S)NRL-, -CRL=N-, -N=CRL, -NRLS(=O)2-, -S(=O)2NRL-, - C(=O)NRLS(=O)2-, -S(=O)2NRLC(=O)-, substituted or unsubstituted C3-C15 cycloalkyl, substituted or unsubstituted Ci-Ci2heterocycloalkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted C1-C30 alkylene, substituted or unsubstituted C2-C3o alkenylene, substituted or unsubstituted C2-C3o alkynylene, or substituted or unsubstituted C1-C30 heteroalkylene, In some embodiments, L3is -O-, -NRL-, - OP(=O)(ORL)O-, -S-, -S(=O)-, -S(=O)2-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, - OC(=O)NRL-, -NRLC(=O)O-, -NRLC(=O)NRL-, -NRLC(=S)NRL-, -NRLS(=O)2-, -S(=O)2NRL-, -C(=O)NRLS(=O)2-, or -S(=O)2NRLC(=O)-. In some embodiments, L3is -O-, -NH-, -S(=O)-, -S(=O)2-, or -C(=O)-. In some embodiments, L3is -C(=O)NH- or -NHC(=O)-. In some embodiments, L3is substituted or unsubstituted C3-C15 cycloalkyl, or substituted or unsubstituted Ci-Ci2heterocycloalkyl. In some embodiments, L3is substituted or unsubstituted aryl or substituted or unsubstituted heteroaryl. In some embodiments, L3is substituted or unsubstituted C1-C30 alkylene. In some embodiments, L3is substituted or unsubstituted C2-C3o alkenylene. In some embodiments, L3is substituted or unsubstituted C1-C30 heteroalkylene. In some embodiments, L3is substituted or unsubstituted C5- C25 heteroalkylene. In some embodiments, L3is substituted or unsubstituted C5-Ci2heteroalkylene. In some embodiments, L3is absent.

[0227] In some embodiments, each of L4is independently -O-, -NRL-, -N(RL)2-, -OP(=O)(ORL)O-, -S-, - S(=O)-, -S(=O)2-, =CH-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, -OC(=O)NRL-, - NRLC(=O)O-, -NRLC(=O)NRL-, -NRLC(=S)NRL-, -CRL=N-, -N=CRL, -NRLS(=O)2-, -S(=O)2NRL-, - C(=O)NRLS(=O)2-, -S(=O)2NRLC(=O)-, substituted or unsubstituted C3-C15 cycloalkyl, substituted or unsubstituted Ci-Ci2heterocycloalkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted C1-C30 alkylene, substituted or unsubstituted C2-C3o alkenylene, substituted or unsubstituted C2-C3o88MEl\59069747.vlAttorney Docket No. 137521-10301 alkynylene, or substituted or unsubstituted C1-C30 heteroalkylene, In some embodiments, L4is -O-, -NRL-, - OP(=O)(ORL)O-, -S-, -S(=O)-, -S(=O)2-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, - OC(=O)NRL-, -NRLC(=O)O-, -NRLC(=O)NRL-, -NRLC(=S)NRL-, -NRLS(=O)2-, -S(=O)2NRL-, -C(=O)NRLS(=O)2-, or -S(=O)2NRLC(=O)-. In some embodiments, L4is -O-, -NH-, -S(=O)-, -S(=O)2-, or -C(=O)-. In some embodiments, L4is -C(=O)NH- or -NHC(=O)-. In some embodiments, L4is substituted or unsubstituted C3-C15 cycloalkyl, or substituted or unsubstituted Ci-Ci2heterocycloalkyl. In some embodiments, L4is substituted or unsubstituted aryl or substituted or unsubstituted heteroaryl. In some embodiments, L4is substituted or unsubstituted C1-C30 alkylene. In some embodiments, L4is substituted or unsubstituted C2-C3o alkenylene. In some embodiments, L4is substituted or unsubstituted C1-C30 heteroalkylene. In some embodiments, L4is substituted or unsubstituted C5- C25 heteroalkylene. In some embodiments, L4is substituted or unsubstituted C5-Ci2heteroalkylene. In some embodiments, L4is absent.

[0228] In some embodiments, each of L5is independently -O-, -NRL-, -N(RL)2-, -OP(=O)(ORL)O-, -S-, - S(=O)-, -S(=O)2-, =CH-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, -OC(=O)NRL-, - NRLC(=O)O-, -NRLC(=O)NRL-, -NRLC(=S)NRL-, -CRL=N-, -N=CRL, -NRLS(=O)2-, -S(=O)2NRL-, - C(=O)NRLS(=O)2-, -S(=O)2NRLC(=O)-, substituted or unsubstituted C3-C15 cycloalkyl, substituted or unsubstituted Ci-Ci2heterocycloalkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted C1-C30 alkylene, substituted or unsubstituted C2-C3o alkenylene, substituted or unsubstituted C2-C3o alkynylene, or substituted or unsubstituted C1-C30 heteroalkylene, In some embodiments, L5is -O-, -NRL-, - OP(=O)(ORL)O-, -S-, -S(=O)-, -S(=O)2-, -C(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -C(=O)NRL-, -NRLC(=O)-, - OC(=O)NRL-, -NRLC(=O)O-, -NRLC(=O)NRL-, -NRLC(=S)NRL-, -NRLS(=O)2-, -S(=O)2NRL-, -C(=O)NRLS(=O)2-, or -S(=O)2NRLC(=O)-. In some embodiments, L5is -O-, -NH-, -S(=O)-, -S(=O)2-, or -C(=O)-. In some embodiments, L5is -C(=O)NH- or -NHC(=O)-. In some embodiments, L5is substituted or unsubstituted C3-C15 cycloalkyl, or substituted or unsubstituted Ci-Ci2heterocycloalkyl. In some embodiments, L5is substituted or unsubstituted aryl or substituted or unsubstituted heteroaryl. In some embodiments, L5is substituted or unsubstituted C1-C30 alkylene. In some embodiments, L5is substituted or unsubstituted C2-C3o alkenylene. In some embodiments, L5is substituted or unsubstituted C1-C30 heteroalkylene. In some embodiments, L5is substituted or unsubstituted C5- C25 heteroalkylene. In some embodiments, L5is substituted or unsubstituted C5-Ci2heteroalkylene. In some embodiments, L5is absent.

[0229] In some embodiments, at least one L1is unsubstituted C3-C2o alkylene.

[0230] In some embodiments, the linker comprises one or more of a substituted or unsubstituted Ce-C aryl, substituted or unsubstituted C5-C9 heteroaryl, a sterol, sulfonamide, phosphate ester, polyethylene glycol, or C3- C2o alkylene, or amino acid residues.

[0231] In some embodiments, the linker comprises one or more selected from AEEA, AEEP, AEEEP, andAEEEEP groups. In some embodiments, the linker comprises o (AEEA). In o some embodiments, the linker comprises(AEEP). In some embodiments,89MEl\59069747.vlAttorney Docket No. 137521-10301the linker comprises o (AEEEA). In some embodiments, the linker comprisesments, the linker is or comprises lysine. In some embodiments, the linker comprises C1-C12 alkylene. In some embodiments, the linker comprises C3-C9 alkylene. In some embodiments, the linker comprises C2-C8 alkylene. In some embodiments, the linker comprises 1 to 10 repeating ethylene glycol units. In some embodiments, the linker comprises 2 to 4 repeating ethylene glycol units. In some embodiments, the linker comprises 5 to 8 repeating ethylene glycol units. In some embodiments, the linker comprises NH2-(CH2)n-COOH, wherein n is 1 to 12. In some embodiments, the linker comprises NH2-(CH2)2-COOH. In some embodiments, the linker comprises NH2-(CH2)3- COOH. In some embodiments, the linker comprises NH2-(CH2)4-COOH. In some embodiments, the linker comprises NH2-(CH2)5-COOH. In some embodiments, the linker comprises NH2-(CH2)6-COOH. In some embodiments, the linker comprises NH2-(CH2)z-COOH. In some embodiments, the linker comprises NH2-(CH2)s- COOH. In some embodiments, the linker comprises NH2-(CH2)IO-COOH. In some embodiments, the linker is absent.

[0233] In some embodiments, a linker of the present disclosure e.g, a linker of Formula (11-1), (ll-1a) or (II-

[0234] In some embodiments, a linker of the present disclosure comprises a structure of

[0235] In some embodiments, a linker of the present disclosure (e.g, a linker of Formula (11-1), (ll-1 a) or (II-90MEl\59069747.vlAttorney Docket No. 137521-1030191MEl\59069747.vlAttorney Docket No. 137521-10301wherein each k1 is independently 0 or an integer from 1 to 20; and each k2 is independently 0 or an integer from1 to 15. In some embodiments, k1 Is O. In some embodiments, k1 is 1. In some embodiments, k1 is 2. In some embodiments, k1 is 3. In some embodiments, k1 is 4. In some embodiments, k1 is 5. In some embodiments, k1 is 6. In some embodiments, k1 is 7. In some embodiments, k1 is 8. In some embodiments, k1 is 9. In some embodiments, k1 is 10. In some embodiments, k1 is 11. In some embodiments, k1 is 12. In some embodiments, k1 is 13. In some embodiments, k1 is 14. In some embodiments, k1 is 15. In some embodiments, k1 is 16. In some embodiments, k1 is 17. In some embodiments, k1 is 18. In some embodiments, k1 is 19. In some embodiments, k1 is 20. In some embodiments, k2 is 0. In some embodiments, k2 is 1. In some embodiments, k2 is 2. In some embodiments, k2 is 3. In some embodiments, k2 is 4. In some embodiments, k2 is 5. In some embodiments, k2 is 6. In some embodiments, k2 is 7. In some embodiments, k2 is 8. In some embodiments, k2 is 9. In some embodiments, k2 is 10. In some embodiments, k2 is 11. In some embodiments, k2 is 12. In some embodiments, k2 is 13. In some embodiments, k2 is 14. In some embodiments, k2 is 15.

[0237] In some embodiments, the linker comprises a structure selected from: H H92MEl\59069747.vlAttorney Docket No. 137521-1030193MEl\59069747.vlAttorney Docket No. 137521-10301

[0239] In some embodiments, the linker is a bond.

[0240] In some embodiments, a conjugate comprises a linker structure selected from the following:94MEl\59069747.vlAttorney Docket No. 137521-1030195MEl\59069747.vlAttorney Docket No. 137521-10301wherein each k1 and k2 is independently 0 or an integer selected from 1 to 20.

[0241] In some embodiments, k1 is selected from 0-12. In some embodiments, k1 is 0. In some embodiments, k1 is 1. In some embodiments, k1 is 2. In some embodiments, k1 is 3. In some embodiments, k1 is 4. In some embodiments, k1 is 5. In some embodiments, k1 is 6. In some embodiments, k1 is 7. In some embodiments, k1 is 8. In some embodiments, k1 is 9. In some embodiments, k1 is 10. In some embodiments, k2 is selected from 0-12. In some embodiments, k2 is 0. In some embodiments, k2 is 1. In some embodiments, k2 is 2. In some embodiments, k2 is 3. In some embodiments, k2 is 4. In some embodiments, k2 is 5. In some embodiments, k2 is 6. In some embodiments, k2 is 7. In some embodiments, k2 is 8. In some embodiments, k2 is 9. In some embodiments, k2 is 10.Preparation of the Peptides and Conjugates

[0242] The compounds used in the reactions described herein are made according to organic synthesis techniques known to those skilled in this art, starting from commercially available chemicals and / or from compounds described in the chemical literature. "Commercially available chemicals” are obtained from standard96MEl\59069747.vlAttorney Docket No. 137521-10301 commercial sources including Acros Organics (Pittsburgh, PA), Aldrich Chemical (Milwaukee, Wl, including Sigma Chemical and Fluka), Apin Chemicals Ltd. (Milton Park, UK), Avocado Research (Lancashire, U.K.), BDH, Inc. (Toronto, Canada), Bionet (Cornwall, U.K.), Chem Service Inc. (West Chester, PA), Crescent Chemical Co. (Hauppauge, NY), Eastman Organic Chemicals, Eastman Kodak Company (Rochester, NY), Fisher Scientific Co. (Pittsburgh, PA), Fisons Chemicals (Leicestershire, UK), Frontier Scientific (Logan, UT), ICN Biomedicals, Inc. (Costa Mesa, CA), Key Organics (Cornwall, U.K.), Lancaster Synthesis (Windham, NH), Maybridge Chemical Co. Ltd. (Cornwall, U.K.), Parish Chemical Co. (Orem, UT), Pfaltz & Bauer, Inc. (Waterbury, CN), Polyorganix (Houston, TX), Pierce Chemical Co. (Rockford, IL), Riedel de Haen AG (Hanover, Germany), Spectrum Quality Product, Inc. (New Brunswick, NJ), TCI America (Portland, OR), Trans World Chemicals, Inc. (Rockville, MD), and Wako Chemicals USA, Inc. (Richmond, VA).

