Compounds for precision conjugation

Abstract

Described herein are compounds and strategies for precision conjugation of polypeptides in a controlled, efficient, reliable, and tailorable fashion for broad and advanced uses. Site-selective modification of polypeptides is desired for the development of biopharmaceutical conjugates for use as novel therapeutics, bioimaging agents, and material sciences. The proximity-triggered conjugation molecules described herein may be relatively small molecules that form stable covalent bonds with specific amino acid residues within the PRM. This chemistry avoids the instability associated with maleimide linkages and ensures that drug payloads remain securely attached to the antibody throughout circulation.

Description

COMPOUNDS FOR PRECISION CONJUGATIONPRIORITY CLAIM

[0001] This patent application claims priority to U.S. provisional patent application no. 63 / 744,675, titled “COMPOUNDS FOR PRECISION CONJUGATION,” and filed on January 13, 2025, which is herein incorporated by reference in its entirety.FIELD OF THE INVENTION

[0002] The prevalence of protein conjugates as therapeutic options is continually increasing due to the unique combination of the biological activity from the protein and the additional therapeutic functionality afforded from the conjugation of additional chemical moieties. Described herein are compounds and strategies for precision conjugation in a controlled, efficient, reliable, and tailorable fashion for broad and advanced applications.BACKGROUND

[0003] The modification of proteins at a single amino acid or specific site with synthetic moieties is a significant challenge, which is due in part to the difficulty of maintaining both protein integrity and function during the chemical bioconjugation process. Among the natural amino acids that compose proteins, only a subset serve as appropriate targets for bioconjugation. Many strategies have been developed to target nucleophilic amino acid residues such as cysteine and lysine residues. Cysteine is a popular choice for bioconjugation due to its nucleophilic properties, and yet its tendency to form disulfide bonds limits its availability for bioconjugation. Further, the most commonly used reagent for conjugation to cystine residues, maleimide, creates a thio-ether Michael-addition product that often exhibits instability in vivo due to the possibility of a retro-Michael-addition reaction. Lysine residues are abundant in native proteins, and this abundance makes site-specific conjugation of lysine residues difficult. Attempts have been made to site-specifically modify only the single N-terminal amine in a protein due to its greater nucleophilicity and easier deprotonation versus the relatively less nucleophilic s-amino group of internal lysines (pKa ~8 vs. pKa ~10, respectively), but pH-selective acylation reactions are difficult to control, and heterogenous product mixtures are often obtained. The most common reagents for amine modification, N-hydroxysuccinimide (NHS) esters, also react with serine, tyrosine, and histidine residues, further affording heterogeneous bioconjugates. A need exists for the precise site-specific-1- SG Docket No.: 14989-700.600bioconjugation of proteins in a controlled, efficient, reliable, and tailorable fashion for broad and advanced therapeutic applications.

[0004] Described herein are multifunctional compounds and strategies for precision conjugation in a controlled, efficient, reliable, and tailorable fashion for broad and advanced applications. The strategies described herein incorporate one or more peptidic recognition motif(s) (PRM) into therapeutic proteins while maintaining both protein integrity and function. Selective noncovalent binding interactions of the compounds described herein with the PRM(s) facilitate a proximity-induced (also called binding-induced) irreversible covalent reaction between lysines and electrophilic warheads within the same compounds. The compounds described herein are highly variable for use in a variety of bioconjugation strategies and uses as described herein.SUMMARY

[0005] Described herein are compositions and methods for bioconjugation of a variety of materials, including but not limited to antibody and antibody fragments. In general, the composition and methods described herein may provide a bioconjugation platform that may address challenges in antibody drug conjugate (ADC) development, including stability, sitespecificity, manufacturing complexity, and therapeutic resistance. Traditional ADCs often rely on cysteine / maleimide or lysine-based chemistry, which can lead to unstable linkages, heterogeneous products, and off-target toxicity. These limitations not only compromise safety and efficacy but also hinder scalability and increase manufacturing costs. The methods and apparatuses may reduce or eliminate these issues by using an engineered Peptidic Recognition Motif (PRM) that may be stably and inoffensively incorporated into proteins, including but not limited to antibodies and antibody-derived molecules (including antibody fragments, antibody constructs, single-domain antibodies, Fc-fusion proteins, bispecific constructs, ADCs, etc.) and may enable precise and predictable conjugation.

[0006] In general, these compositions and methods may include an engineered PRM-targeting molecule, which may also be referred to herein as a proximity-triggered conjugation compound or molecule. The proximity-triggered conjugation molecule may be a relatively small molecule that forms a stable covalent bond with specific amino acid residues within the PRM. This chemistry avoids the instability associated with maleimide linkages and ensures that drug payloads remain securely attached to the antibody throughout circulation. By leveraging natural amino acids and straightforward chemical reactions, these methods and compositions may provide homogeneous ADCs with controlled drug-to-antibody ratios-2- SG Docket No.: 14989-700.600(DAR), reducing aggregation and improving therapeutic performance. These methods and apparatuses generally provide for bioconjugation of distinct drug payloads. These methods may be readily integrated into biopharma pipelines and support rapid conjugation, scalable manufacturing, and flexible design.

[0007] For example, generally described herein are proximity-triggered conjugation compounds or molecules that may be used to site-specificity conjugate one or more cargo molecules onto a polypeptide as will be described in greater detail here. For example, these compounds, which may be referred to herein (for convenience) as proximity-triggered conjugation compounds or molecules, may generally have the formula as shown by Formula (I):PBU-L1-L2-PCU (Formula (I)).

[0008] In general, PBU (“PRM Binding Unit” or “peptidic recognition motif binding unit”) is a moiety capable of non-covalently binding to a peptidic recognition motif (PRM) in a polypeptide. The PRM may comprise a target amino acid comprising lysine (Lys), ornithine (Orn), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys). The PRM may be on the protein to which a cargo is to be conjugated, such as, but not limited to an antibody, antibody fragment, etc. (which may be used herein to refer to any antibody-derived molecule). The PCU (“PRM Conjugation Unit”) is a moiety generally capable of covalently reacting with the target amino acid of the PRM when held at a proximity-induced binding distance to the target amino acid of the PRM. L1may be a cleavable linker, and L2may be a spacer or may be absent. In general, upon non-covalent bonding of the PBU to the PRM in the polypeptide, the PCU is held at the proximity-induced binding distance to the target amino acid of the PRM to cause covalent bonding between the PCU and the target amino acid of the PRM.

[0009] For example, a compound of Formula (I) (e.g., PBU-L1-L2-PCU) may generally include: a PBU comprising a moiety having at least one nitrilotriacetic acid, at least one pyridine, or at least one iminodiacetic acid, or combinations thereof. The PBU may be capable of non-covalently binding to a peptidic recognition motif (PRM) in a polypeptide, wherein the PRM comprises a target amino acid comprising lysine (Lys), ornithine (Om), homolysine (HLys), P2-lysine (P2-Lys), or P3-Iysine (P3-Lys). PCU may generally be an electrophilic moiety and may comprise one or more of a vinyl sulfone group, a vinyl sulfonamide group, a pyrimidine sulfoxide group, a pyrimidine sulfone group, a vinyl pyridine group, or a vinyl pyrimidine group, such that the PCU covalently reacts with the target amino acid of the PRM only when held at a proximity-induced binding distance from-3- SG Docket No.: 14989-700.600the target amino acid of the PRM. As mentioned, LI may be a cleavable linker; and L2 may be an optional spacer that may be absent or included. In general, upon non-covalent bonding of the PBU to the PRM in the polypeptide, the PCU may be held at the proximity -induced binding distance to the target amino acid of the PRM to cause covalent bonding between the PCU and the target amino acid of the PRM.

[0010] In general, the electrophilic moiety that may be used for the PCU may be selected in order to optimize for covalent reaction selectivity, so that the PCU does not react with the target (e.g., Lys) unless and until the proximity is maintained by noncovalently binding to a PRM (via the PBU portion of the molecule to which the PCU is covalently attached) that include the target as part of (or adjacent to, including in some cases immediately adjacent to) the PRM. For example, as described herein, vinyl sulfonamide (an in particular vinyl sulfonamide with an aniline and an N-methyl amide) may be of particular use. Other similar electrophilic moieties, such as those including a vinyl sulfone group, a vinyl sulfonamide group, a pyrimidine sulfoxide group, a pyrimidine sulfone group, a vinyl pyridine group, or a vinyl pyrimidine group, may also be of particular interest. Thus, in general, the PCU includes an electrophile that reacts predominantly with a target (e.g., lysine side chain) under conditions of pH 7-9, in an aqueous solution at about 4-40°C, and at a concentration of about 10-500 pM of the electrophile and the lysine, so that an appreciable reaction with the Lys occurs only if the Lys is a component of a PRM, and so that the electrophile and lysine are held in close proximity via a separate non-covalent binding interaction. The preferred distance between the electrophile and the nearest non-covalent binding component of the PRM may be about 50 angstroms or less (e.g., about 45 angstroms or less, about 40 angstroms or less, about 35 angstroms or less, about 30 angstroms or less, about 25 angstroms or less, about 20 angstroms or less, about 15 angstroms or less, about 10 angstroms or less, etc.). For example, vinyl sulfonamide of one example of a proximity-triggered conjugation molecule described in greater detail herein predominantly reacts with a lysine sidechain only when the lysine is a component of a PRM in the context of a peptide or a protein, respectively. The characteristics of an electrophile that allow it to satisfy the requirement above may include structural, electronic, or other properties.

[0011] For example, in some cases the PCU may comprises an electrophilic warhead moiety that is capable of covalently reacting with the target amino acid only when held within the proximity-induced binding distance to the target amino acid. In some cases the PCU does not substantially covalently react with the target amino acid when the compound is not non--4- SG Docket No.: 14989-700.600covalently bound to the PRM. The PCU may comprise an electrophilic warhead moiety that is capable of covalently reacting with the target amino acid when held within the proximity-induced binding distance to the target amino acid, wherein the electrophilic warhead moiety comprises a vinyl sulfone group, a vinyl sulfonamide group, a pyrimidine sulfoxide group, a pyrimidine sulfone group, a vinyl pyridine group, a vinyl pyrimidine group, or another moiety that satisfies the criterion that covalent reaction with the target residue only occurs upon non-covalent binding of the PBU to the PRM.

[0012] In any of these examples the PBU may comprise at least one nitrilotriacetic acid, at least one iminodiacetic acid, or at least one pyri din-2 -ylmethanamine, or combinations thereof. The PBU may comprise at least one nitrilotriacetic acid, or at least one iminodiacetic acid, or combinations thereof.

[0013] One example of a proximity-triggered conjugation molecule as described herein has the following structure:<wherein:R1is H, halogen, substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, pyrimidine, or -C(=O)OR3; andR2is H, halogen, substituted or unsubstituted alkyl, or -CN; andR3is independently H or substituted or unsubstituted alkyl.

[0014] In some examples, the proximity-triggered conjugation molecule has the following structure:-5- SG Docket No.: 14989-700.600

[0015] Also described herein are methods of selectively modifying a target amino acid of a polypeptide. For example, a method may include: contacting a compound with the polypeptide under conjugation condition, wherein the compound has a formula of: PBU-L1-L2-PCU (Formula (I)) wherein: PBU is a moiety comprising at least one nitrilotriacetic acid, or at least one iminodiacetic acid, or combinations thereof; PCU is an electrophilic moiety comprising on or more of: vinyl sulfone group, a vinyl sulfonamide group, a pyrimidine sulfoxide group, a pyrimidine sulfone group, a vinyl pyridine group, or a vinyl pyrimidine group; L1is a cleavable linker; and L2is an optional spacer that may be present or absent; non-covalently binding the PBU portion of the compound to a peptidic recognition motif (PRM) of the polypeptide, wherein the PRM comprises a target amino acid comprising lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys); covalently bonding the PCU to the target amino acid while the PBU portion of the compound is noncovalently bound to the PRM.

[0016] The polypeptide may generally be a biologically active polypeptide (B). The biologically active peptide may be modified to include the engineered PRM (e.g., a PRM1 or PRM2 amino acid sequence). In some examples, the polypeptide may be an antibody or antibody fragment. In particular, the polypeptide may be engineered to include the PRM within a constant region of the antibody or antibody fragment. The PRM may be on any appropriate portion of the polypeptide being conjugated. For example, the PRM may be on the N-terminus of the polypeptide, on the C-terminus of the polypeptide, or within an internal position of the polypeptide. The PRM may be independently selected from PRM1, PRM2, or combinations thereof, wherein PRM1 comprises: 2 to 4 histidine (His) residues; and a lysine (Lys) residue, ornithine (Om) residue, homolysine (HLys), a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue; wherein PRM2 comprises amino acid residues comprising: at least 2 amino acids selected from an arginine (Arg) residue, a homoarginine (HArg) residue, a citrulline (Cit) residue, norarginine (AGBA) residue, a p2-arginine (P2-Arg), and a p3-arginine-6- SG Docket No.: 14989-700.600(P3-Arg); and a lysine (Lys) residue, an ornithine (Om) residue, a homolysine (HLys) residue, a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue. In some cases the PRM1 comprises at least one of: X-His-His, His-X-His, His-His-X, X-His-His-His, His-X-His-His, His-His-X-His, His-His-His-X, X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, or His-His-His-His-X, wherein X is Lys, Om, HLys, p2-Lys, or p3-Lys. The PRM1 may comprise at least one of: X-His-His, His-X-His, or His-His-X. The PRM1 may comprise at least one of: X-His-His-His, His-X-His-His, His-His-X-His, or His-His-His-X. The PRM1 may comprise at least one of: X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, or His-His-His-His-X. X may be Lys, Orn, HLys, etc. At least one PRM1 may be covalently attached at the N-terminus or C-terminus of the biologically active polypeptide.

[0017] In general, the polypeptide may comprise the amino acid sequence: B-(His)m-X-(His)n or (His)m-X-(His)n-B, wherein, B is the biologically active polypeptide; X is Lys, Om, HLys, P2-Lys, or p3-Lys; m is 0, 1, 2, 3, or 4; and n is 0, 1, 2, 3, or 4, provided that m + n is at least 2. At least one PRM1 may be incorporated in an internal position of the polypeptide. The polypeptide may comprises the amino acid sequence: B1-(Hi s)m-X-(His)n-B2wherein B1and B2are fragments of a biologically active polypeptide B, such that B'B2is the biologically active polypeptide B, and the -(His)m-X-(His)n- sequence is added at an internal position of the biologically active polypeptide B; X is Lys, Om, HLys, p2-Lys, or p3-Lys; m is 0, 1, 2, 3, or 4; and n is 0, 1, 2, 3, or 4, provided that m + n is at least 2.

[0018] For example, as mentioned above, the proximity-triggered conjugation molecule may have the formula: PBU-L1-L2-PCU (Formula (I)), where: PBU is a moiety capable of non-covalently binding to a peptidic recognition motif (PRM) in a polypeptide, wherein the at least one PRM is independently selected from PRM1, PRM2, or combinations thereof; wherein each PRM1 comprises: 2 to 4 histidine (His) residues; and a lysine (Lys) residue, ornithine (Orn) residue, homolysine (HLys), a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue wherein each PRM2 comprises amino acid residues comprising: at least 2 amino acids selected from an arginine (Arg) residue, a homoarginine (HArg) residue, a citrulline (Cit) residue, norarginine (AGBA) residue, a p2-arginine (P2-Arg), and a p3-arginine (P3-Arg); and a lysine (Lys) residue, an ornithine (Om) residue, a homolysine (HLys) residue, a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue.

[0019] PCU is a moiety capable of covalently reacting with the side chain of the at least one Lys, Om, HLys, p2-Lys, or p3-Lys in the PRM upon binding of the PBU with the PRM;-7- SG Docket No.: 14989-700.600L1is a cleavable linker; and L2is a spacer.

[0020] In some embodiments, PBU comprises at least one nitrilotriacetic acid, at least one iminodiacetic acid, or at least one pyridin-2-ylmethanamine, or combinations thereof. In some embodiments, PBU comprises at least one nitrilotriacetic acid, or at least one iminodiacetic acid, or combinations thereof.

[0021] In some embodiments,CH, C-OH, orN.

[0022] In some embodiments, PCU comprises an electrophilic warhead moiety that is empirically shown to be or expected to be by one skilled in the art, capable of covalently reacting with the side chain of the at least one Lys, Om, HLys, p2-Lys, or p3-Lys in the PRM preferentially when the PBU is non-covalently bound to the PRM. In some embodiments, PCU comprises an electrophilic warhead moiety that is capable of covalently reacting with the side chain of the at least one Lys, Om, HLys, p2-Lys, or p3-Lys in the PRM upon binding of the PBU with the PRM, wherein the electrophilic warhead moiety is a vinyl sulfone, a vinyl sulfonamide, a pyrimidine a sulfoxide, a pyrimidine sulfone, a vinyl pyridine, or a vinyl pyrimidine.

[0023] In some embodiments,

[0024] In some embodiments,

[0025] In some embodiments, the compound has the following structure:SG Docket No.: 14989-700.600

[0026] In some embodiments, R1is H; R2is H; and each R3is methyl.

[0027] In some embodiments, L1comprises a cleavable linker. In some embodiments, L1comprises a cleavable linker that upon cleavage provides an electrophilic or nucleophilic moiety.

[0028] In some embodiments, L1is orthogonally cleavable to peptide cleavage. In some embodiments, L1comprises a self-immolative linker. In some embodiments, L1comprises a disulfide group linker, photoreactive linker, fluoride-cleavable linker, alkali-cleavable linker, or oxidation cleavable linker. In some embodiments, L1comprises a nucleophilic cleavable linker, electrophile cleavable linkers, acid cleavable linker, base cleavable linker, linker cleavable with reducing reagents, linker cleavable with oxidizing reagents, or photocleavable linker.

[0029] In some embodiments, L1comprises a dialkoxy silane linker, a 2-cyanoethyl moiety, sulfone linker, ester linker, thiophenylester linker, vinyl sulfide linker, vinyl ether linker, acylhydrazone linker, hydrazone linker, acylsulfonamide linker, disulfide linker, azo linker, ortho-nitrobenzyl linker, phenacyl ester linker, ketal linker, carbamate linker, selenium linker, vicinal diol linker, para-aminobenzyl (PAB)-carbamate linker, trans-cyclooctene carbamate linker.

[0030] In some embodiments, L2is a spacer and is not cleavable. In some embodiments, L2comprises one or more linear structures, one or more non-linear structures, one or more branched structures, one or more cyclic structures, one or more flexible moieties, one or more rigid moieties, or combinations thereof.

[0031] In some embodiments, L1comprises a cleavable linker that upon cleavage provides a moiety capable of forming a linkage with a coupling partner, wherein the cleavable linker is orthogonally cleavable to peptide cleavage; L2further comprises a moiety capable of forming a linkage with a coupling partner; wherein the linkage formation reaction in L1is orthogonal-9- SG Docket No.: 14989-700.600to the linkage formation reaction in L2. In some embodiments, the linkage formation reaction in L1is a coupling reaction between an electrophile and a nucleophile. In some embodiments, the coupling reaction is an amide-forming ligation, ester-forming ligation, ether-forming ligation, imine-forming ligation, oxime-forming ligation, hydrazone-forming ligation, or sulfonamide-forming ligation. In some embodiments, the linkage formation reaction in L2is a cycloaddition reaction between an azide and alkyne to form a triazole, a strain-promoted azide-alkyne cycloaddition (SPAAC), inverse electron-demand Diels-Alder (IEDDA), or sydnone-alkyne cycloaddition (SPSAC).

[0032] In some embodiments, the compound has the following structure:wherein,U1is CH, C-OH, orN.L1is a cleavable linker; L2is a spacer;R3is H or substituted or unsubstituted alkyl;His is histidine;m is 0, 1, 2, 3, or 4; n is 0, 1, 2, 3, or 4, provided that m + n is at least 2;q is 1, 2, or 3;Rais a polypeptide B, and Rbis -OH; or Rais H, and Rbis a polypeptide B; or Rais aSG Docket No.: 14989-700.600fragment (B1) of a polypeptide B, Rbis the other fragment (B2) of the polypeptide B, such that B'-B2is B; or Rais H, and Rbis -OH.

[0034] In another aspect, described herein is a compound having the structure of:wherein,U1is CH, C-OH, orN.L1is a cleavable linker; L2is a spacer;each R3is H or substituted or unsubstituted alkyl;q is 1, 2, or 3;Rais a polypeptide, and Rbis -OH; or Rais H, and Rbis a polypeptide; or Rais a fragment (B1) of a polypeptide B, Rbis the other fragment (B2) of the polypeptide B, such that B^B2is B; or Rais H, and Rbis -OH.

[0035] In another aspect, described herein is a compound having the structure of:wherein,U1is CH, C-OH, orN;L1is a cleavable linker; L2is a spacer;each R3is H or substituted or unsubstituted alkyl;W1is -X^Y-X^Y-, and W2is absent; or W1is absent, and W2is -X^Y-X^Y-; or W1 and W2 are both -Xx-Y-X2-Y-;-11- SG Docket No.: 14989-700.600each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg; or Y is absent;X1comprises 0 to 10 amino acid residues other than Lys, Orn, HLys, and Y;X2comprises 0 to 10 amino acids other than Lys, Orn, HLys, and Y;q is 1, 2, or 3;Rais a polypeptide B, and Rbis -OH; or Rais H, and Rbis a polypeptide B; or Rais a fragment (B1) of a polypeptide B, Rbis the other fragment (B2) of the polypeptide B, such that B'-B2is B; orRais H, and Rbis -OH.

[0036] Also described herein is a method of selectively modifying a lysine residue within a polypeptide comprising: contacting a compound of Formula (I), with a polypeptide in the presence of a nickel (Ni2+) salt, cobalt (Co2+) salt, copper (Cu2+) salt, or zinc (Zn2+) salt, in a suitable solvent; wherein the polypeptide comprises at least one PRM1. In some embodiments, the lysine residue that is selectively modified is the lysine in the at least one PRM1.

[0037] Also described herein is a method of selectively modifying a lysine residue within a polypeptide comprising: contacting a compound of Formula (I), with a polypeptide, in a suitable solvent in the absence of a transition metal salt; wherein the polypeptide comprises at least one PRM2. In some embodiments, the lysine residue that is selectively modified is the lysine in the at least one PRM2.

[0038] Also described herein is a method of selectively modifying at least two lysine residues within a polypeptide comprising: contacting a compound of Formula (I), with a polypeptide, under conjugation condition i) followed by conjugation condition ii), or under conjugation condition ii) followed by conjugation condition i); wherein conjugation condition i) comprises the use of a nickel (Ni2+) salt, cobalt (Co2+) salt, copper (Cu2+) salt, or zinc (Zn2+) salt, in a suitable solvent; wherein conjugation condition ii) comprises a suitable solvent, and the absence of a transition metal salt; wherein the polypeptide comprises at least one PRM1 and at least one PRM2. In some embodiments, conjugation condition i) selectively modifies the lysine in the at least one PRM1. In some embodiments, conjugation condition ii) selectively modifies the lysine in the at least one PRM2. In some embodiments, conjugation condition i) selectively modifies the lysine in PRM1 but not the lysine in PRM2. In some embodiments, conjugation condition ii) selectively modifies the lysine in PRM2 but not the lysine in PRM1.-12- SG Docket No.: 14989-700.600

[0039] As mentioned above, these methods and compositions may be used with any appropriate polypeptide to which a cargo is to be conjugated as the engineered PRM site. Thus, also descried herein are polypeptides comprising: at least one peptidic recognition motif (PRM), and an amino acid sequence of a biologically active polypeptide (B), wherein the at least one PRM is: 1) added to the N-terminus of the biologically active polypeptide, 2) added to the C-terminus of the biologically active polypeptide, or 3) inserted at an internal position of the biologically active polypeptide; wherein the at least one PRM is independently selected from PRM1, PRM2, or combinations thereof: wherein each PRM1 comprises: 2 to 4 histidine (His) residues; and a lysine (Lys) residue, ornithine (Om) residue, homolysine (HLys), a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue wherein each PRM2 comprises amino acid residues comprising: at least 2 amino acids selected from an arginine (Arg) residue, a homoarginine (HArg) residue, a citrulline (Cit) residue, norarginine (AGB A) residue, a p2-arginine (P2-Arg), and a p3-arginine (P3-Arg); and a lysine (Lys) residue, an ornithine (Orn) residue, a homolysine (HLys) residue, a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue.

[0040] In some embodiments, each PRM1 comprises: X-His-His, His-X-His, His-His-X, X-His-His-His, His-X-His-His, His-His-X-His, His-His-His-X, X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, or His-His-His-His-X, wherein X is Lys, Orn, HLys, p2-Lys, or p3-Lys. In some embodiments, each PRM1 comprises: X-His-His, His-X-His, or His-His-X. In some embodiments, eachPRMl comprises: X-His-His-His, His-X-His-His, His-His-X-His, or His-His-His-X. In some embodiments, each PRM1 comprises: X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, or His-His-His-His-X. In some embodiments, X is Lys. In some embodiments, X is Orn. In some embodiments, X is HLys.

[0041] In some embodiments, at least one PRM1 is covalently attached at the N-terminus or C-terminus of the biologically active polypeptide.

[0042] In some embodiments, the polypeptide comprises the amino acid sequence:B-(His)m-X-(His)nor (His)m-X-(His)n-B,wherein,B is the biologically active polypeptide;X is Lys, Orn, HLys, p2-Lys, or p3-Lys;m is 0, 1, 2, 3, or 4; andn is 0, 1, 2, 3, or 4, provided that m + n is at least 2.-13- SG Docket No.: 14989-700.600

[0043] In some embodiments, at least one PRM1 is inserted at an internal position of the polypeptide. In some embodiments, the polypeptide comprises the amino acid sequence:wherein,B1and B2are fragments of a biologically active polypeptide B, such that BXB2is the biologically active polypeptide B, and the -(His)m-X-(His)n- sequence is added at an internal position of the biologically active polypeptide B;X is Lys, Om, HLys, p2-Lys, or p3-Lys;m is 0, 1, 2, 3, or 4; andn is 0, 1, 2, 3, or 4, provided that m + n is at least 2.

[0044] In some embodiments, m + n is at least 3; m + n is at least 4; m + n is at least 5; m + n is at least 6; m is 0 and n is 2; m is 2 and n is 0; m is 0 and n is 4; m is 1 and n is 3; m is 2 and n is 2; m is 3 and n is 1; or m is 4 and n is 0.

[0045] In some embodiments, each PRM2 comprises: X3-X1-Y -X4-X2-Y, wherein: X3or X4is Lys, Orn, HLys, p2-Lys, or p3-Lys; if X3is present, then X4is not present; if X4is present, then X3is not present; each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and p3-Arg; X1comprises 0 to 10 amino acid residues other than X3, X4, and Y; X2comprises 0 to 10 amino acids other than X3, X4, and Y.

[0046] In some embodiments, X3or X4is Lys. In some embodiments, X3or X4is Om. In some embodiments, X3or X4is HLys. In some embodiments, each Y is Arg. In some embodiments, each Y is HArg. In some embodiments, each Y is Cit. In some embodiments, each Y is AGBA. In some embodiments, each amino acid residue of X1is independently selected from alanine (Ala), asparagine (Asn), glutamine (Gin), glycine (Gly), isoleucine(Ile), leucine (Leu), methionine (Met), phenylalanine (Phe), proline (Pro), serine (Ser), threonine (Thr), tryptophan (Trp), tyrosine (Tyr), valine (Vai), glutamic acid (Glu), aspartic acid (Asp), and non-natural amino acids without positively charged or thiol-containing side chains.

[0047] In some embodiments, each amino acid residue of X2is independently selected from alanine (Ala), asparagine (Asn), glutamine (Gin), glycine (Gly), isoleucine(Ile), leucine (Leu), methionine (Met), phenylalanine (Phe), proline (Pro), serine (Ser), threonine (Thr), tryptophan (Trp), tyrosine (Tyr), valine (Vai), glutamic acid (Glu), aspartic acid (Asp), and non-natural amino acids without positively charged or thiol-containing side chains.

[0048] In some embodiments, at least one PRM2 is covalently attached at the N-terminus or C-terminus of the biologically active polypeptide. In some embodiments, the polypeptide-14- SG Docket No.: 14989-700.600comprises the amino acid sequence:B-X3-X1-Y-X4-X2-Y or Y-X2-X4-Y-X1-X3-Bwherein:B is a biologically active polypeptide (B);X3or X4is Lys, Om, HLys, p2-Lys, or p3-Lys;if X3is present, then X4is not present;if X4is present, then X3is not present;each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg; X1comprises 0 to 10 amino acid residues other than X3, X4, and Y; andX2comprises 0 to 10 amino acids other than X3, X4, and Y.

[0049] In some embodiments, at least one PRM2 is inserted at an internal position of the polypeptide. In some embodiments, the polypeptide comprises the amino acid sequence:B^X^-Y-X^-Y-B2wherein,B1and B2are fragments of a biologically active polypeptide B, such that BXB2is the biologically active polypeptide (B), and the -X3-X1-Y-X4-X2-Y-sequence is added at an internal position of B;B1and B2are fragments of a biologically active polypeptide B, such that BXB2is the biologically active polypeptide B and the -X^X^Y-X^X^Y- sequence is added at an internal position of polypeptide B;X3or X4is Lys, Om, HLys, P2-Lys, or p3-Lys;if X3is present, then X4is not present;if X4is present, then X3is not present;each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg; X1comprises 0 to 10 amino acid residues other than X3, X4, and Y; andX2comprises 0 to 10 amino acids other than X3, X4, and Y.

[0050] In some embodiments, the biologically active polypeptide comprises at least one PRM1 and at least one PRM2.

[0051] In some embodiments, the biologically active polypeptide is a therapeutic polypeptide. In some embodiments, the therapeutic polypeptide has a molecular weight of at least about 0.5 kDa. In some embodiments, the therapeutic polypeptide has a molecular weight of about 0.5 kDa to about 5 kDa, about 5 kDa to about 10 kDa, about 10 kDa to about 20 kDa, about 20 kDa to about 30 kDa, about 30 kDa to about 60 kDa, about 60 kDa to about-15- SG Docket No.: 14989-700.600110 kDa, or about 110 kDa to about 200 kDa, or greater than about 200 kDa.

[0052] In some embodiments, the therapeutic polypeptide is an enzyme, receptor or ligand, cell surface receptor, ion channel (i.e., pore-forming membrane protein), kinase, blood protein, chemokine, cytokine, antibody, antibody fragment, antigen, anticoagulant, blood factor, bone morphogenetic protein, growth factor, transcription factor, interferon, interleukin, thrombolytic agent, cytoskeletal protein, or peptide hormone. In some embodiments, the therapeutic polypeptide is a peptide hormone, protein hormone, bacterial / antibiotic peptide, antifungal peptide, invertebrate peptide, venom derived peptide, anticancer peptide, vaccine peptide, anti-inflammatory peptide, neuropeptide, endocrine peptide, gastrointestinal peptide, cardiovascular peptide, renal peptide, respiratory peptide, opioid peptide, neurotrophic peptide. In some embodiments, the therapeutic polypeptide is a monoclonal antibody, polyclonal antibody, single-domain antibody (sdAb, VHH, or nanobody), VHH-Fc, heavy chain only antibody (HCAb), a single chain variable fragment (scFv), di-scFv, tri-scFv, F(ab) fragment antibody, F(ab')2 fragment antibody, bispecific antibody (trifunctional antibody, chemically linked Fab, bi-specific T-cell engager), microantibody, intrabody, affibody molecule, Affilin, Affimer, Affitin, Alphabody, Anticalin, Avimer, DARPin, Kunitz domain peptide, Monobody, nanoCLAMP. In some embodiments, the polypeptide is an IgM antibody, IgD antibody, IgG antibody, IgA antibody, or IgE antibody. In some embodiments, polypeptide is an IgGl antibody, IgG2 antibody, IgG3 antibody, IgG4 antibody, IgAl antibody or IgA2 antibody. In some embodiments, the polypeptide is a full-length cell surface receptor. In some embodiments, the polypeptide is only the extracellular domain of a cell surface receptor. In some embodiments, the cell surface receptor is on the surface of a living or non-living cell. In some embodiments, the cell surface receptor is not associated with a cell.

[0053] Of particular interest herein are antibody or antibody fragment (or more generically, antibody-derived) polypeptides engineered to include one or more PRMs for bioconjugation using the proximity-triggered conjugation molecules described herein.

[0054] For example, an antibody or antibody fragment comprising a polypeptide including a precision conjugation site may include a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide has at least 90% identity with SEQ ID NO. 1, further wherein a sequence corresponding to PRM1 or PRM2 is either at a C-terminal end of the polypeptide or is located between positions 38 to 50 or positions 94-104 of SEQ ID NO. 1, either: in place of any-16- SG Docket No.: 14989-700.600single amino acid in positions 38 to 50 or positions 94-104 of SEQ ID NO. 1, or between any two consecutive amino acids in positions 38 to 50 or positions 94-104 of SEQ ID NO. 1, wherein: the sequence of PRM1 is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Orn), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the sequence of PRM2 is: X3-Xx-Y -X4-X2-Y, wherein X3or X4is Lys, Om, HLys, p2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y. PRM may be a PRM1 or a PRM2. For example, PRM1 may comprise one of: X-His-His, His-X-His, His-His-X, X-His-His-His, His-X-His-His, His-His-X-His, His-His-His-X, X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, or His-His-His-His-X, wherein X is Lys, Orn, HLys, p2-Lys, or p3-Lys. PRM1 may comprise His-Lys-His-His or His-His-Lys-His. In some cases PRM2 comprises one of SEQ ID NO. 32 to SEQ ID NO. 51. PRM2 may comprise Arg-Lys-Arg. The PRM may be covalently attached at a C-terminus of the antibody or antibody fragment. The PRM may be attached to a C-terminus of the antibody or antibody fragment through a linker. For example, the linker may comprise between 1 and 10 Pro- Ala dipeptide units. The linker may comprise between 1 and 10 Gly-Ser dipeptide units. For example, an antibody or antibody fragment comprising a polypeptide including a precision conjugation site may include a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide may consist of: a first amino acid sequence that has at least 95% identity with SEQ ID NO. 2, and a second amino acid sequence that has at least 95% identity with SEQ ID NO. 3 and a PRM1 or PRM2 sequence between the first amino acid sequence and the second amino acid sequence, wherein: the sequence of PRM1 is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the sequence of PRM2 is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Om, HLys, P2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

[0055] An antibody or antibody fragment may comprise a polypeptide including a precision-17- SG Docket No.: 14989-700.600conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a first amino acid sequence that has at least 95% identity with SEQ ID NO. 2, linked to a PRM sequence by some number (e.g., “v”) linking amino acids, where this number (“v”) is between 0 and 13, further wherein the PRM sequence consists of either a PRM1 sequence or a PRM2 sequence, further wherein: the PRM1 sequence is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Orn), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the PRM2 sequence is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Orn, HLys, p2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y. When v is nonzero, the linking amino acids may consist of positions 1 to v of SEQ. ID. NO. 613.