[0243] Suitable reference books and treatises that detail the synthesis of reactants useful in the preparation of compounds described herein, or provide references to articles that describe the preparation, include for example, "Synthetic Organic Chemistry”, John Wiley & Sons, Inc., New York; S. R. Sandler et al., "Organic Functional Group Preparations,” 2nd Ed., Academic Press, New York, 1983; H. 0. House, "Modern Synthetic Reactions”, 2nd Ed., W. A. Benjamin, Inc. Menlo Park, Calif. 1972; T. L. Gilchrist, "Heterocyclic Chemistry”, 2nd Ed., John Wiley & Sons, New York, 1992; J. March, "Advanced Organic Chemistry: Reactions, Mechanisms and Structure”, 4th Ed., Wiley-lnterscience, New York, 1992. Additional suitable reference books and treatises that detail the synthesis of reactants useful in the preparation of compounds described herein, or provide references to articles that describe the preparation, include for example, Fuhrhop, J. and Penzlin G. "Organic Synthesis: Concepts, Methods, Starting Materials”, Second, Revised and Enlarged Edition (1994) John Wiley & Sons ISBN: 3-527-29074-5; Hoffman, R.V. "Organic Chemistry, An Intermediate Text” (1996) Oxford University Press, ISBN 0-19-509618-5; Larock, R. C. "Comprehensive Organic Transformations: A Guide to Functional Group Preparations” 2nd Edition (1999) Wiley-VCH, ISBN: 0-471-19031-4; March, J. "Advanced Organic Chemistry: Reactions, Mechanisms, and Structure” 4th Edition (1992) John Wiley & Sons, ISBN: 0-471-60180-2; Otera, J. (editor) "Modern Carbonyl Chemistry” (2000) Wiley-VCH, ISBN: 3-527-29871-1 ; Patai, S. "Patai's 1992 Guide to the Chemistry of Functional Groups” (1992) Interscience ISBN: 0-471-93022-9; Solomons, T. W. G. "Organic Chemistry” 7th Edition (2000) John Wiley & Sons, ISBN: 0-471-19095-0; Stowell, J.C., "Intermediate Organic Chemistry” 2nd Edition (1993) Wiley-lnterscience, ISBN: 0-471-57456-2; "Industrial Organic Chemicals: Starting Materials and Intermediates: An Ullmann's Encyclopedia” (1999) John Wiley & Sons, ISBN: 3-527-29645-X, in 8 volumes; "Organic Reactions” (1942-2000) John Wiley & Sons, in over 55 volumes; and "Chemistry of Functional Groups” John Wiley & Sons, in 73 volumes.

[0244] Specific and analogous reactants are optionally identified through the indices of known chemicals prepared by the Chemical Abstract Service of the American Chemical Society, which are available in most public and university libraries, as well as on-line. Chemicals that are known but not commercially available in catalogs are optionally prepared by custom chemical synthesis houses, where many of the standard chemical supply houses (e.g., those listed above) provide custom synthesis services. A reference for the preparation and selection of pharmaceutical salts of the compounds described herein is P. H. Stahl & C. G. Wermuth "Handbook97MEl\59069747.vlAttorney Docket No. 137521-10301 of Pharmaceutical Salts”, Verlag Helvetica Chimica Acta, Zurich, 2002.

[0245] In one aspect, described herein is a method of making a conjugate that comprises a cyclic peptide, a non-radionuclide moiety, and optionally a linker. In some embodiments, the conjugate is prepared by one or more of the following steps: (a) synthesizing the peptide sequence by solid phase peptide synthesis; (b) cyclizing the peptide by forming an intramolecular non-peptide bond; and (c) coupling the non-radionuclide moiety to the peptide. In some embodiments, steps (a), (b), and (c) are performed in the recited order. In some embodiments, synthesizing the peptide comprises synthesizing the peptide sequence in a protected form and performing a deprotecting reaction. In some embodiments, cyclizing the peptide comprises forming a non-peptide bond between the N-terminus and the C-terminus of the peptide. In some embodiments, cyclizing the peptide comprises forming a non-peptide bond between the N-terminus and a cysteine or homocysteine of the peptide. In some embodiments, cyclizing the peptide comprises forming a ring closing group selected from -C(=O)-CH2-, -C(=O)- CH2-S-, -S-, -CH=CH-, -NH-, -maleimide-S-, -C(=O)-CH2-NH-, and -C(=O)-CH2-O-. In some embodiments, cyclizing the peptide comprises forming a ring closing group of Table 4B. In some embodiments, cyclizing the peptide comprises reacting a pair of functional groups or amino acids described in Table 4C. In some embodiments, solid phase peptide synthesis can be replaced with other suitable peptide synthesis methods known in the art.Peptide Production

[0246] The peptide of the present technology may be produced by, for example, any known method for producing a peptide, such as the following:• a chemical synthesis method such as a liquid phase method, a solid phase method, a hybrid method combining a liquid phase method and a solid phase method, or the like;• a genetic recombination method; or the like.

[0247] In some of the instances where the peptide of the present technology is produced by a chemical synthesis method, it can be said that the peptide of the present technology is a synthetic peptide.

[0248] In the solid phase method, for example, a hydroxy group of a resin having a hydroxy group and a carboxy group of a first amino acid (normally a C-terminal amino acid of a target peptide) in which an o-amino group is protected by a protecting group are subjected to an esterification reaction. For the esterification catalyst, a known dehydrating and condensing agent such as 1 -mesitylenesulfonyl-3-nitro-1 , 2, 4-triazole (MSNT), dicyclohexylcarbodiimide (DCC), and diisopropylcarbodiimide (DIPCDI) may be used.

[0249] Next, the protecting group of the o-amino group of the first amino acid is removed, a second amino acid in which all functional groups except the carboxy group of the main chain are protected is added, and the carboxy group is activated, binding the first and second amino acids. Furthermore, the o-amino group of the second amino acid is deprotected, a third amino acid in which all functional groups except the carboxy group of the main chain are protected is added, the carboxy group is activated, binding the second and third amino acids. This is repeated, and after a peptide having a target length is synthesized, all of the functional groups are deprotected.

[0250] Examples of the resin for solid-phase synthesis include Merrifield resin, MBHA resin, Cl-Trt resin,98MEl\59069747.vlAttorney Docket No. 137521-10301SASRIN resin, Wang resin, Rink amide resin, HMFS resin, Amino-PEGA resin (Merck KGaA), HMPA-PEGA resin (Merck KGaA), and the like. These resins may be used after being washed using a solvent (dimethylformamide (DMF), 2-propanol, methylene chloride, and the like). A peptide chain can be cleaved from the resin by treating it with an acid such as TFA or hydrogen fluoride (HF).

[0251] Examples of the protecting group of the o-amino group include a benzyloxycarbonyl (Cbz or Z) group, tert-butoxycarbonyl (Boc) group, fluorenylmethoxycarbonyl (Fmoc) group, benzyl group, allyl group, allyloxycarbonyl (Alloc) group, and the like. The Cbz group may be deprotected by a treatment using hydrofluoric acid, hydrogenation, or the like, the Boc group may be deprotected by a treatment using trifluoroacetic acid (TFA), and the Fmoc group may be deprotected by a treatment using pipericine or pyrrolysine.

[0252] Examples, such as methyl ester, ethyl ester, allyl ester, benzyl ester, tert-butyl ester, cyclohexyl ester, and the like may be used to protect the o-carboxy group.

[0253] As other functional groups of an amino acid, the hydroxyl group of serine or threonine can be protected with a benzyl group or a tert-butyl group and the hydroxyl group of tyrosine can be protected with a 2- bromobenzyloxycarbonyl group or a tert-butyl group. The amino group of a lysine side chain or the carboxyl group of glutamic acid or aspartic acid can be protected in a manner similar to the a-amino group or a-carboxyl group.

[0254] Activation of the carboxy group may be performed using a condensing agent. Examples of the condensing agent include dicyclohexylcarbodiimide (DOC), diisopropylcarbodiimide (DIPCDI), 1 -ethyl-3-(3- dimethylaminopropyl)carbodiimide (EDO or WSC), (1 H-benzotriazole-1-yloxy)tris(dimethylamino)phosphonium hexafluorophosphate (BOP), 1-[bis(dimethylamino)methyl]-1 H-benzotriazolium-3-oxide hexafluorophosphate (HBTU), and the like.

[0255] Cleavage of the peptide chain from the resin may be performed by treating the peptide chain using an acid such as TFA, hydrogen fluoride (HF), or the like.

[0256] Production of a peptide by a gene recombination method (translation / synthesis system) may be performed using a nucleic acid encoding the peptide. The nucleic acid encoding the peptide may be DNA or RNA.

[0257] The nucleic acid encoding the peptide may be prepared by a known method or a method equivalent thereto. For example, the peptide may be synthesized by an automated synthesizer. A restriction enzyme recognition site may be added to insert the obtained DNA into a vector. Alternatively, a base sequence encoding an amino acid sequence for splicing a formed peptide chain using an enzyme or the like may be incorporated. The peptide obtained may be converted from a free peptide to a salt thereof or from a salt thereof to a free peptide by a known method or a method based thereon.

[0258] As described above, when the peptide is fused to a cell-penetrating peptide or the like, the nucleic acid also includes a nucleic acid encoding the cell-penetrating peptide.

[0259] A chimeric protein expression method for expressing the target peptide as a chimeric peptide of another peptide may also be used to suppress degradation by a host-derived protease. In this case, a nucleic acid encoding the target peptide and the peptide bound thereto may be used as the nucleic acid.99MEl\59069747.vlAttorney Docket No. 137521-10301

[0260] Subsequently, an expression vector is prepared using the nucleic acid encoding the peptide. The nucleic acid may be digested as is or by a restriction enzyme, and alternatively, the nucleic acid may be inserted downstream of a promotor of the expression vector by adding a linker, or the like. Examples of the vector include an Escherichia coll-dewed plasmid (pBR322, pBR325, pUC12, pUC13, pUC18, pUC19, pUC118, pBluescript II, and the like), a Bacillus subtilis-demed plasmid (pUB110, pTP5, pC1912, pTP4, pE194, pC194, and the like), a yeast-derived plasmid (pSH19, pSH15, YEp, YRp, Yip, YAC, and the like), a bacteriophage (e phage, M13 phage, and the like), a virus (retrovirus, vaccinia virus, adenovirus, adeno-associated virus (AAV), cauliflower mosaic virus, tobacco mosaic virus, baculovirus, and the like), a cosmid, and the like.

[0261] The promoter may be selected appropriately according to the type of host. When the host is an animal cell, for example, a promoter derived from SV40 (simian virus 40) or a promoter derived from CMV (cytomegalovirus) may be used. When the host is Escherichia coll, a trp promoter, a T7 promoter, a lac promoter, or the like may be used.

[0262] The expression vector may incorporate, for example, a DNA replication starting point (ori), a selective marker (antibiotic resistance, auxotrophy, or the like), an enhancer, a splicing signal, a poly-A addition signal, a nucleic acid encoding a tag (FLAG, HA, GST, GFP, or the like), or the like.

[0263] Next, an appropriate host cell is transformed by the expression vector. The host may be appropriately selected in relation to the vector. Examples such as Escherichia coll, Bacillus subtilis {Bacillus), yeast, insects or insect cells, animal cells, or the like may be used as the host. As the animal cells, for example, HEK293T cells, OHO cells, COS cells, myeloma cells, HeLa cells, and Vero cells may be used. Transformation may be carried out according to a known method, such as a lipofection method, a calcium phosphate method, an electroporation method, a microinjection method, a gene gun method, or the like depending on the type of host. The target peptide is expressed by culturing a transformant according to a conventional method.

[0264] As for purification of the peptide from the transformant culture, cultured cells are recovered and then suspended in an appropriate buffer solution, followed by disruption of cells by a method such as sonication, freeze-thawing, or the like, and then a crude extract is obtained by centrifugation or filtration. When the peptide is secreted into the culture solution, a supernatant is recovered.

[0265] Purification of the crude extract or the culture supernatant may also be performed by a known method or a method equivalent thereto (for example, salting-out, dialysis, an ultrafiltration method, gel filtration method, SDS-PAGE method, ion exchange chromatography, affinity chromatography, reversed-phase high-performance liquid chromatography, and the like).

[0266] The obtained peptide may be converted from a free body to a salt or from a salt to a free body by a known method or a method equivalent thereto.

[0267] In one aspect, the translation / synthesis system may be a cell-free translation system. According to the cell-free translation system, a highly pure form of an expression product can generally be obtained without purification. The cell-free translation system includes, for example, a ribosome protein, an aminoacyl-tRNA synthase (ARS), a ribosome RNA, an amino acid, rRNA, GTP, ATP, a translation initiation factor (IF), an elongation factor (EF), a release factor (RF), and a ribosome regeneration factor (RRF), or another factor100MEl\59069747.vlAttorney Docket No. 137521-10301 required for translation. An Escherichia coli extract or a wheat embryo extract may be added to increase expression efficiency. In addition, a rabbit red blood cell extract or an insect cell extract may be added.

[0268] By continuously supplying energy to a system including these using dialysis, a protein of several hundred pg to several mg / mL may be produced in a non-limiting manner. The system may include an RNA polymerase to concurrently perform transcription of genomic DNA. Examples of commercially available cell-free translation systems that may be used include RTS-100 (registered trademark) by Roche Diagnostics K.K., PURE System by GeneFrontier Corporation, PURExpress In vitro Protein Synthesis Kit by New England Biolabs Inc., and the like for a system derived from Escherichia coli, and a system by ZOIGENE, CellFree Sciences Co., Ltd., or the like for a system using wheat embryo extract.

[0269] In the cell translation system, artificial aminoacyl-tRNA may be used and a desired amino acid or hydroxy acid may be linked (acylated) to a tRNA in place of an aminoacyl-tRNA synthesized by a natural aminoacyl-tRNA synthase. The aminoacyl-tRNA may be synthesized using an artificial ribozyme.

[0270] An example of the ribozyme includes a flexizyme (flexizyme) (H. Murakami, H. Saito, and H. Suga, (2003), Chemistry & Biology, Vol. 10, 655-662; and WO 2007 / 066627 and the like), all incorporated herein by reference. Flexizymes are also known under the names of prototype flexizyme (Fx), newly modified dinitrobenzyl flexizyme (dFx), enhanced flexizyme (eFx), aminoflexizyme (aFx), and the like.

[0271] A desired codon may be translated in association with the desired amino acid or hydroxy acid by using the tRNA produced by flexizyme and to which the desired amino acid or hydroxy acid is linked. A specialty amino acid may be used as the desired amino acid. For example, an unnatural amino acid required for the above circularization may also be introduced into the binding peptide by this method.

[0272] Various methods commonly used in the technical field may be used for chemical synthesis of the peptide, including, for example, stepwise solid-phase synthesis, semisynthesis of peptide fragments undergoing conformationally supported religation, and chemical ligation. Synthesis of the peptide is chemical synthesis using various solid phase technologies described in, for example, K. J. Jensen, P. T. Shelton, S. L. Pedersen, Peptide Synthesis and Applications, 2nd Edition, Springer, 2013, and the like. A preferable strategy is based on a combination of an Fmoc group capable of temporarily protecting the o-amino group and being selectively removed using a base, and a protecting group that temporarily protects a side chain functional group and is stable under Fmoc deprotection conditions. Selection of this kind of general peptide side chain is known according to the aforementioned Peptide Synthesis and Applications, 2nd Edition; G. B. Fields, R. L. Noble, Solid Phase Peptide Synthesis Utilizing 9-Fluorenylmethoxycarbonyl Amino Acids, Int. J. Peptide Protein Res. 35, 1990, 161-214, and the like; however, preferable peptide side chain protecting groups include, for example, a benzyl group or a tert-butyl group and a trityl (Trt) group for the hydroxy group of serine or threonine; a 2- bromobenzyloxycarbonyl group or a tert-butyl group for the hydroxy group of tyrosine; a Boc group, a methyltetrazole thiol (Mtt) group, an Alloc group, and an IvDde group for the amino group of the lysine side chain; a Trt group or a Boc group for the imidazole group of histidine; a 2,2,4,6,7-pentamethyldihydrobenzofuran-5- sulfonyl group (Pbf) group for the guanidyl group of arginine; a tert-butyl group, an allyl group, and a 3- methylpentane (Mpe) group for carboxyl groups, such as glutamic acid and aspartic acid; a Trt group for the101MEl\59069747.vlAttorney Docket No. 137521-10301 carboxamide group of glutamine or asparagine; or a Trt group and a monomethoxytrityl (Mmt) group for the thiol group of cysteine.

[0273] The peptide may be synthesized by a stepwise method on the solid-phase resin described above. The C-terminal amino acid to be used and all of the amino acids or peptides to be used for synthesis must be selectively removed during the process of synthesizing the o-amino protecting group. Preferably, the solid-phase resin described above is used, and once a C-terminal carboxyl group of a peptide having its N-terminal properly protected by Fmoc or the like or a C-terminal carboxyl group of an amino acid having its N-terminal protected by Fmoc is made into an activated ester by an appropriate reagent, this is then added to the amino group on the solid-phase resin to start. Subsequent elongation of the peptide chain may be achieved by removing the N- terminal protecting group (Fmoc group) then successively repeating condensation of the protected amino acid derivative according to the amino acid sequence of the target peptide. Note that these may release the target peptide in a final stage. Examples of releasing conditions are given in Teixeira, W. E. Benckhuijsen, P. E. de Koning, A. R. P. M. Valentijn, J. W. Drijfhout, Protein Pept. Lett., 2002, 9, 379-385, and the like, and the peptide may be released in a TFA solution containing water / silyl hydride / thiol as a scavenger in TFA. Typical examples include TFA / Water / TIS / DODT (volume ratio 92.5:2.5:2.5:2.5).

[0274] Synthesis of the peptide described in the present specification may be carried out using a single or multi-channel peptide synthesizer, for example, a Liberty Blue synthesizer from GEM Corporation, a Syro I synthesizer or a successor machine thereof from Biotage Japan, Ltd., or the like.

[0275] Activation of the carboxy group may be performed using a condensing agent. Examples of the condensing agent include dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIPCDI), 1 -ethyl-3-(3- dimethylaminopropyl)carbodiimide (EDC or WSC), (1 H-benzotriazole-1-yloxy)tris(dimethylamino)phosphonium hexafluorophosphate (BOP), 1-[bis(dimethylamino)methyl]-1 H-benzotriazolium-3-oxide hexafluorophosphate (HBTU), and the like.

[0276] Cyclization of the peptide may be carried out according to a known method. In a non-limiting manner, by designing the peptide to comprise two or more cysteine residues, for example, a cyclic structure may be formed by a disulfide bond after translation. Furthermore, according to the method of Goto et al. (Y. Goto, et al. ACS Chem. Biol. 3 120-129 (2008)), a peptide having a chloroacetyl group at its N-terminal may be synthesized by genetic code reprogramming technology and may also be circularized by disposing a cysteine residue containing a sulfur molecule in the peptide. Thus, a mercapto group spontaneously performs a nucleophilic attack on the chloroacetyl group after translation, and the peptide is circularized by thioether binding. Other amino acid combinations that bind to form a ring may be disposed within the peptide and circularized by genetic code reprogramming technology. Alternatively, circularization may be carried out by disposing an L-2- aminoadipic acid residue in the peptide and binding it to the main chain amino group of the N-terminal. In this manner, a known circularization method may be used without any particular limitation.Pharmaceutically acceptable salts

[0277] In some embodiments, the peptides and conjugates described herein exist as their pharmaceutically acceptable salts. In some embodiments, the methods disclosed herein include methods of treating diseases by102MEl\59069747.vlAttorney Docket No. 137521-10301 administering such pharmaceutically acceptable salts. In some embodiments, the methods disclosed herein include methods of treating diseases by administering such pharmaceutically acceptable salts as pharmaceutical compositions. As used herein, a "pharmaceutically acceptable salt” refers to any salt of a compound that is useful for therapeutic purposes of a subject.

[0278] In some embodiments, the peptides and conjugates described herein possess acidic or basic groups and therefore react with any of a number of inorganic or organic bases, and inorganic and organic acids, to form a pharmaceutically acceptable salt. In some embodiments, these salts are prepared in situ during the final isolation and purification of the compounds disclosed herein, or by separately reacting a purified compound in its free form with a suitable acid or base, and isolating the salt thus formed.

[0279] Examples of pharmaceutically acceptable salts include those salts prepared by reaction of the peptides and conjugates described herein with a mineral acid, organic acid, or inorganic base, such salts including acetate, acrylate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, bisulfite, bromide, butyrate, butyn-1 ,4-dioate, camphorate, camphorsulfonate, caproate, caprylate, chlorobenzoate, chloride, citrate, cyclopentanepropionate, decanoate, digluconate, dihydrogenphosphate, dinitrobenzoate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptanoate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hexyne-1,6-dioate, hydroxybenzoate, y-hydroxybutyrate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, iodide, isobutyrate, lactate, maleate, malonate, methanesulfonate, mandelate, metaphosphate, methanesulfonate, methoxybenzoate, methyl benzoate, monohydrogenphosphate, 1- napthalenesulfonate, 2-napthalenesulfonate, nicotinate, nitrate, palmoate, pectinate, persulfate, 3- phenylpropionate, phosphate, picrate, pivalate, propionate, pyrosulfate, pyrophosphate, propiolate, phthalate, phenylacetate, phenylbutyrate, propanesulfonate, salicylate, succinate, sulfate, sulfite, succinate, suberate, sebacate, sulfonate, tartrate, thiocyanate, tosylate, undeconate, and xylenesulfonate.

[0280] Further, the peptides and conjugates described herein can be prepared as pharmaceutically acceptable salts formed by reacting the free base form of the compound with a pharmaceutically acceptable inorganic or organic acid, including, but not limited to, inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, metaphosphoric acid, and the like; and organic acids such as acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, p-toluenesulfonic acid, tartaric acid, trifluoroacetic acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, arylsulfonic acid, methanesulfonic acid, ethanesulfonic acid, 1 ,2-ethanedisulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, 2-naphthalenesulfonic acid, 4-methylbicyclo-[2.2.2]oct-2-ene-1-carboxylic acid, glucoheptonic acid, 4, 4'-methylenebis-(3-hydroxy-2-ene-1 -carboxylic acid), 3-phenylpropionic acid, trimethylacetic acid, tertiary butylacetic acid, lauryl sulfuric acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, and muconic acid.

[0281] In some embodiments, those peptides and conjugates described herein which comprise a free acid group react with a suitable base, such as the hydroxide, carbonate, bicarbonate, or sulfate of a pharmaceutically acceptable metal cation, with ammonia, or with a pharmaceutically acceptable organic primary, secondary,103MEl\59069747.vlAttorney Docket No. 137521-10301 tertiary, or quaternary amine. Representative salts include the alkali or alkaline earth salts, like lithium, sodium, potassium, calcium, and magnesium, and aluminum salts, and the like. Illustrative examples of bases include sodium hydroxide, potassium hydroxide, choline hydroxide, sodium carbonate, N+(C1-4 alkyl)4, and the like.

[0282] Representative organic amines useful for the formation of base addition salts include ethylamine, diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, and the like. It should be understood that the peptides, conjugates, and radiopharmaceuticals described herein also include the quaternization of any basic nitrogen-containing groups they contain. In some embodiments, water or oil-soluble or dispersible products are obtained by such quaternization.Solvates

[0283] In some embodiments, the peptides and conjugates described herein exist as solvates. This disclosure provides for methods of treating diseases by administering such solvates. This disclosure further provides for methods of treating diseases by administering such solvates as pharmaceutical compositions.

[0284] Solvates contain either stoichiometric or non-stoichiometric amounts of a solvent, and, in some embodiments, are formed during the process of crystallization with pharmaceutically acceptable solvents such as water, ethanol, and the like. Hydrates are formed when the solvent is water, or alcoholates are formed when the solvent is alcohol. Solvates of the peptides, conjugates, and radiopharmaceuticals described herein can be conveniently prepared or formed during the processes described herein. In addition, the peptides and conjugates provided herein can exist in unsolvated as well as solvated forms. In general, the solvated forms are considered equivalent to the unsolvated forms for the purposes of the compounds and methods provided herein. Accordingly, one aspect of the present disclosure pertains to hydrates and solvates of peptides and conjugates and / or their pharmaceutical acceptable salts, as described herein, that can be isolated and characterized by methods known in the art, such as, thermogravimetric analysis (TGA), TGA-mass spectroscopy, TGA-Infrared spectroscopy, powder X-ray diffraction (PXRD), Karl Fisher titration, high resolution X-ray diffraction, and the like.Pharmaceutical Compositions

[0285] The peptides and conjugates described herein, including e.g., pharmaceutically acceptable salt or solvate thereof, can be administered per se as a pure chemical or as a component of a pharmaceutically acceptable formulation. In some embodiments, a peptide or conjugate described herein is combined with a pharmaceutically suitable or acceptable carrier selected on the basis of a chosen route of administration and standard pharmaceutical practice as described, for example, in Remington: The Science and Practice of Pharmacy (Gennaro, 21stEd. Mack Pub. Co., Easton, PA (2005)). Provided herein is a pharmaceutical composition comprising at least one peptide or conjugate described herein, or a stereoisomer, pharmaceutically acceptable salt, amide, ester, solvate, or N-oxide thereof, together with one or more pharmaceutically acceptable carriers. The carrier(s) (or excipient(s)) is acceptable or suitable if the carrier is compatible with the other ingredients of the composition and not deleterious to the recipient (i.e., the subject or patient) of the composition.

[0286] In one aspect, the disclosure provides a pharmaceutical composition comprising a herein described peptide or conjugate, or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable excipient or carrier.104MEl\59069747.vlAttorney Docket No. 137521-10301

[0287] The peptides or conjugates, and pharmaceutical compositions thereof of the current disclosure can be administered by any suitable means, including oral, topical (including buccal and sublingual), rectal, vaginal, transdermal, parenteral, subcutaneous, intraperitoneal, intrapulmonary, intradermal, intrathecal and epidural and intranasal, etc.III. METHODS OF TREATMENT

[0288] The present technology also relates to a method for treating a disease or disorder characterized by overexpression or decreased expression of GPC3 by administering the peptides or conjugates thereof of the present technology to a subject.

[0289] The present technology also relates to a use of the peptides or conjugates thereof of the present technology for the treatment of a disease or disorder characterized by overexpression or decreased expression of GPC3.

[0290] The present technology also relates to a use of the peptides or conjugates thereof for manufacturing a pharmaceutical composition for the treatment of a disease or disorder characterized by overexpression or decreased expression of GPC3.

[0291] The present technology also relates to the peptides or conjugates thereof of the present technology for use in a method for treating a disease or disorder characterized by overexpression or decreased expression of GPC3.

[0292] The present technology also relates to a kit for use in a method of diagnosing disease or disorder characterized by overexpression or decreased expression of GPC3 by determination of the expression level of GPC3.

[0293] The present technology also relates to a composition for use in a method of diagnosing disease or disorder characterized by overexpression or decreased expression of GPC3.

[0294] The present technology also relates to use of a peptides or conjugates thereof, or a salt thereof for use in a method of diagnosing disease or disorder characterized by overexpression or decreased expression of GPC3.

[0295] The present technology also provides methods of treating a disease or condition in a subject in need thereof. In some embodiments, the disease or disorder is characterized by overexpression or decreased expression of GPC3 in diseased tissue. In some embodiments, the disease or disorder is characterized by overexpression of GPC3 in diseased tissue. In some embodiments, the disease or disorder is characterized by decreased expression of GPC3 in diseased tissue. In some embodiments, the methods comprise administering a peptide described herein, a conjugate described herein, or a pharmaceutically acceptable salt or solvate thereof described herein, or a pharmaceutical composition comprising the same, to the subject in need thereof. In some embodiments, provided herein is a method of providing a therapeutic and / or prophylactic benefit to a subject in need thereof comprising administering a peptide, a conjugate, or a pharmaceutical composition described herein.

[0296] In some embodiments, the methods comprise administering to a subject a therapeutically effective amount of a peptide, a conjugate, or a pharmaceutically acceptable salt or solvate thereof. In some105MEl\59069747.vlAttorney Docket No. 137521-10301 embodiments, the peptide, conjugate or pharmaceutically acceptable salt or solvate thereof is administered in a pharmaceutical composition. In some embodiments, the subject has cancer. In some embodiments, the cancer is a solid tumor or hematological cancer.