[0056] In some examples an antibody or antibody fragment comprises a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a first amino acid sequence that has at least 95% identity with SEQ ID NO. 4, linked to a PRM sequence by v linking amino acids, where v is between 0 and 13, wherein the PRM sequence consists of either a PRM1 sequence or a PRM2 sequence, further wherein: the PRM1 sequence is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Orn), homolysine (HLys), p2-lysine (P2-Lys), or P3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the PRM2 sequence is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Orn, HLys, P2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and P3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y. When v is nonzero, the linking amino acids may consist of positions 1 to v of SEQ. ID. NO. 613.

[0057] In some cases an antibody or antibody fragment may comprise a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a first amino acid sequence-18- SG Docket No.: 14989-700.600that has at least 95% identity with SEQ ID NO. 1, linked at a C-terminus to a PRM sequence either directly or by a linker, wherein the PRM sequence consists of either a PRM1 sequence or a PRM2 sequence, further wherein: the PRM1 sequence is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the PRM2 sequence is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Orn, HLys, p2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

[0058] The linker may comprise between 1 and 10 Pro- Ala dipeptide units. The linker may comprise between 1 and 10 Gly-Ser dipeptide units.

[0059] For example, an antibody or antibody fragment may include a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide has at least 90% identity with SEQ ID NO. 12, further wherein a sequence corresponding to PRM1 or PRM2 is either at a C-terminal end of the polypeptide or is located between positions 5 to 22 or positions 126 to 136 of SEQ ID NO. 12, either: in place of any single amino acid in positions 5 to 22 or positions 126 to 136 of SEQ ID NO. 12, or between any two consecutive amino acids in positions 5 to 22 or positions 126 to 136 of SEQ ID NO. 12, wherein: the sequence of PRM1 is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the sequence of PRM2 is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Orn, HLys, p2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and P3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y. The PRM may be a PRM1. As mentioned, PRM1 may comprise one of: X-His-His, His-X-His, His-His-X, X-His-His-His, His-X-His-His, His-His-X-His, His-His-His-X, X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, or His-His-His-His-X, wherein X is Lys, Orn, HLys, p2-Lys, or p3-Lys. In some cases PRM1 comprises His-Lys-His-His or His-His-Lys-His. The PRM may be PRM2. For example, PRM2 may comprise one of SEQ ID NO. 32 to SEQ ID-19- SG Docket No.: 14989-700.600NO. 51. PRM2 may comprise Arg-Lys-Arg. The PRM may be covalently attached at a C-terminus of the antibody or antibody fragment. In some cases the PRM may be attached to a C-terminus of the antibody or antibody fragment through a linker; the linker may comprise between 1 and 10 Pro- Ala dipeptide units. The linker may comprise between 1 and 10 Gly-Ser dipeptide units.

[0060] An antibody or antibody fragment comprising a polypeptide including a precision conjugation site may include a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a first amino acid sequence that has at least 95% identity with SEQ ID NO. 15, and a second amino acid sequence that has at least 95% identity with SEQ ID NO.16 and a PRM1 or PRM2 sequence between the first amino acid sequence and the second amino acid sequence, wherein: the sequence of PRM1 is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the sequence of PRM2 is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Orn, HLys, p2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

[0061] In some cases an antibody or antibody fragment may comprise a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a PRM sequence linked to a second amino acid sequence that has at least 95% identity with SEQ ID NO. 14, by v linking amino acids, where v is between 0 and 18, further wherein the PRM sequence consists of either a PRM1 sequence or a PRM2 sequence, further wherein: the PRM1 sequence is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Orn), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the PRM2 sequence is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Om, HLys, p2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y. When v is nonzero, the linking amino acids-20- SG Docket No.: 14989-700.600consist of positions (19-v) to 18 of SEQ. ID. NO. 614.

[0062] Also described herein are antibody or antibody fragment comprising a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a first amino acid sequence that has at least 95% identity with SEQ ID NO. 12, linked at a C-terminus to a PRM sequence either directly or by a linker, wherein the PRM sequence consists of either a PRM1 sequence or a PRM2 sequence, further wherein: the PRM1 sequence is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the PRM2 sequence is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Orn, HLys, p2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y. The linker may comprise between 1 and 10 Pro-Ala dipeptide units. The linker may comprise between 1 and 10 Gly-Ser dipeptide units.

[0063] Also described herein are method of selectively modifying a lysine residue within a polypeptide. In general, these methods may include contacting a compound (e.g. of formula PBU-L1-L2-PCU, as described herein) with a polypeptide (e.g., any one of the antibody or antibody fragments described herein) in the presence of a nickel (Ni2+) salt, cobalt (Co2+) salt, copper (Cu2+) salt, or zinc (Zn2+) salt, in a suitable solvent; wherein the polypeptide comprises at least one PRM1. The lysine residue that is selectively modified may be the lysine in the at least one PRM1.

[0064] A method of selectively modifying a lysine residue within a polypeptide may include: contacting a compound (e.g., of Formula (1)), with a polypeptide (e.g., any one of the antibody or antibody fragments described herein) in a suitable solvent in the absence of a transition metal salt; wherein the polypeptide comprises at least one PRM2. The lysine residue that is selectively modified may be the lysine in the at least one PRM2.

[0065] Other objects, features and advantages of the compounds, methods and compositions described herein will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples, while indicating specific embodiments, are given by way of illustration only, since various changes and modifications within the spirit and scope of the instant disclosure will become-21- SG Docket No.: 14989-700.600apparent to those skilled in the art from this detailed description.INCORPORATION BY REFERENCE

[0066] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.BRIEF DESCRIPTION OF THE FIGURES

[0067] FIG. 1 Illustration of how a test peptide containing two lysines, one being part of PRM1 and the other not, was prepared by oxidative dimerization of two smaller peptides, and possible products upon conjugation with compound 14. The sequence of PRM1 is underlined.

[0068] FIGS. 2A-2B Analysis of the reaction mixture of Example 4 by RP-HPLC and MS. (FIG. 2A) RP-HPLC analysis with UV detection at 220 nm. Starting materials and products from mono and bis conjugation by 14 are labeled. Only mono conjugated test peptide is observed, with no detection of bis conjugated test peptide. (FIG. 2B) Mass spectrum obtained via LC / MS analysis of the sample shown in FIG. 2A. As illustrated in Figure 1, (i) m / z = 4818 corresponds to mono conjugated test peptide, (ii) m / z = 3940 corresponds to unconjugated test peptide. The data shown in FIG. 2 illustrates specificity for PRM1 with a test peptide containing two lysines using one example of a proximity-triggered conjugation molecule, referred to herein as “Compound 14” which is described in greater detail in example 1, below, however the results shown with Compound 14 have been found to apply generally to the proximity-triggered conjugation molecules described herein.

[0069] FIG. 3 Illustration of how the mono conjugated test peptide containing two lysines, one being part of PRM1 and the other not, was cleaved by reduction of the disulfide and the site of conjugation confirmed by MS. The sequence of PRM1 is underlined. Peptide 1 is not conjugated and contains a lysine not associated with PRM1. Peptide 2 is conjugated and contains a lysine associated with PRM1. Peptide 3 is conjugated and contains a lysine not associated with PRM1. Peptide 4 is not conjugated and contains a lysine associated with PRM1. The presence of peptides 1 and 2 indicate conjugation of the lysine associated with PRM1. The presence of peptides 3 and 4 indicates conjugation of the lysine not associated with PRM1. FIG. 3. Illustrates the strategy for reductive cleavage of mono-conjugated test peptide containing PRM1 and identification of site of conjugation of compound 14.-22- SG Docket No.: 14989-700.600

[0070] FIG. 4 Analysis of the reductively cleaved product mixture of Example 4 by LC / MS. (i) LC / MS analysis. Peak assignment: A = TCEP; B = peptide 1 and peptide 2 from Figure 3 co-eluting, (ii) Mass spectrum of peak B from (i) confirming identify as co-eluting peptide 1 and peptide 2. Peptides 2 (m / z 2983) and 1 (m / z 1837) are the only products from reductive cleavage of the conjugated test peptide, indicating the conjugation is exclusively at the lysine associated with PRM1. FIG. 4 illustrates reductive cleavage of mono-conjugated test peptide containing PRM1 and identification of site of conjugation of compound 14.

[0071] FIG. 5 Illustration of how a test peptide containing two lysines, one being part of PRM2 and the other not, was prepared by oxidative dimerization of two smaller peptides, and possible products upon conjugation with compound 14. The sequence of PRM2 is underlined. FIG. 5. Shows an example of the assembly of a test peptide to demonstrate Compound 14 specificity for PRM2 in a single peptide containing two Lys.

[0072] FIGS. 6A-6B Analysis of the reaction mixture of Example 6 by RP-HPLC and MS. (FIG. 6A) RP-HPLC analysis. Starting materials and products from mono and bis conjugation by 14 are labeled as indicated in the figure. The mono conjugation product yield is significantly greater versus that of the bis conjugation product, and the latter is not observed with a reaction time of 16 hours. (FIG. 6B) Mass spectrum obtained via LC / MS analysis of the sample shown in (A). As illustrated in Figure 1, (i) m / z = 4702 corresponds to mono conjugated test peptide, (ii) m / z = 5580 corresponds to bis conjugated test peptide. FIGS. 6A-6B illustrate compound 14’s specificity for PRM2 with a test peptide containing two lysines.

[0073] FIG. 7. Illustration of how the mono conjugated test peptide containing two lysines, one being part of PRM2 and the other not, was cleaved by reduction of the disulfide and the site of conjugation confirmed by MS. The sequence of PRM2 is underlined. Peptide 1 is not conjugated and contains a lysine not associated with PRM2. Peptide 2 is conjugated and contains a lysine associated with PRM2. Peptide 3 is conjugated and contains a lysine not associated with PRM2. Peptide 4 is not conjugated and contains a lysine associated with PRM2. The presence of peptides 1 and 2 indicate conjugation of the lysine associated with PRM2. The presence of peptides 3 and 4 indicate conjugation of the lysine not associated with PRM2. FIG. 7 illustrates a strategy for reductive cleavage of mono-conjugated test peptide containing PRM2 and identification of site of conjugation of compound 14.

[0074] FIG. 8. LC / MS analysis of the reductively cleaved reaction mixture of Example 6 by LC / MS. (i) RP-HPLC analysis. Peak assignment: A = TCEP; B = peptide 4 from Figure 3; C = peptide 2 from Figure 3; D = peptide 1 from Figure 3; E = peptide 3 from Figure 3; F-23- SG Docket No.: 14989-700.600= unreacted compound 14. (ii) Mass spectrum of peak B from (i) confirming identify as peptide 4. (iii) Mass spectrum of peak C from (i) confirming identify as peptide 2. (iv) Mass spectrum of peak D from (i) confirming identify as peptide 1. (v) Mass spectrum of peak E from (i) confirming identify as peptide 3. Peptides 2 (m / z 2867) and 1 (m / z 1837) are by far the major products from reductive cleavage of the conjugated test peptide, indicating the conjugation is primarily at the lysine associated with PRM2. FIG. 8 illustrates reductive cleavage of mono-conjugated test peptide containing PRM2 and identification of site of conjugation.

[0075] FIG. 9. Illustration of how a mixture of 2 test peptides, one containing PRM1 and another containing PRM2, are conjugated to compound 14 as per the protocol described in Example 7, and the identity of the reacting peptide(s) can be determined by MS analysis of the product(s). Peptide 1 contains PRM1, and the sequence of PRM1 is indicated by a line placed over the top of the sequence. Peptide 2 contains PRM2, and the sequence of PRM2 is indicated by underline. FIG. 9. Shows an experiment to demonstrate exclusive conjugation to PRM2 using a mixture of two peptides, one containing PRM1 and the other containing PRM2.

[0076] FIG. 10. Analysis of the product mixture of Example 7 by LC / MS. (i) RP-HPLC analysis. Peak assignment: A = peptide 1, B = peptide 2, C = peptide 4 (peptide sequence identities shown in FIG. 9); D = unreacted compound 14. Chromatogram identities: (a) peptide 1 unreacted, (b) peptide 2 unreacted, (c) peptide 1 treated with compound 14 as described in Example 7, (d) peptide 2 treated with compound 14 as described in Example 7, (e) mixture of peptide 1 and peptide 2 treated with compound 14 as described in Example 7. (ii) Mass spectrum of peak C from (i) confirming the identify as peptide 4. The presence of peptides 3 is not detected, indicating the conjugation of compound 14 occurs exclusively at PRM2, and not at PRM1. FIG. 10 shows compound 14’s Specificity for a peptide containing PRM2 in the presence of a second peptide that contains PRM1.

[0077] FIG. 11. Schematic representation of conjugation of compound 14 to an Fc polypeptide.

[0078] FIG. 12. Mass spectrum of the reaction mixture obtained by treating the Fc polypeptide with compound 14 as described in FIG. 11, Example 9, and Example 10.

[0079] FIG. 13. Peptide Mapping Results of the conjugation of an Fc polypeptide with compound 14 of the invention as described in Example 9. The square symbol (□) indicates the major site of conjugation. The circle symbol (o) indicates lysine residues showing no-24- SG Docket No.: 14989-700.600evidence of conjugation. The triangle symbol (A) indicates minor sites of conjugation.Observed proteolytic fragments are indicated by the rectangles under the Fc sequence. The LC retention time of the fragments is indicated by the value inside the rectangle.

[0080] FIG. 14. Example diagram showing example of selectively modifying a target amino acid of a polypeptide (protein) according to some embodiments.

[0081] FIG. 15. Analysis of the reaction mixtures of Example 11 by RP-HPLC, showing conjugation of compound 14 to different His / Lys sequences. The sequence of test peptides is listed above each chromatogram trace. *, indicates peak corresponding to unconjugated peptide. **, indicates peak corresponding to mono-conjugated peptide. ***, indicates peak corresponding to multi-conjugated peptide. +, indicates peak corresponding to unreacted compound 14. Peak detection by UV at 220nm, and identify confirmed by LC / MS.

[0082] FIGS. 16A-16D. Conjugation of compound 14 to different bis-Arg sequences in the absence of metal ion. FIG. 16A shows sequences of peptides analyzed to understand structure-activity relationship of PRM2 design. FIG. 16B shows Representative RP-HPLC chromatograms of products from conjugation of compound 14 to no peptide, peptide 1, and peptide 10. Peaks corresponding to unreacted peptide, conjugate of peptide and compound 14, and unreacted compound 14 are indicated. FIG. 16C is a graph showing change of ratio of (relative peak area for conjugate peak) / (relative peak are for unreacted peptide peak) over time for peptides 2-7, showing the effect of varying the number of residues between the 2 arginine residues of PRM2. FIG. 16D is a graph showing change of ratio of (relative peak area for conjugate peak) / (relative peak are for unreacted peptide peak) over time for peptides 7-10, showing the effect of varying the number of residues between the lysine and the proximal arginine residues of PRM2. RP-HPLC peak detection was by UV at 220 nm, and peak identify was confirmed by LC / MS.

[0083] FIGS. 17A-17C Analysis of the reaction mixtures of Example 13 by RP-HPLC. FIG. 17A shows chemical structures of compounds 14, 15 and 16. This example shows a comparison of the conjugation efficiencies of proximity-triggered conjugation molecules (compounds 14, 15 and 16) with differing spacer moi eties. FIG. 17B shows RP-HPLC traces of conjugation products after 7 hours reaction time. FIG. 17C shows an overlay of RP-HPLC traces of conjugation products after 24 hours reaction time. *, indicates peak corresponding to unconjugated peptide. **, indicates peak corresponding to conjugated peptide. +, indicates peak corresponding to unreacted compound 14, 15, or 16. Peak detection by UV at 220nm, and identify confirmed by LC / MS chromatograms of reaction products.-25- SG Docket No.: 14989-700.600

[0084] FIG. 18 Diagram showing an example of modifying a protein with two distinct cargo molecules (i.e., dual cargo) using examples of a PRM1 and a PRM2. “X” represents an electrophilic warhead and “Y” represents a functional group left after cleavage of CLink that has subsequently been used to covalently attach a cargo molecule. In this example, the PRM1 include HKHH and the PRM2 includes KXXRXXR, where “X” represents a variable amino acid. In this example, the proximity-triggered conjugation compound is compound 14; however, any proximity conjugating compound may be used.

[0085] FIG. 19A illustrates an example of a polypeptide (e.g., in this example, an IgG light chain constant region) that may be modified to include a PRM insertion site, and in particular in the regions identified. FIG. 19B illustrates modifications of the polypeptide of FIG. 19A to include PRM sites at various locations.

[0086] FIGS. 20A-20B illustrate an example of a polypeptide (e.g., an IgG heavy chain constant region), indicating regions in which a PRM insertion site could be inserted, either by replacing one of the amino acid residues in the identified regions or by inserting between them, with the PRM. The Sequence listing is continued from FIG. 20A to FIG. 20B.DETAILED DESCRIPTION

[0087] Precision conjugation in a controlled, efficient, reliable, and tailorable fashion for broad and advanced therapeutic uses is challenging. Site-selective modification of polypeptides (such as proteins) is desired for the development of biopharmaceutical conjugates for use as novel therapeutics (e.g., antibody-drug conjugates, PEGylation, lipidation, etc.), bioimaging agents, and material sciences. For example, the Drug-to-Antibody Ratio (DAR) is a critical parameter in the development of antibody-drug conjugates (ADCs), a class of targeted cancer therapies. ADCs combine the specificity of monoclonal antibodies with the cytotoxic effects of potent drugs. The DAR quantifies the number of drug molecules attached to an antibody molecule within an ADC. It plays a crucial role in determining the ADC's pharmacokinetics, efficacy, and safety profile. The DAR can impact the ADC's pharmacokinetics (PK), which includes parameters like circulation time in the body, distribution, and elimination. Lower DAR values may result in longer circulation times, while higher DAR values may lead to faster clearance from the bloodstream. The DAR can also influence the efficacy of the ADC. A DAR that is too low may not deliver enough drug to target cells, while a DAR that is too high may lead to increased non-specific toxicity. The optimal DAR seeks to balance these factors.-26- SG Docket No.: 14989-700.600

[0088] Among the approximately 20 primary amino acids that compose proteins, only a subset can serve as appropriate targets for bioconjugation. Many strategies have been developed to target nucleophilic natural amino acid residues, of which cysteine and lysine residues are the most popular labeling sites.

[0089] Cysteine is the most intrinsically reactive amino acid and is capable of being involved in a chemo selective reaction due to its superior nucleophilic properties. Lysine is one of the most prevalent amino acids in the proteome, with ~650,000 lysine residues distributed among ~20,000 human proteins. By contrast, there are only ~260,000 cysteines, many of which are engaged in structural disulfide bonds and therefore are non-nucleophilic.

[0090] Most proteins display multiple copies of the same sidechain at different locations. Reactions that target a particular functional group, e.g., the thiol in Cys or the primary amine in Lys, potentially modify all occurrences of this residue, leading to the formation of a heterogeneous mixture of modified protein. The most commonly used reagent for conjugation to cystine residues, maleimide, creates a thio-ether Michael -addition product that often exhibits instability in vivo due to the possibility of a retro-Michael-addition reaction (P. L Ross, J. Wolfe, Physical and Chemical Stability of Antibody Drug Conjugates: Current Status, J Pharm Set, 2016 Feb;105(2):391-397). This instability often reduces the therapeutic efficacy of protein conjugates and causes off-target toxicities.

[0091] In contrast to the nucleophilicity of thiol in Cys, the primary amine in Lys is less nucleophilic. Further, lysine is ~3 time more prevalent in the human proteome versus cysteine, and this abundance additionally makes site-specific conjugation of lysine residues difficult. Attempts have been made to site-specifically modify only the single N-terminal amine in a protein due to its greater nucleophilicity and easier deprotonation versus the 8-amino group of internal lysines (pKa ~8 vs. pKa ~10, respectively), but pH-selective acylation reactions are difficult to control, and heterogenous product mixtures are often obtained (Rosen, C., Francis, M. Targeting the N terminus for site-selective protein modification, Nat Chem Biol 13, 697-705 (2017)).

[0092] Reactions involving proteins must also proceed in aqueous solutions under mild pH and temperature, and these restrictive conditions greatly limit the type and number of chemical reactions that can be employed for protein conjugation. These factors make it challenging to modify proteins in a regioselective manner. Several strategies have been developed to address this challenge. Some methods rely on genetic code expansion, which allows for the site-specific, ribosomal incorporation of non-canonical amino acids that exhibit-27- SG Docket No.: 14989-700.600suitable functionalities for bioorthogonal chemistry ((e.g., see., F. Tian, et al., A general approach to site-specific antibody drug conjugates, Proc. Natl. Acad. Set. U.S.A. Ill (5) 1766-1771, (2014); Paresh Agarwal and Carolyn R. Bertozzi, Site-Specific Antibody-Drug Conjugates: The Nexus of Bioorthogonal Chemistry, Protein Engineering, and Drug Development, Bioconjugate Chem. 2015, 26, 2, 176-192). While scientifically elegant, genetic code expansion methods are complicated to carry out, lower yielding versus standard protein expression methods, and often problematic for large-scale manufacture. Other methods rely on site-directed mutagenesis to incorporate unpaired cysteine residues that can be used to conjugate thiol -reactive electrophiles (e.g., maleimide). R.G.E. Coumans, et al., A Platform for the Generation of Site-Specific Antibody-Drug Conjugates That Allows for Selective Reduction of Engineered Cysteines, Bioconjugate Chemistry 202031 (9), 2136-2146). When used for proteins that contain naturally occurring disulfide bonds, these methods require partial reduction of the protein after initial expression to selectively reduce only the engineered cysteine residues so they alone will react with the conjugation reagent. These methods require addition of an additional step to the manufacturing process, and partial reduction can be difficult to control depending on the redox reactivity of the naturally occurring disulfides. Other methods developed for regiospecific modification of proteins involve the use of enzymes (S. S. van Berkel, F. L. van Delft, Enzymatic strategies for (near) clinical development of antibody-drug conjugates, Drug Discovery Today: Technologies, Volume 30, December 2018, Pages 3-10). In these methods, conjugation of a protein occurs at a specific position of the proteins that is recognized by the enzyme. Some enzymes recognize discreet peptide sequences (e.g., sortase, tubulin tyrosine ligase, and microbial transglutaminase) while others recognize certain glycan residues attached to the protein (e.g., glycosyltransferase). Enzymatic reactions are often difficult to optimize for high yield since the enzyme also catalyzes the reverse reaction, often requiring that the enzyme be separated from the initially formed product, and so application to manufacture can be problematic. The length of the peptide recognition sequences for the enzymes that catalyze conjugation reactions to peptides is relatively long (5-13 amino acids), and so conjugation is restricted to protein N- or C-termini. Enzymes that catalyze conjugation reactions to glycans are obviously restricted to use with glycoproteins, as are chemical methods that attach synthetic compounds to glycans after glycan oxidation. Finally, another method has been developed that makes use of a peptide ligand to a binding site on an IgG protein to direct the reaction of an NHS-ester to specific solvent-exposed lysine residue (Q. Zhou etal., Site-Specific-28- SG Docket No.: 14989-700.600Antibody-Drug Conjugation through Glycoengineering, Bioconjugate Chem. 2014, 25, 3, 510-520). The lysine residue is conjugated to a linker that comprises a thiol, and this is used for further synthetic elaboration. This method requires partial reduction of the IgG to reduce only the conjugated cysteine and is only applicable to IgG proteins. Selective chemical methods applicable to proteins consisting of only canonical amino acids are an attractive alternative to methods requiring genetic code expansion of unnatural amino acids, as these proteins can be produced using standard, higher-yielding expression methods. Further, methods that are non-enzymatic, do not require partial reduction of the protein to facilitate reaction of electrophiles with thiols, do not require acylation of amines, and are not restricted to application with a limited number of proteins are preferred for the manufacture of a broad range of therapeutic proteins.

[0093] Described herein are compounds and strategies for the precise and site-specific conjugation of canonical amino acids within polypeptides, specifically lysine residues. The strategies described herein take advantage of a conjugation compound that comprises a weakly electrophilic group whose amine reactivity is sufficiently attenuated such that it reacts with the 8-amine of a lysine residue only when it is held in close proximity to the lysine by a proximal non-covalent binding interaction with the protein. The incorporation of suitable biocompatible amino acid sequences comprising canonical amino acids, one of which is lysine (i.e., peptidic recognition motifs (PRMs)), into proteins ensures little to no negative impact on the properties of the protein (e.g., biocompatibility, no immunogenicity, no reduction in activity, solubility, reactivity, etc.) due to non-specific conjugation of the conjugation compound to non-PRM-proximal lysine residues. Such PRMs facilitate non-covalent binding interactions with PRM binding units (PBUs) within the compounds described herein, thereby placing PRM conjugation units (PCUs) within the same compounds near the PRM lysine(s) and enabling proximity-induced (i.e., binding-induced) irreversible covalent reaction between PRM lysines and the suitable electrophile(s) of the conjugation compound.

[0094] Polyhistidine tags (His tags) are widely used for protein purification by immobilized-metal ion affinity chromatography (see Block, H. et al. Immobilized-metal affinity chromatography (IMAC): a review. Methods Enzymol. 463, 439-473 (2009)).Strategies have been described that incorporate a lysine into such His tags (See, e.g., C. Kofoed et al., Chem. Eur. J. 2022, 28, e202200147; M. C. Martos-Maldonado, et al., Nature Communications | (2018) 9:3307). However, such strategies have disadvantages such as,-29- SG Docket No.: 14989-700.600incorporation of tags at the N-terminus of peptides, only reactions limited to reactive acylating electrophiles (which also result in non-selective reactions with Cys residues and Lys residues). At least 6 His residues are used and the presence of multiple basic imidazole rings of the histidines adjusts the nucleophilicity and basicity of the Lys residue and provides limited selective Lys modification.

[0095] FIG. 14 shows a diagram illustrating an example of selectively modifying a target amino acid of a polypeptide (protein) according to some embodiments. A polypeptide (protein) has been modified to include a Peptide Recognition Motif (PRM), which includes a select string of amino acids. A proximity-triggered conjugation molecule includes a PRM Binding Unit (PBU) that is linked to a PRM Conjugation Unit (PCU) via a Cleaveable Linker (CLink). In some examples, the CLink includes a cleavable linker portion (referred to herein as L1) and a spacer linker portion (referred to herein as L2). When the proximity-triggered conjugation molecule comes into contact with the PRM modified polypeptide (protein) under certain conditions, the PBU non-covalently binds with the PRM. In this example, the PRM is a PRM1 type and the presence of a transition metal salt (e.g., Ni2+) enables the PBU to selectively form the non-covalent bond via transition metal complex with side chain histidines in the PRM1. The non-covalent binding of the PBU situates the PCU in close enough proximity to a target amino acid (e.g., lysine) of the PRM1 to cause covalent bonding between the PCU and an amino group (NH2) of the target amino acid. This proximity-induced bonding can prevent the PCU from interacting with other amino acid residues other than the target amino acid, thereby providing selective modification of the polypeptide (protein). After the PCU is covalently bonded to the PRM, the Cleaveable Linker (CLink) may be cleaved to release the PBU. Cargo (e.g., drug) or a click handle may then be attached to the PCU modified polypeptide (protein).

[0096] As discussed herein, the composition of each of the proximity-triggered conjugation molecule (i.e., each of the PCU, PBU and CLink) and the PRM may be chosen based on providing selective and efficient conjugation of the polypeptide via proximity-induced bonding. For example, the chemical composition of the PCU, CLink, PBU and PRM may be chosen based on the ability to selectively bond the PCU with the PRM only when the PBU is non-covalently bound with the PRM and when the PCU is held within a proximity-induced binding distance. In some examples, the proximity-induced binding distance is equal to or less than 50 angstroms (e.g., between 1-50 angstroms), equal to or less than 40 angstroms (e.g., between 1-40 angstroms), equal to or less than 30 angstroms (e.g., between-30- SG Docket No.: 14989-700.6001-30 angstroms), equal to or less than 20 angstroms, or equal to or less than 10 angstroms (e.g., between 1-10 angstroms).

[0097] In the example of FIG. 14, the PRM is a first type of PRM (PRM1), which includes a PBU that is configured to non-covalently bond with the PRM via a transition metal complex. Also described herein is a second type of PRM (PRM2), which includes a PBU that is configured to non-covalently bond with a PRM via an ionic bond (salt bridge). A PRM2 may enable non-covalent bonding with a PRM without the present of a metal catalyst.

[0098] In some examples, structure-activity relationship (SAR) analysis techniques are used as a basis to determine effective and optimal combinations of PCU, PBU, CLink and PRM compositions. For instance, FIGS. 15, 16 and 17 show experimental results indicating the extent of conjugation of various peptide sequences to His / Lys sequences. One or more of the amino acid sequences identified as having efficient conjugation results may be integrated into a PRM, which is engineered into the polypeptide, thereby providing site specificity within the polypeptide. This site specificity allows for superior control as to the location and number of cargo molecules added onto the polypeptide compared to techniques involving the use of naturally occurring or recombinant amino acid clusters. A detailed description of the results shown in FIGS. 15-17, as well as results of other analyses, are presented herein.

[0099] In FIG. 14, the proximity-triggered conjugation molecule is compound 14, described herein. However, the proximity-triggered conjugation molecules described herein are not limited to compound 14. For example, as described herein, the composition of each of the PBU, the CLink (including each of cleavable linker portion (L1) and spacer linker portion (L2)), and the PCU may vary. For instance, the compounds 15 and 16, shown in FIG. 17 A, are examples of proximity-triggered conjugation molecules having different spacer linker portions (L2).

[0100] In FIGS. 16C-16D, reactions were analyzed at t=0, 24 hr, 48 hr, and 72 hr. FIG. 16B shows representative RP-HPLC chromatograms of conjugations with no peptide, peptide 1, and peptide 10 at t=24 hr. As shown, the conjugation of peptide 1 (containing the PRM1 sequence HKHH) produced no conjugate with compound 14 due to the absence of metal ion, while the conjugation of peptide 10 (containing the PRM2 sequence RKR) did produce conjugate with compound 14. The ratio (relative peak area for conjugate peak) / (relative peak are for unreacted peptide peak) was used to quantitate the progress of conjugations of compound 14 to peptides 2-10. Fig. 16C shows the increase in the ratio (relative peak area for conjugate peak) / (relative peak are for unreacted peptide peak) over time for peptides 2-7.-31- SG Docket No.: 14989-700.600These peptides have progressively fewer residues between the two arginine residues of the PRM2. From this series, peptide 7 with sequence Ac-KAWRRGS-NH2 (two arginines immediately adjacent) proceeded most efficiently. FIG. 16D shows the increase in the ratio (relative peak area for conjugate peak) / (relative peak are for unreacted peptide peak) over time for peptides 7-10. These peptides all have no residues between the 2 arginine residues of PRM2 and have progressively fewer residues between the Lys residue of PRM2 and the proximal Arg. From this series, peptide 10 with sequence Ac-AWRKRGS-NH2 (Lys between the 2 arginines of PRM2) proceeded most efficiently.

[0101] Described herein is a versatile approach for the precise labelling of Lys residues in proteins that proceeds in high yield. As discussed above, the strategies disclosed herein utilize at least two different PRMs, each of which reacts with the compounds disclosed herein under different reaction conditions. As such, proteins can be labelled with at least two distinct moieties providing access to modified proteins in a rapid and high yielding manner.

[0102] Further, the described approach can be used to attach multiple copies of a compound to a protein by incorporating multiple copies of the PRM. For example, multiple PRMls and / or multiple PRM2s may be incorporated into a protein. The FIG. 18 shows an example of modifying a protein with a dual cargo using examples of a PRM1 and a PRM2. In this example, alkylation of PRM1 does not occur in the absence of Ni2+and alkylation of PRM2 may occur at a very low level in the presence of Ni2+(e.g. 40hr). In this example, PRM2 can be alkylated first in the absence of Ni2+so then PRM2 is masked during subsequent +Ni2+PRM1 alkylation.Biologically Active Polypeptides

[0103] Polypeptides described herein are biologically active polypeptides. Any therapeutic polypeptide is compatible with the compounds and strategies described herein.

[0104] The terms “polypeptide,” “peptide” and “protein” are used interchangeably herein to refer to a polymer of amino acid residues linked together by peptide bonds. That is, a description directed to a polypeptide applies equally to a description of a peptide and a description of a protein, and vice versa. The terms apply to naturally occurring amino acid polymers as well as amino acid polymers in which one or more amino acid residues is a nonnatural amino acid. Additionally, such “polypeptides,” “peptides” and “proteins” include amino acid chains of any length, including full length proteins, wherein the amino acid residues are linked by covalent peptide bonds. The amino acids may either be naturally occurring or non-naturally occurring; as will be appreciated by those in the art, any structure-32- SG Docket No.: 14989-700.600for which a set of rotamers is known or can be generated can be used as an amino acid. The side chains may be in either the (R) or the (S) configuration. In an embodiment, the amino acids are in the (S) or L- configuration.