[0297] In one aspect, provided herein is a method of treating cancer by administering a herein described peptide, conjugate or a pharmaceutically acceptable salt or solvate thereof to a subject in need thereof. According to a further aspect of the disclosure, there is provided a peptide, or a drug conjugate thereof as defined herein, for use in preventing, suppressing or treating a disease or disorder characterized by overexpression or decreased expression of GPC3 in diseased tissue. In some embodiments, the disease or disorder is characterized by overexpression of GPC3 in diseased tissue (such as a tumor). In one embodiment, the GPC3 is mammalian GPC3. In a further embodiment, the mammalian GPC3 is human GPC3.

[0298] In one aspect, provided herein is a method of preventing, suppressing or treating a disease or disorder characterized by overexpression or decreased expression of GPC3 in diseased tissue. In some embodiments, the disease or disorder is characterized by overexpression of GPC3 in diseased tissue (such as a tumor), which comprises administering to a patient in need thereof a peptide or a conjugate described herein. In some embodiments, the disease or disorder characterized by overexpression of GPC3 in diseased tissue is a cancer.

[0299] Non-limiting examples of cancers to be treated by the methods of the present disclosure can include hematological cancer or solid tumor, such as melanoma (e.g., metastatic malignant melanoma), renal cancer (e.g., clear cell carcinoma), prostate cancer (e.g., hormone refractory prostate adenocarcinoma), pancreatic adenocarcinoma, breast cancer, colon cancer, lung cancer (e.g., non-small cell lung cancer), esophageal cancer, squamous cell carcinoma of the head and neck, liver cancer, ovarian cancer, cervical cancer, thyroid cancer, glioblastoma, glioma, leukemia, lymphoma, and other neoplastic malignancies. In some embodiments, a subject or population of subjects to be treated with a pharmaceutical composition of the present disclosure have a solid tumor. In some embodiments, a solid tumor is a melanoma, renal cell carcinoma, lung cancer, bladder cancer, breast cancer, cervical cancer, colon cancer, gall bladder cancer, laryngeal cancer, liver cancer, thyroid cancer, stomach cancer, salivary gland cancer, prostate cancer, pancreatic cancer, or Merkel cell carcinoma. In some embodiments, a subject or population of subjects to be treated with a pharmaceutical composition of the present disclosure have a hematological cancer. In some embodiments, the subject has a hematological cancer such as Diffuse large B cell lymphoma ("DLBCL”), Hodgkin's lymphoma ("HL”), Non-Hodgkin's lymphoma ("NHL”), Follicular lymphoma ("FL”), acute myeloid leukemia ("AML”), or Multiple myeloma ("MM”). In some embodiments, a subject or population of subjects to be treated having the cancer selected from the group consisting of ovarian cancer, lung cancer, and melanoma. In some embodiments, the cancer is selected from hepatocellular carcinoma, squamous cell carcinoma of the lung, lung adenocarcinoma, germ cell tumors, hepatoblastoma, wilms tumor, malignant rhabdoid tumors, rhabdomyosarcoma, liposarcoma, thyroid cancers, pancreatic cancer, small bowel cancer, small cell neuroendocrine carcinoma (SONG), castration resistant prostatic adenocarcinoma, ovarian cancer, gastric cancer, esophageal carcinoma and malignant melanoma. In some embodiments, the castration resistant prostatic adenocarcinoma is hormonally treated. In some embodiments, the cancer ICs neuroendocrine prostate cancer (NEPC).106MEl\59069747.vlAttorney Docket No. 137521-10301

[0300] In some embodiments, provided herein are methods and compositions for treating a disease or condition. Exemplary disease or condition includes refractory or recurrent malignancies whose growth may be inhibited using the methods of treatment of the present disclosure. In some embodiments, the disease or condition is a cancer. In some embodiments, the cancer is breast cancer, head and neck squamous cell carcinoma, non-small cell lung cancer, hepatocellular cancer, bladder cancer, colorectal cancer, gastric adenocarcinoma, ovarian cancer, melanoma, or advanced cancer. In some embodiments, a cancer to be treated by the methods of treatment of the present disclosure is selected from the group consisting of carcinoma, squamous carcinoma, adenocarcinoma, sarcomata, endometrial cancer, breast cancer, ovarian cancer, cervical cancer, fallopian tube cancer, primary peritoneal cancer, colon cancer, colorectal cancer, squamous cell carcinoma of the anogenital region, melanoma, renal cell carcinoma, lung cancer, non-small cell lung cancer, squamous cell carcinoma of the lung, stomach cancer, bladder cancer, gall bladder cancer, liver cancer, thyroid cancer, laryngeal cancer, salivary gland cancer, esophageal cancer, head and neck cancer, glioblastoma, glioma, squamous cell carcinoma of the head and neck, prostate cancer, pancreatic cancer, mesothelioma, sarcoma, hematological cancer, leukemia, lymphoma, neuroma, and combinations thereof. In some embodiments, a cancer to be treated by the methods of the present disclosure include, for example, carcinoma, squamous carcinoma (for example, cervical canal, eyelid, tunica conjunctiva, vagina, lung, oral cavity, skin, urinary bladder, tongue, larynx, and gullet), and adenocarcinoma (for example, prostate, small intestine, endometrium, cervical canal, large intestine, lung, pancreas, gullet, rectum, uterus, stomach, mammary gland, and ovary). In some embodiments, the cancer is neuroendocrine prostate cancer (NEPC). In some embodiments, a cancer to be treated by the methods of the present disclosure further include sarcomata (for example, myogenic sarcoma), leukosis, neuroma, melanoma, and lymphoma. In some embodiments, a cancer to be treated by the methods of the present disclosure is breast cancer. In some embodiments, a cancer to be treated by the methods of treatment of the present disclosure is triple negative breast cancer (TNBC). In some embodiments, a cancer to be treated by the methods of treatment of the present disclosure is pancreatic cancer. In some embodiments, a cancer to be treated by the methods of the present disclosure is non-small cell lung cancer, ovarian cancer, or bladder cancer. In some embodiments, a cancer to be treated by the methods of the present disclosure is non-small cell lung cancer. In some embodiments, a cancer to be treated by the methods of the present disclosure is bladder cancer. In some embodiments, a cancer to be treated by the methods of the present disclosure is ovarian cancer. In some embodiments, a cancer to be treated by the methods of the present disclosure is hepatocellular carcinoma, squamous cell carcinoma of the lung, lung adenocarcinoma, germ cell tumors, hepatoblastoma, wilms tumor, malignant rhabdoid tumors, rhabdomyosarcoma, liposarcoma, thyroid cancers, pancreatic cancer, small bowel cancer, small cell neuroendocrine carcinoma (SCNC), hormonally treated, castration resistant prostatic adenocarcinoma, ovarian cancer, gastric cancer, esophageal carcinoma, or malignant melanoma.

[0301] Further examples of cancers (and their benign counterparts) which may be treated include, but are not limited to tumors of epithelial origin (adenomas and carcinomas of various types including adenocarcinomas, squamous carcinomas, transitional cell carcinomas and other carcinomas) such as carcinomas of the bladder107MEl\59069747.vlAttorney Docket No. 137521-10301 and urinary tract, breast, gastrointestinal tract (including the esophagus, stomach (gastric), small intestine, colon, rectum and anus), liver (hepatocellular carcinoma), gall bladder and biliary system, exocrine pancreas, kidney, lung (for example adenocarcinomas, small cell lung carcinomas, non-small cell lung carcinomas, bronchioalveolar carcinomas and mesotheliomas), head and neck (for example cancers of the tongue, buccal cavity, larynx, pharynx, nasopharynx, tonsil, salivary glands, nasal cavity and paranasal sinuses), ovary, fallopian tubes, peritoneum, vagina, vulva, penis, cervix, myometrium, endometrium, thyroid (for example thyroid follicular carcinoma), adrenal, prostate, skin and adnexae (for example melanoma, basal cell carcinoma, squamous cell carcinoma, keratoacanthoma, dysplastic naevus); hematological malignancies ( / .e. leukemias, lymphomas) and premalignant hematological disorders and disorders of borderline malignancy including hematological malignancies and related conditions of lymphoid lineage (for example acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia, B-cell lymphomas such as diffuse large B-cell lymphoma, follicular lymphoma, Burkitt's lymphoma, mantle cell lymphoma, T-cell lymphomas and leukemias, natural killer cell lymphomas, Hodgkin's lymphomas, hairy cell leukemia, monoclonal gammopathy of uncertain significance, plasmacytoma, multiple myeloma, and post-transplant lymphoproliferative disorders), and hematological malignancies and related conditions of myeloid lineage (for example acute myelogenous leukemia, chronic myelogenous leukemia, chronic myelomonocytic leukemia, hypereosinophilic syndrome, myeloproliferative disorders such as polycythemia vera, essential thrombocythemia and primary myelofibrosis, myeloproliferative syndrome, myelodysplastic syndrome, and promyelocytic leukemia); tumors of mesenchymal origin, for example sarcomas of soft tissue, bone or cartilage such as osteosarcomas, fibrosarcoma's, chondrosarcomas, rhabdomyosarcomas, leiomyosarcomas, liposarcomas, angiosarcomas, Kaposi's sarcoma, Ewing's sarcoma, synovial sarcomas, epithelioid sarcomas, gastrointestinal stromal tumors, benign and malignant histiocytomas, and dermatofibrosarcoma protuberans; tumors of the central or peripheral nervous system (for example astrocytoma's, gliomas and glioblastomas, meningiomas, ependymomas, pineal tumors and schwannomas); endocrine tumors (for example pituitary tumors, adrenal tumors, islet cell tumors, parathyroid tumors, carcinoid tumors and medullary carcinoma of the thyroid); ocular and adnexal tumors (for example retinoblastoma); germ cell and trophoblastic tumors (for example teratomas, seminomas, dysgerminomas, hydatidiform moles and choriocarcinomas); and pediatric and embryonal tumors (for example medulloblastoma, neuroblastoma, Wilms tumor, and primitive neuroectodermal tumors); or syndromes, congenital or otherwise, which leave the patient susceptible to malignancy (for example Xeroderma Pigmentosum).

[0302] In some embodiments, the cancer is selected from glioblastoma, prostate cancer, lung cancer, breast cancer, gastric cancer, ovarian cancer, gladder cancer, colon cancer, esophageal cancer, multiple myeloma and fibrosarcoma. In some embodiments, the cancer is selected from: breast cancer, lung cancer, gastric cancer, pancreatic cancer, prostate cancer, liver cancer, glioblastoma and angiogenesis. In some embodiments, the cancer is selected from: prostate cancer, lung cancer (such as non-small cell lung carcinomas (NSCLC)), breast cancer (such as triple negative breast cancer), gastric cancer, ovarian cancer, esophageal cancer, multiple myeloma and fibrosarcoma. In some embodiments, the cancer is prostate cancer. In some embodiments, the conjugate is useful for preventing, suppressing or treating solid tumors such as fibrosarcoma's and breast, and108MEl\59069747.vlAttorney Docket No. 137521-10301 non-small cell lung carcinomas. In some embodiments, the cancer is selected from lung cancer, such as nonsmall cell lung carcinomas (NSCLC). In some embodiments, the cancer is breast cancer. In some embodiments, the breast cancer is triple negative breast cancer. In some embodiments, the breast cancer is Herceptin resistant breast cancer. In some embodiments, the subject has failed to respond to Herceptin. In some embodiments, the cancer is gastric cancer. In some embodiments, the cancer is ovarian cancer. In some embodiments, the cancer is esophageal cancer. In some embodiments, the cancer is multiple myeloma. In some embodiments, the cancer is fibrosarcoma.

[0303] In some embodiments, provided herein are methods for killing a cell comprising contacting the cell with a peptide, a conjugate, or a pharmaceutically acceptable salt or solvate thereof. In some embodiments, the cell expresses GPC3. In some embodiments, the cell over-expresses GPC3. In some embodiments, the conjugate or pharmaceutically acceptable salt or solvate thereof binds to a structure on the cell, wherein the structure is an GPC3.

[0304] After contacting a cell, the described peptide or conjugate can be internalized by the cell. The internalization can be mediated by cell receptors, cell membrane endocytosis, etc. In some embodiments, the described peptide or conjugate is internalized by a cell through GPC3. In some embodiments, rapid internalization rate into cancer cells accompanied by a slow externalization rate can offer therapeutic benefit.

[0305] In one aspect, the disclosed peptide, conjugate or a pharmaceutically acceptable salt or solvate thereof is configured to treat cancer by ablating tumor cells. In some embodiments, the peptide, conjugate or a pharmaceutically acceptable salt or solvate thereof does not modulate the biology of the tumor cell and / or the surrounding stroma. In some embodiments, the conjugate or a pharmaceutically acceptable salt or solvate thereof does not modulate immune cells. In some embodiments, the ablating of tumor cells can lead to a downstream immunological cascade.

[0306] In addition to the methods of treatment described above, the conjugates and compositions described herein can be used to image, and / or as part of a treatment for diseases.

[0307] In one aspect, described herein is a method of treatments that comprises administering a first conjugate and a second conjugate. The first conjugate can be used as companion diagnostics and the second conjugate can be used for therapeutics. In some embodiments, the first conjugate and the second conjugate have the same structure except for the non-radionuclide moiety.

[0308] In one aspect, described herein is a method of diagnosing or imaging a cancer in a subject in need thereof, comprising administering to the subject a peptide, a conjugate or a pharmaceutical composition described herein.