[0001] Exemplary polypeptides include, but are not limited to therapeutic proteins, enzymes, receptors or ligands, cell surface receptors, ion channels (i.e., pore-forming membrane protein), kinases, blood proteins, chemokines, cytokines, antibodies, antibody fragments, antigens, anticoagulants, blood factors, bone morphogenetic proteins, growth factors, transcription factors, interferons, interleukins, thrombolytic agents, cytoskeletal proteins, and hormones. Suitable classes of enzymes include, but are not limited to, hydrolases such as proteases, carbohydrases, lipases; isomerases, such as racemases, epimerases, tautomerases, or mutases; transferases, kinases, oxidoreductases, and phosphatases. Suitable enzymes are listed in the Swiss-Prot enzyme database.

[0105] The term “biologically active polypeptide” refers to any polypeptide substance which can affect any physical or biochemical properties of a biological system, pathway, molecule, or interaction relating to an organism, including but not limited to, viruses, bacteria, bacteriophage, transposon, prion, insects, fungi, plants, animals, and humans. As used herein, biologically active polypeptides include but are not limited to any substance intended for diagnosis, cure, mitigation, treatment, or prevention of disease in humans or other animals, or to otherwise enhance physical or mental well-being of humans or animals. Classes of biologically active polypeptides that are suitable for use as therapeutics include, but are not limited to, imaging agents, antibiotics, fungicides, anti-virals, anti-inflammatory agents, anti-tumor agents, cardiovascular agents, anti-anxiety agents, hormones, growth factors, microbially derived toxins, and the like.

[0106] Proteins with their structures and sequences can be found at the Worldwide Protein Data Bank organization (wwPDB; wwpdb.org), which includes the RCSB Protein Data Bank (RCSB PDB; rcsb.org), the Protein Data Bank Japan (PDBj; pdbj.org), the Protein Data Bank in Europe (PDBe; pdbe.org), and BioMagResBank (BMRB; www.bmrb.wisc.edu).

[0107] In some embodiments, if polypeptides are described in the art and contemplated for use in the methods described herein then such polypeptides contemplated for use with the compounds described herein have at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the polypeptides known in the art.

[0108] As described herein, the term “percent (%) sequence identity,” and terms related thereto, in the context of amino acid sequences or nucleic acid sequences, is the percentage of-33- SG Docket No.: 14989-700.600amino acid residues or nucleic acid residues in a candidate sequence that are identical with the amino acid residues or nucleic acid residues, respectively, in a selected sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity. Alignment for purposes of determining percent amino acid sequence identity or percent nucleic acid identity can be achieved in various ways that are within the skill in the art, for instance, using publicly available computer software such as Clustal Omega, BLAST, BLAST-2, ALIGN, ALIGN-2 or Megalign (DNASTAR) software, with BLAST being the alignment algorithm of preference. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms needed to achieve maximal alignment over the full-length of the sequences being compared, although for simplicity it maybe preferred to use default parameters.

[0109] In some embodiments, the polypeptide comprises at least 10 amino acids. In some embodiments, the polypeptide comprises 10 or more, 15 or more, 20 or more, 25 or more, 30 or more, 35 or more, 40 or more, 45 or more, 50 or more, 55 or more, 60 or more, 65 or more, 70 or more, 75 or more, 80 or more, 85 or more, 90 or more, 95 or more, 100 or more amino acids.

[0110] In some embodiments, the polypeptide is 25 or more amino acids in length. In embodiments, the polypeptide is about 25 to about 2000 or more amino acids in length. In embodiments, the polypeptide is about or at least about 25, 30, 35, 40, 45, 50, 100, 150, 200, 250, 300, 350, 400, 450, 475, 500, 525, 550, 575, 600, 625, 650, 700, 750, 800, 850, 900, 950, 1000, 1200, 1400, 1600, 1800, or 2000 amino acids in length. In embodiments, the polypeptide is about: 25 to about 2000, 25 to about 1000, 25 to about 500, 25 to about 250, 25 to about 100, or 25 to about 50, amino acids in length.

[0111] In some embodiments, the therapeutic polypeptide has a molecular weight of at least about 1 kDa. In some embodiments, the therapeutic polypeptide has a molecular weight of about 0.5 kDa to about 5 kDa, about 5 kDa to about 10 kDa, about 10 kDa to about 20 kDa, about 20 kDa to about 30 kDa, about 30 kDa to about 60 kDa, about 60 kDa to about 110 kDa, or about 110 kDa to about 200 kDa, or greater than about 200 kDa.

[0112] In embodiments, the molecular weight of the polypeptide is about 0.5 kDa, 1 kDa, about 2 kDa, about 3 kDa, about 4 kDa, about 5 kDa, about 6 kDa, about 7 kDa, about 8 kDa, about 9 kDa, about 10 kDa, about 11 kDa, about 12 kDa, about 13 kDa, about 14 kDa, about 15 kDa, about 16 kDa, about 17 kDa, about 18 kDa, about 19 kDa, about 20 kDa, about 30-34- SG Docket No.: 14989-700.600kDa, about 40 kDa, about 50 kDa, about 60 kDa, about 70 kDa, about 80 kDa, about 90 kDa, about 100 kDa, about 150 kDa, about 200 kDa, about 250 kDa, about 300 kDa, about 350 kDa, about 400 kDa, about 450 kDa, about 500 kDa, or more. In embodiments, the molecular weight of the polypeptide is about 1 to about 10 kDA, about 1 to about 20 kDA, about 1 to about 30 kDA, about 1 to about 40 kDA, about 1 to about 50 kDA, about 1 to about 60 kDA, about 1 to about 70 kDA, about 1 to about 80 kDA, about 1 to about 90 kDA, about 1 to about 100 kDA, about 1 kDa to about 200 kDa, about 1 kDa to about 300 kDa, about 1 kDa to about 400 kDa, about 1 kDa to about 500 kDa, about 2 to about 10 kDA, about 2 to about 20 kDA, about 2 to about 30 kDA, about 2 to about 40 kDA, about 2 to about 50 kDA, about 2 to about 60 kDA, about 2 to about 70 kDA, about 2 to about 80 kDA, about 2 to about 90 kDA, about 2 to about 100 kDA, about 2 kDa to about 200 kDa, about 2 kDa to about 300 kDa, about 2 kDa to about 400 kDa, about 2 kDa to about 500 kDa, about 3 to about 10 kDA, about 3 to about 20 kDA, about 3 to about 30 kDA, about 3 to about 40 kDA, about 3 to about 50 kDA, about 3 to about 60 kDA, about 3 to about 70 kDA, about 3 to about 80 kDA, about 3 to about 90 kDA, about 3 to about 100 kDA, about 3 kDa to about 200 kDa, about 3 kDa to about 300 kDa, about 3 kDa to about 400 kDa, or about 3 kDa to about 500 kDa.

[0113] In some embodiments, the therapeutic polypeptide has a molecular weight between about 0.1 kDa to about 100 kDa. In some embodiments, the therapeutic polypeptide has a molecular weight between about 100 Da and about 100,000 Da or more. In some embodiments, the therapeutic polypeptide has a molecular weight between about 1,000 Da and about 200,000 Da, including but not limited to, about 100,000 Da, about 95,000 Da, about 90,000 Da, about 85,000 Da, about 80,000 Da, about 75,000 Da, about 70,000 Da, about 65,000 Da, about 60,000 Da, about 55,000 Da, about 50,000 Da, about 45,000 Da, about 40,000 Da, about 35,000 Da, about 30,000 Da, about 25,000 Da, about 20,000 Da, about 15,000 Da, about 10,000 Da, about 9,000 Da, about 8,000 Da, about 7,000 Da, about 6,000 Da, about 5,000 Da, about 4,000 Da, about 3,000 Da, about 2,000 Da, and about 1,000 Da. In some embodiments, the therapeutic polypeptide has a molecular weight between about 1,000 Da and about 40,000 Da. In some embodiments, the therapeutic polypeptide has a molecular weight between about 2,000 to about 50,000 Da. In some embodiments, the therapeutic polypeptide has a molecular weight between about 5,000 Da and about 40,000 Da. In some embodiments, the therapeutic polypeptide has a molecular weight between about 10,000 Da and about 40,000 Da.-35- SG Docket No.: 14989-700.600

[0114] In some embodiments, the therapeutic polypeptide is peptide hormone, protein hormone, bacterial / antibiotic peptide, antifungal peptide, invertebrate peptide, venom derived peptide, anticancer peptide, vaccine peptide, anti-inflammatory peptide, neuropeptide, endocrine peptide, gastrointestinal peptide, cardiovascular peptide, renal peptide, respiratory peptide, opioid peptide, neurotrophic peptide.

[0115] In some embodiments, the therapeutic polypeptide is a cytokine. Non-limiting examples of cytokine may include 4-1BBL, acylation stimulating protein, adipokine, albinterferon, APRIL, Arh, BAFF, Bcl-6, CCL1, CCL1 / TCA3, CCL11, CCL12 / MCP-5, CCL13 / MCP-4, CCL14, CCL15, CCL16, CCL17 / TARC, CCL18, CCL19, CCL2, CCL2 / MCP-1, CCL20, CCL21, CCL22 / MDC, CCL23, CCL24, CCL25, CCL26, CCL27, CCL28, CCL3, CCL3L3, CCL4, CCL4L1 / LAG-1, CCL5, CCL6, CCL7, CCL8, CCL9, CCR10, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CD153, CD154, CD178, CD40LG, CD70, CD95L / CD178, Cerberus (protein), chemokines, CLCF1, CNTF, colony-stimulating factor, common b chain (CD131), common g chain (CD132), CX3CL1, CX3CR1, CXCL1, CXCL10, CXCL11, CXCL12, CXCL13, CXCL14, CXCL15, CXCL16, CXCL17, CXCL2, CXCL2 / MIP-2, CXCL3, CXCL4, CXCL5, CXCL6, CXCL7, CXCL9, CXCR3, CXCR4, CXCR5, EDA-A1, Epo, erythropoietin, FAM19A1, FAM19A2, FAM19A3, FAM19A4, FAM19A5, Flt-3L, FMS-like tyrosine kinase 3 ligand, Foxp3, GATA-3, GcMAF, G-CSF, GITRL, GM-CSF, granulocyte colony-stimulating factor, granulocyte-macrophage colonystimulating factor, hepatocyte growth factor, IFNA1, IFNA10, IFNA13, IFNA14, IFNA2, IFNA4, IFNA5 / IFNaG, IFNA7, IFNA8, IFNB1, IFNE, IFNG, IFNZ, IFN-a, IFN-P, IFN-y, IFNco / IFNWl, IL-1, IL-10, IL-10 family, IL-10-like, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-17 family, IL-17A-F, IL-18, IL-18BP, IL-19, IL-1A, IL-1B, IL-1F10, IL-1F3 / IL-1RA, IL-1F5, IL-1F6, IL-1F7, IL-1F8, IL-1F9, IL-l-like, IL-IRA, IL-1RL2, IL-la, IL-ip, IL-2, IL-20, IL-21, IL-22, IL-23, IL-24, IL-28 A, IL-28B, IL-29, IL-3, IL-31, IL-33, IL-35, IL-4, IL-5, IL-6, IL-6-like, IL-7, IL-8 / CXCL8, IL-9, inflammasome, interferome, interferon, interferon beta-la, interferon beta-lb, interferon gamma, interferon type I, interferon type II, interferon type III, interferons, interleukin, interleukin 1 receptor antagonist, Interleukin 8, IRF4, Leptin, leukemia inhibitory factor (LIF), leukocytepromoting factor, LIGHT, LTA / TNFB, LT-P, lymphokine, lymphotoxin, lymphotoxin alpha, lymphotoxin beta, macrophage colony-stimulating factor, macrophage inflammatory protein, macrophage-activating factor, M-CSF, MHC class III, miscellaneous hematopoietins, monokine, MSP, myokine, myonectin, nicotinamide phosphoribosyltransferase, oncostatin M-36- SG Docket No.: 14989-700.600(OSM), oprelvekin, OX40L, platelet factor 4, promegapoietin, RANKL, SCF, STAT3, STAT4, STAT6, stromal cell-derived factor 1, TALL-1, TBX21, TGF-a, TGF-P, TGF-P 1, TGF-P2, TGF-P3, TNF, TNFSF10, TNFSF11, TNFSF12, TNFSF13, TNFSF14, TNFSF15, TNFSF4, TNFSF8, TNF-a, TNF-P, Tpo, TRAIL, TRANCE, TWEAK, vascular endothelial growth inhibitor, XCL1, or XCL2. Example of immune checkpoint protein may include VISTA, PD-L1, CTLA-4, PD-L2, B7-1 (CD80), B7-2 (CD86), B7-H3 (CD276), B7-H2, B7-H4 (VTCN1), HVEM (CD270, TNFRSF14), Galectin 9, Galectin3, CEACAM1 (CD66a), OX-2 (CD200), PVR (CD155), PVRL2 (Nectin-2, CD112), FGL-1, PEC AM-1, TSG-6, CD47, Stabilin-1 (Clever- 1), Neuropilin 1, Neuropilin 2, CD 158 (family), IGSF2 (CD 101), CD155, GITRL, CD137L, OX40L, LIGHT, CD70, PD-1, RGMB, CTLA-4 (CD152), BTLA, CD160, Tim-3, CD200R, TIGIT, CD112R (PVRIG), LAG-3 (CD223), PECAM-1, CD44, SIRP alpha (CD 172a), or IGSF11.

[0116] In embodiments, the polypeptide is selected from: Human Antihemophilic Factor; Human Antihemophilic Factor-von Willebrand Factor Complex; Recombinant Antihemophilic Factor (Turoctocog Alfa); Ado-trastuzumab emtansine; Albiglutide;Alglucosidase Alpha; Human Alpha- 1 Proteinase Inhibitor; Botulinum Toxin Type B (Rimabotulinumtoxin B); Coagulation Factor IX Fc Fusion; Recombinant Coagulation factor IX; Recombinant Coagulation factor Vila; Recombinant Coagulation factor XIII A-subunit; Human Coagulation Factor Vlll-von Willebrand Factor Complex; Collagenase Clostridium Histolyticum; Human Platelet-derived Growth Factor (Cecaplermin); Abatacept; Abciximab; Adalimumab; Aflibercept; Agalsidase Beta; Aldesleukin; Alefacept; Alemtuzumab;Alglucosidase Alfa; Alteplase; Anakinra; Octocog Alfa; Recombinant Human Antithrombin; Azficel-T; Basiliximab; Belatacept; Belimumab; Bevacizumab; Botulinum Toxin Type A; Brentuximab Vedotin; Recombinant Cl Esterase Inhibitor; Canakinumab; Certolizumab Pegol; Cetuximab; Nonacog Alfa; Daclizumab; Darbepoetin Alfa; Denosumab; Digoxin Immune Fab; Dornase Alfa; Ecallantide; Eculizumab; Etanercept; Fibrinogen; Filgrastim; Galsulfase; Golimumab; Ibritumomab Tiuxetan; Idursulfase; Infliximab; Interferon Alfa; Interferon Alfa-2b; Interferon Alfacon-1; Interferon Alfa-2a; Interferon Alfa-n3; Interferon Beta-la; Interferon Beta-lb; Interferon Gamma-lb; Ipilimumab; Laronidase; Epoetin Alfa; Moroctocog Alfa; Murom onab-CD3; Natalizumab; Ocriplasmin; Ofatumumab; Omalizumab; Oprelvekin; Palifermin; Palivizumab; Panitumumab; Pegfilgrastim; Pertuzumab; Human Papilloma Virus (HPV) Types 6;11 ; 16; 18-L1 viral protein Virus like Particles (VLP); HPV Type 16 and 18 LI protein VLPs; Ranibizumab; Rasburicase; Raxibacumab; RecombinantSG Docket No.: 14989-700.600Factor IX; Reteplase; Rilonacept; Rituximab; Romiplostim; Sargramostim; Tenecteplase; Tocilizumab; Trastuzumab; Ustekinumab; Abarelix; Cetrorelix; Desirudin; Enfuvirtide; Exenatide; Follitropin Beta; Ganirelix; Degarelix; Hyaluronidase; Insulin Aspart; Insulin Degludec; Insulin Detemir; Insulin Glargine; Recombinant Insulin Glulisine; Human Insulin; Insulin Lispro; Recombinant Insulin Lispro Protamine; Recombinant Insulin Lispro;Lanreotide; Liraglutide; Surfaxin (Lucinactant; Sinapultide); Mecasermin; Insulin like Growth Factor; Nesiritide; Pramlintide; Recombinant Teduglutide; Tesamorelin Acetate; Goserelin; AbobotulinumtoxinA; Agalsidase Alfa; Alipogene Tiparvovec; Ancestim;Anistreplase; Ardeparin Sodium; Avian TB Vaccine; Batroxobin; Bivalirudin; Buserelin (Gonadotropin-releasing Hormone Agonist); Cabozantinib S-Malate; Carperitide;Catumaxomab; Ceruletide; Coagulation Factor VIII; Coccidiosis Vaccine; Dalteparin Sodium; Deferiprone; Defibrotide; Dibotermin Alfa; Drotrecogin Alfa; Edotreotide;Efalizumab; Enoxaparin Sodium; Epoetin Delta; Eptifibatide; Eptotermin Alfa; Follitropin Alfa for Injection; Fomivirsen; Gemtuzumab ozogamicin; Gonadorelin; Recombinant Chorionic Human Gonadotropin; Histrelin Acetate (gonadotropin releasing hormone agonist); HVT IBD vaccine; Imiglucerase; Insulin Isophane; Lenograstim (Granulocyte-Colony Stimulating Factor); Lepirudin; Leptospira Vaccine for Dogs; Leuproprelin;Linaclotide; Lipegfilgrastim; Lixisenatide; Lutropin Alfa (human leutinizing hormone); Mepolizumab; Mifamurtide; Mipomersen Sodium; Mirimostim (macrophage-colony stimulating factor); Mogamulizumab; Molgramostim (granulocyte macrophage-colony stimulating factor); Monteplase; Nadroparin calcium; Nafarelin; Nebacumab; Octreotide; Pamiteplase; Pancrelipase; Pamaparin sodium; Pasireotide daspartate; Peginesatide acetate; Pegvisomant; Pentetreotide; Poractant alfa; Pralmorelin (growth hormone releasing peptide); Protirelin; PTH 1-84; rhBMP-2; rhBMP-7; Eptortermin Alfa; Romurtide; Sermorelin;Somatostatin; Somatrem; Vassopressin; Desmopressin; Taliglucerase Alfa; Taltirelin (thyrotropin-releasing hormone analog); Tasonermin; Taspoglutide; Thromobomodulin Alfa; Thyrotropin Alfa; Traf ermin; Triptorelin Pamoate; Urofollitropin for Injection; Urokinase; Velaglucerase Alfa; Cholera Toxin B; Recombinant Antihemophilic Factor (Efraloctocog Alfa); Human Alpha- 1 Proteinase Inhibitor; Asparaginase Erwinia Chrysanthemi; Capromab; Denileukin Diftitox; Ovine Digoxin Immune Fab; Elosulfase Alfa; Epoetin Alfa; Factor IX Complex; Factor XIII Concentrate; Technetium (Fanolesomab); Fibrinogen; Thrombin; Influenza Hemagglutinin and Neuraminidase; Glucarpidase; Hemin for Injection; Hep B Surface Antigen; Human Albumin; Incobotulinumtoxin; Nofetumomab; Obinutuzumab; L--38- SG Docket No.: 14989-700.600asparaginase (from Escherichia. coir, Erwinia sp:, Pseudomonas sp.,' etc.); Pembrolizumab; Protein C Concentrate; Ramucirumab; Siltuximab; Tbo-Filgrastim; Pertussis Toxin Subunits A-E; Topical Bovine Thrombin; Topical Human Thrombin; Tositumomab; Vedolizumab; Ziv-Aflibercept; Glucagon; Somatropin; Plasmodium falciparum or a Plasmodium vivax Antigen (e g., CSP, CelTOS, TRAP, Rh5, AMA-1, LSA-1, LSA-3, Pfs25, MSP-1, MSP-3, STARP, EXP1, pb9, GLURP). The sequences of these polypeptides, including variations, are available in the literature and known to those of skill in the art. Any known sequence of any of the polypeptides listed is contemplated for use in the methods of the present invention.

[0117] In some embodiments, the therapeutic polypeptide is a monoclonal antibody, polyclonal antibody, single-domain antibody (sdAb, VHH, or nanobody), VHH-Fc, heavy chain only antibody (HCAb), a single chain variable fragment (scFv), di-scFv, tri-scFv, F(ab) fragment antibody, F(ab')2 fragment antibody, bispecific antibody (trifunctional antibody, chemically linked Fab, bi-specific T-cell engager), microantibody, intrabody, affibody molecule, Aflfilin, Affimer, Affitin, Alphabody, Anticalin, Avimer, DARPin, Kunitz domain peptide, Monobody, nanoCLAMP.

[0118] In some embodiments, the polypeptide is an IgM antibody, IgD antibody, IgG antibody, IgA antibody, or IgE antibody.

[0119] In some embodiments, the polypeptide is an IgGl antibody, IgG2 antibody, IgG3 antibody, IgG4 antibody, IgAl antibody or IgA2 antibody.

[0120] In some embodiments, the polypeptide is an antibody fragment. The term “antibody fragment” refers to any form of an antibody other than the full-length form. Antibody fragments herein antibodies that are smaller components that exist within full-length antibodies, and antibodies that have been engineered, such as antibody variants. Antibody fragments include but are not limited to Fv, Fc, Fab, and (Fab')2, single chain Fv (scFv), diabodies, triabodies, tetrabodies, bifunctional hybrid antibodies, CDR1, CDR2, CDR3, combinations of CDRs, variable regions, framework regions, constant regions, heavy chains, single-domain antibodies (sdAb, VHH, or nanobodies), light chains, and variable regions, and alternative scaffold non-antibody molecules, bispecific antibodies, and the like (Maynard & Georgiou, Annu. Rev. Biomed. Eng. 2:339-76, 2000; Hudson, Curr. Opin. Biotechnol. 9:395-402, 1998). Another functional substructure is a single chain Fv (scFv), comprised of the variable regions of the immunoglobulin heavy and light chain, covalently connected by a peptide linker (Hu et al., Cancer Research, 56, 3055-3061, 1996). These small proteins generally retain specificity and affinity for antigen in a single polypeptide and can provide a-39- SG Docket No.: 14989-700.600convenient building block for larger, antigen-specific molecules.

[0121] The term “antigen” is well understood in the art and includes substances which are immunogenic, i.e., immunogens, as well as substances which induce immunological unresponsiveness, or anergy, i.e., anergens. Where the antigen is a polypeptide, it may be a transmembrane molecule (e.g., receptor) or ligand such as a growth factor. Exemplary antigens include molecules, such as renin; a growth hormone, including human growth hormone and bovine growth hormone; growth hormone releasing factor; parathyroid hormone; thyroid stimulating hormone; lipoproteins; alpha- 1 -antitrypsin; insulin A-chain; insulin B-chain; proinsulin; follicle stimulating hormone; calcitonin; luteinizing hormone; glucagon; clotting factors, such as factor VIIIC, factor IX, tissue factor (TF), and von Willebrands factor; anti- clotting factors such as Protein C; atrial natriuretic factor; lung surfactant; a plasminogen activator, such as urokinase or human urine or tissue-type plasminogen activator (t-PA); bombesin; thrombin; hemopoietic growth factor; tumor necrosis factor-alpha and -beta; enkephalinase; RANTES (regulated on activation normally T-cell expressed and secreted); human macrophage inflammatory protein (MIP-I -alpha); a serum albumin such as human serum albumin; Muellerian-inhibiting substance; relaxin A-chain; relaxin B-chain; prorelaxin; mouse gonadotropin-associated peptide; a microbial protein, such as beta-lactamase; DNase; IgE; a cytotoxic T-lymphocyte associated antigen (CTLA), such as CTLA-4; inhibin; activin; vascular endothelial growth factor (VEGF); receptors for hormones or growth factors; protein A or D; rheumatoid factors; a neurotrophic factor such as bone-derived neurotrophic factor (BDNF), neurotrophin-3, -4, - 5, or -6 (NT-3, NT-4, NT-5, or NT-6), or a nerve growth factor such as NGF-P; platelet derived growth factor (PDGF); fibroblast growth factor such as aFGF and bFGF; epidermal growth factor (EGF); transforming growth factor (TGF), such as TGF-alpha and TGF-beta, including TGF-pi, TGF-PP2, TGF-PP3, TGF-PP4, or TGF-PP5; insulin-like growth factor-I and -II (IGF-I and IGF-II); des(l-3)-IGF-I (brain IGF-I), insulin-like growth factor binding proteins; CD proteins, such as CD3, CD4, CD8, CD 19 and CD20; erythropoietin; osteoinductive factors; immunotoxins; a bone morphogenetic protein (BMP); an interferon such as interferon-alpha, -beta, and -gamma; colony stimulating factors (CSFs), e.g., M-CSF, GM- CSF, and G-CSF; interleukins (ILs), e.g., IL-Ito IL-10; superoxide dismutase; T-cell receptors; surface membrane proteins; decay accelerating factor; viral antigen, such as, for example, a portion of the AIDS envelope; transport proteins; homing receptors; addressins; regulatory proteins; integrins, such as CD Ila, CD lib, CD lie, CD 18, an ICAM, VLA-4 and VCAM; a tumor-40- SG Docket No.: 14989-700.600associated antigen, such as HER2, HER3 or HER4 receptor; and fragments of any of the above-listed polypeptides.

[0122] Complementarity determining regions (CDRs) generally include amino acids (e.g., non-contiguous or contiguous) within antibody variable regions that confer antigen specificity and / or binding affinity. In general, there are three CDRs in each heavy chain variable region (CDR-H1, CDR-H2, CDR-H3) and three CDRs in each light chain variable region (CDR-L1, CDR-L2, CDR-L3). Framework regions (FRs) generally refer to and / or include non-CDR regions of the heavy and light chain variable regions. In general, there are four FRs in each full-length heavy chain variable region (FR-H1, FR-H2, FR-H3, and FR-H4), and four FRs in each full-length light chain variable region (FR-L1, FR-L2, FR-L3, and FR-L4).

[0123] Variable regions (also referred to as variable domains) generally refer to and / or include the domain of an antibody heavy or light chain that is involved in binding the antibody to antigen (e.g., a single variable domain comprises a CDR 1, CDR 2, and CDR 3). The variable domains of the heavy chain and light chain (VH and VL, respectively) of a native antibody generally have similar structures, with each domain comprising four conserved framework regions (FRs) and three CDRs. In certain instances, a single VH or VL domain can be sufficient to confer antigen-binding specificity.

[0124] Fc region generally encompasses and / or refers to a C-terminal region of an immunoglobulin heavy chain that contains at least a portion of the constant region. Generally, the Fc region includes an immunoglobulin CH2 and CH3 domain (e.g., an IgG CH2 and CH3 domain). The term includes native sequence Fc regions and variant Fc regions. In certain embodiments, the Fc region include IgG and sub-classes thereof (e.g., IgGl and IgG4), IgM, IgE, IgA, and / or IgD heavy chain constant regions and / or heavy chain constant regions derived from IgG and sub-classes thereof (e.g., IgGl and IgG4), IgM, IgE, IgA, and IgD.

[0125] Unless otherwise specified herein, numbering of amino acid residues in the Fc region or constant region is according to the EU numbering system, also called the EU index, as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD, 1991.

[0126] A nanobody is the variable domain of heavy-chain-only antibody (HcAbs) that was first isolated from the serum of Camelidae family. The nomenclature of “nanobody” originally adopted by the Belgian company Ablynx® stemmed from its nanometric size, i.e., 4 nm in length, 2.5 nm in width, and only 15 kD in molecular weight, which was attributed to-41- SG Docket No.: 14989-700.600the lack of the light chains (L) and heavy chain constant domain (CH) in contrast to the conventional monoclonal antibodies (mAbs) (Revets H, et al., Wolfson W. Ablynx makes nanobodies from llama bodies. Chem Biol. 2006;13(12): 1243-4; Muyldermans S, etal., Sequence and structure of VH domain from naturally occurring camel heavy chain immunoglobulins lacking light chains. Protein Eng. 1994;7(9): 1129-35). Nanobodies as novel agents for cancer therapy. Expert opinion on biological therapy. 2005;5(l): 111-24). The antigen-binding capacity of nanobodies, however, remains similar to that of conventional antibodies. The complementarity-determining region 3 (CDR3) of nanobodies is similar or even longer than that of human VH domain (variable domain of heavy immunoglobulin chain).Nanobodies can form finger-like structures to recognize cavities or hidden epitopes that are not available to mAbs, and exhibit excellent stability, hydrophilicity, and water solubility that help maintain their binding affinity across different conditions, which can be further reinforced by mutating key AAs in the framework region.

[0127] In some embodiments, the polypeptide is an antigen-binding fragment. The term “antigen-binding fragment,” as used herein, refers to one or more fragments of an antibody that retain the ability to bind to an antigen. Antigen-binding fragments generally refer to and / or include antibody-derived proteins that comprise a functional set of CDRs (e.g., a CDR-H1-3 and CDR-L1-3) and have a molecule weight less than a full-length IgG antibody (e.g., a molecular weight less than -150,00 Daltons). It has been shown that the antigenbinding function of an antibody can be performed by fragments of an intact antibody.Examples of binding fragments encompassed within the term “antigen-binding fragment” of an antibody include (i) a Fab fragment, a monovalent fragment consisting of the VL, VH, CL and CHI domains; (ii) a F(ab')2 fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; (iii) a Fd fragment consisting of the VH and CHI domains; (iv) a Fv fragment consisting of the VL and VH domains of a single arm of an antibody, (v) a dAb fragment (Ward et al., Nature 341:544-546, 1989), which consists of a VH domain; (vi) an isolated complementarity determining region (CDR), e.g., VH CDR3 comprising or not additional sequence (linker, framework region(s) etc.) and (v) a combination of two to six isolated CDRs comprising or not additional sequence (linker, framework region(s) etc.). Furthermore, although the two domains of the Fv fragment, VL and VH, are coded for by separate genes, they can be joined, using recombinant methods, by a synthetic linker that enables them to be made as a single polypeptide chain in which the VL and VH regions pair to form monovalent molecules (known as single chain Fv (scFv); see-42- SG Docket No.: 14989-700.600e.g., Bird et al., Science 242:423-426, 1988); and (Huston et al., Proc. Natl. Acad. Sci. USA 85:5879-5883, 1988). Such single chain antibodies are also intended to be encompassed within the term “antigen-binding fragment” of an antibody. Furthermore, the antigen-binding fragments include binding-domain immunoglobulin fusion proteins comprising (i) a binding domain polypeptide (such as a heavy chain variable region, a light chain variable region, or a heavy chain variable region fused to a light chain variable region via a linker peptide) that is fused to an immunoglobulin hinge region polypeptide, (ii) an immunoglobulin heavy chain CH2 constant region fused to the hinge region, and (iii) an immunoglobulin heavy chain CH3 constant region fused to the CH2 constant region. The hinge region may be modified by replacing one or more cysteine residues with serine residues to prevent dimerization. These antibody fragments are obtained using conventional techniques known to those with skill in the art, and the fragments are screened for utility in the same manner as are intact antibodies.

[0128] In some embodiments, the polypeptide is an antibody, antibody fragment or variant thereof that binds to an antigen, e.g., a tumor-associated antigen. In some embodiments, the tumor-associated antigen is selected from the group consisting of PSMA, CD70, CD3, HER2, HER3, TROP2, PD-I, PDL-1, VEGFR, EGFR, c-Met (HGFR), CD 19, CD22, CD24, CD25 (IL-2R alpha), CD30, CD33, CD37, CD38, CD44, CD46, CD47, CD48, CD52, CD56 (NCAM-1), CD71 (Transferrin R), CD74, CD79b, CD96, CD97, CD99, CD123 (IL-3R alpha), CD138 (syndecan-1), CD142, CD166 (ALCAM), CD179, CD203c (ENPP3), TIMI, CD205 (LY75), CD221 (IGF-1R), CD223, CD262 (TRAIL R2), CD276 (B7-H3), mesothelin, EpCAM, MUCI, MUC16 (CA-125), GPC3, CEA, CEACAM5, CEACAM6, CA9, DLL3, ROR1, ROR2, GPNMB, GCC, GUCY2c, NaPi2b, Flt-1, Flt-3, FOLR1 (folate receptor alpha), Tissue Factor (TF), CA6, BCMA, SLAMF7 (CS1), TIMI, CanAg, Ckit (CD117), EphA2, Nectin4, SLTRK6, FGFR2, LYPD3 (C4.4a), Cadherin 3, Cadherin 6, 5T4 (TPBG), STEAP1, PTK7, Ephrin-A4, SLC34A2, LIV-1 (SLC39A6 orZIP6), SLC1A5, TENB2, ETBR, integrin v3, Cripto, AGS-5 (SLC44A4), LY6E, SLITRK6, AXL, LAMP1, LRRC15, TNF-alpha, CTLA-4 and MN / CA IX antibody, antibody fragment or variant.

[0129] In some embodiments, the polypeptide comprises genetically engineered versions of naturally occurring human proteins. In some embodiments, the polypeptide comprises Adrenocorticotropic hormone (ACTH), Adropin, Amylin, Angiotensin, Atrial natriuretic peptide (ANP), Calcitonin, Cholecystokinin (CCK), Gastrin, Ghrelin, Glucagon, Glucosedependent insulinotropic polypeptide (GIP), Glucagon-like peptide- 1 (GLP-1), Growth hormone, Follicle-stimulating hormone (FSH), Insulin, Leptin, Luteinizing hormone (LH),SG Docket No.: 14989-700.600Melanocyte-stimulating hormone (MSH), Oxytocin, Parathyroid hormone (PTH), Prolactin, Renin, Somatostatin, Thyroid-stimulating hormone (TSH), Thyrotropin-releasing hormone (TRH), Vasopressin, also called arginine vasopressin (A VP) or anti-diuretic hormone (ADH), Vasoactive intestinal peptide (VIP), Somatotropin (GH1), Gonadotropin Releasing Hormone 1 (GNRH1), Gonadotropin Releasing Hormone 2 (GNRH2), Growth Hormone Releasing Hormone (GHRH), Parathyroid Hormone Like Hormone (PTHLH), Corticotropin Releasing Hormone (CRH), Anti-Mullerian Hormone (AMH), Chorionic Somatomammotropin Hormone 1 (CSH1), Chorionic Somatomammotropin Hormone 2 (CSH2), Pro-Melanin Concentrating Hormone (PMCH), Resistin (RETN), or genetically engineered versions thereof.