[0309] In some embodiments, the subject is 1 to 100 years old. In some embodiments, the subject is 5 to 10, 5 to 15, 5 to 18, 5 to 25, 5 to 35, 5 to 45, 5 to 55, 5 to 65, 5 to 75, 10 to 15, 10 to 18, 10 to 25, 10 to 35, 10 to 45,10 to 55, 10 to 65, 10 to 75, 15 to 18, 15 to 25, 15 to 35, 15 to 45, 15 to 55, 15 to 65, 15 to 75, 18 to 25, 18 to 35,18 to 45, 18 to 55, 18 to 65, 18 to 75, 25 to 35, 25 to 45, 25 to 55, 25 to 65, 25 to 75, 35 to 45, 35 to 55, 35 to 65,35 to 75, 45 to 55, 45 to 65, 45 to 75, 55 to 65, 55 to 75, or 65 to 75 years old. In some embodiments, the subject is at least 5, 10, 15, 18, 25, 35, 45, 55, or 65 years old. In some embodiments, the subject is at most 10, 15, 18,109MEl\59069747.vlAttorney Docket No. 137521-1030125, 35, 45, 55, 65, or 75 years old.

[0310] The amount of peptide or conjugate described herein or a pharmaceutically acceptable salt or stereoisomer thereof and / or pharmaceutical compositions administered can be sufficient to deliver a therapeutically effective dose of the particular subject. In some embodiments, conjugate dosages can be between about 0.1 pg and about 50 mg per kilogram of body weight. In some embodiments, the dose of the conjugate or a pharmaceutically acceptable salt or stereoisomer thereof described herein for the described methods is about 0.001 mg to about 1000 mg per dose for the subject being treated.

[0311] Although the present disclosure and its advantages have been described in detail, it should be understood that various changes, substitutions and alterations can be made herein without departing from the spirit and scope of the disclosure as defined in the appended claims.

[0312] The present disclosure is further illustrated in the following Examples which are given for illustration purposes only and are not intended to limit the disclosure in any way.EXAMPLESAbbreviations:

[0313] Unless otherwise stated in the present specification, the following abbreviations are used according to the following meanings:Alloc allyloxycarbonyl aq. aqueousBiotin-OSu biotin N-hydroxysuccinimide ester (CAS 35013-72-0) Boc fert-butyloxycarbonylCIAcOH chloroacetic acidCIAcOSu N-succinimidyl 2-chloroacetate (CAS 27243-15-8)DCM dichloromethane (CAS 75-09-2)DIG N,N' -diisopropylcarbodiimide (CAS 693-13-0)DIPEA, DIEA N,N-diisopropylethylamine (CAS 7087-68-5)DMEM Dulbecco's Modified Eagle MediumDMF N,N-dimethylformamide (CAS 68-12-2)DODT 2,2'-(ethylenedioxy)diethanethiol (CAS 14970-87-7)EDCI-HCI / V-(3-dimethylaminopropyl)-A / '-ethylcarbodiimide hydrochloride(CAS 25952-53-8)EDTA ethylenediaminetetraacetic acidEMEM Eagle's minimal essential medium eq equivalentEt ethylEt3N, TEA triethylamine (CAS 121-44-8)FBS fetal bovine serum110MEl\59069747.vlAttorney Docket No. 137521-10301Fmoc 9-fluorenylmethoxycarbonyl hr hour HATU 1-[bis(dimethylamino)methylene]-1 / 7-1J2J3-triazolo[4,5-b]pyridinium-3-oxide hexafluorophosphate (CAS 148893-10-1)HOSu N-hydroxysuccinimide (CAS 6066-82-6) iPrOH, IPA isopropanol M molar min minutes NHS N-hydroxysuccinimide (CAS 6066-82-6) NMP W-methy Ipy rrolidone (CAS 872-50-4) Pd(PPh3)4tetrakis(triphenylphosphine)palladium(0) (CAS 14221-01-3) PG protecting group Ph phenyl RP reverse phase rpm rotations per minute rt room temperature SAPE Streptavidin, R-Phycoerythrin conjugate SPPS solid phase peptide synthesis Su succinimidyl SulfoCy5- sulfo Cyanine5 (CAS 2791287-13-1) SulfoCy5-OSu, (CAS 146368-14-1) SulfoCy5-NHS see SulfoCy5-OSu tert tertiary TFA trifluoroacetic acid (CAS 76-05-1) TIS triisopropylsilane (CAS 6485-79-6) tr retention time Trt trityl. PyAOP (7-azabenzotriazol-1-yloxy)trispyrrolidinophosphonium hexafluorophosphate(CAS 156311-83-0)MeCN acetonitrile AcOH acetic acid Et2O diethyl ether AA amino acid Me methyl HFIP 1 ,1 , 1 ,3,3,3-hexafluoro-2-propanol (CAS 920-66-1) PhSIH3phenylsilane (CAS 694-53-1) tBu tertiary butyl111MEl\59069747.vlAttorney Docket No. 137521-10301DMSO dimethylsulfoxideMpe 3-Methyl-pent-3-yl

[0314] The present technology is described in detail below based on examples, but the present technology is not limited to these examples. A person having ordinary skill in the art is capable of easily adding modifications and changes to the present technology based on the description of the present specification, and these are included in the technical scope of the present technology.

[0315] Abbreviation for non-natural amino acids used in this invention will be described here.

[0316] Analytical methods, materials, and instruments.

[0317] Unless otherwise noted, reagents and solvents were used as received from commercial suppliers. If not otherwise specified, purity and low-resolution mass spectral data were measured using a Shimadzu Nexera XR system coupled to a LCMS-2020 MS (ESI). Methods are specified below.Method A-1 (HPLC-MS): Phenomenex Kinetex EVO C182.6um, 2.1 ID x 150mm, 100A (with a guard cartridge 2.1mmlD); 60 °C; 0.5 mL / min; (A) H2O + 0.025% TFA / (B) MeCN + 0.025% TFA; gradient: from 5 to 45% B in 7.2 min. Detection: electrospray mass spectra (+) and PDA-UV at UV of 225 nm.Method A-2 (HPLC-MS): Phenomenex Kinetex EVO C182.6um, 2.1 ID x 150mm, 100A (with a guard cartridge 2.1mmlD); 60 °C; 0.5 mL / min; (A) H2O + 0.025% TFA / (B) MeCN + 0.025% TFA; gradient: from 20 to 60% B in 7.2 min. Detection: electrospray mass spectra (+) and PDA-UV at UV of 225 nm.

[0318] General preparative HPLC purification method.

[0319] Crude containing the desired polypeptide were purified by preparative RP-HPLC using DAD-UV chromatogram at UV 220 nm and electrospray mass spectra (+) for the detection and employing commercially available RP-HPLC columns such as Waters XBridge C18, Waters XSelect C18, Phenomenex Kinetex EVO C18, and Interchim US5C18HQ-100 / 300. Otherwise specified, (A) H2O + 0.1% TFA and (B) MeCN + 0.1% with varying gradients were employed as eluents. Product-containing fractions were collected and lyophilized to obtain the purified product.

[0320] General synthetic aspects.

[0321] Macrocyclic peptides type-A can be synthesized as outlined in Scheme-1 .Scheme-1112MEl\59069747.vlAttorney Docket No. 137521-10301

[0322] Step-A1 : Peptide elongation on resin.

[0323] Th peptide elongations in this invention were performed in automated peptide synthesizers employing conditions as described in Method-S1 , Method-S2, and Method-S3 in accordance with the manufacturer's instruction. In addition, the peptide coupling with certain Fmoc-AAs was performed by the synthesizer or by manually as described in the table S1-2. In detail, Fmoc-Sieber amide resin was suspended in the solvent (e.g., DMF or DCM) and then loaded onto the peptide synthesizer. After Fmoc removal of the resin, Fmoc-AA coupling and subsequent Fmoc removal as one cycle was performed employing conditions suitable for each Fmoc-AA as described below. The peptide elongation on resin was continued until the desired linear polypeptide sequence 1a was obtained. Below described the AA coupling and Fmoc deprotection conditions.

[0324] Method S1 employing Liberty BLUE HT 12TM (GEM. Inc.).

[0325] General AA coupling conditionsTable S1-1Fmoc-AA Reagents Coupling conditions50°C, 1-2x 15-20 min; or 40°C,Fmoc-MeC(Trt)-OH1 -2x 30 min; or 25°C, 1 -2x 30 minDIG / Oxyma Pure® in DMF orFmoc-H(Trt)-OH 50°C, 2x 30 minNMP90°C, 1-2x 3-10 minOther than listed above or 75°C, 1-2x 10-30 min

[0326] Fmoc deprotection on resin.Table S1 -2Fmoc-AA on peptide Reagents Deprotection conditionsFmoc-AA coupled with pyrrolidine / DMF (1 / 9)25°C, 1 -2x 1-5 min or 25°C, 5 min then 25°C, 15 minN-Me-AA or piperidine / DMF (1 / 4)Other than listed above 90°C, 1 min or 75°C, 3 min or 50°C, 1 .5 min113MEl\59069747.vlAttorney Docket No. 137521-10301

[0327] Method S2 employing Liberty Prime (CEM. Inc.).

[0328] General AA coupling on resin.Table S2-1Fmoc-AA Reagents Coupling conditionsFmoc-MeC(Trt)-OH 50°C, 1 -2x 15-20 min or 40°C, 1 -2x 30 minDIG / Oxyma Pure® 105°C, 1 -2x 1-2 min; or 90°C, 1 -2x 3-10 min;Other than listed above in DMF or NMP or 75°C, 2x 20-30 min

[0329] General method for Fmoc deprotectionTable S2-2Fmoc-AA on peptide Reagents Deprotection conditionsFmoc-AA coupled with pyrrolidine / DMF (1 / 9) 25°C, 1 -2x 1-5 minN-Me-AAOther than listed above 83 mM Oxyma Pure® in pyrrolidine / DMF (4 / 96) 110°C, 1-1.5 min

[0330] Method S3 employing Gyro I (Biotage. Inc.).

[0331] General AA coupling

[0332] Table S3-1Fmoc-AA Reagents Coupling conditionsFmoc-MeC(Trt)-OH 25°C, 40 minHATU / DIEA in DMF or NMPOther than listed above 75°C, 2x 20-30 min

[0333] General Fmoc deprotection

[0334] Table S3-2Fmoc-AA on peptide Reagents Deprotection conditionsAll AAs piperidine / DMF (1 / 4) 25°C, 5 min then 25°C, 15 min

[0335] Method S4: General peptide coupling without automated synthesizers.

[0336] The peptide coupling with the AAs listed below were performed manually without microwave employing conditions in Table S4. (Note: the AAs listed below can be introduced by the peptide synthesizer employing above methods as well).

[0337] Table S4Fmoc-AA Reagents Coupling conditionsFmoc-MeNal27N-OH HATU / DIEA (3-4.2 / 3-4 / 6-12) in NMP 25°C, 1x 30-60 minFmoc-SR3Et-OH HATU / DIEA (3 / 3 / 6) in DMF 25°C, 1x 40 minFmoc-MeA4paa(tBu)-OH DIG / Oxyma Pure® (3 / 4 / 8) in DMF 75°C, 1x 30 min

[0338] Step A2: Chloroacetylation.

[0339] The obtained polypeptide 1a on resin was transferred in a syringe with a flit. The resin was shaken in a114MEl\59069747.vlAttorney Docket No. 137521-10301 reagent mixture (see below) at room temperature for 0.5-2h. The solution was then drained through the frit. The resin was washed successively a few times with DMF, DCM, and Et2O to afford the linear peptides 1b on resin.Reagent mixture: resin / CIAcOH in DMF(0.087-0.25M) / HATU in DMF(0.087-0.25M) / DIPEA in DMF (0.17-0.5M)= 1 eq / 4.2-5 eq / 4-5 eq / 4.2-10 eq.

[0340] Step A3: Cleavage of polypeptides from the resin along with the global deprotection of side chain protecting groups (PGs).

[0341] Polypeptide 1b on resin was shaken at rt for 5-90 min in the cleavage cocktail (TFA / TIS / DODT / H2O = 92.5 / 2.5 / 2.5 / 2.5). The resin was filtered and rinsed with the cocktail used. Combined filtrates were poured into cold diethyl ether. The resulting suspension was centrifuged (9000 rpm, 1 min at 0°C), and then the supernatant was decanted out. The precipitate was suspended in cold ether, vortexed briefly, and then centrifuged. This process was repeated a few times when appropriate and necessary. The crude containing polypeptide 1c was dried under reduced pressure.

[0342] Step A4: Peptide cyclization and purification.

[0343] The crude containing polypeptide 1c was dissolved in MeCN / H2O (1 / 1). To the solution was added TEA (5-20 eq), and then the mixture was stirred for 1 hr to 18 hrs until completion of the reaction. The resulting mixture was rendered acidic by AcOH, and then reaction mixture was concentrated by Genevac EZ-2 Elite or HT-12. The resulting residue was purified by RP-HPLC to afford Macrocyclic peptide type-A.

[0344]

[0345] Macrocyclic peptides type-B in the present invention can be synthesized as outlined in Scheme 2. Note that the following methods can afford desired sidechain branched polypeptide at any AA position by appropriate order of Fmoc-AA used.

[0346] Scheme 2115MEl\59069747.vlAttorney Docket No. 137521-10301

[0347] Step B1 : Polypeptides type 2a on-resin were synthesized in the fashion analogous to Step-A1 in Scheme-1 but employing appropriately protected AA building blocks such as Alloc-K(Fmoc)-OH. Subsequently, treatment of 2a with one of the Fmoc deprotection conditions above, followed by the coupling with the Boc- protected AAs employing the described condition in Table S5 afforded 2b.

[0348] Table S5Boc-protected AAs (the side chain protected Reagents Coupling conditions properly116MEl\59069747.vlAttorney Docket No. 137521-10301Boc-df-OH HATU / DIEA (3 / 3 / 6) in DMF 25°C, 30 min (coupling manually)Boc-H(Trt)-OH DIG / Oxyma Pure® (4.2 / 8 / 4) in DMF 50°C, 2x 30 min (Liberty Blue)Boc-de(tBu)-OH HATU / DIEA (3 / 3 / 6) in DMF 75°C, 2x 30 min (Liberty Blue)Step B3: Deprotection of the Alloc protecting groupPolypeptides 2b on resin were swollen in DOM. The resin was shaken with Pd(PPh3)4 (0.2-0.25 eq) and PhSi H3 (10-15 eq) at room temperature for 1 h. The resin was washed with DMF. Then the resin was washed by DOM followed by DMF to afford polypeptides 2c on resin.

[0349] Step B4, -B5, and -B6:Subsequently, polypeptide 2c on resin were transformed to Macrocyclic peptides type-B by similar fashion analogous to the preparation in Scheme-1.

[0350] Macrocyclic peptides type-C in the present invention can be synthesized as outlined in Scheme-3.

[0351] Scheme-3117MEl\59069747.vlAttorney Docket No. 137521-10301118MEl\59069747.vlAttorney Docket No. 137521-10301

[0352] Step C1, C2, and C3:Polypeptides 3c were synthesized by methods analogously to the preparation above employing appropriate Fmoc-AA building blocks.

[0353] Step C4: elongation of the side chain with Boc-PEG12c-OH.

[0354] Polypeptides 3c on resin were suspended in DMF. The resin was shaken with Boc-PEG12c-OH (4.2- 5.3 eq) / HATU (4-5 eq) / DIEA (8-10 eq) at room temperature for 30 min; or Boc-PEG12c-OH (4.2-5.3 eq) / PyAOP (4-5 eq) / DIEA (8-10 eq) at 40°C for 30 min. The resin was washed with DMF. Then the resin was washed by DOM followed by DMF to provide polypeptides 3d on resin.

[0355] Step C5 and C6:Polypeptides 3f were synthesized from polypeptides 3d on resin in the fashion analogous to Scheme-1.

[0356] Step C7: Coupling reaction with Biotin-OSu

[0357] To the polypeptides 3f in DMSO were added Biotin-OSu (approx. 1 eq) and DIEA (3- 5 eq). The mixture was shaken at room temperature for 0.2-1 h. The reaction mixture was directly purified by preparative RP-HPLC to afford Macrocyclic peptides type-C.

[0358] Macrocyclic peptide-conjugates type-D, when one of AA sidechains contains a nucleophilic amine, in the present invention can be synthesized outlined in scheme-4.

[0359] Scheme-4119MEl\59069747.vlAttorney Docket No. 137521-10301Cleavage Deprotection CyclizationRc= H or Me Step D2

[0360] Polypeptides 4a on resin were synthesized by methods described above. Subsequently, the cleavage from the resin along with the global deprotection, followed by the cyclization and then RP-HPLC purification were performed similarly by the methods described above to afford polypeptide 4b. Finally, treatment of 4b with sulfoCy5-NHS, followed by the RP-HPLC purification afforded Macrocyclic peptide-conjugates type-D.

[0361] Alternatively, the crude containing 4b was treated with sulfoCy5-NHS, followed by RP-HPLC afforded Macrocyclic peptide-conjugates type-D.

[0362] Macrocyclic peptide-conjugates type-E can be synthesized via appropriate AA sequencing as shown in Scheme-5.

[0363] Scheme-5120MEl\59069747.vlAttorney Docket No. 137521-10301Synthesis cycle side chain SPPS then chloroacetylationStep E1Cleavage 5aDeprotectionCyclization

[0364] Polypeptides 5b were synthesized by methods described above via polypeptides 5a on resin. After the purification of 5b, treatment with sulfoCy5-NHS, followed by the RP-HPLC purification afforded Macrocyclic peptide-conjugates type-E.

[0365] Macrocyclic peptide type-F can be synthesized as outlined in Scheme-6.

[0366] Scheme-6 (SEQ ID NOs: 626-632, respectively, in order of appearance)121MEl\59069747.vlAttorney Docket No. 137521-10301122MEl\59069747.vlAttorney Docket No. 137521-103016f123MEl\59069747.vlAttorney Docket No. 137521-10301

[0367] Polypeptides type 6b via 6a on-resin were synthesized in the fashion analogous to Step-A1 inScheme-1 but employing appropriately protected AA building blocks such as Alloc-K(Fmoc)-OH. Subsequently, Alloc deprotection and the chloroacetiylation described in Step-B3 and Step-A2 respectively can afford linear polypeptide type 6d on resin. Cleavage from the resin along with the removal of the acid labile protecting groups by methods described in Step-A3 can afford 6e, which was then treated for the macrocyclization by methods described in such as Step-A4 can afford 6f. Removal of the Fmoc group of 6f can be achieved by treatment with suitable bases such as pyrrolidine in solvent such as DMF to afford Macrocyclic peptide type 6 (SEQ ID NO: 42).

[0368] Example-1 A:

[0369] The desired polypeptide was synthesized as outlined in Scheme-1 . The linear polypeptide was synthesized on Fmoc-Sieber amide resin (0.25 mmol) using Fmoc-AAs listed in below table. Method S-1 was employed for the peptide elongation. The obtained peptide on the resin was subjected to the chloroacetylation described in Step-A2 (CIAcOH (1.25 mmol) / HATU / DIEA (2.5 mmol) in DMF (12.5 mL), 30 min), followed by treating with the cleavage cocktail (12 mL) for 5 min as outlined in Step-A3 to furnish the linear peptide. The crude containing the linear peptide (0.25 mmol as theoretical based on the resin used) was dissolved in CH3CN / H2O (100 mL), and then added EtaN (2.5 mmol, 348 pL), and then shaken for at room temperature for 18 hrs, and then concentrated as outlined in Step-A4. The resulting residue was purified by preparative RP-HPLC.124MEl\59069747.vlAttorney Docket No. 137521-10301Pure fractions were combined and lyophilized to afford the title macrocyclic peptide (SEQ ID NO: 45). HPLC-MS(Method A-1): 4.67 min, ESI-MS m / z: 890.7 [M+2H]2+.

[0370] Table S6 order of Fmoc-AA used in AA position in the cyclic Fmoc-AA the SPPS peptide1 X12 Fmoc-MeC(Trt)-OH2 Xu Fmoc-3Py6Ph-OH3 X10 Fmoc-G-OH4 X9Fmoc-MeNal27N-OH5 X8Fmoc-Y(tBu)-OH6 X7Fmoc-I-OH7 X6Fmoc-TMe-OH8 X5Fmoc-F4COO(tBu)-OH9 X4Fmoc-Me(Trt)-OH10 x3Fmoc-D(Mpe)-OH11 x2Fmoc-MeHseMe-OH12 Xi Fmoc-MeK(Boc)-OH

[0371] Example-1 B: Synthesis of the conjugated form consisting of the GPC3 binding peptide in this disclosure and biotin as a payload.(SEQ ID NO: 633)

[0372] The desired polypeptide was synthesized as outlined in Scheme-3 on Fmoc-Sieber amide resin (0.100 mmol). Elongation of the linear peptide was achieved by Method S-1 employing Fmoc-AAs listed in Table E1 B below. The obtained peptide on the resin was treated with the reagent mixture (4.5 mL) and performed the chloroacetylation as described in Step-C2 at room temperature for 30 min. Subsequently, the resin was shaken with Pd(PPhs)4 (23.1 mg, 0.2 eq) and PhSIFL (123 pL, 10 eq) in DCM (5 mL) at room temperature for 1 hr. The obtained peptide on the resin was shaken with Boc-PEG12c-OH (0.53 mmol) in DMF (2.5 mL) / PyAOP (0.50125MEl\59069747.vlAttorney Docket No. 137521-10301 mmol) and DIEA (1.0 mmol) in DMF (2 mL) at 40 °C for 30 min as outlined in step-C3. The polypeptide on the resin was treated with the cleavage cocktail (4.5 mL) for 60 min as outlined in step-C4 to furnish the linear peptide, which was dissolved in MeCN / FW (20mL). To the mixture was added Et3N (279 pL), and then the mixture was shaken for at room temperature for 15 hrs, and then concentrated as outlined in Step-C5. The reaction mixture was concentrated in Genevac EZ-2 Elite, and then the resulting residue was dissolved in DMSO (4 mL). To the solution were added Biotin-OSu (34.1 mg, 0.1 mmol) and DIEA (52.3 pL). The mixture was shaken at room temperature for 1 hr as outlined in Step-06. To the mixture was added AcOH. The mixture was directly purified by preparative RP-HPLC. Pure fractions were combined and lyophilized to afford the title macrocyclic peptide (Conjugate ID NO.: PLB-1). HPLC-MS (Method A-2): 4.13 min, ESI-MS m / z: 868.0 [M+3H]3+.Table E1 B

[0373] Example-10: The following conjugates were prepared analogously to Example-1 B.Table E1C126MEl\59069747.vlAttorney Docket No. 137521-10301

[0374] Example-1 D: Synthesis of the conjugated form consisting of the GPC3 binding peptide in this disclosure and biotin as a payload.127MEl\59069747.vlAttorney Docket No. 137521-10301(SEQ ID NO: 634)

[0375] The desired polypeptide was synthesized as outlined in Scheme-1 on Fmoc-Sieber amide resin (0.050 mmol). Elongation of the linear peptide except for X9 was achieved by Method S-1 employing Fmoc-AAs listed in Table E1D below. Coupling reaction of Fmoc-MeNal27N-OH was performed manually as described in Method S4. The obtained peptide on the resin was treated with the reagent mixture (6 mL) and performed the chloroacetylation as described in Step-A2 at room temperature for 30 min. The polypeptide on the resin was treated with the cleavage cocktail (3 mL) for 30 min as outlined in Step-A3 to furnish the linear peptide, which was dissolved in MeCN / FW (10 mL). To the mixture was added EtsN (69.7 pL), and then the mixture was shaken for at room temperature for 3 hrs, and then concentrated as outlined in Step-A4. The resulting residue was purified by preparative RP-HPLC. Pure fractions were combined and lyophilized to afford the title macrocyclic peptide (Conjugate ID No.: PLB-60). HPLC-MS (Method A-2): 3.75 min, ESI-MS m / z: 914.3 [M+3H]3+.Table E1D

[0376] Example-1 E: Synthesis of the conjugated form consisting of the GPC3 binding peptide in this disclosure and sulfoCy5 as a payload.128MEl\59069747.vlAttorney Docket No. 137521-10301

[0377] The desired polypeptide was synthesized as outlined in Scheme-4. To a solution of the GPC binding peptide in Example-1A (10.0 mg, 4.71 pimol) in DMF (150 piL) at room temperature were added triethylamine (6.6 piL, 47.1 pimol) and SulfoCy5-NHS (4.6 mg, 6.13 pimol). The mixture was stirred at room temperature for 18 hrs as outlined in step-D4. The resulting mixture was purified by preparative RP-HPLC. Pure fractions were combined and lyophilized to afford the title macrocyclic peptide (Conjugate ID No.: PLD-45). HPLC-MS (Method A-2): 3.80 min, ESI-MS m / z: 1209.6 [M+2H]2+.

[0378] Example-1 F: The following compounds were prepared by analogous method described above.Table E1F129MEl\59069747.vlAttorney Docket No. 137521-10301

[0379] Example-1 G: The following compounds were prepared by analogous method described above.

[0380] The desired macrocyclic peptide was synthesized as outlined in Scheme-6. Starting with Fmoc-Sieber amide resin (2x0.250 mmol), elongation of the linear peptide except for X9 was achieved by Method S-1 employing Fmoc-AAs listed in Table E1 G below. Coupling reaction of Fmoc-MeNal27N-OH for X9 was performed manually as described in Method S4. The obtained peptide on the resin was treated with Fmoc-datb- OH (0.75 mmol) / HATU (0.75 mmol) / DIEA (1.5 mmol) in DMF (10mL) at room temperature for 30 min and then washed with DMF. Subsequently, the resin was shaken with Pd(PPh3)4 (57.8 mg, 0.2 eq) and PhSi H3 (308 pL, 10 eq) in DCM / AcOH (99 / 1 , 5 mL) at room temperature for 1 hr, and then washed successively with DCM and DMF. The resin was treated with the reagent mixture (CIAcOH (1.25 mmol) / HATU (1.25 mmol) / DIEA (2.5 mmol) in DMF (10 mL). The whole mixture was shaken at room temperature for 30 min. The polypeptide on resin was successively washed with DMF, DCM, and then Et20. The polypeptide on resin was dried under reducing pressure. The polypeptide on the resin was treated with the cleavage cocktail (10 mL) for 90 min as outlined in Step-A3 to furnish the linear peptide, which was dissolved in DMSO / MeCN / FW (1 / 1 / 1 , 100 mL). The mixture was split into four portions, and each mixture was then added TEA in each mixture (4x 174 L for the whole crude). The mixture was shaken at room temperature for 2 hrs, and then concentrated in Genevac EZ-2 Elite. The resulting residue was dissolved in DMSO (15 mL). To the solution was added pyrrolidine (310 L), and then the solution was shaken at room temperature for 1 hr. The reaction mixture was rendered acidic by AcOH, and then directly purified by preparative RP-HPLC. Pure fractions were combined and lyophilized to afford the title130MEl\59069747.vlAttorney Docket No. 137521-10301 macrocyclic peptide (SEQ ID NO: 42). HPLC-MS (Method A-2): 3.67 min, ESI-MS m / z: 654.4 [M+3H]3+.Table E1G

[0381] Example-1 H: The following compounds were prepared by analogous method described above.