[0130] In some embodiments, the polypeptide is or binds to or targets adrenocorticotropic hormone (ACTH), amylin, angiotensin, atrial natriuretic peptide (ANP), calcitonin, cholecystokinin (CCK), gastrin, ghrelin, glucagon, growth hormone, follicle-stimulating hormone (FSH), insulin, leptin, melanocyte-stimulating hormone (MSH), oxytocin, parathyroid hormone (PTH), prolactin, renin, somatostatin, thyroid-stimulating hormone (TSH), thyrotropin-releasing hormone (TRH), vasopressin, or vasoactive intestinal peptide.

[0131] In some embodiments, the polypeptide is or binds to or targets angiotensin receptor; apelin receptor; bombesin receptor; bradykinin receptor; calcitonin receptor; chemokine receptor; cholecytokinin receptor; corticotropic-releasing factor receptor; galanin receptor; ghrelin receptor; glucagon receptor; glycoprotein hormone receptor; gonadotropin-releasing hormone receptor; kisspeptin receptor; melanocortin receptor; motilin receptor; neuromedin U receptor; neuropeptide FF / AF receptor; neuropeptide S receptor; neuropeptide W7B receptor; neuropeptide Y receptor; opioid receptor; orexin receptor; parathyroid hormone receptor; prokineticin receptor; prolactin-releasing peptide receptor; QRFP receptor; relaxin family peptide receptor; somatostatin receptor; tachykinin receptor; thyrotropin-releasing hormone receptor; urotensin receptor; vasopressin and oxytocin receptor; VIP and PACAP receptor; or combinations thereof.

[0132] In some embodiments, the polypeptide is or binds to or targets angiotensin receptors (e.g. AGTR1, AGTR2); apelin receptor (APLNR); bombesin receptors (BB1 / NMBR, BB2 / GRPR, BRS3); bradykinin receptors (BDKRB1, BDKRB2); calcitonin receptor (e.g. CALCR, CALCRL); chemokine receptors (e.g. CCR1-10, CXCR1-6, etc); cholecytokinin receptors (CCKAR, CCKBR); corticotropic-releasing factor receptors (CRHR1, CRHR2); galanin receptors (GALR1-3); ghrelin receptor (GHSR); glucagon-44- SG Docket No.: 14989-700.600receptor family (e g. GHRHR, GIPR, GLP1R, GLP2R, GCGR, SCTR); glycoprotein hormone receptors (FSHR, LHCGR, TSHR); gonadotropin-releasing hormone receptor (GNRHR, GNRHR2); kisspeptin receptor (KISS1R); melanocortin receptors (MC1-5R); motilin receptor (MLNR); neuromedin U receptors (NMUR1, NMUR1-2); neuropeptide FF / AF receptors (NPFFR1, NPFFR2); neuropeptide S receptors (NPSR1); neuropeptide W / B receptors (NPBWR1, NPBWR2); neuropeptide Y receptors (NPY1-6R); opioid receptors (0PRD1, 0PRK1, 0PRM1); orexin receptors (HCRTR1, HCRTR2); parathyroid hormone receptors (PTH1R, PTH2R); prokineticin receptors (PR0KR1, PR0KR2); prolactin-releasing peptide receptor (PRLHR); QRFP receptor (QRFPR); relaxin family peptide receptors (RXFP1-4); somatostatin receptors (SSTR1-5); tachykinin receptors (TACR1-3); thyrotropinreleasing hormone receptor (TRHR1, TRHR2); urotensin receptor (UTS2R); vasopressin and oxytocin receptors (AVPR1 A, AVPR1B, AVPR2, OXTR); or VIP and PACAP receptors (ADCYAP1R1, VIPR1, VIPR2).

[0133] In some embodiments, the polypeptide is or binds to or targets formyl peptide receptor (FPR1, FPR2, or FPR3), platelet activating factor receptor (PAFR), activated complement component 5a receptor (C5aR); or a chemokine GPCR that is binds to a CC chemokine (P-chemokine), CXC chemokine (a-chemokine), C chemokine (y chemokine), or CX3C chemokine (d-chemokine).

[0134] In some embodiments, the polypeptide binds to or targets CCR1, CCR2, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CCR9, CCR10, CXCR1, CXCR2, CXCR3, CXCR4, CXCR5, CXCR6, CXCR7, CX3CR1, or XCR1.

[0135] In some embodiments, the therapeutic polypeptide is or binds to or targets CD20, HER2, CD19, EGFR, IL-23, PD-1, SARS-CoV-2, TNF, VEGF, CGRP, IL-17, IL-6R, or POLL In some embodiments, the polypeptide is or binds to or targets F-Box and WD Repeat Domain Containing 7 (FBXW7); Kirsten rat sarcoma virus (KRAS); Isocitrate dehydrogenase 1 (IDH1); guanine nucleotide-binding protein alpha subunit (GNAS); DNA methyltransferase 3a (DNMT3a); Fibroblast growth factor receptor 3 (FGFR3); tumor protein p53 (p53); or Catenin beta-1 (CTNNB1).

[0136] In some embodiments, the polypeptide is or binds to or targets cytoplasmic tyrosine kinase (CTK), a serine / threonine kinase (S / T Kinase), a lipid kinase (LK), or a receptor tyrosine kinase (RTK). In some embodiments, the disease protein is Sphingosine kinase 1 (SKI), phosphoinositide 3 -kinase (PI3K), Protein kinase Ci (PKCi), mammalian target of rapamycin (mTOR), cyclin-dependent kinases (CDKs), Ataxia telangiectasia mutated-45- SG Docket No.: 14989-700.600(ATM), protein kinase B (Akt), ribosomal protein S6 kinase (S6K), Serine / threonine kinase 11 or liver kinase B1(STK11 / LKB1), Polo-like kinases (PLKs), B-Raf proto-oncogene (b-Raf), Aurora Kinase A & B (Aur A & B), Mitogen Activated Protein (MAP) kinase kinase kinase (MAP3K), Mitogen-activated protein kinase 3 (MAPK3), Mitogen-activated protein kinase 1 (MAPK1), Apoptosis signal-regulating kinase 1 (ASK1), Mitogen-activated protein kinase kinase (MEK), Proto-oncogene tyrosine-protein kinase Src (c-SRC), c-Yes protooncogene (c-YES), Abelson murine leukemia viral oncogene homolog 1 (Abl), Janus kinase 1 (JAK1), Janus kinase 2 (JAK2), Janus kinase 3 (JAK3), Tyrosine kinase 2 (TYK2), Recepteur d’Origine Nantais (RON), Fibroblast growth factor receptor 1 (FGFR 1), FGFR 2, FGFR 3, FGFR 4, c-MET proto-oncogene (c-Met), c-RET proto-oncogene (c-Ret), Insulinlike growth factor 1 receptor (IGF-IR), Epidermal growth factor receptor (EGFR), Platelet-derived growth factor receptor a (PDGFR-a), Platelet-derived growth factor receptor p (PDGFR-P), proto-oncogene c-Kit or Mast / stem cell growth factor receptor (c-Kit), vascular endothelial growth factor receptor 1 (VEGFR-1), vascular endothelial growth factor receptor 2 (VEGFR-2), Fms-like tyrosine kinase 3 (Flt3), Fms-like tyrosine kinase 4 (Flt-4), Anaplastic lymphoma kinase (ALK), human epidermal growth factor receptor-2 (HER2), ribosomal S6 kinase 2 (RSK2), Glycogen synthase kinase-3 beta (GSK3P), or Bruton's tyrosine kinase (BTK), the small G protein is Rat sarcoma virus (Ras), or Ras homolog enriched in brain (RHEB). In some embodiments, the small G protein is Kirsten rat sarcoma virus (KRAS), Harvey rat sarcoma virus (HRAS), or neuroblastoma rat sarcoma virus (NRAS).

[0137] In some embodiments, the polypeptide is or binds to or targets protein tyrosine phosphatase (PTP), protein tyrosine kinases (PTK), protease, stimulator of interferon genes (STING), small G protein, GTPase, GTPase-activating protein, transcription activator, cofactor, tumor suppressor, regulatory protein, transporter protein, enzyme, nuclear receptor protein, methyltransferase, or growth factor ligand.

[0138] In some embodiments, the polypeptide is or binds to or targets a protease that is Cathepsin A, Cathepsin B, Cathepsin C, Cathepsin D, Cathepsin E, Cathepsin F, Cathepsin G, Cathepsin H, Cathepsin K, Cathepsin LI, Cathepsin L2, Cathepsin O, Cathepsin S, Cathepsin W, or Cathepsin Z.

[0139] In some embodiments, the polypeptide is or binds to one or more targets.

[0140] Any of the aforementioned polypeptides / proteins can be modified with the compounds described herein to provide a route to their site-specific conjugation with diverse-46- SG Docket No.: 14989-700.600moieties. Selective conjugation of polypeptides includes, can be used for: PEGylation (e.g., see Harris, J. M. & Chess, R. B. Effect of pegylation on pharmaceuticals.Nat. Rev. Drug Discov. 2,214-221 (2003), PASylation (e.g., see M. Schlapschy et al. “PASylation: a biological alternative to PEGylation for extending the plasma half-life of pharmaceutically active proteins”, Protein Engineering Design And Selection, vol. 26, no. 8, 1 Aug. 1, 2013), glycosylation (Sola, R. J. & Griebenow, K. Glycosylation of therapeutic proteins. BioDrugs 24,9-21 (2010)), lipidation (Menacho-Melgar, R., et al., J. Control. Release 295,1-12 (2019)), protein fusion (Leader, B., et al., Protein therapeutics: a summary and pharmacological classification. Nat. Rev. Drug Discov. 7,21-39 (2008)), peptide-drug conjugates, antibody-drug conjugates (ADCs), antibody-antisense oligonucleotide conjugates, bispecific antibodies, trispecific antibodies, multi-specific antibodies, etc.

[0141] By controlling both the stoichiometry and site of conjugation of a PEG, lipid, proline / alanine-rich sequences (PAS), glycosyl unit(s), drug, etc. on a protein, one can optimize both half-life and potency in a manner that is difficult to achieve with less specific chemistries.

[0142] Derivatizations of polypeptides / proteins with non-proteinaceous moieties are especially useful for improving or restoring biological properties of the polypeptides / proteins. For example, PEG modification of polypeptides / proteins can alter the stability, in vivo circulating half-life, binding affinity, solubility and resistance to proteolysis. The moieties suitable for derivatization of the polypeptides / proteins may be water soluble polymers. Nonlimiting examples of water soluble polymers may include, but are not limited to, polyethylene glycol (PEG), copolymers of ethylene glycol / propylene glycol, carboxymethylcellulose, dextran, polyvinyl alcohol, polyvinyl pyrrolidone, poly-1,3 -di oxolane, poly-1, 3, 6-trioxane, ethylene / maleic anhydride copolymer, polyamino acids (either homopolymers or random copolymers), and dextran or poly(n-vinyl pyrrolidone)polyethylene glycol, propropylene glycol homopolymers, polypropylene oxide / ethylene oxide co-polymers, polyoxyethylated polyols (e.g., glycerol), polyvinyl alcohol, and mixtures thereof. Polyethylene glycol propionaldehyde may have advantages in manufacturing due to its stability in water. The polymer may be of any molecular weight and may be branched or unbranched. The number of polymers attached to the antibody may vary, and if more than one polymer is attached, they may be the same or different molecules. In general, the number and or type of polymers used for derivatization may be determined based on considerations including, but not limited to, the particular properties or functions of the polypeptides / proteins to be improved, whether-47- SG Docket No.: 14989-700.600the polypeptide / protein derivative will be used in a therapy under defined conditions.

[0143] Lipidation occurs on numerous proteins and regulates many aspects of physiology. The effects of protein lipidation on cellular function are achieved by regulating proteinmembrane interactions, protein-protein interactions, protein stability, and enzymatic activities. The lipid moieties added to proteins can be either fatty acyl or polyisoprenyl groups, and the modifications typically occur on the nucleophilic side chains of proteins (e.g., cysteine, serine, and lysine).

[0144] As described above, any appropriate polypeptide may be used with the compositions and methods described herein, including the addition of one or more PRM sequence to the polypeptide. For example, in some cases the polypeptide may be an antibody, antibody fragment, or any other antibody-derived molecule, to which a cargo may be linked via the inclusion of a PRM and the use of a proximity-triggered conjugation compound as described. In general, the polypeptide may be modified to include an appropriate PRM sequence (including but not limited to those described, e.g., in SEQ ID NO. 27-31 (PRM1 examples) and SEQ ID NO. 32-51 (PRM2 examples).

[0145] The identification of regions of polypeptides where the PRMs described herein can be added to / inserted can be achieved by methods known in the art. For example, a useful method for identification of certain residues or regions of the antibody that are preferred locations for mutagenesis is called “alanine scanning mutagenesis” as described by Cunningham and Wells (1989) Science, 244: 108 1-1085. Here, a residue or group of target residues is identified (e.g., charged residues such as Arg, Asp, His, Lys, and GIu) and replaced by a neutral or negatively charged amino acid (for example alanine or polyalanine) to affect the interaction of the amino acids with antigen. Those amino acid locations demonstrating functional sensitivity to the substitutions may then be refined by introducing further or other variants at, or for, the sites of substitution. Thus, while the site for introducing an amino acid sequence variation is predetermined, the nature of the mutation per se need not be predetermined. For example, to analyze the performance of a mutation at a given site, alanine scanning or random mutagenesis is conducted at the target codon or region and the expressed antibodies are screened for the desired activity. Amino acid sequence insertions include amino- and / or carboxyl-terminal fusions ranging in length from one residue to polypeptides containing a hundred or more residues, as well as intrasequence insertions of single or multiple amino acid residues. Non-limiting examples of terminal insertions include an antibody with an N-terminal methionyl residue or the antibody fused to a cytotoxic-48- SG Docket No.: 14989-700.600polypeptide. Other insertional variants of the antibody molecule include the fusion to the N-or C-terminus of the antibody to an enzyme (e.g., for ADEPT) or a polypeptide which increases the serum half-life of the antibody. Another type of variant is an amino acid substitution variant. These variants have at least one amino acid residue in the antibody molecule replaced by a different residue. The sites of greatest interest for substitutional mutagenesis include the hypervariable regions, but FR alterations are also contemplated.

[0146] Substantial modifications in the biological properties of the polypeptides / proteins are accomplished by selecting substitutions that differ significantly in their effect on maintaining: (a) the structure of the polypeptide backbone in the area of the substitution, for example, as a sheet or helical conformation, (b) the charge or hydrophobicity of the molecule at the target site, or (c) the bulk of the side chain. Naturally occurring residues are divided into groups based on common side-chain properties:(1) hydrophobic: norleucine, Met, Ala, Vai, Leu, lie;(2) neutral hydrophilic: Cys, Ser, Thr;(3) acidic: Asp, GIu;(4) basic: Asn, Gin, His, Lys, Arg;(5) residues that influence chain orientation: Gly, Pro; and(6) aromatic: Tip, Tyr, Phe.

[0147] Non-conservative substitutions may entail exchanging a member of one of these classes for another class.

[0148] In some embodiments, the methods described herein are used to prepare an Fc fusion protein, e.g., comprising the Fc domain of IgG fused together with a protein or peptide of interest, or any other antibody fragment described in the art, e.g., in U.S. Pat. No.5,648,237, “Expression of Functional Antibody Fragments,” incorporated by reference herein in its entirety.

[0149] FIGS. 19A-19B and 20A-20B illustrate examples of an IgG polypeptides that may be used with any of the compositions and methods described herein. FIGS. 19A-19B and 20A-20B illustrate IgGl / kappa constant regions that may be modified to include one or more PRM (e.g., PRM1 or PRM2). FIG. 19A shows the sequence (SEQ ID NO. 1) for an IgGl Light chain constant region with two runs of sequence (e.g., residues 38-50 and 94-104) as well as the C-terminal end, illustrating where PRMs may be added to the sequence. Three locations / residues are highlighted in this example, the K at position 43, the D at position 45 and the V at position 99, as regions of particular interest. In practice a PRM1 or PRM2 may-49- SG Docket No.: 14989-700.600replace the indicated residue in 38 to 50 or 94-104, or the PRM may be inserted between any two adjacent residues in this region. Alternatively or additionally a PRM1 or PRM2 may be added to the C-terminus of the polypeptide, either by directly replacing the final residue (e.g., C at position 108) with the PRM or by adding the PRM immediately after the residue; in either case a linker may connect the PRM to the C-terminus. Any appropriate linker may be used, such as 1-10 Pro- Ala or Gly-Ser dipeptide units. Alternatively in some cases no linker is used or needed. FIG. 19B schematically illustrates the various IgG Light chain variations including a PRM at the indicated locations.

[0150] Similarly, FIGS. 20A-20B illustrate the amino acid sequence listing (SEQ ID NO.12) For the IgG Heavy Chain constant region shown. Two regions (and the C-terminus) are identified as candidates for insertion of a PRM, between position 5-22 (highlighting positions 10 (Ala), 14 (Gly), 15 (Pro), and 17 (Vai)) and 126-136 (highlighting position 131 (Ser)). In general, a polypetide including a target may be generated as described herein by replacing an amino acid with and / or inserting a PRM, either a PRM1 or PRM2. Example sequences have been generated and analyzed, and are shown in the tables below, referencing the corresponding sequence listing:-50- SG Docket No.: 14989-700.600

[0151] The polypeptides comprising the PRMs described herein may be produced by recombinant DNA technology, e.g. by cultivating a cell comprising the described nucleic acid molecule or vectors which encode the polypeptides comprising the PRMs and isolating said polypeptides comprising the PRMs from the culture. The polypeptides comprising the PRMs may be produced in any suitable cell-culture system including prokaryotic cells, e.g. E. coli (e.g. BL21, W3110, or JM83), P. fluorescens, o Bacillus sublihis: or eukaryotic cells, e.g.-51- SG Docket No.: 14989-700.600Pichia pastoris yeast strain X-33 or CHO cells. Further suitable cell lines known in the art are obtainable from cell line depositories, like the American Type Culture Collection (ATCC). The term “prokaryotic” is meant to include bacterial cells while the term “eukaryotic” is meant to include yeast, higher plant, insect and mammalian cells. The transformed hosts can be grown in fermenters and cultured according to techniques known in the art to achieve optimal cell growth. Further examples of methods, vectors, and translation and transcription elements, and other elements useful in the methods herein are described in, e.g.: U.S. Pat. No.5,055,294 to Gilroy and U.S. Pat. No. 5,128,130 to Gilroy et al.; U.S. Pat. No. 5,281,532 to Rammler et al.; U.S. Pat. Nos. 4,695,455 and 4,861,595 to Barnes et al.; U.S. Pat. No.4,755,465 to Gray et al.; and U.S. Pat. No. 5,169,760 to Wilcox.Peptidic Recognition Motifs (PRMs)

[0152] The therapeutic polypeptides described herein comprise:at least one peptidic recognition motif (PRM), and- the amino acid sequence of a biologically active polypeptide (B), wherein the at least one PRM is:1) added to the N-terminus of the biologically active polypeptide,2) added to the C-terminus of the biologically active polypeptide, or3) inserted at an internal position of the biologically active polypeptide;wherein the at least one PRM is independently selected from PRM1, PRM2, or combinations thereof:wherein each PRM1 comprises:• 2 to 4 histidine (His) residues; and• a lysine (Lys) residue, ornithine (Orn) residue, homolysine (HLys), a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residuewherein each PRM2 comprises:• at least 2 amino acids selected from an arginine (Arg) residue, a homoarginine (HArg) residue, a citrulline (Cit) residue, norarginine (AGBA) residue, a p2- arginine (P2-Arg), and a p3-arginine (P3-Arg); and• a lysine (Lys) residue, an ornithine (Orn) residue, a homolysine (HLys) residue, a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue.

[0153] In some embodiments, the therapeutic polypeptides described herein comprise: at least one peptidic recognition motif (PRM), and the amino acid sequence of a biologically active polypeptide (B), wherein the at least one PRM is: 1) added to the N-terminus of the-52- SG Docket No.: 14989-700.600biologically active polypeptide, 2) added to the C-terminus of the biologically active polypeptide, or 3) inserted at an internal position of the biologically active polypeptide; wherein the at least one PRM is independently selected from PRM1, PRM2, or combinations thereof; wherein each PRM1 comprises: at least 2 histidine (His) residues; and a lysine (Lys) residue, ornithine (Om) residue, homolysine (HLys), a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue; wherein each PRM2 comprises: at least 2 amino acids selected from an arginine (Arg) residue, a homoarginine (HArg) residue, a citrulline (Cit) residue, norarginine (AGBA) residue, a p2-arginine (P2-Arg), and a p3-arginine (P3-Arg); and a lysine (Lys) residue, an ornithine (Orn) residue, a homolysine (HLys) residue, a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue.

[0154] The PRMs described herein are biosynthetically incorporated into polypeptides. The term “biosynthetically,” as used herein, refers to any method utilizing a translation system (cellular or non-cellular), including use of at least one of the following components: a polynucleotide, a codon, a tRNA, and a ribosome. Methods for incorporating amino acids into polypeptides are well known in the art, e.g., C. L. Young, et al., Recombinant protein expression and purification, Biotechnol. J. 2012, 7, 620-634; Block, H. et al. Immobilized-metal affinity chromatography (IMAC): a review, Methods Enzymol. 463, 439-473 (2009); Hochuli, E., et al. Genetic Approach to Facilitate Purification of Recombinant Proteins with a Novel Metal Chelate Adsorbent. Nat Biotechnol 6, 1321- 1325 (1988); Skerra A. A general vector, pASK84, for cloning, bacterial production, and single-step purification of antibody Fab fragments. Gene, 141(1), 79-84 (1994); Petty, K J., Metal-chelate affinity chromatography. Current protocols in neuroscience vol. Chapter 5 (2001); Carson, M. etal., His-tag impact on structure. Acta crystallographica. Section D, Biological crystallography vol. 63, Pt 3 (2007); Chen, X., et al., Fusion protein linkers: property, design and functionality Adv. Drug Delivery Rev. 65, 1357- 1369 (2013); Bomhorst, J. A. & Falke, J. J. Purification of proteins using polyhistidine affinity tags. Methods in enzymology, 326, 245-254 (2000).PRM1

[0155] In some embodiments, eachPRMl comprises:X-His-His, His-X-His, His-His-X, X-His-His-His, His-X-His-His, His-His-X-His, His-His-His-X,X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, orHis-His-His-His-X,-53- SG Docket No.: 14989-700.600wherein X is Lys, Om, HLys, p2-Lys, or p3-Lys.

[0156] In some embodiments, each PRM1 comprises X-His-His, His-X-His, or His-His-X. In some embodiments, each PRM1 comprises X-His-His. In some embodiments, each PRM1 comprises His-X-His. In some embodiments, each PRM1 comprises His-His-X.

[0157] In some embodiments, each PRM1 comprises X-His-His-His, His-X-His-His, His-His-X-His, or His-His-His-X. In some embodiments, each PRM1 comprises X-His-His-His. In some embodiments, each PRM1 comprises His-X-His-His. In some embodiments, each PRM1 comprises His-His-X-His. In some embodiments, each PRM1 comprises His-His-His-X.

[0158] In some embodiments, each PRM1 comprises X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, or His-His-His-His-X. In some embodiments, each PRM1 comprises X-His-His-His-His. In some embodiments, each PRM1 comprises His-X-His-His-His. In some embodiments, each PRM1 comprises His-His-X-His-His. In some embodiments, each PRM1 comprises His-His-His-His-X. In some embodiments, each PRM1 comprises His-His-His-His-X.

[0159] In some embodiments, X is Lys. In some embodiments, X is Om. In some embodiments, X is HLys.

[0160] In some embodiments, at least one PRM1 is covalently attached at the N-terminus or C-terminus of the biologically active polypeptide.

[0161] In some embodiments, the polypeptide comprises the amino acid sequence:B-(His)m-X-(His)nor (His)m-X-(His)n-B,wherein,B is the biologically active polypeptide;X is Lys, Om, HLys, p2-Lys, or p3-Lys;m is 0, 1, 2, 3, or 4; andn is 0, 1, 2, 3, or 4, provided that m + n is at least 2.

[0162] In some embodiments, at least one PRM1 is inserted at an internal position of the polypeptide.

[0163] In some embodiments, the polypeptide comprises the amino acid sequence:wherein,B1and B2are fragments of a biologically active polypeptide B, such that BXB2is the biologically active polypeptide B, and the -(His)m-X-(His)n- sequence is added at-54- SG Docket No.: 14989-700.600an internal position of the biologically active polypeptide B;X is Lys, Om, HLys, p2-Lys, or p3-Lys;m is 0, 1, 2, 3, or 4; andn is 0, 1, 2, 3, or 4, provided that m + n is at least 2.

[0164] In some embodiments, m is 0 and n is 4; m is 1 and n is 3; m is 2 and n is 2; m is 3 and n is 1; or m is 4 and n is 0. In some embodiments, m is 0 and n is 4. In some embodiments, m is 1 and n is 3. In some embodiments, m is 2 and n is 20. In some embodiments, m is 3 and n is 1. In some embodiments, m is 4 and n is 0.

[0165] In some embodiments, m + n is at least 2. In some embodiments, m + n is at least 3. In some embodiments, m + n is at least 4. In some embodiments, m + n is at least 5.

[0166] In some embodiments, PRM1 comprises at least 2 His residues. In some embodiments, PRM1 comprises at least 3 His residues. In some embodiments, PRM1 comprises at least 4 His residues. In some embodiments, PRM1 comprises at least 5 His residues.

[0167] In some embodiments, the biologically active polypeptide comprises at least one PRM1 that is covalently attached at the N-terminus or C-terminus of the polypeptide. In some embodiments, the biologically active polypeptide comprises at least one PRM1 that is covalently attached at the N-terminus. In some embodiments, the biologically active polypeptide comprises at least one PRM1 that is covalently attached at the C-terminus. In some embodiments, the biologically active polypeptide comprises at least one PRM1 that is covalently attached to the polypeptide through any one of the amino acid residues of the polypeptide.

[0168] In some embodiments, at least one PRM1 is attached at the N-terminus or C-terminus of the polypeptide with an optional linker.PRM2

[0169] In some embodiments, each PRM2 comprises:X3-X1-Y -X4-X2-Ywherein:X3or X4are Lys, Orn, HLys, p2-Lys, or p3-Lys;if X3is present, then X4is not present;if X4is present, then X3is not present;each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg; XIcomprises 0 to 10 amino acid residues other than X3, X4, and Y;-55- SG Docket No.: 14989-700.600X2comprises 0 to 10 amino acids other than X3, X4, and Y.

[0170] In some embodiments, X3or X4is Lys. In some embodiments, X3or X4is Om. In some embodiments, X3or X4is HLys.

[0171] In some embodiments, each Y is Arg. In some embodiments, each Y is HArg. In some embodiments, each Y is Cit. In some embodiments, each Y is AGBA. In some embodiments, the two Y residues are different and independently selected from Arg, HArg, Cit, and AGBA.

[0172] In some embodiments, each amino acid residue of X1is independently selected from alanine (Ala), asparagine (Asn), glutamine (Gin), glycine (Gly), isoleucine(Ile), leucine (Leu), methionine (Met), phenylalanine (Phe), proline (Pro), serine (Ser), threonine (Thr), tryptophan (Trp), tyrosine (Tyr), valine (Vai), glutamic acid (Glu), aspartic acid (Asp), and non-natural amino acids without positively charged or thiol-containing side chains. In some embodiments, X1does not comprise a His residue.

[0173] In some embodiments, X1comprises 0 amino acids.

[0174] In some embodiments, X1comprises 1 amino acid.

[0175] In some embodiments, X1comprises 2 amino acids.

[0176] In some embodiments, X1comprises 3 amino acids.

[0177] In some embodiments, X1comprises 4 amino acids.

[0178] In some embodiments, X1comprises 5 amino acids.

[0179] In some embodiments, X1comprises 6 amino acids.

[0180] In some embodiments, X1comprises 7 amino acids.

[0181] In some embodiments, X1comprises 8 amino acids.

[0182] In some embodiments, X1comprises 9 amino acids.

[0183] In some embodiments, X1comprises 10 amino acids.

[0184] In some embodiments, each amino acid residue of X2is independently selected from alanine (Ala), asparagine (Asn), glutamine (Gin), glycine (Gly), isoleucine(Ile), leucine (Leu), methionine (Met), phenylalanine (Phe), proline (Pro), serine (Ser), threonine (Thr), tryptophan (Trp), tyrosine (Tyr), valine (Vai), glutamic acid (Glu), aspartic acid (Asp), and non-natural amino acids without positively charged or thiol-containing side chains. In some embodiments, X2does not comprise a His residue.

[0185] In some embodiments, X2comprises 0 amino acids.

[0186] In some embodiments, X2comprises 1 amino acid.

[0187] In some embodiments, X2comprises 2 amino acids.SG Docket No.: 14989-700.600

[0188] In some embodiments, X2comprises 3 amino acids.

[0189] In some embodiments, X2comprises 4 amino acids.

[0190] In some embodiments, X2comprises 5 amino acids.

[0191] In some embodiments, X2comprises 6 amino acids.

[0192] In some embodiments, X2comprises 7 amino acids.

[0193] In some embodiments, X2comprises 8 amino acids.

[0194] In some embodiments, X2comprises 9 amino acids.

[0195] In some embodiments, X2comprises 10 amino acids.

[0196] In some embodiments, PRM2 does not comprise a His residue.

[0197] In some embodiments, at least one PRM2 is covalently attached at the N-terminus or C-terminus of the biologically active polypeptide.

[0198] In some embodiments, the polypeptide comprises the amino acid sequence:B-X3-X1-Y-X4-X2-Y or Y-X2-X4-Y-X1-X3-Bwherein:B is a biologically active polypeptide (B);X3or X4is Lys, Om, HLys, p2-Lys, or p3-Lys;if X3is present, then X4is not present;if X4is present, then X3is not present;each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg; X1comprises 0 to 10 amino acid residues other than X3, X4, and Y;X2comprises 0 to 10 amino acids other than X3, X4, and Y.

[0199] In some embodiments, at least one PRM2 is inserted at an internal position of the polypeptide.

[0200] In some embodiments, the polypeptide comprises the amino acid sequence:B^X^-Y-X^-Y-B2wherein,B1and B2are fragments of a biologically active polypeptide B, such that BXB2is the biologically active polypeptide (B), and the -X3-X1-Y-X4-X2-Y- sequence is added at an internal position of B;B1and B2are fragments of a biologically active polypeptide B, such that BXB2is the biologically active polypeptide B and the -X^X^Y-X^X^Y- sequence is added at an internal position of polypeptide B;-57- SG Docket No.: 14989-700.600X3or X4is Lys, Om, HLys, p2-Lys, or p3-Lys;if X3is present, then X4is not present;if X4is present, then X3is not present;each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg; X1comprises 0 to 10 amino acid residues other than X3, X4, and Y;X2comprises 0 to 10 amino acids other than X3, X4, and Y.

[0201] In some embodiments, the biologically active polypeptide comprises at least one PRM2 that is covalently attached at the N-terminus or C-terminus of the polypeptide. In some embodiments, the biologically active polypeptide comprises at least one PRM2 that is covalently attached at the N-terminus. In some embodiments, the biologically active polypeptide comprises at least one PRM2 that is covalently attached at the C-terminus. In some embodiments, the biologically active polypeptide comprises at least one PRM2 that is covalently attached to the polypeptide through any one of the internal amino acid residues of the polypeptide.

[0202] In some embodiments, at least one PRM2 is attached at the N-terminus or C-terminus of the polypeptide with an optional linker.

[0203] In some embodiments, the biologically active polypeptide comprises at least one PRM1 and at least one PRM2.

[0204] In some embodiments, the biologically active polypeptide comprises at least one PRM1 and at least one PRM2, wherein at least one PRM is covalently attached at the N-terminus or C-terminus of the polypeptide. In some embodiments, the biologically active polypeptide comprises at least one PRM1 and at least one PRM2, wherein at least one PRM is covalently attached at the N-terminus. In some embodiments, the biologically active polypeptide comprises at least one PRM1 and at least one PRM2, wherein at least one PRM is covalently attached at the C-terminus. In some embodiments, the biologically active polypeptide comprises at least one PRM1 and at least one PRM2, wherein at least one PRM is covalently attached to the polypeptide through any one of the internal amino acid residues of the polypeptide. In some embodiments, at least one PRM1 is attached at the N-terminus or C-terminus of the polypeptide with an optional linker. In some embodiments, at least one PRM2 is attached at the N-terminus or C-terminus of the polypeptide with an optional linker.

[0205] In some embodiments, the biologically active polypeptide comprises at least one PRM1 and at least one PRM2, wherein at least one PRM1 is inserted at an internal position of the polypeptide, and at least one PRM2 is inserted at an internal position of the polypeptide.-58- SG Docket No.: 14989-700.600

[0206] In some embodiments, the biologically active polypeptide comprises at least one PRM1 and at least one PRM2, wherein at least one PRM1 is inserted at an internal position of the polypeptide, and at least one PRM2 is covalently attached at the N-terminus. In some embodiments, the biologically active polypeptide comprises at least one PRM1 and at least one PRM2, wherein at least one PRM1 is inserted at an internal position of the polypeptide, and at least one PRM2 is covalently attached at the C-terminus.