[0382] The desired macrocyclic peptide was synthesized as outlined in Scheme-2 on Fmoc-Sieber amide resin (0.50 mmol). Elongation of the linear peptide except for X9 was achieved by Method S-1 employing Fmoc- AAs listed in Table E1 H below. Coupling reaction of Fmoc-MeNal27N-OH was performed manually as described in Method S4. The obtained peptide on the resin was treated with Boc-de(tBu)-OH (2.1 mmol) in DMF (10 mL) / DIG (4.0 mmol) in DMF (4 mL) / Oxyma pure® (2.0 mmol) in DMF (4 mL) at 75 °C for 30 min as outlined in Step- 62. Subsequently, the resin was shaken in a mixture of Pd(PPh3)4 (115.6 mg, 0.2 eq) and PhSIFL (616 pL, 10 eq) in DOM (25 mL) at room temperature for 1 hr as outlined in Step-B3. The resin was treated with the reagent mixture (25 mL) and performed the chloroacetylation as described in Step-B4 at room temperature for 30 min. The polypeptide on the resin was treated with the cleavage cocktail (30 mL) for 10 min as outlined in Step-B5 to furnish the linear peptide, which was dissolved in MeCN / FW (100 mL). To the mixture was added TEA (697 L) and then the mixture was shaken for at room temperature for 14 hrs, and then concentrated as outlined in Step- 66. The resulting residue was purified by preparative RP-HPLC. Pure fractions were combined and lyophilized to131MEl\59069747.vlAttorney Docket No. 137521-10301 afford the title macrocyclic peptide (SEQ ID NO: 33). HPLC-MS (Method A-1): 5.74 min, ESI-MS m / z: 982.9 [M+2H]2+.Table E1 H

[0383] Example 2: The following peptides were synthesized by similar methods described above.

[0384] Table S7132MEl\59069747.vlAttorney Docket No. 137521-10301133MEl\59069747.vlAttorney Docket No. 137521-10301134MEl\59069747.vlAttorney Docket No. 137521-10301135MEl\59069747.vlAttorney Docket No. 137521-10301136MEl\59069747.vlAttorney Docket No. 137521-10301137MEl\59069747.vlAttorney Docket No. 137521-10301138MEl\59069747.vlAttorney Docket No. 137521-10301139MEl\59069747.vlAttorney Docket No. 137521-10301140MEl\59069747.vlAttorney Docket No. 137521-10301141MEl\59069747.vlAttorney Docket No. 137521-10301142MEl\59069747.vlAttorney Docket No. 137521-10301143MEl\59069747.vlAttorney Docket No. 137521-10301144MEl\59069747.vlAttorney Docket No. 137521-10301145MEl\59069747.vlAttorney Docket No. 137521-10301146MEl\59069747.vlAttorney Docket No. 137521-10301147MEl\59069747.vlAttorney Docket No. 137521-10301148MEl\59069747.vlAttorney Docket No. 137521-10301149MEl\59069747.vlAttorney Docket No. 137521-10301150MEl\59069747.vlAttorney Docket No. 137521-10301151MEl\59069747.vlAttorney Docket No. 137521-10301152MEl\59069747.vlAttorney Docket No. 137521-10301153MEl\59069747.vlAttorney Docket No. 137521-10301154MEl\59069747.vlAttorney Docket No. 137521-10301155MEl\59069747.vlAttorney Docket No. 137521-10301156MEl\59069747.vlAttorney Docket No. 137521-10301157MEl\59069747.vlAttorney Docket No. 137521-10301158MEl\59069747.vlAttorney Docket No. 137521-10301159MEl\59069747.vlAttorney Docket No. 137521-10301160MEl\59069747.vlAttorney Docket No. 137521-10301161MEl\59069747.vlAttorney Docket No. 137521-10301162MEl\59069747.vlAttorney Docket No. 137521-10301163MEl\59069747.vlAttorney Docket No. 137521-10301164MEl\59069747.vlAttorney Docket No. 137521-10301165MEl\59069747.vlAttorney Docket No. 137521-10301

[0385] Example-3: Synthesis of the conjugated form consisting of the GPC binding peptide in this invention and sulfoCy5 as a payload.

[0386] The desired polypeptide was synthesized as outlined in Scheme-4. To a solution of the GPC binding peptide in Example-1 (10.0 mg, 4.71 pimol) in DMF (150 piL) at room temperature were added triethylamine (6.6 piL, 47.1 pimol) and SulfoCy5-NHS (4.6 mg, 6.13 pimol). The mixture was stirred at room temperature for 18 hrs as outlined in step-D4. The resulting mixture was purified by preparative RP-HPLC. Pure fractions were combined and lyophilized to afford the title macrocyclic peptide. HPLC-MS (Method A-2): 3.80 min, ESI-MS m / z: 1209.6 [M+2H]2+.

[0387] Example-4: The following compounds were prepared by analogous method described above.

[0388] Table S8166MEl\59069747.vlAttorney Docket No. 137521-10301

[0389] Biological example-1.

[0390] Macrocyclic peptides in this invention were tested for binding to Glypican-3 protein by SPR.

[0391] Method B-1.

[0392] A CM3 sensor chip (Cytiva) was inserted into a Biacore T200 (Cytiva), primed three times with a running buffer: HBS-P+, pH 7.4 (Cytiva), and equilibrated at a flow rate of 30 pL / min. Ligand immobilization was performed at a flow rate of 10 pL / min. 50 pL each of 60 mM EDC solution (Cytiva) and 650 mM NHS solution (Cytiva) were mixed, and then reacted at a flow rate of 10 pL / min for 420 seconds. 150 pL of 0.2 pM human167MEl\59069747.vlAttorney Docket No. 137521-10301Glypican3 (R&D systems, catalog number: 2119-GP) solution which was prepared by diluting with 10 mM acetic acid solution (pH 5.0), was reacted at a flow rate of 10 pL / min for 420 seconds, and Glypican3 was immobilized on the CM3 sensor chip. After immobilization, 1.0 M ethanolamine aqueous solution (Cytiva) was allowed to react for 420 seconds at a flow rate of 10 pL / min for capping. A peptide lysate prepared to 10 mM in DMSO solution was diluted with running buffer to a final concentration of 10 pM peptide lysate. After that, 5 serial dilution series (100 nM, 50 nM, 25 nM, 10 nM, 5 nM) of peptide solutions were prepared. Using the above samples, peptide kinetics for Gly pican3 was obtained by SPR measurement. Kinetics evaluation of Gly pican3 was performed by SPR measurements at a flow rate of 10 pL / min, setting the association time to 120 seconds and the disassociation time to 600 seconds.

[0393] Method B-2.

[0394] A CM3 sensor chip (Cytiva) was inserted into a Biacore T200 (Cytiva), primed three times with a running buffer: HBS-P+, pH 7.4 (Cytiva), and equilibrated at a flow rate of 30 pL / min. Using the Anti-Fc Capture Kit (Cytiva), 50 pL each of 60 mM EDC solution (Cytiva) and 650 mM NHS solution (Cytiva) were mixed, and then reacted at a flow rate of 10 pL / min for 420 seconds. 150 pL of 20 pL / mL Anti-human IgG (Fc) Antibody solution was prepared by diluting with 10 mM acetic acid solution (pH 5.0), reacted at a flow rate of 10 pL / min for 420 seconds, and immobilized on a CM3 sensor chip. After immobilization, 1.0 M ethanolamine aqueous solution (Cytiva) was allowed to react for 420 seconds at a flow rate of 10 pL / min for capping. Human Glypican3-Fc (AGRO Biosystems, GP3-H5258) was allowed to react for 60 seconds at a flow rate of 10 pL / min. After capturing human Glypican3-Fc, a peptide solution prepared to 10 mM in DMSO solution was diluted with a running buffer to a final concentration of 10 pM. After that, 5 serial dilution series (100 nM, 50 nM, 25 nM, 10 nM, 5 nM) of peptide solutions were prepared. Using the above samples, peptide kinetics for Gly pican3 was obtained by SPR measurement. Kinetics evaluation of Glypican3 was performed by SPR measurements at a flow rate of 10 pL / min, setting the association time to 120 seconds and the disassociation time to 600 seconds.

[0395] Method B-3.

[0396] A Protein A sensor chip (Cytiva) was inserted into a Biacore T200 (Cytiva), priming was performed three times with a running buffer: HBS-P+, pH 7.4 (Cytiva), and equilibration was carried out at a flow rate of 30 pL / min. Glypican3-Fc was allowed to react for 60 seconds at a flow rate of 10 pL / min. After capturing human Glypican3 -Fc, a peptide solution prepared to 10 mM in DMSO solution was diluted with a running buffer to a final concentration of 10 pM. After that, 5 serial dilution series (100 nM, 50 nM, 25 nM, 10 nM, 5 nM) of peptide solutions were prepared. Using the above samples, peptide kinetics for Gly pican3 was obtained by SPR measurement. Kinetics evaluation of Glypican3 was performed by SPR measurements at a flow rate of 10 pL / min, setting the association time to 120 seconds and the disassociation time to 600 seconds.

[0397] Method B-4

[0398] A Protein A sensor chip (Cytiva) was inserted into a Biacore T200 (Cytiva), priming was performed three times with a running buffer: HBS-P+, pH 7.4 (Cytiva), and equilibration was carried out at a flow rate of 30 pL / min. Glypican3-Fc was allowed to react for 60 seconds at a flow rate of 10 pL / min. After capturing human Glypican3 -Fc, a peptide solution prepared to 1 mM in DMSO solution was diluted with a running buffer to a final168MEl\59069747.vlAttorney Docket No. 137521-10301 concentration of 1 pM. After that, the peptide solution was diluted to 40 nM in a similar manner. Using the above samples, peptide kinetics for Glypican3 was obtained by SPR measurement. Kinetics evaluation of Glypican3 was performed by SPR measurements at a flow rate of 30 pL / min, setting the association time to 120 seconds and the disassociation time to 600 seconds.

[0399] The obtained data by Method B-1 , Method B-2, Method B-3, and B4 were analyzed by follow. Single Cycle Kinetics was used as the kinetics evaluation model, and curve fitting was performed using Biacore T200 Evaluation Software Version 3.0 (Cytiva). The obtained sensorgram was subjected to curve fitting by the method of least squares and obtained KD values of the macrocyclic peptides in this invention for Glypican 3 protein are shown in Biological Table-1.Biological Table-1.169MEl\59069747.vlAttorney Docket No. 137521-10301170MEl\59069747.vlAttorney Docket No. 137521-10301

[0400] Furthermore, SPR KD data obtained by Method B-1 , Method B-2, and Method B-3 were analyzed and are shown in the table below. Single Cycle Kinetics was used as the kinetics evaluation model, and curve fitting was performed using Biacore T200 Evaluation Software Version 3.0 (Cytiva). The obtained sensorgram was subjected to curve fitting by the method of least squares and obtained KD values of the macrocyclic peptides in this disclosure for Glypican 3 protein are shown in Biological Table-2. The Conjugate IDs such as M-76, match the conjugates as set forth in Tables 1-9B.Biological Table-1171MEl\59069747.vlAttorney Docket No. 137521-10301172MEl\59069747.vlAttorney Docket No. 137521-10301173MEl\59069747.vlAttorney Docket No. 137521-10301

[0401] Certain linker-added peptides have identical or substantially the same peptide sequences compared to certain conjugates comprising such linker-added peptides, or "naked” peptides without the linker. Although the GPC3 binding affinity for such linker-added peptides have not been directly tested, they likely bind to GPC3 with similar avidity compared to that of the conjugates comprising such linker-added peptides, or that of the "naked” peptides without the linker portion.

[0402] The present technology may be used in bio-related industries and the pharmaceutical industry.

[0403] All references cited in this specification, and their references, are incorporated by reference herein in their entirety where appropriate for teachings of additional or alternative details, features, and / or technical background.

[0404] While the disclosure has been particularly shown and described with reference to particular embodiments, it will be appreciated that variations of the above-disclosed and other features and functions, or alternatives thereof, may be desirably combined into many other different systems or applications. Also, that various presently unforeseen or unanticipated alternatives, modifications, variations, or improvements therein may be subsequently made by those skilled in the art which are also intended to be encompassed by the following claims.174MEl\59069747.vl

Claims

1. Attorney Docket No. 137521-10301CLAIMSWe claim:A peptide that has avidity for Glypican 3 (GPC3), or a pharmaceutically acceptable salt thereof, wherein the peptide comprises an amino acid sequence of Formula I:Formula I, wherein R is -CH2-CO-; and,(i)Xi is MeLys, MeLysAc, MeAla, MeGlu, MeGIn, or CrpG, wherein the N-terminal nitrogen of Xi is attached to the C(O) of R;X2 is Melle, MeGIn, MeLys, MeHseMe, MeLysCOpipzaa, or MeGlu;X3 is Asp, MeAsp, Hgn, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, Hva, or (S)Mor(2CO);X4 is MeGIn, MeLys, MeAspapCOpipzaa, MeGlu, or CrpG;X5is F4COO, MeF4C00, Y, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A;Xe is TMe, altMe, lie, or Leu;X7 is TMe, lie, Vai, Leu, Gcpe, or Eva;Xs is Tyr, Cha, F3OH, F4aao, F4COO, F4aa, Yae, YaeCOpipzaa, F4F, F4U, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A;X9is MeNal27N, MeF3Me4C, MeF34dOMe, or MeW7N;X10 is Gly, MeGly, D-Ala, D-Ser, or D-Pro;X11 is 3Py6Ph, 3Py6Pyrazol1 , F41 Me4Pyz, 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61ap4, F41 apPyz, F42Py, F44Py, or Bph4C; and,X12 is MeCys or Cys, wherein sulfur of the X12 side chain is covalently attached to R, and carboxylic acid of X12 is replacedand, wherein Xi is MeGlu or CrpG; or X2 is MeLys; or X3 is MeAsp, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, (S)Mor(2CO), or Hva; or X4 is MeGlu, or CrpG; or X5 is MeF4C00, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A; or Xe is alTMe or Leu; or X7 is TMe, Vai, Leu, Gcpe, or Eva; or X8is Cha, F3OH, F4F, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A; or X9 is MeF3Me4C, MeF34dOMe, or MeW7N; or X10 is MeGly, D-Ala, D-Ser, or D-Pro; or Xu is175MEl\59069747.vlAttorney Docket No. 137521-103013Py6Pyrazol1 , 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61ap4, F41apPyz, F42Py, F44Py, or Bph4C; or X12 is Cys; or(ii) Xi to X12 have a sequence from Table 1, 2, 3, 4, 9A, or 20A.