[0207] In some embodiments, the biologically active polypeptide comprises at least one PRM1 and at least one PRM2, wherein at least one PRM2 is inserted at an internal position of the polypeptide, and at least one PRM1 is covalently attached at the N-terminus. In some embodiments, the biologically active polypeptide comprises at least one PRM1 and at least one PRM2, wherein at least one PRM2 is inserted at an internal position of the polypeptide, and at least one PRM1 is covalently attached at the C-terminus.

[0208] In some embodiments, the optional linker is a flexible linker, rigid linker, or in vivo cleavable linker. In some embodiments, a flexible linker comprises glycine and serine residues (“GS” linker), glycine residues, alkylene chains, or PEG chains, or combinations thereof. In some embodiments, a rigid linker comprises a -helix-forming sequences, prolinerich sequences, or combinations thereof. In some embodiments, an in vivo cleavable linker comprises a disulfide, a carbonate, or an ester.

[0209] In some embodiments, the optional linker comprises one or more amino acids. In some embodiments, the linker comprises 1 to 3, 1 to 5, 1 to 10, 5 to 10, or 5 to 20 amino acids. In some embodiments, the linker comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids. In some embodiments, the linker comprises 1, 2, 3, 4, or 5 amino acids. In some embodiments, one or more amino acids of the linker are unnatural amino acids. In some embodiments, the linker comprises L-amino acids and / or D-amino acids.

[0210] In some embodiments, the optional linker comprises the amino acid sequence Gly-Gly-Gly or Gly-Ser-Gly.

[0211] In some embodiments, the optional linker is hydrophilic. In some embodiments, the optional linker is hydrophobic.

[0212] In some embodiments, at least one PRM is inserted at an internal position of the polypeptide.

[0213] In some embodiments, the polypeptide comprises a loop region and at least one PRM is inserted at or near the loop region of the polypeptide.Precision Labels - Compound of Formula (I)-59- SG Docket No.: 14989-700.600

[0214] Described herein are multifunctional compounds and strategies for precision conjugation in a controlled, efficient, reliable, and tailorable fashion for broad and advanced applications. The strategies described herein incorporate one or more peptidic recognition motif(s) (PRM) into therapeutic peptides while maintaining both peptide integrity and function. Selective noncovalent binding interactions of the compounds described herein with the PRM(s) facilitate a proximity-induced (also called binding-induced) irreversible covalent reaction between lysines and electrophilic warheads within the same compounds. The proximity -triggered conjugation compounds described herein are highly variable for use in a variety of bioconjugation strategies and uses as described herein.

[0215] In some embodiments, described herein is a compound of Formula (I):PBU-L1-L2-PCU(Formula (I))wherein:PBU is a moiety capable of non-covalently binding to a peptidic recognition motif (PRM) in a polypeptide, wherein PRM comprises:a) 2 to 4 histidine (His) residues; anda lysine (Lys) residue, ornithine (Orn) residue, homolysine (HLys), a p2- lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue; or b) amino acid residues comprising:at least 2 amino acids selected from an arginine (Arg) residue, a homoarginine (HArg) residue, a citrulline (Cit) residue, norarginine (AGBA) residue, a p2-arginine (P2-Arg), and a p3-arginine (P3-Arg); and a lysine (Lys) residue, an ornithine (Orn) residue, a homolysine (HLys) residue, a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue;PCU is a moiety capable of covalently reacting with the side chain of the at least one Lys, Orn, HLys, P2-Lys, or p3-Lys in the PRM upon binding of the PBU with the PRM;L1is a cleavable linker; andL2is a spacer.

[0216] In some embodiments, described herein is a compound of Formula (I): PBU-L1-!?-PCU (Formula (I)), wherein: PBU is a moiety capable of non-covalently binding to a peptidic recognition motif (PRM) in a polypeptide, wherein PRM comprises: at least 2 histidine (His) residues; and a lysine (Lys) residue, ornithine (Orn) residue, homolysine (HLys), a p2-lysine-60- SG Docket No.: 14989-700.600(P2-Lys) residue, or p3-lysine (P3-Lys) residue; or amino acid residues comprising: at least 2 amino acids selected from an arginine (Arg) residue, a homoarginine (HArg) residue, a citrulline (Cit) residue, norarginine (AGBA) residue, a p2-arginine (P2-Arg), and a p3-arginine (P3-Arg); and a lysine (Lys) residue, an ornithine (Om) residue, a homolysine (HLys) residue, a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue; PCU is a moiety capable of covalently reacting with the side chain of the at least one Lys, Om, HLys, p2-Lys, or p3-Lys in the PRM upon binding of the PBU with the PRM; L1is a cleavable linker; and L2is a spacer.PRM Non-Covalent Binding Units (PBU)

[0217] In some embodiments, PBU is a moiety capable of forming a transition metal complex with the side chains of histidines in PRM1 and a divalent transition metal.

[0218] In some embodiments, the divalent transition metal is nickel (Ni2+), cobalt (Co2+), copper (Cu2+), or zinc (Zn2+). In some embodiments, the divalent transition metal is nickel (Ni2+). In some embodiments, the divalent transition metal is cobalt (Co2+). In some embodiments, the divalent transition metal is copper (Cu2+). In some embodiments, the divalent transition metal is zinc (Zn2+).

[0219] In some embodiments, PBU is a moiety capable of non-covalently binding to the arginine (Arg) residue(s), homoarginine (HArg) residue(s), citrulline (Cit) residue(s), norarginine (AGBA) residue(s), p2-arginine (P2-Arg) residue(s), and / or p3-arginine (P3-Arg) residue(s) in PRM2.

[0220] In some embodiments, PBU comprises at least one nitrilotriacetic acid, at least one iminodiacetic acid, or at least one pyridin-2-ylmethanamine, or combinations thereof.

[0221] In some embodiments, PBU comprises at least one moiety that isome embodiments, PBUcomprises at least one moiety thatsome embodiments, PBUSG Docket No.: 14989-700.600comprises at least one moiety that

[0222] In some embodiments, PBU comprises one nitrilotriacetic acid. In some embodiments, PBU comprises two nitrilotriacetic acids. In some embodiments, PBU comprises three nitrilotriacetic acids. In some embodiments, PBU comprises one iminodiacetic acid. In some embodiments, PBU comprises two iminodiacetic acids. In some embodiments, PBU comprises three iminodiacetic acids.

[0223] In some embodiments, PBU comprises two nitrilotriacetic acids. In some embodiments, PBU comprises three nitrilotriacetic acids. In some embodiments, PBU comprises one iminodiacetic acid. In some embodiments, PBU comprises two iminodiacetic acids. In some embodiments, PBU comprises three iminodiacetic acids.and U1is CH, C-OH, or N. In some embodiments, U1is CH. In some embodiments, U1is C-OH. In some embodiments, U1is N.SG Docket No.: 14989-700.600,SG Docket No.: 14989-700.600

[0230] In some embodiments,

[0231] In some embodiments,

[0232] In some embodiments,

[0233] In some embodiments,

[0234] In some embodiments,PRM Conjugation Units (PCUs)

[0235] In some embodiments, PCU comprises an electrophilic warhead moiety that is empirically shown to be or expected to be by one skilled in the art, capable of covalently reacting with the at least one Lys, Om, or HLys of the PRM only when held in close proximity to the Lys, Om, or HLys of the PRM by binding of the PBU to the PRM.

[0236] In some embodiments, PCU comprises an electrophilic warhead moiety that is empirically shown to be, or expected to be by one skilled in the art, capable of covalently reacting with the at least one Lys, Orn, or HLys of the PRM only when held in closeSG Docket No.: 14989-700.600proximity to the Lys, Om, or HLys of the PRM by binding of the PBU to the PRM, wherein the electrophilic warhead moiety isa vinyl sulfone, a vinyl sulfonamide, a pyrimidine sulfoxide, a pyrimidine sulfone, a vinyl pyridine, or a vinyl pyrimidine.

[0237] In some embodiments, the electrophilic warhead is.; R1is H, halogen, substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, or -C(=O)OR3; R2is H, halogen, substituted or unsubstituted alkyl, or -CN; and each R3is independently H or substituted or unsubstituted alkyl.

[0238] In some embodiments, the electrophilic warhead is". n some emoments, te eectropc warea s . nsome embodiments, the electrophilic warhead issomeembodiments, the electrophilic warheadsome embodiments, theelectrophilic warhead is. In some embodiments, the electrophilic warhead isSG Docket No.: 14989-700.600In some embodiments, the electrophilic warhead is

[0239] In some embodiments, R1is H, halogen, methyl, ethyl, propyl, n-butyl, sec-butyl,tert-butyl, n-pentyl, neopentyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl,

[0240] In some embodiments, R2is H, halogen, methyl, ethyl, propyl, n-butyl, sec-butyl, tert-butyl, n-pentyl, neopentyl, or -CN. In some embodiments, R2is H, halogen, methyl, ethyl, or -CN. In some embodiments, R2is H.

[0241] In some embodiments, each R3is independently H, methyl, ethyl, propyl, n-butyl, sec-butyl, tert-butyl, n-pentyl, or neopentyl. In some embodiments, each R3is independently H, methyl, or ethyl. In some embodiments, each R3is independently H, or methyl. In some embodiments, each R3is methyl.

[0242] In some embodiments, R2is H, halogen, methyl, ethyl, propyl, n-butyl, sec-butyl, tert-butyl, n-pentyl, neopentyl, or -CN; and each R3is independently H, methyl, ethyl, propyl, tert-butyl, n-pentyl, or neopentyl.

[0243] In some embodiments, R1is H; R2is H; and each R3is methyl.

[0244] In some embodiments, PCU further comprises an optionally substituted aryl or an optionally substituted heteroaryl, and the electrophilic warhead is covalently attached to the optionally substituted aryl or the optionally substituted heteroaryl. In some embodiments,, substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, or -C(=O)OR3; R2is H, halogen, substituted or unsubstituted alkyl, or -CN; and each R3is independently H or substituted orSG Docket No.: 14989-700.600unsubstituted alkyl.

[0245] In some embodiments, PCU is

[0246] In some embodiments,some embodiments, PCU is, some embodiments, PCUisIn some embodiments,some embodiments,some embodiments,Compounds of Formula (I)

[0247] In some embodiments, the compound of Formula (I) has the following structure:

[0248] In some embodiments, the compound of Formula (I) has the following structure:SG Docket No.: 14989-700.600

[0249] In some embodiments, the compound of Formula (I) has the following structure:

[0250] In some embodiments, R1is H, halogen, methyl, ethyl, propyl, n-butyl, sec-butyl,tert-butyl, n-pentyl, neopentyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl,

[0251] In some embodiments, R2is H, halogen, methyl, ethyl, propyl, n-butyl, sec-butyl, tert-butyl, n-pentyl, neopentyl, or -CN. In some embodiments, R2is H, halogen, methyl, ethyl, or -CN. In some embodiments, R2is H.

[0252] In some embodiments, R3is H, methyl, ethyl, propyl, n-butyl, sec-butyl, tert-butyl, n-pentyl, or neopentyl. In some embodiments, R3is H, methyl, or ethyl. In some embodiments, R3is H or methyl. In some embodiments, R3is methyl.

[0253] In some embodiments, R2is H, halogen, methyl, ethyl, propyl, n-butyl, sec-butyl, tert-butyl, n-pentyl, neopentyl, or -CN; and each R3is independently H, methyl, ethyl, propyl, tert-butyl, n-pentyl, or neopentyl.

[0254] In some embodiments, R1is H; R2is H; and R3is methyl.Cleavable Linker (L1)

[0255] Compounds described herein include a linker connecting the PBU moiety, which is capable of non-covalently binding to a peptidic recognition motif (PRM) in a polypeptide,-68- SG Docket No.: 14989-700.600and the PCU moiety, which is capable of covalently reacting with the side chain of the at least one Lys, Om, HLys, p2-Lys, or p3-Lys in the PRM upon binding of the PBU with the PRM unit. Non-covalent binding of the PRM to the PBU is needed to hold the PCU electrophilic warhead moiety in close enough proximity to trigger covalent reaction and attachment of the PCU to the side chain of the at least one Lys, Om, HLys, p2-Lys, or p3-Lys. The linker connecting PBU to PCU comprises two components, L1, which comprises a cleavable linker, and L2, which comprises a spacer.

[0256] In some embodiments, L1comprises a cleavable linker. In some embodiments, L1comprises a cleavable linker that upon cleavage provides an electrophilic or nucleophilic moiety.

[0257] In some embodiments, the cleavable linker is orthogonally cleavable to peptide cleavage.

[0258] As used herein, “orthogonally cleavable to peptide cleavage” refers to cleavable linkers that are compatible with the biological molecules and systems. Cleavable linkers that are orthogonally cleavable to peptide cleavage can be cleaved without breaking peptide bonds in therapeutic peptides. Such cleavable linkers comprise chemically cleavable linkers.

[0259] Such cleavable linkers can be cleaved with nucleophilic / basic reagents such as, but not limited to halogen nucleophiles (e.g., fluoridolyzable linkers (e.g., dialkoxy silane, 2-cyanoethyl moiety); oxygen nucleophiles (e.g., hydroxyl anions) (e.g., sulfone linkers (which could be cleaved at basic pH), ester linkers (which could be cleaved by saponification)); thiols nucleophiles (e.g., thiophenylester linkers, vinyl sulfide linkers); nitrogen nucleophiles (cleavage by aminolysis or exchange reaction) (e.g., ester linker cleavage by hydroxyl-amine or hydrazine (e.g., cleavage of levulinoyl esters with hydrazine), acylhydrazone linkers and hydrazone linkers, which are cleavable through transimination in mildly acidic medium). See Leriche et al., Cleavable linkers in chemical biology, Bioorganic & Medicinal Chemistry, Geoffray, Volume 20, Issue 2, 15 January 2012, Pages 571-582, which is incorporated by reference in its entirety for such linkers, including the references disclosed therein describing such linkers.

[0260] Such cleavable linkers include safety-catch linkers that are sensitive to nucleophiles (cleavable) upon activation. Safety-catch linkers allow greater control over the timing of the bond breakage because the linker will remain stable until it is activated for cleavage by a chemical modification. Such cleavable linkers include acylsulfonamide linkers (activated by alkylation (e.g., cyanomethylation with ICH2CN) and cleaved with ammonium hydroxide; N--69- SG Docket No.: 14989-700.600alkylation using sodium iodoacetate following treatment with ammonium hydroxide).1. alkylation2. hydrolysis

[0261] Such cleavable linkers can be cleaved with reducing reagents (e.g., disulfide linkers, azo linkers). By way of example reducing agents include, but are not limited to, dithiothreitol (DTT), 2-mercaptoethanol, dithioerythritol, cysteine, cysteamine (2-aminoethanethiol), and reduced glutathione.

[0262] Such cleavable linkers include photocleavable linkers. The term “photocleavable linker,” as used herein, refers to a linker that breaks upon exposure to light. Photocleavable linkers include, but are not limited to:• ortho-nitrobenzyl linker cleaved at 300-365nm (X is NH or O);• phenacyl ester linker cleaved at 254 nm. The dashed line indicates bond that is cleaved.

[0263] Such cleavable linkers include electrophile / acid cleavable linkers (e.g., acidic cleavable linkers that are sensitive to proton sources, and alkyl 2-(diphenylphosphino)benzoate derivatives sensitive to azide compounds). Acid cleavable linkers include:

[0264] Acid-labile linkers include chemically cleavable linkers that respond to low pH. In some embodiments, the linker is pH-cleavable, i.e., sensitive to hydrolysis at certain pH values. For example, the pH-cleavable linker can be hydrolyzable under acidic conditions (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, but are unstable below pH 7.0, such as pH 6.5 to 4.5. See, e.g., Jan Anderl, et al. Linker Technologies for Antibody-Drug Conjugates. Antibody-Drug Conjugates, 2013:71-77; Alain-70- SG Docket No.: 14989-700.600Beck, et al. The next generation of antibody-drug conjugates comes of age. Discov Med.2010 Oct; 10(53):329-39.

[0265] Such cleavable linkers include oxidation cleavable linkers (e.g., vicinal diols).

[0266] Oxidation cleavable linkers also include selenium-based linkers:

[0267] In contrast, disulfide linkers breakdown under reducing environments. See, e.g., Jan Anderl, et al. Linker Technologies for Antibody-Drug Conjugates. Antibody -Drug Conjugates, 2013:77-80; Jessica R, et al. Antibody Drug Conjugates: Design and Selection of Linker, Payload and Conjugation Chemistry. AAPSJ. 2015 Mar; 17(2)339-51. Wu AM and Senter PD. Arming antibodies: prospects and challenges for immunoconjugates. Nat Biotechnol. 2005 Sep; 23(9): 1137-46.

[0268] Such cleavable linkers include self-immolative linkers. See Edupuganti et al., Self-immolative Linkers in Prodrugs and Antibody Drug Conjugates in Cancer Treatment. Recent Vat. Anticancer Drug Discov. 2021; 16(4): 479-497; Hamann etal. (2005) Expert Opin. Ther.Patents (2005) 15: 1087-103; Yam B. Poudel, et al., ACS Medicinal Chemistry Letters 2020 11 (11), 2190-2194.

[0269] Self-immolative linkers include para-aminobenzyl (PAB)-linkers. For example, cleavage of the imine formed with an aldehyde and para-aminobenzyl carbamate results in self-immolation:

[0270] Examples of self-immolative linkers further include, but are not limited to, aromatic compounds that are electronically similar to p-aminobenzyl alcohol (see, e.g., US 2005 / 0256030 Al), such as 2-aminoimidazol-5-methanol derivatives (Hay et al. (1999) Bioorg. Med. Chem. Lett. 9: 2237) and ortho- or para-aminobenzylacetals.

[0271] Cleavable linkers may include linkers formed with trans-cyclooctene:SG Docket No.: 14989-700.600

[0272] Linkers can be used that undergo cyclization upon amide bond hydrolysis, such as substituted and unsubstituted 4-aminobutyric acid amides (Rodrigues et al., Chemistry Biology, 1995, 2, 223); substituted bicyclo[2.2.1] and bicyclo[2.2.2] ring systems (Storm, et al., J. Amer. Chem. Soc., 1972, 94: 5815); and 2-aminophenylpropionic acid amides (Amsberry, et al., J. Org. Chem., 1990, 55: 5867).

[0273] In some embodiments, the linker is enzymatically cleavable. For example, P-glucuronide linkers can be broken down by P -glucuronidase without breaking peptide bonds. See, e.g., Jan Anderl, et al. Linker Technologies for Antibody-Drug Conjugates. Antibody-Drug Conjugates, 2013:85-86; Jessica R, et al. Antibody Drug Conjugates: Design and Selection of Linker, Payload and Conjugation Chemistry. AAPS J. 2015 Mar; 17(2):339-51 ; Scott Jeffrey. Beta-glucuronide-linker drug conjugates. Patent EP1912671 A2.SG Docket No.: 14989-700.600R5R6; R5is substituted or unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aryl or substituted or unsubstituted heteroaryl; and R6is H or substituted or unsubstituted alkyl; or R5and R6are taken together with the carbon atom to which they are attached to form a substituted or unsubstituted cycloalkyl.

[0276] In some embodiments, R5is sec-butyl, tert-butyl, n-pentyl, neopentyl, cyclopropyl,cyclobutyl, cyclopentyl, cyclohexyl, adamantyl, phenyl,Spacers

[0278] In some embodiments, the linker L2is a spacer and is not cleavable. In someSG Docket No.: 14989-700.600embodiments, the linker L2is flexible. In some embodiments, the linker is rigid. In some embodiments, the linker comprises a linear structure. In some embodiments, the linker comprises a non-linear structure. In some embodiments, the linker comprises a branched structure. In some embodiments, the linker comprises a cyclic structure. In some embodiments, the linker comprises one or more linear structures, one or more non-linear structures, one or more branched structures, one or more cyclic structures, one or more flexible moieties, one or more rigid moieties, or combinations thereof.

[0279] In some embodiments, the linker L2comprises one or more amino acid residues. In some embodiments, the linker comprises 1, 2, 3, 4, 5, 1 to 3, 1 to 4, or 1 to 5, amino acid residues. In some embodiments, one or more amino acids of the linker are unnatural amino acids.

[0280] In some embodiments the linker L2has 1 to 10 atoms in length. In some embodiments, the linker has 1 to 20 atoms in length.

[0281] In some embodiments, the linker L2comprises 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, proline-rich sequences, and regions rich in double and / or triple bonds.

[0282] In some embodiments, the linker L2comprises one or more of unsubstituted or substituted alkylene, unsubstituted or substituted cycloalkylene, unsubstituted or substituted heterocycloalkylene, unsubstituted or substituted arylene, and unsubstituted or substituted heteroarylene.

[0283] In some embodiments, the linker L2comprises:a bond, -alkylene-, -(alkylene-O)n-alkylene-, -alkylene-C(=O)-, -T-C(=O)-, -(alkylene-O)n-alkylene-C(=O)-, -alkylene-(alkylene-O)n-C(=O)-, -alkylene-T-alkylene-, -alkylene-NH-, -(alkylene-O)n-alkylene-NH-, -C(=O)-alkylene-NH-, -C(=O)-(alkylene-O)n-alkylene-NH-, -(alkylene-O)n-alkylene-Q-, -alkylene-Q-alkylene-, -alkylene-(alkylene-O)n-alkylene-C(O)-, -alkylene-Q-(alkylene-O)n-alkylene-, -(alkylene-O)n-alkylene-Q-alkylene, -Q-(alkylene-O)n-alkylene-, -Q-(alkylene-O)n-(alkylene-O)n-alkylene-, -(alkylene-O)n-alkylene-Q-(alkylene-O)n-alkylene-Q-, -C(=O)-AA-NH-alkylene-, -(alkylene-O)n-alkylene-C(=O)-AA-NH-alkylene-C(O)-, -(alkylene-O)n-alkylene-C(O)-AA-NH-alkylene-, -C(O)-alkylene-C(=O)-AA-NH-(alkylene-O)n-alkylene-, -alkylene-C(=O)-AA-NH-(alkylene-O)n-alkylene-, -alkylene-NH-AA-C(=O)-alkylene-, -(CH2)I-6--74- SG Docket No.: 14989-700.600substituted with one to three groups independently selected from the group consisting of -OH, -NH2, C1-C3 alkyl, C1-C3 alkoxy and C3-C6 cycloalkyl;wherein:each AA is independently an amino acid;each Q is independently:each T is independently cycloalkylene, heterocycloalkylene, arylene, or heteroarlyene; each alkylene is independently selected from the group consistingof: -(CH2)-, -(CH2)2-, -(CH2)3-, -(CH2)4-, -(CH2)5-, -(CH2)6-, -(CH2)7-, -(CH2)8-, -(CH2, -(CH2)IO-, -(CH2)H- and -(CH2)I2-; andeach n is independently an integer from 1 to 10;optionally, cycloalkylene, is a monocyclic cycloalkylene,optionally, heterocycloalkylene, is a monocyclic heterocycloalkylene,optionally heteroarlyene is a monocyclic heteroarlyene,optionally arylene is phenylene.

[0284] In some embodiments, the linker L2is absent or comprises one or more amino acids, PEG groups, -L3-, -L4-, -L5-, -L6-, -L7-, -L3-L4-, -L3-L4-L5-, -L3-L4-L5-L6-, -L3-L4-L5-L6-L7-, or a combination thereof;each L3is independently absent, -O-, -S-, -S(=O)-, -S(=O)2., -NH-, -CH(OH)-, -C(=O)-, - NHC(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -OC(=O)NH-, -NHC(=O)NH-, - NHC(=O)O-, -(CH2)P-, -C(=O)-(CH2CH2X)P-, or -(CH2CH2X)P-, each p is independently 1, 2, 3, 4, 5, or 6;each L4is independently absent, unsubstituted or substituted alkylene, unsubstituted or substituted heteroalkylene, unsubstituted or substituted alkenylene, unsubstituted or substituted alkynylene, unsubstituted or substituted cycloalkylene, unsubstituted or substituted heterocycloalkylene, unsubstituted or substituted arylene, unsubstituted or substituted heteroarylene, one or more amino acids,each L5is independently absent, -O-, -S-, -S(=O)-, -S(=O)2., -NH-, -CH(OH)-, -NHC(=O)-, - C(=O)O-, -OC(=O)-, -OC(=O)NH-, -NHC(=O)NH-, -NHC(=O)O-, -(CH2)P-, -C(=O)-, - (CH2)P-C(=O)-, -C(=O)-(CH2)P-, -(CH2)P-C(=O)-(CH2)P-, -C(=O)NH-, -C(=O)NH- (CH2)P-, -(CH2)q-C(=O)NH-, -(CH2)PC(=O)NH-(CH2)P-, -NHC(=O)-, -NHC(=O)-(CH2)P-, -(CH2)q-NHC(=O)-, -(CH2)PNHC(=O)-(CH2)P-, -NHC(=O)NH-, -NHC(=O)NH-(CH2)P-, --75- SG Docket No.: 14989-700.600(CH2)q-NHC(=O)NH-, -(CH2)pNHC(=O)NH-(CH2)p-, -(CH2)P-, -C(=O)-(CH2CH2X)P-, or -(CH2CH2X)P-, each p is independently 1, 2, 3, 4, 5, or 6;each L6is independently absent, unsubstituted or substituted alkylene, unsubstituted or substituted heteroalkylene, unsubstituted or substituted alkenylene, unsubstituted or substituted alkynylene, unsubstituted or substituted cycloalkylene, unsubstituted or substituted heterocycloalkylene, unsubstituted or substituted arylene, unsubstituted or substituted heteroarylene, one or more amino acids, -(CH2)q-, -(CH2CH2X)q-, or - (XCH2CH2)q-, each q is independently 1, 2, 3, 4, 5, or 6;each L7is independently absent, -O-, -S-, -S(O)-, -S(O)2., -NH-, -CH(OH)-, -C(=O)-, - C(=O)NH-, -NHC(=O)-, -C(=O)O-, -OC(=O)-, -OC(=O)O-, -OC(=O)NH-, - NHC(=O)NH-, or -NHC(=O)O-;each X is independently selected from O, S, and NRX; and each Rxis independently selectedSG Docket No.: 14989-700.600substituted or unsubstituted alkyl.substituted or unsubstituted alkyl.

[0287] In some embodiments, L2comprisessubstituted or unsubstituted alkyl.

[0288] In some embodiments, R4is H, methyl, ethyl, propyl, n-butyl, sec-butyl, tert-butyl, n-pentyl, or neopentyl. In some embodiments, R4is H or methyl. In some embodiments, R4is

[0289] For additional details regarding linkers and their use in the compounds described herein, see: Wu, A.M.; Senter, P.D. Arming antibodies: prospects and challenges for immunoconjugates. Nat. Biotechnol. 2005, 23(9): 1137-1146; Beck, A.; et al. The next generation of antibody-drug conjugates comes of age. Discov. Med. 2010, 10(53): 329-339; Nolting, B.; et al. Linker technologies for antibody-drug conjugates. Methods. Mol. Biol. 2013, 1045: 71-100; Jain, N.; et al. Current ADC linker chemistry. Pharm. Res. 2015, 32: 3526-3540; McCombs, J.R.; Owen, S.C. Antibody drug conjugates: design and selection of linker, payload and conjugation chemistry. AAPSJ. 2015, 17(2): 339-351; Jun Lu, etal., Linkers Having a Crucial Role in Antibody-Drug Conjugates, IntJMol Set. 2016 Apr; 17(4): 561; each of which is incorporated by reference for such linker disclosures.

[0290] In some embodiments, L1comprises a cleavable linker that upon cleavage provides a moiety capable of forming a linkage with a coupling partner, wherein the cleavable linker is orthogonally cleavable to peptide cleavage; L2further comprises a moiety capable of forming a linkage with a coupling partner; wherein the linkage formation reaction in L1is orthogonal to the linkage formation reaction in L2.SG Docket No.: 14989-700.600

[0291] In some embodiments, the linkage formation reaction in L1is a coupling reaction between an electrophile and a nucleophile.

[0292] In some embodiments, the coupling reaction is an amide-forming ligation, ester-forming ligation, ether-forming ligation, imine-forming ligation, oxime-forming ligation, hydrazone-forming ligation, or sulfonamide-forming ligation.

[0293] In some embodiments, the linkage formation reaction in L2is a cycloaddition reaction between an azide and alkyne to form a triazole, a strain-promoted azide-alkyne cycloaddition (SPAAC), inverse electron-demand Diels-Alder (IEDDA), or sydnone-alkyne cycloaddition (SPSAC).

[0294] In some embodiments, the IEDDA comprises an inverse electron demand [4 + 2] cycloaddition between a 1,2,4,5-tetrazine and a strained alkene dienophile or a strained alkyne dienophile.

[0295] In some embodiments, the dienophile group comprises a norbornene, transcyclooctene (TCO), cyclopropene or bicyclo[6.1.0]non-4-yn-9-ylmethanol (BCN).

[0296] In some embodiments, the SPAAC reaction comprises a reaction between an azide group and a strained alkyne.

[0297] In some embodiments, the strained alkyne is azadib enzocy cl ooctyne (ADIBO, DIB AC or DBCO), tetramethyldibenzocyclooctyne (TMDIBO), cyclooctyne (OCT), aryl-less cyclooctyne (ALO), monofluorocyclooctyne (MOFO), difluorocyclooctyne (DIFO), dibenzocyclooctyne (DIBO), dimethoxyazacyclooctyne (DIMAC), biarylazacyclooctynone (BARAC), bicyclononyne (BCN), tetramethylthiepinium (TMTH), difluorobenzocyclooctyne (DIFBO), carboxymethylmonobenzocyclooctyne (COMBO) or benzocyclononyne.

[0298] In some embodiments, -L^L2- is:

[0299] In some embodiments, the compound has the following structure:-78- SG Docket No.: 14989-700.600

[0300] In some embodiments, the compound has the following structure:R5is substituted or unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aryl or substituted or unsubstituted heteroaryl; and R6is H or substituted or unsubstituted alkyl; orR5and R6are taken together with the carbon atom to which they are attached to form a substituted or unsubstituted cycloalkyl;L2is a linker; andRcis azide or a substituted or unsubstituted 1,2,4,5-tetrazine.

[0301] In some embodiments, R5is sec-butyl, tert-butyl, n-pentyl, neopentyl, cyclopropyl,cyclobutyl, cyclopentyl, cyclohexyl, adamantyl, phenyl,

[0302] In some embodiments, L2is an alkyl linker or a PEG linker.

[0303] In some embodiments, L2is -(CH2)t- or -(CH2CH2O)v-CH2CH2-; t is 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12; and v is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12.

[0304] In some embodiments, described herein is a compound having the structure (Compound 14):SG Docket No.: 14989-700.600

[0305] In some embodiments, described herein is a compound having the structure (Compound 15):

[0306] In some embodiments, described herein is a compound having the structure (Compound 16):

[0307] In some embodiments, described herein is a compound having the structure:SG Docket No.: 14989-700.600wherein,U1is CH, C-OH, orN.L1is a cleavable linker; L2is a spacer;R3is H or substituted or unsubstituted alkyl;His is histidine;m is 0, 1, 2, 3, or 4; n is 0, 1, 2, 3, or 4, provided that m + n is at least 2; q is 1, 2, or 3;Rais a polypeptide B, and Rbis -OH; or Rais H, and Rbis a polypeptide B; or Rais a fragment (B1) of a polypeptide B, Rbis the other fragment (B2) of the polypeptide B, such that B^B2is B; orRais H, and Rbis -OH.

[0308] In some embodiments, described herein is a compound having the structure:wherein,U1is CH, C-OH, orN.L1is a cleavable linker; L2is a spacer;each R3is H or substituted or unsubstituted alkyl;q is 1, 2, or 3;Rais a polypeptide, and Rbis -OH; or Rais H, and Rbis a polypeptide; or Rais a fragment (B1) of a polypeptide B, Rbis the other fragment (B2) of the polypeptide B, such that B^B2is B; orRais H, and Rbis -OH.

[0309] In some embodiments, described herein is a compound having the structure:-81- SG Docket No.: 14989-700.600wherein,U1is CH, C-OH, orN;L1is a cleavable linker; L2is a spacer;each R3is H or substituted or unsubstituted alkyl;W1is a -X^Y-X^Y-, and W2is absent; or W1is absent, and W2is -X^Y-X^Y-; or W1and W2are both -X1-Y-X2-Y-;each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg; X1comprises 1 to 6 amino acid residues other than Lys, Orn, HLys, and Y;X2comprises 1 to 6 amino acids other than Lys, Orn, HLys, and Y;q is 1, 2, or 3;Rais a polypeptide B, and Rbis -OH; or Rais H, and Rbis a polypeptide B; or Rais a fragment (B1) of a polypeptide B, Rbis the other fragment (B2) of the polypeptide B, such that B^B2is B; orRais H, and Rbis -OH.

[0310] In some embodiments, described herein is a method of selectively modifying a lysine residue within a polypeptide comprising: contacting a compound of Formula (I) described herein with a polypeptide described herein in the presence of a nickel (Ni2+) salt, cobalt (Co2+) salt, copper (Cu2+) salt, or zinc (Zn2+) salt, in a suitable solvent, wherein the polypeptide comprises at least one PRM1. In some embodiments, the lysine residue that is selectively modified is the lysine in the at least one PRM1.

[0311] In some embodiments, described herein is a method of selectively modifying a lysine residue within a polypeptide comprising: contacting a compound of Formula (I), with a polypeptide, in a suitable solvent in the absence of a transition metal salt; wherein the polypeptide comprises at least one PRM2. In some embodiments, the lysine residue that is selectively modified is the lysine in the at least one PRM2.

[0312] In some embodiments, described herein is a method of selectively modifying at least-82- SG Docket No.: 14989-700.600two lysine residues within a polypeptide comprising: contacting a compound of Formula (I), with a polypeptide, under conjugation condition i) followed by conjugation condition ii), or under conjugation condition ii) followed by conjugation condition i); wherein conjugation condition i) comprises the use of a nickel (Ni2+) salt, cobalt (Co2+) salt, copper (Cu2+) salt, or zinc (Zn2+) salt, in a suitable solvent; wherein conjugation condition ii) comprises a suitable solvent, and the absence of a transition metal salt; wherein the polypeptide comprises at least one PRM1 and at least one PRM2. In some embodiments, conjugation condition i) selectively modifies the lysine in the at least one PRM1. In some embodiments, conjugation condition ii) selectively modifies the lysine in the at least one PRM2.

[0313] The methods of selective conjugation described herein provide a straightforward and high yielding method for controlling the identity, number, and location of conjugated moieties to polypeptides by controlling the number of PRM1 and / or PRM2 that are added to the polypeptides. In some embodiments, the polypeptide comprises at least one PRM1, at least one PRM2, or at least one PRM1 and at least one PRM2, wherein at least one PRM is 1) added to the N-terminus of the biologically active polypeptide, 2) added to the C-terminus of the biologically active polypeptide, and / or 3) inserted at an internal position of the biologically active polypeptide. In some embodiments, the cleavable linker of the compound of Formula (I) that reacts with PRM1 is different than the cleavable linker of the compound of Formula (I) that reacts with PRM2. In some embodiments, the cleavable linker of the compound of Formula (I) that reacts with PRM1 is the same as the cleavable linker of the compound of Formula (I) that reacts with PRM2. In some embodiments, the cleavable linker of the compound of Formula (I) that reacts with PRM1 is different than the cleavable linker of the compound of Formula (I) that reacts with PRM2, and the cleavable linker of the compound of Formula (I) that reacts with PRM1 is cleaved under conditions that do not cleave the cleavable linker of the compound of Formula (I) that reacts with PRM2. In some embodiments, the cleavable linker of the compound of Formula (I) that reacts with PRM1 is cleaved under conditions that do not cleave the cleavable linker of the compound of Formula (I) that reacts with PRM2. In some embodiments, the cleavable linker of the compound of Formula (I) that reacts with PRM2 is cleaved under conditions that do not cleave the cleavable linker of the compound of Formula (I) that reacts with PRM1. In some embodiments, the cleavable linker of the compound of Formula (I) that reacts with PRM1 is cleaved under conditions that do not cleave the cleavable linker of the compound of Formula (I) that reacts with PRM2; and the cleavable linker of the compound of Formula (I) that reacts-83- SG Docket No.: 14989-700.600with PRM2 is cleaved under conditions that do not cleave the cleavable linker of the compound of Formula (I) that reacts with PRM1. In some embodiments, the polypeptide comprises one PRM1, one PRM2, or one PRM1 and one PRM2. In some embodiments, the polypeptide comprises one, two or three PRM1, one, two or three PRM2, or one, two or three PRM1 and one, two or three PRM2.Further Forms of Compounds

[0314] In one aspect, compounds described herein are in the form of salts. In addition, the compounds described herein can exist in unsolvated as well as solvated forms with solvents such as water, ethanol, and the like. The solvated forms of the compounds presented herein are also considered to be disclosed herein.

[0315] Salts typically are more soluble than the non-salt forms. In some embodiments, salts are obtained by reacting a compound disclosed herein with an acid. In some embodiments, the compound disclosed herein (i.e., free base form) is basic and is reacted with an organic acid or an inorganic acid. Inorganic acids include, but are not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid, and metaphosphoric acid.Organic acids include, but are not limited to, l-hydroxy-2-naphthoic acid; 2,2-dichloroacetic acid; 2-hydroxyethanesulfonic acid; 2-oxoglutaric acid; 4-acetamidobenzoic acid; 4-aminosalicylic acid; acetic acid; adipic acid; ascorbic acid (L); aspartic acid (L); benzenesulfonic acid; benzoic acid; camphoric acid (+); camphor- 10-sulfonic acid (+); capric acid (decanoic acid); caproic acid (hexanoic acid); caprylic acid (octanoic acid); carbonic acid; cinnamic acid; citric acid; cyclamic acid; dodecylsulfuric acid; ethane-l,2-disulfonic acid; ethanesulfonic acid; formic acid; fumaric acid; galactaric acid; gentisic acid; glucoheptonic acid (D); gluconic acid (D); glucuronic acid (D); glutamic acid; glutaric acid; glycerophosphoric acid; glycolic acid; hippuric acid; isobutyric acid; lactic acid (DL); lactobionic acid; lauric acid; maleic acid; malic acid (- L); malonic acid; mandelic acid (DL); methanesulfonic acid; naphthalene-l,5-disulfonic acid; naphthalene-2-sulfonic acid; nicotinic acid; oleic acid; oxalic acid; palmitic acid; pamoic acid; phosphoric acid; proprionic acid; pyroglutamic acid (- L); salicylic acid; sebacic acid; stearic acid; succinic acid; sulfuric acid; tartaric acid (+ L); thiocyanic acid; toluenesulfonic acid ( ); and undecylenic acid.

[0316] In some embodiments, salts are obtained by reacting a compound disclosed herein with a base. In some embodiments, the compound disclosed herein is acidic and is reacted with a base. In such situations, an acidic proton of the compound disclosed herein is replaced by a metal ion, e.g., lithium, sodium, potassium, magnesium, calcium, or an aluminum ion. In-84- SG Docket No.: 14989-700.600some cases, compounds described herein coordinate with an organic base, such as, but not limited to, ethanolamine, diethanolamine, triethanolamine, tromethamine, meglumine, N-methylglucamine, dicyclohexylamine, tris(hydroxymethyl)methylamine. In other cases, compounds described herein form salts with amino acids such as, but not limited to, arginine, lysine, and the like. Acceptable inorganic bases used to form salts with compounds that include an acidic proton, include, but are not limited to, aluminum hydroxide, calcium hydroxide, potassium hydroxide, sodium carbonate, potassium carbonate, sodium hydroxide, lithium hydroxide, and the like. In some embodiments, the compounds provided herein are prepared as a sodium salt, calcium salt, potassium salt, magnesium salt, meglumine salt, N-methylglucamine salt or ammonium salt.

[0317] It should be understood that a reference to a salt includes the solvent addition forms. In some embodiments, solvates contain either stoichiometric or non-stoichiometric amounts of a solvent, and 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 compounds described herein are conveniently prepared or formed during the processes described herein. In addition, the compounds provided herein optionally exist in unsolvated as well as solvated forms.

[0318] In another embodiment, the compounds described herein are labeled isotopically (e.g. with a radioisotope) or by another other means, including, but not limited to, the use of chromophores or fluorescent moieties, bioluminescent labels, or chemiluminescent labels.

[0319] Compounds described herein include isotopically-labeled compounds, which are identical to those recited in the various formulae and structures presented herein, but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature. Examples of isotopes that can be incorporated into the present compounds include isotopes of hydrogen, carbon, nitrogen, oxygen, sulfur, fluorine chlorine, iodine, phosphorus, such as, for example,2H,3H,13C,14C,15N,18O,170,35S,18F,36C1,123I,124I,125I,1311,32P and33P. In one aspect, isotopically-labeled compounds described herein, for example those into which radioactive isotopes such as3H and14C are incorporated, are useful in drug and / or substrate tissue distribution assays. In one aspect, substitution with isotopes such as deuterium affords certain therapeutic advantages resulting from greater metabolic stability, such as, for example, increased in vivo half-life or reduced dosage requirements.-85- SG Docket No.: 14989-700.600Certain Terminology

[0320] Unless otherwise stated, the following terms used in this application have the definitions given below. The use of the term “including” as well as other forms, such as “include”, “includes,” and “included,” is not limiting.

[0321] As used herein, the term “about,” in the context of a given value or range, includes and / or refers to a value or range that is within 10% of the given value or range.

[0322] As used herein, Ci-Cxincludes 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, zso-propyl, / / -butyl, zso-butyl, .scc-butyl, and / -butyl.

[0323] The term “acyl,” as used herein refers to the group -C(=O)-R, where R is alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroaryl, heterocycle, or any other moiety were the atom attached to the carbonyl is carbon. An “acetyl” group refers to a -C(=O)CH3 group.

[0324] The term “alkenyl,” as used herein refers to a straight-chain or branched-chain hydrocarbon radical having one or more double bonds and containing from 2 to 20 carbon atoms. In certain embodiments, alkenyl includes 2 to 6 carbon atoms. The term “alkenylene” refers to a divalent alkenyl. In some embodiments, an alkenyl is selected from ethenyl ( / .< ., vinyl), propenyl (i.e., allyl), butenyl, pentenyl, pentadienyl, and the like. Non-limiting examples of an alkenyl group include -CH=CH2, -C(CH3)=CH2, -CH=CHCH3, -C(CH3)=CHCH3, and -CH2CH=CH2.

[0325] The term “alkoxy” refers to a (alkyl)-O- group, where alkyl is as defined herein. In some embodiments, the alkoxy group is a Ci-Cealkoxy, which refers to a (Ci-C6alkyl)-O-group. Examples of alkyl groups include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, iso-butoxy, sec-butoxy, tert-butoxy, and the like.

[0326] An “alkyl” group refers to an aliphatic hydrocarbon group. In some embodiments, the alkyl is a straight-chain or branched-chain aliphatic hydrocarbon group containing from 1 to 20 carbon atoms. In certain embodiments, alkyl includes 1 to 10 carbon atoms. In further embodiments, the alkyl includes 1 to 8 carbon atoms. Examples of alkyl radicals include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, iso-amyl, hexyl, octyl, nonyl, and the like. In some embodiments, an alkyl is a Ci-Cealkyl. In one aspect the alkyl is methyl, ethyl, propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, or t-butyl.-86- SG Docket No.: 14989-700.600The term “alkylene” refers to a divalent alkyl, such as methylene (-CH2-). In some embodiments, an alkylene is a Ci-Cealkylene. In other embodiments, an alkylene is a Ci-C4alkylene. Typical alkylene groups include, but are not limited to, -CH2-, -CH2CH2-, -CH2CH2CH2-, -CH2CH2CH2CH2-, and the like.

[0327] The term “amino,” as used herein refers to -NRR , wherein R and R are independently selected from hydrogen, alkyl, acyl, heteroalkyl, aryl, cycloalkyl, heteroaryl, and heterocycloalkyl, any of which may themselves be optionally substituted. Additionally, R and R’ may combine to form heterocycloalkyl, either of which may be optionally substituted. In one aspect, “amino” as used herein refers to an -NH2 group.

[0328] The term “alkynyl,” as used herein refers to a straight-chain or branched chain hydrocarbon radical having one or more triple bonds and containing from 2 to 20 carbon atoms. In certain embodiments, said alkynyl comprises from 2 to 6 carbon atoms. In further embodiments, said alkynyl comprises from 2 to 4 carbon atoms. In one embodiment, an alkenyl group has the formula -C=C-R, wherein R refers to the remaining portions of the alkynyl group. In some embodiments, R is H or an alkyl. In some embodiments, an alkynyl is selected from ethynyl, propynyl, butynyl, pentynyl, hexynyl, and the like. Non-limiting examples of an alkynyl group include -C=CH, -OCCH3 -OCCH2CH3, -CH2OCH. The term “alkynylene” refers to a carbon-carbon triple bond attached at two positions such as ethynylene (-C=C-). Examples of alkynyl radicals include ethynyl, propynyl, hydroxypropynyl, butyn-l-yl, butyn-2-yl, pentyn-l-yl, 3-methylbutyn-l-yl, hexyn-2-yl, and the like. Unless otherwise specified, the term “alkynyl” may include “alkynylene” groups.

[0329] The term “aromatic” refers to a planar ring having a delocalized ^-electron system containing 4n+271 electrons, where n is an integer. The term “aromatic” includes both carbocyclic aryl (“aryl”, e.g., phenyl) and heterocyclic aryl (or “heteroaryl” or “heteroaromatic”) groups (e.g., pyridine). The term includes monocyclic or fused-ring polycyclic (i.e., rings which share adjacent pairs of carbon atoms) groups.

[0330] The term “carbocyclic” or “carbocycle” refers to a ring or ring system where the atoms forming the backbone of the ring are all carbon atoms. The term thus distinguishes carbocyclic from “heterocyclic” rings or “heterocycles” in which the ring backbone contains at least one atom which is different from carbon. In some embodiments, at least one of the two rings of a bicyclic carbocycle is aromatic. In some embodiments, both rings of a bicyclic carbocycle are aromatic. Carbocycles include aryls and cycloalkyls.

[0331] The term “aryl” as used herein means a carbocyclic aromatic system containing one,-87- SG Docket No.: 14989-700.600two or three rings wherein such polycyclic ring systems are fused together. The term "aryl" embraces aromatic groups such as phenyl, naphthyl, anthracenyl, and phenanthryl. In one aspect, aryl is phenyl or a naphthyl. In some embodiments, an aryl is a phenyl. In some embodiments, an aryl is a phenyl, naphthyl, indanyl, indenyl, or tetrahyodronaphthyl. In some embodiments, an aryl is a Ce-Cioaryl. Depending on the structure, an aryl group is a monoradical or a diradical (i.e., an arylene group).

[0332] The terms “benzo” and “benz,” as used herein refer to fused bicyclic or polyclic ring system that is formed with benzene as one of the rings. Examples include benzofuran, benzothiophene, and benzimidazole.

[0333] The term “cycloalkyl,” as used herein refers to a saturated or partially saturated monocyclic, bicyclic or tricyclic alkyl group wherein each cyclic moiety contains from 3 to 12 carbon atom ring members and which may optionally be a benzo fused ring system which is optionally substituted as defined herein. In some embodiments, cycloalkyl groups include groups having from 3 to 10 ring atoms. In certain embodiments, said cycloalkyl will comprise from 5 to 7 carbon atoms. In certain embodiments, said cycloalkyl will comprise from 3 to 6 carbon atoms. Examples of such cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, tetrahydronaphthyl, indanyl, octahydronaphthyl, 2,3-dihydro-lH-indenyl, adamantly, and the like. “Bicyclic” and “tricyclic” as used herein are intended to include both fused ring systems, such as decahydronaphthalene, octahydronaphthalene as well as the multicyclic (multi centered) saturated or partially unsaturated type. The latter type of isomer is exemplified in general by, bicyclo[l,l,l]pentane, camphor, adamantane, and bicyclo[3,2,l]octane. In some embodiments, a cycloalkyl is a Cs-Cecycloalkyl. In some embodiments, a cycloalkyl is a C3-C4cycloalkyl.

[0334] The term "heterocycle" or “heterocyclic” refers to heteroaromatic rings (also known as heteroaryls) and heterocycloalkyl rings containing one to four heteroatoms in the ring(s), where each heteroatom in the ring(s) is selected from O, S and N, wherein each heterocyclic group has from 3 to 10 atoms in its ring system, and with the proviso that any ring does not contain two adjacent O or S atoms. Non-aromatic heterocyclic groups (also known as heterocycloalkyls) include rings having 3 to 10 atoms in its ring system and aromatic heterocyclic groups include rings having 5 to 10 atoms in its ring system. The heterocyclic groups include benzo-fused ring systems. Examples of non-aromatic heterocyclic groups are pyrrolidinyl, tetrahydrofuranyl, dihydrofuranyl, tetrahydrothienyl, oxazolidinonyl,-88- SG Docket No.: 14989-700.600tetrahydropyranyl, dihydropyranyl, tetrahydrothiopyranyl, piperidinyl, morpholinyl, thiomorpholinyl, thioxanyl, piperazinyl, aziridinyl, azetidinyl, oxetanyl, thietanyl, homopiperidinyl, oxepanyl, thiepanyl, oxazepinyl, diazepinyl, thiazepinyl, 1, 2,3,6-tetrahydropyridinyl, pyrrolin-2-yl, pyrrolin-3-yl, indolinyl, 2H-pyranyl, 4H-pyranyl, dioxanyl, 1,3-dioxolanyl, pyrazolinyl, dithianyl, dithiolanyl, dihydropyranyl, dihydrothienyl, dihydrofuranyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, 3-azabicyclo[3.1.0]hexanyl, 3-azabicyclo[4.1.0]heptanyl, 3H-indolyl, indolin-2-onyl, isoindolin-l-onyl, isoindoline-1, 3-dionyl, 3,4-dihydroisoquinolin-l(2H)-onyl, 3,4-dihydroquinolin-2(lH)-onyl, isoindoline-1, 3-dithionyl, benzo[d]oxazol-2(3H)-onyl, lH-benzo[d]imidazol-2(3H)-onyl, benzo[d]thiazol-2(3H)-onyl, and quinolizinyl. Examples of aromatic heterocyclic groups are pyridinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, quinolinyl, isoquinolinyl, indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl, indolizinyl, phthalazinyl, pyridazinyl, triazinyl, isoindolyl, pteridinyl, purinyl, oxadiazolyl, thiadiazolyl, furazanyl, benzofurazanyl, benzothiophenyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinoxalinyl, naphthyridinyl, and furopyridinyl. The foregoing groups are either C-attached (or C-linked) or TV-attached where such is possible. For instance, a group derived from pyrrole includes both pyrrol-l-yl (TV-attached) or pyrrol-3-yl (C-attached). Further, a group derived from imidazole includes imidazol-l-yl or imidazol-3-yl (both TV-attached) or imidazol-2-yl, imidazol-4-yl or imidazol-5-yl (all C-attached). The heterocyclic groups include benzo-fused ring systems. Nonaromatic heterocycles are optionally substituted with one or two oxo (=0) moieties, such as pyrrolidin-2-one. In some embodiments, at least one of the two rings of a bicyclic heterocycle is aromatic. In some embodiments, both rings of a bicyclic heterocycle are aromatic.

[0335] The terms “heteroaryl” or, alternatively, “heteroaromatic” refers to an aryl group that includes one or more ring heteroatoms selected from nitrogen, oxygen and sulfur. In some embodiments, the term "heteroaryl," as used herein refers to a 3 to 15 membered unsaturated heteromonocyclic ring, or a fused monocyclic, bicyclic, or tricyclic ring system in which at least one of the fused rings is aromatic, which contains at least one atom selected from N, O, and S. In certain embodiments, said heteroaryl will comprise from 1 to 4 heteroatoms as ring members. In further embodiments, said heteroaryl will comprise from 1 to 2 heteroatoms as ring members. In certain embodiments, said heteroaryl will comprise from 5 to 7 atoms. The term also embraces fused polycyclic groups wherein heterocyclic rings are fused with aryl rings, wherein heteroaryl rings are fused with other heteroaryl rings,-89- SG Docket No.: 14989-700.600wherein heteroaryl rings are fused with heterocycloalkyl rings, or wherein heteroaryl rings are fused with cycloalkyl rings. Examples of heteroaryl groups include pyrrolyl, imidazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazinyl, triazolyl, furyl, thienyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, thiadiazolyl, isothiazolyl, indolyl, isoindolyl, indolizinyl, benzimidazolyl, quinolyl, isoquinolyl, quinoxalinyl, quinazolinyl, indazolyl, benzotri azolyl, benzodi oxolyl, benzopyranyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, benzothiadiazolyl, benzofuranyl, benzothienyl, chromonyl, coumarinyl, benzopyranyl, tetrahydroquinolinyl, tetrazolopyridazinyl, tetrahydroisoquinolinyl, thienopyridinyl, furopyridinyl, pyrrolopyridinyl, and the like. Exemplary tricyclic heterocyclic groups include carbazolyl, benzidolyl, phenanthrolinyl, dibenzofuranyl, acridinyl, phenanthridinyl, xanthenyl, and the like. In some embodiments, a heteroaryl contains 0-4 N atoms in the ring. In some embodiments, a heteroaryl contains 1-4 N atoms in the ring. In some embodiments, a heteroaryl contains 1 O atom in the ring. In some embodiments, a heteroaryl contains 1 S atom in the ring. In some embodiments, a heteroaryl contains 1-4 N atoms, 0-1 0 atoms, and 0-1 S atoms in the ring. In some embodiments, heteroaryl is a Ci-Cgheteroaryl. In some embodiments, monocyclic heteroaryl is a Ci-Csheteroaryl. In some embodiments, monocyclic heteroaryl is a 5-membered or 6-membered heteroaryl. In some embodiments, bicyclic heteroaryl is a Ce-Cgheteroaryl.

[0336] A “heterocycloalkyl” group refers to a cycloalkyl group that includes at least one heteroatom selected from nitrogen, oxygen and sulfur. In some embodiments, the term “heterocycloalkyl” as used herein each refer to a saturated, partially unsaturated, or fully unsaturated (but nonaromatic) monocyclic, bicyclic, or tricyclic heterocyclic group containing at least one heteroatom as a ring member, wherein each said heteroatom may be independently selected from nitrogen, oxygen, and sulfur. In certain embodiments, said hetercycloalkyl will comprise from 1 to 4 heteroatoms as ring members. In further embodiments, said hetercycloalkyl will comprise from 1 to 2 heteroatoms as ring members. In certain embodiments, said hetercycloalkyl will comprise from 3 to 8 ring members in each ring. In further embodiments, said hetercycloalkyl will comprise from 3 to 7 ring members in each ring. In yet further embodiments, said hetercycloalkyl will comprise from 5 to 6 ring members in each ring. “Heterocycloalkyl” and “heterocycle” are intended to include sulfones, sulfoxides, N-oxides of tertiary nitrogen ring members, and carbocyclic fused and benzo fused ring systems; additionally, both terms also include systems where a heterocycle ring is fused to an aryl group, as defined herein, or an additional heterocycle group. Examples of-90- SG Docket No.: 14989-700.600heterocycle groups include aziridinyl, azetidinyl, 1,3-benzodioxolyl, dihydroisoindolyl, dihydroisoquinolinyl, dihydrocinnolinyl, dihydrobenzodioxinyl, dihydrofl, 3]oxazolo[4, 5-b]pyridinyl, benzothiazolyl, dihydroindolyl, dihydropyridinyl, 1,3-dioxanyl, 1,4-dioxanyl, 1,3-dioxolanyl, isoindolinyl, morpholinyl, piperazinyl, pyrrolidinyl, tetrahydropyridinyl, piperidinyl, thiomorpholinyl, and the like. The heterocycle groups may be optionally substituted unless specifically prohibited. In one aspect, a heterocycloalkyl is a C2-Cioheterocycloalkyl. In another aspect, a heterocycloalkyl is a C4-Cioheterocycloalkyl. In some embodiments, a heterocycloalkyl is monocyclic or bicyclic. In some embodiments, a heterocycloalkyl is monocyclic and is a 3, 4, 5, 6, 7, or 8-membered ring. In some embodiments, a heterocycloalkyl is monocyclic and is a 3, 4, 5, or 6-membered ring. In some embodiments, a heterocycloalkyl is monocyclic and is a 3 or 4-membered ring. In some embodiments, a heterocycloalkyl contains 1-2 N atoms in the ring. In some embodiments, a heterocycloalkyl contains 1-20 atoms. In some embodiments, a heterocycloalkyl contains 1 S atom. In some embodiments, a heterocycloalkyl contains 0-2 N atoms, 0-2 O atoms and 0-1 S atoms in the ring.

[0337] The term “carbamate,” as used herein refers to an ester of carbamic acid (-NHCOO-) which may be attached to the parent molecular moiety from either the nitrogen or acid end, and which may be optionally substituted as defined herein.

[0338] The term “carboxyl” or “carboxy,” as used herein, refers to -C(=O)OH or the corresponding “carboxylate” anion, such as is in a carboxylic acid salt.

[0339] The term “cyano,” as used herein refers to -CN.

[0340] The term “ester,” as used herein refers to a carboxy group bridging two moieties linked at carbon atoms.

[0341] The term “ether,” as used herein refers to an oxy group bridging two moieties linked at carbon atoms.

[0342] The term “halo,” or “halogen,” as used herein refers to fluorine, chlorine, bromine, or iodine. In some embodiments, halo is fluoro, chloro, or bromo.

[0343] The term “haloalkyl,” as used herein refers to an alkyl radical having the meaning as defined above wherein one or more hydrogens are replaced with a halogen. Specifically embraced are monohaloalkyl, dihaloalkyl and polyhaloalkyl radicals. A monohaloalkyl radical, for one example, may have an iodo, bromo, chloro or fluoro atom within the radical. Dihalo and polyhaloalkyl radicals may have two or more of the same halo atoms or a combination of different halo radicals. Examples of haloalkyl radicals include fluoromethyl,-91- SG Docket No.: 14989-700.600difluoromethyl, trifluoromethyl, chloromethyl, dichloromethyl, trichloromethyl, pentafluoroethyl, heptafluoropropyl, difluorochloromethyl, dichlorofluoromethyl, difluoroethyl, difluoropropyl, di chloroethyl and di chloropropyl. “Haloalkylene” refers to a haloalkyl group attached at two or more positions. Examples include fluoromethylene (-CFH-), difluoromethylene (-CF2 -), chloromethylene (-CHC1-), and the like. In one aspect, a haloalkyl is a Ci-Cehaloalkyl. In another aspect, a haloalkyl is a Ci-C4haloalkyl.

[0344] The term “haloalkoxy,” as used herein refers to a haloalkyl group attached to the parent molecular moiety through an oxygen atom. In one aspect, the haloalkoxy is a Ci-Cehaloalkoxy, which refers to a (Ci-C6haloalkyl)-O- group. In another aspect, the haloalkoxy is a Ci-C4haloalkoxy, which refers to a (Ci-C4haloalkyl)-O- group.

[0345] The term “heteroalkyl” refers to an alkyl wherein 1 or more carbon atoms are replaced with a heteroatom. In some embodiments, “heteroalkyl” refers to an alkyl wherein 1 or more carbon atoms are replaced with one or more heteroatoms that are independently selected from NH, -N(alkyl), O, S, S(=O) and S(=O)2. The attachment of the heteroatom(s) to the remainder of the compound is at a carbon atoms of the heteroalkyl. In some embodiments, up to two heteroatoms may be consecutive, such as, for example, -CH2-NH-OCH3. In some embodiments, “heteroalkyl” is an “alkoxyalkyl”, “alkylthioalkyl”, or “alkylaminoalkyl”. “Alkoxyalkyl” refers to an alkyl in which one hydrogen atom is replaced by an alkoxy group, as defined herein. In some embodiments, an alkoxyalkyl is a (Ci-C6alkoxy)-Ci-C6alkyl. Typical alkoxyalkyl groups include, but are not limited to, -CH2OCH3, -CH2CH2OCH3, -CH2CH2CH2OCH3, -CH2CH2CH2CH2OCH3, -CH2OCH2CH3, -CH2CH2OCH2CH3, -CH2CH2CH2OCH2CH3, -CH2CH2CH2CH2OCH2CH3, and the like.“Alkylthioalkyl” refers to an alkyl in which one hydrogen atom is replaced by an alkylthio group, as defined herein. In some embodiments, an alkoxyalkyl is a (Ci-Ce alkylthio)-Ci-Cealkyl. Typical alkoxyalkyl groups include, but are not limited to, -CH2SCH3, -CH2CH2SCH3, -CH2CH2CH2SCH3, -CH2CH2CH2CH2SCH3, -CH2SCH2CH3, -CH2CH2SCH2CH3, -CH2CH2CH2SCH2CH3, -CH2CH2CH2CH2SCH2CH3, and the like.“Alkylaminoalkyl” refers to an alkyl in which one hydrogen atom is replaced by an alkylamino group, as defined herein. In some embodiments, an alkoxyalkyl is a (Ci-C6alkylamino)-Ci-C6alkyl. Typical alkoxyalkyl groups include, but are not limited to, -CH2NHCH3, -CH2CH2NHCH3, -CH2CH2CH2NHCH3, -CH2CH2CH2CH2NHCH3, -CH2NHCH2CH3, -CH2CH2NHCH2CH3, -CH2CH2CH2NHCH2CH3, -CH2CH2CH2CH2NHCH2CH3, and the like.SG Docket No.: 14989-700.600

[0346] The term “hydroxy,” or “hydroxyl,” as used herein refers to -OH.

[0347] The term “hydroxyalkyl,” as used herein refers to a hydroxy group attached to the parent molecular moiety through an alkyl group. In some embodiments, a hydroxyalkyl is a Ci-C4hydroxy alkyl. Typical hydroxyalkyl groups include, but are not limited to, -CH2OH, -CH2CH2OH, -CH2CH2CH2OH, -CH2CH2CH2CH2OH, and the like.

[0348] The phrase “linear chain of atoms” refers to the longest straight chain of atoms independently selected from carbon, nitrogen, oxygen and sulfur.

[0349] The term “nitro,” as used herein refers to -NO2.

[0350] The term “oxo,” as used herein refers to =0.

[0351] The terms “sulfonate,” “sulfonic acid,” and “sulfonic,” as used herein refer the -SO3H group and its anion as the sulfonic acid is used in salt formation.

[0352] The term “sulfanyl,” as used herein refers to -S-.

[0353] The term “sulfinyl,” as used herein refers to -S(=O)-.

[0354] The term “sulfonyl,” as used herein refers to a -S(=O)2-, -S(=O)2R, or -S(=O)2R-group, with R as defined herein.

[0355] Any definition herein may be used in combination with any other definition to describe a composite structural group. By convention, the trailing element of any such definition is that which attaches to the parent moiety. For example, the composite group alkylamido would represent an alkyl group attached to the parent molecule through an amido group, and the term alkoxyalkyl would represent an alkoxy group attached to the parent molecule through an alkyl group.

[0356] When a group is defined to be “null,” what is meant is that said group is absent.

[0357] In some embodiments, the term “optionally substituted” or “substituted” means that the referenced group is optionally substituted with one or more additional group(s) individually and independently selected from 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 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, C3-C6cycloalkyl, Ci-C4fluoroalkyl, Ci-C4heteroalkyl, Ci-C4alkoxy, Ci-C4fluoroalkoxy, -SCi-C4alkyl, --93- SG Docket No.: 14989-700.600S(=0)Ci-C4alkyl, and -S(=O)2Ci-C4alkyl. In some embodiments, optional substituents are independently selected from halogen, -CN, -NH2, -OH, -NH(CH3), -N(CHs)2, -CH3, -CH2CH3, -CHF2, -CF3, -OCH3, -OCHF2, 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 atom (acyclic or cyclic) includes oxo (=0).

[0358] The term “bond” refers to a covalent linkage between two atoms, or two moieties when the atoms joined by the bond are considered to be part of larger substructure. A bond may be single, double, or triple unless otherwise specified. A dashed line between two atoms in a drawing of a molecule indicates that an additional bond may be present or absent at that position.

[0359] The term “amino acid” as used herein refers to both natural and non-natural amino acids. The term “non-natural amino acid” as used herein refers to an amino acid other than the 20 amino acids that occur naturally in protein. The term “amino acid analog” as used herein refers to a molecule which is structurally similar to an amino acid and which can be substituted for an amino acid in a peptide disclosed herein. Amino acid analogs include, without limitation, P-amino acids and amino acids where the amino or carboxy group is substituted by a similarly reactive group (e.g., substitution of the primary amine with a secondary or tertiary amine, or substitution of the carboxy group with an ester).

[0360] 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 non-natural amino acids.

[0361] As used herein, amino acid residue refers to an amino acid formed upon chemical digestion (hydrolysis) of a polypeptide at its peptide linkages. The amino acid residues described herein are, in certain embodiments, in the “L” isomeric form. Residues in the “D” isomeric form can be substituted for any “L” amino acid residue, as long as the desired functional property is retained by the polypeptide. In addition, the phrase “amino acid residue” is broadly defined to include the amino acids listed in Table A and modified and unusual amino acids, such as those listed in Table B and referred to in 37 C.F.R. §§1.821-1.822 and incorporated herein by reference.Table A: Amino Acids and Abbreviations-94- SG Docket No.: 14989-700.600Table B: Non-Natural Amino Acids or Amino Acid Analogs and Abbreviations-95- SG Docket No.: 14989-700.600"SG Docket No.: 14989-700.600SG Docket No.: 14989-700.600EXAMPLES

[0362] The following examples are provided for illustrative purposes only and not to limit the scope of the claims provided herein.Example 1: Synthesis of proximity-triggered conjugation molecule (Compound 14) Scheme 1

[0363] To a cooled (ice bath) solution of compound 1 (207 mg) and TEA (0.42 mL) in DCM (5 mL) was added compound 2 (243 mg) and the mixture was stirred at room temperature. After 30 min, the crude product was purified directly by RP-HPLC to give compound 3 (252 mg) as a white powder after lyophilization, which was treated with TFA / DCM (6 mL, 1 / 1, v / v) at room temperature for 1 h. The solvents were removed under reduced pressure and the residue was dissolved in DMF (3 mL). DIEA (0.3 mL) was added, followed by PFTU (0.4 g), and the mixture was stirred at room temperature for 20 min. The crude product was purified by RP-HPLC to give compound 5 (280 mg) as a white powder.SG Docket No.: 14989-700.600Scheme 2

[0364] To a stirred solution of compound 6 (123 mg, 1 mmol) and compound 7 (610 mg, 2 mmol) in DMF (6 mL) was added DIEA (0.7 mL), followed by PyAOP (1.1 g), and the reaction mixture was stirred at room temperature for 1 h. The crude product was purified by RP-HPLC to give compound 8 as an off-white solid after lyophilization (510 mg, TFA salt).

[0365] Compound 9 (172 mg) and compound 10 (216 mg) were dissolved in DCM (4 mL). DIEA (0.18 mL) was added and the mixture was stirred at room temperature for 48 h. The volatiles were removed under reduced pressure and the residue was purified by RP-HPLC to give compound 11 as a colorless syrup (0.32 g)

[0366] To a stirred solution of compound 8 (80 mg) and compound 11 (40 mg) in DMF (2 mL) was added DIEA (0.07 mL), followed by PyAOP (60 mg), and the reaction mixture was stirred at room temperature for 2 h. The crude product was purified by RP-HPLC to give compound 12 as a white solid after lyophilization (77 mg).

[0367] Compound 12 (53 mg) was treated with TFA / DCM (5 mL, 1 / 1, v / v) at room temperature for 2 h. The solvents were removed under reduced pressure and the residue was dissolved in MeOH (3 mL). MeONa (4 M in MeOH, 0.25 mL) was added. After stirring at room temperature for 20 min, HC1 (4N in dioxane, 0.18 mL) was added and the mixture was evaporated to dryness under reduced pressure. The residue was suspended in DMF (2 mL) and DIEA (0.06 mL) was added, followed by the addition of compound 5 (20 mg). The mixture was stirred at room temperature for 24 h and the crude was purified by RP-HPLC to-99- SG Docket No.: 14989-700.600give compound 14 as white powder after lyophilization (23 mg). MS: m / z 879.4 [M+H]+.Example 2: Linker Conjugation to HKHH (PRM1)Scheme 3.