2. The peptide or pharmaceutically acceptable salt thereof of claim 1 , comprising a peptide of Formula I, wherein R is -CH2-CO-; and(i) Xi is MeLys, MeLysAc, MeAla, MeGlu, MeGIn, or CrpG, wherein the N-terminal nitrogen of Xi is attached to the 0(0) of R; andX2 is Melle, MeGIn, MeLys, MeHseMe, MeLysCOpipzaa, or MeGlu; andX3 is Asp, MeAsp, Hgn, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, Hva, or (S)Mor(2CO); andX4 is MeGIn, MeLys, MeAspapCOpipzaa, MeGlu, or CrpG; andX5is F4C00, MeF4C00, Y, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A; andXe is TMe, altMe, lie, or Leu; andX7 is TMe, lie, Vai, Leu, Gcpe, or Eva; andXs is Tyr, Cha, F30H, F4aao, F4C00, F4aa, Yae, YaeCOpipzaa, F4F, F4U, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A; andX9is MeNal27N, MeF3Me4C, MeF34dOMe, or MeW7N; andX10 is Gly, MeGly, D-Ala, D-Ser, or D-Pro; andX11 is 3Py6Ph, 3Py6Pyrazol1, F41 Me4Pyz, 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61ap4, F41apPyz, F42Py, F44Py, or Bph4C; andX12 is MeCys or Cys, wherein sulfur of the X12 side chain is covalently attached to R, and carboxylic acid of X12 is replaced with -C(0)NH2 or; and wherein Xi is MeGlu or CrpG; or X2 is MeLys; or X3 is MeAsp, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, (S)Mor(2CO), or Hva; or X4 is MeGlu, or CrpG; or X5 is MeF4C00, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A; or Xe is alTMe or Leu; or X7 is TMe, Vai, Leu, Gcpe, or Eva; or X8is Cha, F30H, F4F, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A; or X9 is MeF3Me4C, MeF34dOMe, or MeW7N; or X10 is MeGly, D-Ala, D-Ser, or D-Pro; or Xu is 3Py6Pyrazol1, 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61ap4, F41apPyz, F42Py, F44Py, or Bph4C; or X12 is Cys; or(ii) Xi to X12 have a sequence from Table 1, 2, 3, 4, or 9A.

3. The peptide or pharmaceutically acceptable salt thereof of claim 1 , wherein Xi to X12 of the peptide of Formula (I) have a sequence from Table 1, 2, 3, 4, 9A, or 20A.

4. The peptide or pharmaceutically acceptable salt thereof of claim 2, wherein Xi to X12 of the peptide of Formula (I) have a sequence from Table 1, 2, 3, 4, or 9A.

5. A peptide that has avidity for Glypican 3 (GPC3), or a pharmaceutically acceptable salt thereof, wherein176MEl\59069747.vlAttorney Docket No. 137521-10301 the peptide comprises an amino acid sequence of Formula II:Formula II wherein:R* is -CH2-S-CH2-CO-; each Z is independently selected from H and Me; and,(i)A1 is the side chain of Lys, LysAc, Ala, Glu, Gin, or CrpG;A2 is the side chain of lie, Gin, Lys, HseMe, LysCOpipzaa, or Glu;A3 is the side chain of Asp, Hgn, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, Hva, or (S)Mor(2CO);A4 is the side chain of Gin, Lys, AspapCOpipzaa, Glu, or CrpG;A5is the side chain of F4COO, Y, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A;Ae is the side chain of TMe, altMe, lie, or Leu;Az is the side chain of Tme, lie, Vai, Leu, Gcpe, or Eva;As is the side chain of Tyr, Cha, F3OH, F4aao, F4COO, F4aa, Yae, YaeCOpipzaa, F4F, F4U, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A;A9 is the side chain of Nal27N, F3Me4C, F34dOMe, or W7N;Aw is the side chain of Gly, D-Ala, D-Ser, or D-Pro;An is the side chain of 3Py6Ph, 3Py6Pyrazol1 , F41 Me4Pyz, 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61 ap4, F41 apPyz, F42Py, F44Py, or Bph4C; and, wherein A1 is the side chain of Glu or CrpG; or A2 is the side chain of Lys; or A3 is the side177MEl\59069747.vlAttorney Docket No. 137521-10301 chain of Asp, Ser, Thr, Ala, or Lys, Glu, Asn, diMeDap, (S)Mor(2CO), or Hva; or A4 is the side chain of Glu or CrpG; or A5 is the side chain of F4C00, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A; or Ae is the side chain of alTMe or Leu; or Az is the side chain of Tme, Vai, Leu, Gcpe, or Eva; or As is the side chain of Cha, F3OH, F4F, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A; or A9 is the side chain of F3Me4C, MeF34dOMe, or MeW7N; or Aw is the side chain of Gly, D-Ala, D-Ser, or D-Pro; or An is the side chain of 3Py6Pyrazol1 , 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61 ap4, F41 apPyz, F42Py, F44Py, or Bph4C; or(ii) A1 to An correspond to the side chains of Xi to Xu of a sequence from Table 1 , 2, 3, 4, 9A, or 20A.

6. The peptide or pharmaceutically acceptable salt thereof of claim 3, comprising a peptide of Formula II: wherein:R* is -CH2-S-CH2-CO-; each Z is independently selected from H and Me; and,(i)A1 is the side chain of Lys, LysAc, Ala, Glu, Gin, or CrpG;A2 is the side chain of lie, Gin, Lys, HseMe, LysCOpipzaa, or Glu;A3 is the side chain of Asp, Hgn, Ser, Thr, Ala, Lys, Glu, Asn, diMeDap, Hva, or (S)Mor(2CO); A4 is the side chain of Gin, Lys, AspapCOpipzaa, Glu, or CrpG;A5is the side chain of F4COO, Y, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A;Ae is the side chain of TMe, altMe, lie, or Leu;Az is the side chain of Tme, lie, Vai, Leu, Gcpe, or Eva;As is the side chain of Tyr, Cha, F3OH, F4aao, F4COO, F4aa, Yae, YaeCOpipzaa, F4F, F4U, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A;A9 is the side chain of Nal27N, F3Me4C, F34dOMe, or W7N;Aw is the side chain of Gly, D-Ala, D-Ser, or D-Pro;An is the side chain of 3Py6Ph, 3Py6Pyrazol1 , F41 Me4Pyz, 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61 ap4, F41 apPyz, F42Py, F44Py, or Bph4C; and, wherein A1 is the side chain of Glu or CrpG; or A2 is the side chain of Lys; or A3 is the side chain of Asp, Ser, Thr, Ala, or Lys, Glu, Asn, diMeDap, (S)Mor(2CO), or Hva; or A4 is the side chain of Glu or CrpG; or A5 is the side chain of F4COO, 4Py, 3Py6NH2, 3Py6COO, F4(SO3H), or (PyrrCOO)A; or As is the side chain of alTMe or Leu; or Az is the side chain of Tme, Vai, Leu, Gcpe, or Eva; or As is178MEl\59069747.vlAttorney Docket No. 137521-10301 the side chain of Cha, F3OH, F4F, Aph, 3Py6NH2, 4Py, F4(C0NMe0H), F4C0NHMe, F4CONMe2, or (PyrrCOO)A; or A9 is the side chain of F3Me4C, MeF34dOMe, or MeW7N; or Aw is the side chain of Gly, D-Ala, D-Ser, or D-Pro; or An is the side chain of 3Py6Pyrazol1 , 3Py6O4pip1 Ms, 3Py6O4pip1 Ac, 3Py6O4thp, F43Py5H, F44Pdo, 3Py63Py5, 3Py63Py, 3Py61 ap4, F41 apPyz, F42Py, F44Py, or Bph4C; or(ii) A1 to An correspond to the side chains of Xi to Xu of a sequence from Table 1 , 2, 3, 4, or 9A.

7. The peptide or pharmaceutically acceptable salt thereof of claim 3, wherein Xi to X12 of the peptide of Formula (I) have a sequence from Table 1 , 2, 3, 4, 9A, or 20A.

8. The peptide or pharmaceutically acceptable salt thereof of claim 6, wherein Xi to X12 of the peptide of Formula (I) have a sequence from Table 1 , 2, 3, 4, or 9A.

9. The peptide or pharmaceutically acceptable salt thereof of any one of claims 1-8, wherein Xi to X12 of the peptide of Formula (I) have the sequence of SEQ ID NO: 226, 520, or 521.

10. The peptide or pharmaceutically acceptable salt thereof of any one of claims 1-9, wherein the peptide is not any of SEQ ID Nos: 1-72.

11. A conjugate comprising the peptide or pharmaceutically acceptable salt thereof according to any one of claims 1-10, covalently linked to a non-radionuclide moiety.

12. The conjugate or pharmaceutically acceptable salt thereof of claim 11 , wherein the non-radionuclide moiety is a substance or a payload molecule selected from the group consisting of: a nucleotide, a small molecule (e.g., with a M.W. of less than about 1000 Da), a medium sized molecule (e.g., with a M.W. of about 1 ,000-2,500 Da), a large sized molecule (e.g., with a M.W. of >2,500 Da), a polymer compound, a protein, a peptide, a tag, a biological fragment, a carrier including pharmaceutical compound, or a combination thereof.

13. The conjugate or pharmaceutically acceptable salt thereof of claim 12, wherein the peptide comprises the seqeunce of any one of SEQ ID NOs: 226, 520, and 521 .

14. The conjugate of pharmaceutically acceptable salt thereof of any one of claims 11-13, wherein the nonradionuclide moiety is covalently attached to the side chain of the first amino acid of the peptide (Xi, A1), the side chain of the second amino acid of the peptide (X2, A2), the side chain of the third amino acid of the peptide (X3, A3), the side chain of the fourth amino acid of the peptide (X4, A4), the side chain of the eighth amino acid of the peptide (Xs, As), the C-terminus of the C-terminal amino acid of the peptide (X12, Q, when Q is -NH2); or the side chain of the thirteenth amino acid (X13, Q, when Q is D-Lys) when present.

15. The conjugate or pharmaceutically acceptable salt thereof of any one of claims 11-14, further comprising a linker that covalently connects the peptide to the non-radionuclide moiety.

16. The conjugate or pharmaceutically acceptable salt thereof of claim 15, wherein the linker comprises 3 to179MEl\59069747.vlAttorney Docket No. 137521-1030110 intervening non-hydrogen, organic atoms between the non-radionuclide moiety and the peptide.The conjugate or pharmaceutically acceptable salt thereof of any one of claims 11-14, wherein the peptide is directly attached to the non-radionuclide moiety.

18. The conjugate or pharmaceutically acceptable salt thereof of any one of claims 11-14, wherein the180MEl\59069747.vlAttorney Docket No. 137521-1030119. The conjugate or pharmaceutically acceptable salt thereof of any one of claims 11-18, wherein the conjugate is not a conjugate of Table B, or wherein the conjugate is a conjugate from Table 5, 6, 7, 8, 9B, or 20B.

20. A pharmaceutical composition comprising the peptide or pharmaceutically acceptable salt thereof of any one of claims 1-10, and a pharmaceutically acceptable excipient or carrier.21 . A pharmaceutical composition comprising the conjugate or pharmaceutically acceptable salt thereof of any one of claims 11-19, and a pharmaceutically acceptable excipient or carrier.

22. A method of treating a disease or disorder characterized by overexpression of glypican 3 (GPC3) in a subject in need of treatment, the method comprising administering to the subject a therapeutically effective amount of the peptide or pharmaceutically acceptable salt thereof of any one of claims 1-10, the conjugate or pharmaceutically acceptable salt thereof of any one of claims 11-19, or the pharmaceutical composition of claim 20 or 21 .

23. A method of diagnosing a disease or disorder characterized by overexpression of GPC3 in a subject in need thereof, comprising administering to the subject, or contacting a sample from the subject in vitro, the peptide or pharmaceutically acceptable salt thereof of any one of claims 1-10, the conjugate or pharmaceutically acceptable salt thereof of any one of claims 11-19, or the pharmaceutical composition of claim 20 or 21; and detecting binding to GPC3 by the peptide or the conjugate, thereby determining expression level of GPC3 in the subject.

24. The method of claim 22 or 23, wherein the disease or disorder is cancer, such as a solid tumor or a hematological cancer.

25. The method of claim 24, wherein the cancer is hepatocellular carcinoma, squamous cell carcinoma of the lung, lung adenocarcinoma, germ cell tumors, hepatoblastoma, wilms tumor, malignant rhabdoid tumors, rhabdomyosarcoma, liposarcoma, thyroid cancers, pancreatic cancer, small bowel cancer,181MEl\59069747.vlAttorney Docket No. 137521-10301 small cell neuroendocrine carcinoma (SCNC), hormonally treated, castration resistant prostatic adenocarcinoma, ovarian cancer, gastric cancer, esophageal carcinoma, or malignant melanoma.

26. A kit, tester, or composition for determining expression level of GPC3 in a sample, wherein the kit, tester, or composition comprises the peptide or pharmaceutically acceptable salt thereof of any one of claims 1-10, the conjugate or pharmaceutically acceptable salt thereof of any one of claims 11-19, or the pharmaceutical composition of claim 20 or 21 .

27. The kit, tester, or composition of claim 26, adapted for use in a method of diagnosing a disease or disorder characterized by an overexpression or a decreased expression of GPC3.

28. The kit, tester, or composition of claim 26 or 27, wherein the sample is from a subject suspected of having a disease or disorder characterized by an overexpression or a decreased expression of GPC3.

29. Use of the peptide, conjugate, or pharmaceutically acceptable salt thereof according to any one of claims 1-19, in the manufacture of a medicament for diagnosing and / or treating a disease or disorder characterized by an overexpression or a decreased expression of GPC3.

30. The peptide, conjugate, or pharmaceutically acceptable salt thereof according to any one of claims 1- 19, for use in diagnosing and / or treating a disease or disorder characterized by an overexpression or a decreased expression of GPC3.182MEl\59069747.vl

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