[0368] To a 1.5 mL Eppendorf tube charged with NaHCCh (aq., 10 mM, 215 pL, pH 8.0) was added compound 14 (1 mM solution in ACN / water (6 / 4), 10 pL), followed by compound15 (1 mM solution in water, 5 pL) and NiCh (1 mM in water, 20 pL). The tube was sealed and agitated gently at room temperature for 16 h. The formation of compound 16 was monitored by LC / MS. MS: m / z 1954.1 [M+H]+.Example 3: Periodate Oxidation of Peptide-Linker Conjugate

[0369] To the above-described sample of unpurified compound 16 (20 pM) was addedNaICU (10 mM solution in water, 2 pL), and after 30 mins at room temperature, ISHzOMe(0.1 M solution in water, 5 pL) was added. After 2 h, the formation of compound 17 and compound 18 was confirmed by LC / MS, m / z 1454.9 [M+H]+; 555.3 [M+H]+.Example 4: Preparation of Peptide Dimer, PRM1 Linker Conjugation., and Reductive Cleavage to Identify Site of Conjugation-100- SG Docket No.: 14989-700.600Scheme 4

[0370] To a solution of compound 19 (10 mg) in ACN / water (6 / 4, v / v) was added 2,2’ -dithiodi pyridine (5 mg). After standing at room temperature for 10 min, the crude was-101- SG Docket No.: 14989-700.600purified by RP-HPLC to give compound 20 as a white solid (TFA salt, 9 mg).

[0371] Compound 20 (3 mg) was dissolved in water (0.5 mL) and to this solution compound 21 (5 mg) was added, followed by NaHCO3 (sat., aq., 2 pL). The mixture was stirred at room temperature for 1 h and the crude was purified by RP-HPLC to givecompound 22 as a white powder after lyophilization (4 mg, TFA salt). MS: m / z 985.9 [M+4H+] / 4Scheme 5-102- SG Docket No.: 14989-700.600

[0372] To a 1.5 mL Eppendorf tube charged with NaHCCh (aq., 10 mM, 210 pL) was added compound 14 (1 mM solution in ACN / water (6 / 4), 10 pL), followed by compound 22(1 mM solution in water, 10 pL) and NiCh (1 mM in water, 20 pL). The tube was sealed and agitated gently at room temperature for 16 h. The formation of compound 23 was monitored by LC / MS. MS: m / z 1205.6 [M+4H+] / 4.

[0373] To the abovementioned tube was added TCEP (10 mM solution in water, 3 pL).After standing at room temperature for 10 min, the formation of compound 19 and compound24 was confirmed by LC / MS, m / z 919.8 [M+2H+] / 2; 995.5 [M+3H+] / 3.Example 5: Linker Conjugation to PRM2Scheme 6

[0374] To a solution of compound 25 (10 mg) in ACN / water (6 / 4, v / v) was added 2,2’ -dithiodi pyridine (5 mg). After standing at room temperature for 10 min, the crude was-103- SG Docket No.: 14989-700.600purified by RP-HPLC to give compound 26 as a white solid (TFA salt, 8 mg). MS: m / z 1049.9 [M+2H+] / 2.

[0375] To a 1.5 mL Eppendorf tube charged with NaHCCh (aq., 10 mM, 235 pL) wasadded compound 14 (1 mM solution in ACN / water (6 / 4), 10 pL), followed by compound 26(1 mM solution in water, 5 pL). The tube was sealed and agitated gently at room temperature for 40 h. The formation of compound 27 was monitored by LC / MS. MS: m / z 993.4 [M+3H+] / 3.Example 6: Preparation of Peptide Dimer, PRM2 Linker Conjugation., and Reductive Cleavage to Identify Site of ConjugationScheme 7-104- SG Docket No.: 14989-700.600

[0376] Compound 26 (3 mg) was dissolved in water (0.5 mL) and to this solution compound 19 (3 mg) was added, followed by NaHCCh (sat., aq., 2 pL). The mixture was stirred at room temperature for 1 h and the crude was purified by RP-HPLC to give compound 28 as a white powder after lyophilization (4 mg, TFA salt). MS: m / z 1275.8 [M+3H+] / 3.-105- SG Docket No.: 14989-700.600Scheme 8-106- SG Docket No.: 14989-700.600

[0377] To a 1.5 mL Eppendorf tube charged with NaHCCh (aq., 10 mM, 235 pL) was added compound 14 (1 mM solution in ACN / water (6 / 4), 10 pL), followed by compound 28 (1 mM solution in water, 5 pL). The tube was sealed and agitated gently at room temperature for 40 h. The formation of compound 29 was monitored by LC / MS. MS: m / z 1568.9 [M+3H+] / 3.

[0378] To the same tube was added N-Ac-L-Cysteine (10 mM solution in water, 2 pL).After standing at room temperature for 20 min, TCEP (10 mM solution in water, 4 pL) was added, and the tube was agitated gently at room temperature for 10 min. The formation of compound 30 and compound 19 was monitored by LC / MS. MS: m / z 957.0 [M+3H+] / 3, 919.4[M+2H+] / 2.Example 7: Demonstration of Specific Conjugation to a peptide containing PRM2 in the presence of a second peptide containing PRM1-107- SG Docket No.: 14989-700.600

[0379] To a 1.5 mL Eppendorf tube charged with NaHCCh (aq., 10 mM, 230 pL) wasadded compound 14 (1 mM solution in ACN / water (6 / 4), 10 pL), followed by compound 31(1 mM solution in water, 5 pL) and compound 15 (1 mM solution in water, 5 pL). The tube was sealed and agitated gently at room temperature for 20 h. The formation of compound 32 was confirmed by LC / MS. MS: m / z 962.5 [M+2H+] / 2. No formation of compound 16 was detected by LC / MS.Example 8: Preparation of Fc-PRMl Polypeptide

[0380] Immunoglobulin Fc Fusions: mammalian expression vectors were constructed to express PRM1 linked to an Fc-fragment consisting of the CH2 and CH3 domains of theheavy chain and hinge regions of human IgG2. Each vector contained a strong constitutive promoter (CMV) and a signal peptide sequence for secretion of the fusion protein into the culture media. Vectors were designed to enable PRM1 to be fused to the C-terminus of the immunoglobulin protein and to incorporate a (Pro-Ala)io linker between PRM1 and IgG.

[0381] Fusion proteins were transiently expressed in discoCHO™ cells (ATUM) by transfecting plasmid DNA into the cells using Lipofectamine 2000 (Thermo Fisher).Transfected cells were grown in discoCHO™ expression medium in shaker flasks in a 37°C humidified CO2 incubator on an orbital shaker rotating at 125 rpm. Cultures were harvested 96 h post-transfection by centrifugation and the secreted fusion proteins were purified from the supernatants using protein A affinity chromatography.-108- SG Docket No.: 14989-700.600

[0382] Protein A agarose resin was mixed with culture supernatant and incubated at 25°C for several hours. The resin was then washed three times with PBS and bound Fc-PRMl polypeptide was eluted with 0.1 M glycine buffer (pH 2.8). Eluates were neutralized with IM Tris buffer and quantified by measuring absorbance at 280 nm using a NanoDrop® spectrophotometer. Protein concentrations were determined using calculated extinction coefficients derived from the primary sequence of the protein. Correct protein molecular weight was confirmed by mass spectrometry of the reduced and deglycosylated expression product. MS: m / z 27778.6.

[0383] The full amino acid sequence of recombinant Fc-PRMl polypeptide is shown below:

[0384] ERKSSVECPPCPAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPE VQFNWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNK GLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESN GQPENNYKTTPPMLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQAS LSLSPGAPAPAPAPAPAPAPAPAPAPAHKHH.

[0385] The molecular weight of the reduced and deglycosylated protein is MW = 27778.Example 9: Conjugation Protocol

[0386] A 100 mM solution of NiCh was made by dissolving 12.6 mg of the anhydrous salt in water with vortexing, and this was diluted 100-fold in water to make a 1 mM solution of NiCh. Separately, a 1 mM solution of compound 14 was made by dissolving 3.0 mg in 3.41 mL of 1 : 1 acetonitrile / water. 200 pL of the 1 mM 14 solution was mixed with 400 pL of the 1 mM NiCh solution, and then 270 mL of this mixture was mixed with 980 mL of a solution of Fc-PRMl polypeptide dissolved in water buffered with 12.75 mM NaHCCh, pH 8. The reaction mixture was allowed to incubate at ambient room temperature for 18 hours and was stopped by freezing.Example 10: Conjugation Analysis

[0387] The products from the reaction of 14 with Fc-PRMl polypeptide in Example 9 were characterized by LC / MS on a Waters Xevo G2 QTOF coupled with a Waters Acquity UPLC after reduction with TCEP and deglycosylation using N-peptidyl galactosidase (PNGase) (see FIG. 12).

[0388] The site of conjugation was determined by peptide mapping, which involved tryptic fragmentation of the deglycosylated / reduced sample of conjugated and unconjugated Fc-PRMl polypeptide, and comparison of the masses for each fragment of the conjugated and-109- SG Docket No.: 14989-700.600unconjugated Fc-PRMl polypeptide samples by analysis on a Thermo Orbitrap Eclipse Tribrid Mass Spectrometer coupled with a Waters Acquity UPLC. Peptide fragments exhibiting an increase of mass equal to the molecular weight of 14 over that predicted for the unconjugated versions were identified and the common lysine conjugation site identified. When multiple lysines were present in a group of overlapping peptide fragments, MS / MS and analysis software were used to predict the likely lysine that was conjugated.

[0389] Results from peptide mapping of the reaction mixture shown in FIG. 13. For peptide mapping, the proteins in the reaction mixture were deglycosylated and reduced, subjected to trypsin digestion and then analyzed by MS / MS. The instrument software identifies peptide fragments having the additional molecular weight of the conjugated linker. When multiple lysine residues are in a peptide fragment, the software identifies most likely sites of lysine conjugation. The square symbol (□) indicates the major site of conjugation. The circle symbol (o) indicates lysine residues showing no evidence of conjugation. The triangle symbol (A) indicates minor sites of conjugation. Observed proteolytic fragments are indicated by the rectangles under the Fc-PRMl polypeptide sequence. The LC retention time of the fragments is indicated by the value inside the rectangle.

[0390] Three lysine residues are identified as likely sites of linker conjugation.

[0391] The Fc-PRMl polypeptide sequence number for each lysine is listed along with the sum of the percentage abundance of all peptide fragments containing the indicated lysine and having the added linker molecular weight. The percentage abundance is relative to total MS / MS fragments. The percentage abundance numbers can only be used for qualitative comparison due to different MS response factors.Example 11: Linker conjugation to peptides containing different His / Lys sequences

[0392] Using a conjugation protocol as described in Example 2, compound 14 was conjugated to peptides containing His / Lys sequences shown in FIG. 15. Reaction products were analyzed by RP-HPLC (220 nm) and peak identify confirmed by LC / MS. As shown in FIG. 15, the sequence HKH gave incomplete conjugation, the sequence HKHH gave-110- SG Docket No.: 14989-700.600complete conjugation with no multi -conjugation products, and the sequences HHHK, HHKHH, HHHHK, and HHHHHHK gave varying amounts of multi-conjugation products.Example 12: Linker conjugation to peptides having different bis-Arg sequences

[0393] Using a conjugation protocol as described in Example 5, compound 14 was conjugated to the peptides containing bis-Arg sequences shown in FIG 16A. Conjugation products were analyzed by RP-HPLC (220 nm) and peak identify confirmed by LC / MS. Reactions were analyzed at t=0, 24 hr, 48 hr, and 72 hr. FIG. 16B shows representative RP-HPLC chromatograms of conjugations with no peptide, peptide 1, and peptide 10 at t=24 hr. As shown, the conjugation of peptide 1 (containing the PRM1 sequence HKHH) produced no conjugate with compound 14 due to the absence of metal ion, while the conjugation of peptide 10 (containing the PRM2 sequence RKR) did produce conjugate with compound 14. The ratio (relative peak area for conjugate peak) / (relative peak are for unreacted peptide peak) was used to quantitate the progress of conjugations of compound 14 to peptides 2-10. FIG. 16C shows the increase in the ratio (relative peak area for conjugate peak) / (relative peak are for unreacted peptide peak) over time for peptides 2-7. These peptides have progressively fewer residues between the two arginine residues of the PRM2. From this series, peptide 7 with sequence Ac-KAWRRGS-NH2 (two arginines immediately adjacent) proceeded most efficiently. FIG. 16D shows the increase in the ratio (relative peak area for conjugate peak) / (relative peak are for unreacted peptide peak) over time for peptides 7-10. These peptides all have no residues between the 2 arginine residues of PRM2 and have progressively fewer residues between the Lys residue of PRM2 and the proximal Arg. From this series, peptide 10 with sequence Ac-AWRKRGS-NH2 (Lys between the 2 arginines of PRM2) proceeded most efficiently.Example 13. Comparison of the conjugation efficiencies of proximity-triggered conjugation molecules with different spacer moieties (compounds 14. 15, and 16)

[0394] Using a conjugation protocol as described in Example 2, compounds 14, 15, and 16, shown in FIG. 17A, were conjugated to a peptide with the sequence Ac-WGSGSHKHH-OH. Reaction products were analyzed by RP-HPLC (220 nm) and peak identify confirmed by LC / MS. As shown in FIGS. 17B and 17C, conjugations with compounds 15 and 16 proceeded more slowly than the conjugation with compound 14. However, compounds 15 and 16 do show significant conjugation, and therefore may be used for selective conjugation. These results show how different spacers between the diol and the vinyl sulfonamide may affect the conjugation reaction.-111- SG Docket No.: 14989-700.600

[0395] When a feature or element is herein referred to as being “on” another feature or element, it can be directly on the other feature or element or intervening features and / or elements may also be present. In contrast, when a feature or element is referred to as being “directly on” another feature or element, there are no intervening features or elements present. It will also be understood that, when a feature or element is referred to as being “connected”, “attached” or “coupled” to another feature or element, it can be directly connected, attached or coupled to the other feature or element or intervening features or elements may be present. In contrast, when a feature or element is referred to as being “directly connected”, “directly attached” or “directly coupled” to another feature or element, there are no intervening features or elements present. Although described or shown with respect to one embodiment, the features and elements so described or shown can apply to other embodiments. It will also be appreciated by those of skill in the art that references to a structure or feature that is disposed “adjacent” another feature may have portions that overlap or underlie the adjacent feature.

[0396] Terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. For example, as used herein, the singular forms “a”, “an” and “the” are intended to include the plural forms as well, unless the context clearly indicates otherwise. It will be further understood that the terms “comprises” and / or “comprising,” when used in this specification, specify the presence of stated features, steps, operations, elements, and / or components, but do not preclude the presence or addition of one or more other features, steps, operations, elements, components, and / or groups thereof. As used herein, the term “and / or” includes any and all combinations of one or more of the associated listed items and may be abbreviated as “ / ”.

[0397] Spatially relative terms, such as “under”, “below”, “lower”, “over”, “upper” and the like, may be used herein for ease of description to describe one element or feature's relationship to another element(s) or feature(s) as illustrated in the figures. It will be understood that the spatially relative terms are intended to encompass different orientations of the device in use or operation in addition to the orientation depicted in the figures. For example, if a device in the figures is inverted, elements described as “under” or “beneath” other elements or features would then be oriented “over” the other elements or features. Thus, the exemplary term “under” can encompass both an orientation of over and under. The device may be otherwise oriented (rotated 90 degrees or at other orientations) and the spatially relative descriptors used herein interpreted accordingly. Similarly, the terms “upwardly”,-112- SG Docket No.: 14989-700.600“downwardly”, “vertical”, “horizontal” and the like are used herein for the purpose of explanation only unless specifically indicated otherwise.

[0398] Although the terms “first” and “second” may be used herein to describe various features / elements (including steps), these features / elements should not be limited by these terms, unless the context indicates otherwise. These terms may be used to distinguish one feature / element from another feature / element. Thus, a first feature / element discussed below could be termed a second feature / element, and similarly, a second feature / element discussed below could be termed a first feature / element without departing from the teachings of the present invention.

[0399] Throughout this specification and the claims which follow, unless the context requires otherwise, the word “comprise”, and variations such as “comprises” and “comprising” means various components can be co-jointly employed in the methods and articles (e.g., compositions and apparatuses including device and methods). For example, the term “comprising” will be understood to imply the inclusion of any stated elements or steps but not the exclusion of any other elements or steps.

[0400] In general, any of the apparatuses and methods described herein should be understood to be inclusive, but all or a sub-set of the components and / or steps may alternatively be exclusive, and may be expressed as “consisting of’ or alternatively “consisting essentially of’ the various components, steps, sub-components or sub-steps.

[0401] As used herein in the specification and claims, including as used in the examples and unless otherwise expressly specified, all numbers may be read as if prefaced by the word “about” or “approximately,” even if the term does not expressly appear. The phrase “about” or “approximately” may be used when describing magnitude and / or position to indicate that the value and / or position described is within a reasonable expected range of values and / or positions. For example, a numeric value may have a value that is + / - 0.1% of the stated value (or range of values), + / - 1% of the stated value (or range of values), + / - 2% of the stated value (or range of values), + / - 5% of the stated value (or range of values), + / - 10% of the stated value (or range of values), etc. Any numerical values given herein should also be understood to include about or approximately that value, unless the context indicates otherwise. For example, if the value “10” is disclosed, then “about 10” is also disclosed. Any numerical range recited herein is intended to include all sub-ranges subsumed therein. It is also understood that when a value is disclosed that “less than or equal to” the value, “greater than or equal to the value” and possible ranges between values are also disclosed, as appropriately-113- SG Docket No.: 14989-700.600understood by the skilled artisan. For example, if the value “X” is disclosed the “less than or equal to X” as well as “greater than or equal to X” (e.g., where X is a numerical value) is also disclosed. It is also understood that the throughout the application, data is provided in a number of different formats, and that this data, represents endpoints and starting points, and ranges for any combination of the data points. For example, if a particular data point “10” and a particular data point “15” are disclosed, it is understood that greater than, greater than or equal to, less than, less than or equal to, and equal to 10 and 15 are considered disclosed as well as between 10 and 15. It is also understood that each unit between two particular units are also disclosed. For example, if 10 and 15 are disclosed, then 11, 12, 13, and 14 are also disclosed.

[0402] Although various illustrative embodiments are described above, any of a number of changes may be made to various embodiments without departing from the scope of the invention as described by the claims. For example, the order in which various described method steps are performed may often be changed in alternative embodiments, and in other alternative embodiments one or more method steps may be skipped altogether. Optional features of various device and system embodiments may be included in some embodiments and not in others. Therefore, the foregoing description is provided primarily for exemplary purposes and should not be interpreted to limit the scope of the invention as it is set forth in the claims.

[0403] The examples and illustrations included herein show, by way of illustration and not of limitation, specific embodiments in which the subject matter may be practiced. As mentioned, other embodiments may be utilized and derived there from, such that structural and logical substitutions and changes may be made without departing from the scope of this disclosure. Such embodiments of the inventive subject matter may be referred to herein individually or collectively by the term “invention” merely for convenience and without intending to voluntarily limit the scope of this application to any single invention or inventive concept, if more than one is, in fact, disclosed. Thus, although specific embodiments have been illustrated and described herein, any arrangement calculated to achieve the same purpose may be substituted for the specific embodiments shown. This disclosure is intended to cover any and all adaptations or variations of various embodiments. Combinations of the above embodiments, and other embodiments not specifically described herein, will be apparent to those of skill in the art upon reviewing the above description.-114- SG Docket No.: 14989-700.600SEQUENCE LISTINGSEQ ID NO: 1IgGl / kappa Light Chain Constant Region KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 2IgGl / kappa Light Chain Constant Region - upstream fragment KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPRESEQ ID NO: 3IgGl / kappa Light Chain Constant Region - downstream fragment GNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 4IgGl / kappa Light Chain Constant Region - upstream fragment 2 KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQSEQ ID NO: 5IgGl / kappa Light Chain Constant Region - downstream fragment 2RGECSEQ ID NO: 6IgGl / kappa Light Chain Constant Region (swap for K in position 43) - upstream fragment KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWSEQ ID NO: 7IgGl / kappa Light Chain Constant Region (swap for K in position 43) - downstream fragment VDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC-148- SG Docket No.: 14989-700.600SEQ ID NO: 8IgGl / kappa Light Chain Constant Region (swap for D in position 45) - upstream fragment KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVSEQ ID NO: 9IgGl / kappa Light Chain Constant Region (swap for D in position 45) - downstream fragment NALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 10IgGl / kappa Light Chain Constant Region (swap for V in position 99) - upstream fragment KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKSEQ ID NO: 11IgGl / kappa Light Chain Constant Region (swap for V in position 99) - downstream fragment DNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 12IgGl / kappa Heavy Chain Constant Region QGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFGSVLQSSGL YSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCGSPELLGGPSVFLFPPKPKDTL MISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEY KCKVSNKALGSPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPEN NYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKSEQ ID NO: 13IgGl / kappa Heavy Chain Constant Region - upstream fragment 1QGTLSEQ ID NO: 14IgGl / kappa Heavy Chain Constant Region - downstream fragment 1-149- SG Docket No.: 14989-700.600SSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFGSVLQSSGLYSLSSVVTVPSSSLGTQTYICN VNHKPSNTKVDKRVEPKSCDKTHTCPPCGSPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPE VKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALGSPIEKTISKAKG QPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLT VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKSEQ ID NO: 15IgGl / kappa Heavy Chain Constant Region - upstream fragment 2 QGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFGSVLQSSGL YSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCGSPESEQ ID NO: 16IgGl / kappa Heavy Chain Constant Region - downstream fragment 2 PKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQD WLNGKEYKCKVSNKALGSPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWES NGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKSEQ ID NO: 17IgGl / kappa Heavy Chain Constant Region (swap for A in position 10) - upstream fragment QGTLVTVSSSEQ ID NO: 18IgGl / kappa Heavy Chain Constant Region (swap for A in position 10) - downstream fragment STKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFGSVLQSSGLYSLSSVVTVP SSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCGSPELLGGPSVFLFPPKPKDTLMISRTPEVTC VVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKAL GSPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLD SDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKSEQ ID NO: 19-150- SG Docket No.: 14989-700.600IgGl / kappa Heavy Chain Constant Region (swap for G in position 14) - upstream fragment QGTLVTVSSASTKSEQ ID NO: 20IgGl / kappa Heavy Chain Constant Region (swap for G in position 14) - downstream fragment PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFGSVLQSSGLYSLSSVVTVPSSSLG TQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCGSPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVD VSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALGSPIE KTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGS FFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKSEQ ID NO: 21IgGl / kappa Heavy Chain Constant Region (swap for P in position 15) - upstream fragment QGTLVTVSSASTKGSEQ ID NO: 22IgGl / kappa Heavy Chain Constant Region (swap for P in position 15) - downstream fragment SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFGSVLQSSGLYSLSSVVTVPSSSLGT QTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCGSPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDV SHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALGSPIEKT ISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFF LYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKSEQ ID NO: 23IgGl / kappa Heavy Chain Constant Region (swap for V in position 17) - upstream fragment QGTLVTVSSASTKG PSSEQ ID NO: 24-151- SG Docket No.: 14989-700.600IgGl / kappa Heavy Chain Constant Region (swap for V in position 17) - downstream fragment FPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFGSVLQSSGLYSLSSVVTVPSSSLGTQ TYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCGSPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVS HEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALGSPIEKTI SKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKSEQ ID NO: 25IgGl / kappa Heavy Chain Constant Region (swap for S in position 131) - upstream fragment QGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFGSVLQSSGL YSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCGSPELLGGPSSEQ ID NO: 26IgGl / kappa Heavy Chain Constant Region (swap for S in position 131) - downstream fragment VFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVL HQDWLNGKEYKCKVSNKALGSPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAV EWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKSEQ ID NO: 27PRM1 sequenceKHHHHSEQ ID NO: 28PRM1 sequenceHKHHHSEQ ID NO: 29PRM1 sequenceHKHHH-152- SG Docket No.: 14989-700.600SEQ ID NO: 30PRM1 sequenceHHKHHSEQ ID NO: 31PRM1 sequenceHHHHKSEQ ID NO: 32PRM2 sequence - K-[X]n-R-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)KXRXRSEQ ID NO: 33PRM2 sequence - K-[X]n-R-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)KRXRSEQ ID NO: 34PRM2 sequence - K-[X]n-R-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)KXRRSEQ ID NO: 35PRM2 sequence [X]n-R-K-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)XRKXRSEQ ID NO: 36-153- SG Docket No.: 14989-700.600PRM2 sequence [X]n-R-K-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)XRKRSEQ ID NO: 37PRM2 sequence [X]n-R-K-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)RKXRSEQ ID NO: 38PRM2 sequence - K-[X]n-R-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)KXXRXRSEQ ID NO: 39PRM2 sequence - K-[X]n-R-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)KXRXXRSEQ ID NO: 40PRM2 sequence - K-[X]n-R-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)KXXRXXRSEQ ID NO: 41PRM2 sequence - K-[X]n-R-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)KXXRRSEQ ID NO: 42-154- SG Docket No.: 14989-700.600PRM2 sequence - K-[X]n-R-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)KXXXRRSEQ ID NO: 43PRM2 sequence - K-[X]n-R-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)KRXXRSEQ ID NO: 44PRM2 sequence - K-[X]n-R-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)KRXXXRSEQ ID NO: 45PRM2 sequence [X]n-R-K-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)XXRKXRSEQ ID NO: 46PRM2 sequence [X]n-R-K-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)XRKXXRSEQ ID NO: 47PRM2 sequence [X]n-R-K-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)XXRKXXRSEQ ID NO: 48-155- SG Docket No.: 14989-700.600PRM2 sequence [X]n-R-K-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)XXRKRSEQ ID NO: 49PRM2 sequence [X]n-R-K-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)XXXRKRSEQ ID NO: 50PRM2 sequence [X]n-R-K-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)RKXXRSEQ ID NO: 51PRM2 sequence [X]n-R-K-[X]n-R, where X is any amino acid residue except R or K, and where n is between 0 and 10 (and each X can be independently selected)RKXXXRSEQ ID NO: 52IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREHHKHKVQWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 53IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAHHKHKVQWKVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 54IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH-156- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAHHKHVQWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 55IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKHHKHVQWKVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 56IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKHHKHQWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 57IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVHHKHQWKVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC SEQ ID NO: 58IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVHHKHWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 59IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQHHKHWKVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 60IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQHHKHKVDNALQSGNSQESVTEQDSKDSTY SLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC-157- SG Docket No.: 14989-700.600SEQ ID NO: 61IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWHHKHKVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 62IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWHHKHVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 63IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKHHKHVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 64IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKHHKHDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 65IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVHHKHDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 66IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVHHKHNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC-158- SG Docket No.: 14989-700.600SEQ ID NO: 67IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDHHKHNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 68IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDHHKHALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 69IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNHHKHALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 70IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNHHKHLQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 71IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNAHHKHLQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 72IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNAHHKHQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 73-159- SG Docket No.: 14989-700.600IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALHHKHQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 74IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALHHKHSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 75IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQHHKHSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 76IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQHHKHGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 77IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSHHKHGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 78IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQHHKHLSSPVTKSFNRGECSEQ ID NO: 79IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH-160- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGHHKHLSSPVTKSFNRGECSEQ ID NO: 80IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGHHKHSSPVTKSFNRGECSEQ ID NO: 81IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLHHKHSSPVTKSFNRGECSEQ ID NO: 82IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLHHKHSPVTKSFNRGECSEQ ID NO: 83IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSHHKHSPVTKSFNRGECSEQ ID NO: 84IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSHHKHPVTKSFNRGECSEQ ID NO: 85IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH-161- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSHHKHPVTKSFNRGECSEQ ID NO: 86IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSHHKHVTKSFNRGECSEQ ID NO: 87IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPHHKHVTKSFNRGECSEQ ID NO: 88IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPHHKHTKSFNRGECSEQ ID NO: 89IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVHHKHTKSFNRGECSEQ ID NO: 90IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVHHKHKSFNRGECSEQ ID NO: 91IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH-162- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTHHKHKSFNRGECSEQ ID NO: 92IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTHHKHSFNRGECSEQ ID NO: 93IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKHHKHSFNRGECSEQ ID NO: 94IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKHHKHFNRGECSEQ ID NO: 95IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSHHKHFNRGECSEQ ID NO: 96IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSHHKHNRGECSEQ ID NO: 97IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH-163- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFHHKHNRGECSEQ ID NO: 98IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFHHKHRGECSEQ ID NO: 99IgGl / kappa Light Chain Constant Region with PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNHHKHRGECSEQ ID NO: 100IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEHHKHSEQ ID NO: 101IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECHHKHSEQ ID NO: 102IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAHHKHSEQ ID NO: 103-164- SG Docket No.: 14989-700.600IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAHHKHSEQ ID NO: 104IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAHHKHSEQ ID NO: 105IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAHHKHSEQ ID NO: 106IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAHHKHSEQ ID NO: 107IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAHHKHSEQ ID NO: 108-165- SG Docket No.: 14989-700.600IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAHHKHSEQ ID NO: 109IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAHHKHSEQ ID NO: 110IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAHHKHSEQ ID NO: 111IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAHHKHSEQ ID NO: 112IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAHHKHSEQ ID NO: 113-166- SG Docket No.: 14989-700.600IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAHHKHSEQ ID NO: 114IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPAHHKHSEQ ID NO: 115IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPAHHKHSEQ ID NO: 116IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPAPAHHKHSEQ ID NO: 117IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPAPAHHKHSEQ ID NO: 118-167- SG Docket No.: 14989-700.600IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPAPAPAHHKHSEQ ID NO: 119IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPAPAPAHHKHSEQ ID NO: 120IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPAPAPAPAHHKHSEQ ID NO: 121I IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPAPAPAPAHHKHSEQ ID NO: 122IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSHHKHSEQ ID NO: 123-168- SG Docket No.: 14989-700.600IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSHHKHSEQ ID NO: 124IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSHHKHSEQ ID NO: 125IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSHHKHSEQ ID NO: 126IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSHHKHSEQ ID NO: 127IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSHHKHSEQ ID NO: 128-169- SG Docket No.: 14989-700.600IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSHHKHSEQ ID NO: 129IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSHHKHSEQ ID NO: 130IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSHHKHSEQ ID NO: 131IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSHHKHSEQ ID NO: 132IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSGSHHKHSEQ ID NO: 133-170- SG Docket No.: 14989-700.600IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSGSHHKHSEQ ID NO: 134IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSGSGSHHKHSEQ ID NO: 135IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSGSGSHHKHSEQ ID NO: 136IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSGSGSGSHHKHSEQ ID NO: 137IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSGSGSGSHHKHSEQ ID NO: 138-171- SG Docket No.: 14989-700.600IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSGSGSGSGSHHKHSEQ ID NO: 139IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSGSGSGSGSHHKHSEQ ID NO: 140IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSGSGSGSGSGSHHKHSEQ ID NO: 141IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HHKH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSGSGSGSGSGSHHKHSEQ ID NO: 142IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREHKHHKVQWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 143IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH-172- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAHKHHKVQWKVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 144IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAHKHHVQWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 145IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKHKHHVQWKVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 146IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKHKHHQWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 147IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVHKHHQWKVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC SEQ ID NO: 148IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVHKHHWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 149IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQHKHHWKVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC-173- SG Docket No.: 14989-700.600SEQ ID NO: 150IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQHKHHKVDNALQSGNSQESVTEQDSKDSTY SLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 151IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWHKHHKVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 152IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWHKHHVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 153IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKHKHHVDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 154IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKHKHHDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 155IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVHKHHDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC-174- SG Docket No.: 14989-700.600SEQ ID NO: 156IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVHKHHNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 157IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDHKHHNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 158IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDHKHHALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 159IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNHKHHALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 160IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNHKHHLQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 161IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNAHKHHLQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 162-175- SG Docket No.: 14989-700.600IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNAHKHHQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 163IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALHKHHQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 164IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALHKHHSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 165IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQHKHHSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 166IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQHKHHGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 167IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSHKHHGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 168IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH-176- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQHKHHLSSPVTKSFNRGECSEQ ID NO: 169IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGHKHHLSSPVTKSFNRGECSEQ ID NO: 170IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGHKHHSSPVTKSFNRGECSEQ ID NO: 171IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLHKHHSSPVTKSFNRGECSEQ ID NO: 172IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLHKHHSPVTKSFNRGECSEQ ID NO: 173IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSHKHHSPVTKSFNRGECSEQ ID NO: 174IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH-177- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSHKHHPVTKSFNRGECSEQ ID NO: 175IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSHKHHPVTKSFNRGECSEQ ID NO: 176IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSHKHHVTKSFNRGECSEQ ID NO: 177IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPHKHHVTKSFNRGECSEQ ID NO: 178IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPHKHHTKSFNRGECSEQ ID NO: 179IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVHKHHTKSFNRGECSEQ ID NO: 180IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH-178- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVHKHHKSFNRGECSEQ ID NO: 181IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTHKHHKSFNRGECSEQ ID NO: 182IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTHKHHSFNRGECSEQ ID NO: 183IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKHKHHSFNRGECSEQ ID NO: 184IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKHKHHFNRGECSEQ ID NO: 185IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSHKHHFNRGECSEQ ID NO: 186IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH-179- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSHKHHNRGECSEQ ID NO: 187IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFHKHHNRGECSEQ ID NO: 188IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFHKHHRGECSEQ ID NO: 189IgGl / kappa Light Chain Constant Region with PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNHKHHRGECSEQ ID NO: 190IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEHKHHSEQ ID NO: 191IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECHKHHSEQ ID NO: 192IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH-180- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAHKHHSEQ ID NO: 193IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAHKHHSEQ ID NO: 194IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAHKHHSEQ ID NO: 195IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAHKHHSEQ ID NO: 196IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAHKHHSEQ ID NO: 197IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH-181- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAHKHHSEQ ID NO: 198IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAHKHHSEQ ID NO: 199IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAHKHHSEQ ID NO: 200IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAHKHHSEQ ID NO: 201IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAHKHHSEQ ID NO: 202IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH-182- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAHKHHSEQ ID NO: 203IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAHKHHSEQ ID NO: 204IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPAHKHHSEQ ID NO: 205IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPAHKHHSEQ ID NO: 206IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPAPAHKHHSEQ ID NO: 207IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH-183- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPAPAHKHHSEQ ID NO: 208IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPAPAPAHKHHSEQ ID NO: 209IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPAPAPAHKHHSEQ ID NO: 210IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPAPAPAPAHKHHSEQ ID NO: 211I IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker (PA)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPAPAPAPAHKHHSEQ ID NO: 212IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH-184- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSHKHHSEQ ID NO: 213IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSHKHHSEQ ID NO: 214IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSHKHHSEQ ID NO: 215IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSHKHHSEQ ID NO: 216IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSHKHHSEQ ID NO: 217IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH-185- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSHKHHSEQ ID NO: 218IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSHKHHSEQ ID NO: 219IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSHKHHSEQ ID NO: 220IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSHKHHSEQ ID NO: 221IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSHKHHSEQ ID NO: 222IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH-186- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSGSHKHHSEQ ID NO: 223IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSGSHKHHSEQ ID NO: 224IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSGSGSHKHHSEQ ID NO: 225IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSGSGSHKHHSEQ ID NO: 226IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSGSGSGSHKHHSEQ ID NO: 227IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH-187- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSGSGSGSHKHHSEQ ID NO: 228IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSGSGSGSGSHKHHSEQ ID NO: 229IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSGSGSGSGSHKHHSEQ ID NO: 230IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSGSGSGSGSGSGSGSGSGSHKHHSEQ ID NO: 231IgGl / kappa Light Chain Constant Region with C-term PRM1 sequence - with linker Gly-Ser (GS)n (n=0-10) HKHH KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSGSGSGSGSGSGSGSGSGSHKHHSEQ ID NO: 232IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPRERKRKVQWKVDNALQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC-188- SG Docket No.: 14989-700.600SEQ ID NO: 233IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREARKRKVQWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 234IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREARKRVQWKVDNALQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 235IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKRKRVQWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 236IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKRKRQWKVDNALQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 237IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVRKRQWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC SEQ ID NO: 238IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVRKRWKVDNALQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 239-189- SG Docket No.: 14989-700.600IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQRKRWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 240IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQRKRKVDNALQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 241IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWRKRKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 242IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWRKRVDNALQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 243IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKRKRVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 244IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKRKRDNALQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 245IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR-190- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVRKRDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 246IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVRKRNALQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 247IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDRKRNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 248IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDRKRALQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 249IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNRKRALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 250IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNRKRLQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 251IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR-191- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNARKRLQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 252IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNARKRQSGNSQESVTEQDSKDSTY SLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 253IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALRKRQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 254IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALRKRSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 255IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQRKRSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 256IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQRKRGNSQESVTEQDSKDSTY SLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 257IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR-192- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSRKRGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECSEQ ID NO: 258IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQRKRLSSPVTKSFNRGECSEQ ID NO: 259IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGRKRLSSPVTKSFNRGECSEQ ID NO: 260IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGRKRSSPVTKSFNRGECSEQ ID NO: 261IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLRKRSSPVTKSFNRGECSEQ ID NO: 262IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLRKRSPVTKSFNRGECSEQ ID NO: 263IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR-193- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSRKRSPVTKSFNRGECSEQ ID NO: 264IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSRKRPVTKSFNRGECSEQ ID NO: 265IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSRKRPVTKSFNRGECSEQ ID NO: 266IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSRKRVTKSFNRGECSEQ ID NO: 267IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPRKRVTKSFNRGECSEQ ID NO: 268IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPRKRTKSFNRGECSEQ ID NO: 269IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR-194- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVRKRTKSFNRGECSEQ ID NO: 270IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVRKRKSFNRGECSEQ ID NO: 271IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTRKRKSFNRGECSEQ ID NO: 272IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTRKRSFNRGECSEQ ID NO: 273IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKRKRSFNRGECSEQ ID NO: 274IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKRKRFNRGECSEQ ID NO: 275IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR-195- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSRKRFNRGECSEQ ID NO: 276IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSRKRNRGECSEQ ID NO: T!IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFRKRNRGECSEQ ID NO: 278IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFRKRRGECSEQ ID NO: 279IgGl / kappa Light Chain Constant Region with PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRKRRGECSEQ ID NO: 280IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGERKRSEQ ID NO: 281IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - RKR-196- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECRKRSEQ ID NO: 282IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPARKRSEQ ID NO: 283IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPARKRSEQ ID NO: 284IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPARKRSEQ ID NO: 285IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPARKRSEQ ID NO: 286IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR-197- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPARKRSEQ ID NO: 287IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPARKRSEQ ID NO: 288IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPARKRSEQ ID NO: 289IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPARKRSEQ ID NO: 290IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPARKRSEQ ID NO: 291IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR-198- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPARKRSEQ ID NO: 292IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPARKRSEQ ID NO: 293IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPARKRSEQ ID NO: 294IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPARKRSEQ ID NO: 295IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPARKRSEQ ID NO: 296IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR-199- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPAPARKRSEQ ID NO: 297IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPAPARKRSEQ ID NO: 298IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPAPAPARKRSEQ ID NO: 299IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPAPAPARKRSEQ ID NO: 300IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEPAPAPAPAPAPAPAPAPAPARKRSEQ ID NO: 301I IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker (PA)n (n=0-10) RKR-200- SG Docket No.: 14989-700.600KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECPAPAPAPAPAPAPAPAPAPARKRSEQ ID NO: 302IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker Gly-Ser (GS)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEGSRKRSEQ ID NO: 303IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker Gly-Ser (GS)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECGSRKRSEQ ID NO: 304IgGl / kappa Light Chain Constant Region with C-term PRM2 sequence - with linker Gly-Ser (GS)n (n=0-10) RKR KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQW...

Claims

CLAIMSWHAT IS CLAIMED IS:

1. A compound of F ormula (I) :PBU-L1-L2-PCU (Formula (I))wherein:PBU is a moiety capable of non-covalently binding to a peptidic recognition motif (PRM) in a polypeptide, wherein the PRM comprises a target amino acid comprising lysine (Lys), ornithine (Orn), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys);PCU is a moiety capable of covalently reacting with the target amino acid of the PRM when held at a proximity-induced binding distance to the target amino acid of the PRM;L1is a cleavable linker; andL2is a spacer or is absent;wherein upon non-covalent bonding of the PBU to the PRM in the polypeptide, the PCU is held at the proximity-induced binding distance to the target amino acid of the PRM to cause covalent bonding between the PCU and the target amino acid of the PRM.

2. The compound of claim 1, wherein the PRM is PRM1, wherein the PRM1 further comprises 2 to 4 histidine (His) residues.

3. The compound of any one of claims 1 and 2, wherein the PRM is PRM2, wherein the PRM2 further comprises at least 2 amino acids independently selected from arginine (Arg), homoarginine (HArg), citrulline (Cit), norarginine (AGBA), p2-arginine (P2-Arg), and p3-arginine (P3-Arg).

4. The compound of any one of claims 1-3, wherein the PCU comprises an electrophilic warhead moiety that is capable of covalently reacting with the target amino acid only when held within the proximity-induced binding distance to the target amino acid.

5. The compound of claim 4, wherein the proximity -induced binding distance is within 50 angstroms.

6. The compound of any one of claims 1-5, wherein the PCU does not substantially-115- SG Docket No.: 14989-700.600covalently react with the target amino acid when the compound is not non-covalently bound to the PRM.

7. The compound of any one of claims 1-6, wherein the PCU comprises an electrophilic warhead moiety that is capable of covalently reacting with the target amino acid when held within the proximity-induced binding distance to the target amino acid, wherein the electrophilic warhead moiety comprises a vinyl sulfone group, a vinyl sulfonamide group, a pyrimidine sulfoxide group, a pyrimidine sulfone group, a vinyl pyridine group, or a vinyl pyrimidine group.

8. The compound of any one of claims 1-7, wherein the PBU comprises at least one nitrilotriacetic acid, at least one iminodiacetic acid, or at least one pyridin-2- ylmethanamine, or combinations thereof.

9. The compound of any one of claims 1-8, wherein the PBU comprises at least one nitrilotriacetic acid, or at least one iminodiacetic acid, or combinations thereof.

10. The compound of any one of claims 1-8, wherein the PBU comprises at least one moiety that is:

11. The compound of any one of claims 1-8, wherein the PBU comprises at least one moiety that is12. The compound of any one of claims 1-8, wherein the PBU comprises:-116- SG Docket No.: 14989-700.600U1is CH, C-OH, orN13. The compound of any one of claims 1-8, wherein the PBU isU1is CH, C-OH, orN.

14. The compound of any one of claims 1-8, wherein the PBU comprisesU1is CH, C-OH, orN15. The compound of any one of claims 1-8, wherein the PBU is-117- SG Docket No.: 14989-700.600U1is CH, C-OH, orN.

16. The compound of any one of claims 1-15, wherein:L1comprises a nucleophilic cleavable linker, electrophile cleavable linkers, acid cleavable linker, base cleavable linker, linker cleavable with reducing reagents, linker cleavable with oxidizing reagents, or photocleavable linker.

17. The compound of any one of claims 1-16, wherein L2is not cleavable.

18. The compound claim 1, wherein the compound has the following structure:wherein:R1is H, halogen, substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, pyrimidine, or -C(=O)OR3; andR2is H, halogen, substituted or unsubstituted alkyl, or -CN; andR3is independently H or substituted or unsubstituted alkyl.

19. The compound of claim 1, wherein the compound has the following structure:-118- SG Docket No.: 14989-700.60020. The compound of claim 1, wherein the compound has the following structure:

21. The compound of claim 1, wherein the compound has the following structure:

22. The compound of claim 1, wherein the compound has the following structure:-119- SG Docket No.: 14989-700.60023. A compound having the structure of:

24. A compound having the structure of:

25. A compound having the structure of:-HO- SG Docket No.: 14989-700.60026. The compound of claim 4, wherein the proximity -induced binding distance is within 10 angstroms.

27. A compound of Formula (I):PBU-L'-L2-PCU (Formula (I))wherein:PBU is a moiety comprising at least one nitrilotriacetic acid, or at least one iminodiacetic acid, or combinations thereof, wherein the PBU is capable of non- covalently binding to a peptidic recognition motif (PRM) in a polypeptide, wherein the PRM comprises a target amino acid comprising lysine (Lys), ornithine (Orn), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys);PCU is an electrophilic moiety comprising one or more of: vinyl sulfone group, a vinyl sulfonamide group, a pyrimidine sulfoxide group, a pyrimidine sulfone group, a vinyl pyridine group, or a vinyl pyrimidine group, such that the PCU covalently reacts with the target amino acid of the PRM only when held at a proximity-induced binding distance from the target amino acid of the PRM;L1is a cleavable linker; andL2is a spacer or is absent;wherein upon non-covalent bonding of the PBU to the PRM in the polypeptide, the PCU is held at the proximity-induced binding distance to the target amino acid of the PRM to cause covalent bonding between the PCU and the target amino acid of the PRM.

28. A method of selectively modifying a target amino acid of in a polypeptide, the method comprising:contacting a compound with the polypeptide under conjugation condition, wherein the compound has a formula of:-121- SG Docket No.: 14989-700.600PBU-L'-L2-PCU (Formula (I))wherein:PBU is a moiety comprising at least one nitrilotriacetic acid, or at least one iminodiacetic acid, or combinations thereof;PCU is an electrophilic moiety comprising on or more of: vinyl sulfone group, a vinyl sulfonamide group, a pyrimidine sulfoxide group, a pyrimidine sulfone group, a vinyl pyridine group, or a vinyl pyrimidine group;L1is a cleavable linker; andL2is a spacer or is absent;non-covalently binding the PBU portion of the compound to a peptidic recognition motif (PRM) of the polypeptide, wherein the PRM comprises a target amino acid comprising lysine (Lys), ornithine (Orn), homolysine (HLys), p2-lysine (P2- Lys), or p3-lysine (P3-Lys);covalently bonding the PCU to the target amino acid while the PBU portion of the compound is noncovalently bound to the PRM.

29. The method of claim 28, wherein the polypeptide is a biologically active polypeptide (B).

30. The method of claim 28, wherein the polypeptide is an antibody or antibody fragment, further wherein the PRM is within a constant region of the antibody or antibody fragment.

31. The method of any one of claims 28-30, wherein the PRM is1) on the N-terminus of the polypeptide,2) on a C-terminus of the polypeptide, or3) within an internal position of the polypeptide;wherein the PRM is independently selected from PRM1, PRM2, or combinations thereof:wherein PRM1 comprises:2 to 4 histidine (His) residues; anda lysine (Lys) residue, ornithine (Orn) residue, homolysine (HLys), a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue;wherein PRM2 comprises amino acid residues comprising:-122- SG Docket No.: 14989-700.600at least 2 amino acids selected from an arginine (Arg) residue, a homoarginine (HArg) residue, a citrulline (Cit) residue, norarginine (AGBA) residue, a P2-arginine (P2-Arg), and a p3-arginine (P3-Arg); and a lysine (Lys) residue, an ornithine (Orn) residue, a homolysine (HLys) residue, a p2-lysine (P2-Lys) residue, or p3-lysine (P3-Lys) residue.

32. The method of claim 31, wherein PRM1 comprises at least one of:X-His-His, His-X-His, His-His-X,X-His-His-His, His-X-His-His, His-His-X-His, His-His-His-X,X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, or His-His-His-His-X,wherein X is Lys, Om, HLys, p2-Lys, or p3-Lys.

33. The method of claim 31, wherein PRM1 comprises at least one of: X-His-His, His-X- His, or His-His-X.

34. The method of claim 31, wherein PRM1 comprises at least one of: X-His-His-His, His- X-His-His, His-His-X-His, or His-His-His-X.

35. The method of claim 31, wherein PRM1 comprises at least one of: X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, or His-His-His-His-X.

36. The method of any one of claims 32-35, wherein X is Lys.

37. The method of any one of claims 32-35, wherein X is Orn.

38. The method of any one of claims 32-35, wherein X is HLys.

39. The method of any one of claims 28-38, wherein at least one PRM1 is covalently attached at the N-terminus or C-terminus of the biologically active polypeptide.

40. The method of any one of claims 28-39, wherein the polypeptide comprises the amino acid sequence:B-(His)m-X-(His)nor (His)m-X-(His)n-B,wherein,B is the biologically active polypeptide;X is Lys, Om, HLys, p2-Lys, or p3-Lys;-123- SG Docket No.: 14989-700.600m is 0, 1, 2, 3, or 4; andn is 0, 1, 2, 3, or 4, provided that m + n is at least 2.

41. The method of any one of claims 21-40, wherein at least one PRM1 is incorporated in an internal position of the polypeptide.

42. The method of any one of claims 28-41, wherein the polypeptide comprises the amino acid sequence:wherein,B1and B2are fragments of a biologically active polypeptide B, such that BXB2is the biologically active polypeptide B, and the -(His)m-X-(His)n- sequence is added at an internal position of the biologically active polypeptide B;X is Lys, Om, HLys, p2-Lys, or p3-Lys;m is 0, 1, 2, 3, or 4; andn is 0, 1, 2, 3, or 4, provided that m + n is at least 2.

43. The method of claim 40 or 42, wherein:m + n is at least 4;m + n is at least 5;m + n is at least 6;m is 0 and n is 2;m is 2 and n is 0;m is 0 and n is 4;m is 1 and n is 3;m is 2 and n is 2;m is 3 and n is 1; orm is 4 and n is 0.

44. The method of any one of claims 28-43, wherein PRM2 comprises:X3-X1-Y-X4-X2-Yfurther wherein:X3or X4is Lys, Om, HLys, P2-Lys, or p3-Lys;if X3is present, then X4is not present;if X4is present, then X3is not present;-124- SG Docket No.: 14989-700.600each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg; X1comprises 0 to 10 amino acid residues other than X and Y;X2comprises 0 to 10 amino acids other than X3, X4, and Y.

45. The method of claim 44, wherein X3or X4is Lys.

46. The method of claim 44, wherein X3or X4is Om.

47. The method of claim 44, wherein X3or X4is HLys.

48. The method of any one of claims 44-47, wherein each Y is Arg.

49. The method of any one of claims 44-47, wherein each Y is HArg.

50. The method of any one of claims 44-47, wherein each Y is Cit.

51. The method of any one of claims 44-47, wherein each Y is AGBA.

52. The method of any one of claims 44-47, wherein each amino acid residue of X1is independently selected from alanine (Ala), asparagine (Asn), glutamine (Gin), glycine (Gly), isoleucine(Ile), leucine (Leu), methionine (Met), phenylalanine (Phe), proline (Pro), serine (Ser), threonine (Thr), tryptophan (Trp), tyrosine (Tyr), valine (Vai), glutamic acid (Glu), aspartic acid (Asp), and non-natural amino acids without positively charged or thiol-containing side chains.

53. The method of any one of claims 44-52, wherein X1comprises 0 amino acids.

54. The method of any one of claims 44-52, wherein X1comprises 1 amino acid.

55. The method of any one of claims 44-52, wherein X1comprises 2 amino acids.

56. The method of any one of claims 44-52, wherein X1comprises 3 amino acids.

57. The method of any one of claims 44-52, wherein X1comprises 4 amino acids.

58. The method of any one of claims 44-52, wherein X1comprises 5 amino acids.

59. The method of any one of claims 44-52, wherein X1comprises 6 amino acids.

60. The method of any one of claims 44-52, wherein X1comprises 7 amino acids.-125- SG Docket No.: 14989-700.60061. The method of any one of claims 44-52, wherein X1comprises 8 amino acids.

62. The method of any one of claims 44-52, wherein X1comprises 9 amino acids.

63. The method of any one of claims 44-52, wherein X1comprises 10 amino acids.

64. The method of any one of claims 44-60, wherein each amino acid residue of X2is independently selected from alanine (Ala), asparagine (Asn), glutamine (Gin), glycine (Gly), isoleucine(Ile), leucine (Leu), methionine (Met), phenylalanine (Phe), proline (Pro), serine (Ser), threonine (Thr), tryptophan (Trp), tyrosine (Tyr), valine (Vai), glutamic acid (Glu), aspartic acid (Asp), and non-natural amino acids without positively charged or thiol-containing side chains.

65. The method of any one of claims 44-63, wherein X2comprises 0 amino acids.

66. The method of any one of claims 44-63, wherein X2comprises 1 amino acid.

67. The method of any one of claims 44-63, wherein X2comprises 2 amino acids.

68. The method of any one of claims 44-63, wherein X2comprises 3 amino acids.

69. The method of any one of claims 44-63, wherein X2comprises 4 amino acids.

70. The method of any one of claims 44-63, wherein X2comprises 5 amino acids.

71. The method of any one of claims 44-63, wherein X2comprises 6 amino acids.

72. The method of any one of claims 44-63, wherein X2comprises 7 amino acids.

73. The method of any one of claims 44-63, wherein X2comprises 8 amino acids.

74. The method of any one of claims 44-63, wherein X2comprises 9 amino acids.

75. The method of any one of claims 44-63, wherein X2comprises 10 amino acids.

76. The method of any one of claims 44-75, wherein at least one PRM2 is covalently attached at the N-terminus or C-terminus of the biologically active polypeptide.

77. The method of any one of claims 44-76, wherein the polypeptide comprises the amino acid sequence:-126- SG Docket No.: 14989-700.600B-X3-X1-Y-X4-X2-Y or X2-Y-X1-X-Bwherein:B is a biologically active polypeptide (B);X3or X4is Lys, Om, HLys, p2-Lys, or p3-Lys;if X3is present, then X4is not present;if X4is present, then X3is not present;each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg; X1comprises 0 to 10 amino acid residues other than X3, X4, and Y;X2comprises 0 to 10 amino acids other than X3, X4, and Y.

78. The method of any one of claims 28-77, wherein at least one PRM2 is inserted at an internal position of the polypeptide.

79. The method of any one of claims 28-78, wherein the polypeptide comprises the amino acid sequence:B^-X^Y-X^-B2wherein,B1and B2are fragments of a biologically active polypeptide B, such that BXB2is the biologically active polypeptide (B), and the -X3-X1-Y-X4-X2-Y- sequence is added at an internal position of B;B1and B2are fragments of a biologically active polypeptide B, such that BXB2is the biologically active polypeptide B and the -X^X^Y-X^X^Y- sequence is added at an internal position of polypeptide B;X3or X4is Lys, Om, HLys, p2-Lys, or p3-Lys;if X3is present, then X4is not present;if X4is present, then X3is not present;each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and p3-Arg; X1comprises 0 to 10 amino acid residues other than X3, X4, and Y; andX2comprises 0 to 10 amino acids other than X3, X4, and Y.

80. The method of any one of claims 28-79, wherein the polypeptide comprises at least one PRM1 and at least one PRM2.

81. The method of any one of claims 28-80, wherein the polypeptide is a therapeutic polypeptide.-127- SG Docket No.: 14989-700.60082. The method of claim 81, wherein the therapeutic polypeptide has a molecular weight of at least about 0.5 kDa.

83. The method of claim 81, wherein the therapeutic polypeptide has a molecular weight of about 0.5 kDa to about 5 kDa, about 5 kDa to about 10 kDa, about 10 kDa to about 20 kDa, about 20 kDa to about 30 kDa, about 30 kDa to about 60 kDa, about 60 kDa to about 110 kDa, or about 110 kDa to about 200 kDa, or greater than about 200 kDa.

84. The method of any one of claims 81-83, wherein the therapeutic polypeptide is an enzyme, receptor or ligand, cell surface receptor, ion channel (i.e., pore-forming membrane protein), kinase, blood protein, chemokine, cytokine, antibody, antibody fragment, antigen, anticoagulant, blood factor, bone morphogenetic protein, growth factor, transcription factor, interferon, interleukin, thrombolytic agent, cytoskeletal protein, or peptide hormone.

85. The method of any one of claims 81-83, wherein the therapeutic polypeptide is a peptide hormone, protein hormone, bacterial / antibiotic peptide, antifungal peptide, invertebrate peptide, venom derived peptide, anticancer peptide, vaccine peptide, anti-inflammatory peptide, neuropeptide, endocrine peptide, gastrointestinal peptide, cardiovascular peptide, renal peptide, respiratory peptide, opioid peptide, neurotrophic peptide.

86. The method of any one of claims 80-83, wherein the therapeutic polypeptide is a monoclonal antibody, polyclonal antibody, single-domain antibody (sdAb, VHH, or nanobody), VHH-Fc, heavy chain only antibody (HCAb), a single chain variable fragment (scFv), di-scFv, tri-scFv, F(ab) fragment antibody, F(ab')2 fragment antibody, bispecific antibody (trifunctional antibody, chemically linked Fab, bi-specific T-cell engager), microantibody, intrabody, affibody molecule, Affilin, Affimer, Affitin, Alphabody, Anticalin, Avimer, DARPin, Kunitz domain peptide, Monobody, nanoCLAMP87. The method of any one of claims 28-80, wherein the polypeptide is an IgM antibody, IgD antibody, IgG antibody, IgA antibody, or IgE antibody.

88. The method of any one of claims 28-80, wherein polypeptide is an IgGl antibody, IgG2 antibody, IgG3 antibody, IgG4 antibody, IgAl antibody or IgA2 antibody.-128- SG Docket No.: 14989-700.60089. The method of any one of claims 28-80, wherein the polypeptide is a full-length cell surface receptor.

90. The method of any one of claims 28-80, wherein the polypeptide is only the extracellular domain of a cell surface receptor.

91. The method of claim 89 or claim 90, wherein the cell surface receptor is on the surface of a living or non-living cell.

92. The method of claim 89 or claim 90, wherein the cell surface receptor is not associated with a cell.

93. A compound of Formula (I):PBU-I^-l PCU (Formula (I))wherein:PBU is a moiety capable of non-covalently binding to a peptidic recognition motif (PRM) in a polypeptide, wherein the at least one PRM is independently selected from PRM1, PRM2, or combinations thereof;wherein each PRM1 comprises:• 2 to 4 histidine (His) residues; and• a lysine (Lys) residue, an ornithine (Orn) residue, a homolysine (HLys) residue, a p2-lysine (P2-Lys) residue, or a p3-lysine (P3-Lys) residue;: wherein each PRM2 comprises:• at least 2 amino acids selected from an arginine (Arg) residue, a homoarginine (HArg) residue, a citrulline (Cit) residue, a norarginine (AGBA) residue, a p2- arginine (P2-Arg), and a p3-arginine (P3-Arg) residue; and• a lysine (Lys) residue, an ornithine (Orn) residue, a homolysine (HLys) residue, a p2-lysine (P2-Lys) residue, or a p3-lysine (P3-Lys) residue;PCU is a moiety capable of covalently reacting with a side chain of the at least one Lys, Orn, HLys, p2-Lys, or p3-Lys in the PRM when held at a proximity -induced binding distance to the side chain of the at least one Lys, Orn, HLys, p2-Lys, or p3- Lys,;L1is a cleavable linker; andL2is a spacer;wherein upon non-covalent bonding of the PBU to the PRM of the polypeptide, the-129- SG Docket No.: 14989-700.600PCU is held at the proximity-induced binding distance of the side chain of the at least one Lys, Om, HLys, p2-Lys, or p3-Lys.

94. The compound of claim 93, wherein the PBU comprises at least one nitrilotriacetic acid, at least one iminodiacetic acid, or at least one pyridin-2-ylmethanamine, or combinations thereof.

95. The compound of claim 93, wherein the PBU comprises at least one nitrilotriacetic acid, or at least one iminodiacetic acid, or combinations thereof.

96. The compound of claim 93, wherein:

97. The compound of claim 93, wherein:the PBU comprises at least one moiety that i98. The compound of claim 93, wherein:-ISO- SG Docket No.: 14989-700.600U1is CH, C-OH, orN99. The compound of claim 93, wherein:U1is CH, C-OH, orN.

100. The compound of any one of claims 93-99, wherein PCU comprises an electrophilic warhead 5moiety that is capable of covalently reacting with the side chain of the at least one Lys, Om, HLys, p2-Lys, or p3-Lys in the PRM only when held within the proximity- induced binding distance to the at least one Lys, Om, HLys, p2-Lys, or p3-Lys in the PRM.

101. The compound of any one of claims 93-99, wherein the PCU comprises an electrophilic warhead moiety that is capable of covalently reacting with the side chain of the at least one Lys, Om, HLys, p2-Lys, or p3-Lys in the PRM upon binding of the PBU with the PRM, wherein the electrophilic warhead moiety is a vinyl sulfone group, a vinyl sulfonamide group, a pyrimidine sulfoxide group, a pyrimidine sulfone group, a vinyl pyridine group, or a vinyl pyrimidine group.

102. The compound of any one of claims 93-101, wherein, the PCU comprises an electrophilic warhead moiety that is capable of covalently reacting with the side chain of the at least one Lys, Om, HLys, p2-Lys, or p3-Lys in the PRM upon binding of the PBU with the PRM, wherein the electrophilic warhead is:-131- SG Docket No.: 14989-700.600"wherein:R1is H, halogen, substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, or -C(=O)OR3; andR2is H, halogen, substituted or unsubstituted alkyl, or -CN.

103. The compound of any one of claims 93-101, wherein:wherein:R1is H, halogen, substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, pyrimidine, or -C(=O)OR3; andR2is H, halogen, substituted or unsubstituted alkyl, or -CN; andeach R3is independently H or substituted or unsubstituted alkyl.

104. The compound of any one of claims 93-101, wherein:thewherein:R1is H, halogen, substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, pyrimidine, or -C(=O)OR3; andR2is H, halogen, substituted or unsubstituted alkyl, or -CN; andeach R3is independently H or substituted or unsubstituted alkyl.

105. The compound of claim 93, wherein the compound has the following structure:-132- SG Docket No.: 14989-700.600wherein:R1is H, halogen, substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, pyrimidine, or -C(=O)OR3; andR2is H, halogen, substituted or unsubstituted alkyl, or -CN; andR3is independently H or substituted or unsubstituted alkyl.

106. The compound of any one of claims 102-105, wherein:R1is H, halogen, methyl, ethyl, propyl, n-butyl, sec-butyl, tert-butyl, n-pentyl,107. The compound of any one of claims 103-106, wherein:R2is H, halogen, methyl, ethyl, propyl, n-butyl, sec-butyl, tert-butyl, n-pentyl, neopentyl, or -CN; andeach R3is independently H, methyl, ethyl, propyl, tert-butyl, n-pentyl, or neopentyl.

108. The compound of any one of claims 103-106, wherein:R1is H;R2is H; andeach R3is methyl.

109. The compound of any one of claims 93-108, wherein L1comprises a cleavable linker-133- SG Docket No.: 14989-700.600that upon cleavage provides an electrophilic or nucleophilic moiety.

110. The compound of any one of claims 93-108, wherein L1is orthogonally cleavable to peptide cleavage.

111. The compound of any one of claims 93-108, wherein L1comprises a self-immolative linker.

112. The compound of any one of claims 93-108, wherein L1comprises a disulfide group linker, photoreactive linker, fluoride-cleavable linker, alkali-cleavable linker, or oxidation cleavable linker.

113. The compound of any one of claims 93-108, wherein:L1comprises a nucleophilic cleavable linker, electrophile cleavable linkers, acid cleavable linker, base cleavable linker, linker cleavable with reducing reagents, linker cleavable with oxidizing reagents, or photocleavable linker.

114. The compound of any one of claims 93-108, wherein:L1comprises a dialkoxy silane linker, a 2-cyanoethyl moiety, sulfone linker, ester linker, thiophenylester linker, vinyl sulfide linker, vinyl ether linker, acylhydrazone linker, hydrazone linker, acylsulfonamide linker, disulfide linker, azo linker, orthonitrobenzyl linker, phenacyl ester linker, ketal linker, carbamate linker, selenium linker, vicinal diol linker, para-aminobenzyl (PAB)-carbamate linker, transcyclooctene carbamate linker.

115. The compound of any one of claims 93-108, wherein:-134- SG Docket No.: 14989-700.600116. The compound of any one of claims 93-115, whereinL2is not cleavable.

117. The compound of any one of claims 93-115, wherein:L2comprises one or more linear structures, one or more non-linear structures, one or more branched structures, one or more cyclic structures, one or more flexible moieties, one or more rigid moieties, or combinations thereof.

118. The compound of any one of claims 93-115, wherein:-135- SG Docket No.: 14989-700.600stituted alkyl.

119. The compound of any one of claims 93-108, wherein:L1comprises a cleavable linker that upon cleavage provides a moiety capable of forming a linkage with a coupling partner, wherein the cleavable linker is orthogonally cleavable to peptide cleavage;L2further comprises a moiety capable of forming a linkage with a coupling partner; wherein the linkage formation reaction in L1is orthogonal to the linkage formation reaction in L2.

120. The compound of claim 119, wherein:the linkage formation reaction in L1is a coupling reaction between an electrophile and a nucleophile.

121. The compound of claim 120, wherein the coupling reaction is an amide-forming ligation, ester-forming ligation, ether-forming ligation, imine-forming ligation, oxime-forming ligation, hydrazone-forming ligation, or sulfonamide-forming ligation.

122. The compound of any one of claims 119-121, wherein:the linkage formation reaction in L2is a cycloaddition reaction between an azide and alkyne to form a triazole, a strain-promoted azide-alkyne cycloaddition (SPAAC), inverse electron-demand Diels-Alder (IEDDA), or sydnone-alkyne cycloaddition (SPSAC).

123. The compound of claim 93, wherein the compound has the following structure:-136- SG Docket No.: 14989-700.600124. A compound having the structure of:wherein,U1is CH, C-OH, or N.L1is a cleavable linker; L2is a spacer;R3is H or substituted or unsubstituted alkyl;His is histidine;m is 0, 1, 2, 3, or 4; n is 0, 1, 2, 3, or 4, provided that m + n is at least 2; q is 1, 2, or 3;Rais a polypeptide B, and Rbis -OH; or Rais H, and Rbis a polypeptide B; or Rais a fragment (B1) of a polypeptide B, Rbis the other fragment (B2) of the polypeptide B, such that B^B2is B; orRais H, and Rbis -OH.

125. A compound having the structure of:-137- SG Docket No.: 14989-700.600wherein,U1is CH, C-OH, orN.L1is a cleavable linker; L2is a spacer;each R3is H or substituted or unsubstituted alkyl;q is 1, 2, or 3;Rais a polypeptide, and Rbis -OH; or Rais H, and Rbis a polypeptide; orRais a fragment (B1) of a polypeptide B, Rbis the other fragment (B2) of the polypeptide B, such that B^B2is B; orRais H, and Rbis -OH.

126. A compound having the structure of:wherein,U1is CH, C-OH, orN;L1is a cleavable linker; L2is a spacer;each R3is H or substituted or unsubstituted alkyl;W1is a -X^Y-X^Y-, and W2is absent; or W1is absent, and W2is -X^Y-X^Y-; or W1and W2are both -X1-Y-X2-Y-;;each Y is independently selected from Arg, HArg, Cit, AGBA, p2-Arg, and p3-Arg, or-138- SG Docket No.: 14989-700.600Y is absent;X1comprises 0 to 10 amino acid residues other than Lys, Om, HLys, and Y;X2comprises 0 to 10 amino acids other than Lys, Orn, HLys, and Y;q is 1, 2, or 3;Rais a polypeptide B, and Rbis -OH; or Rais H, and Rbis a polypeptide B; or Rais a fragment (B1) of a polypeptide B, Rbis the other fragment (B2) of the polypeptide B, such that B^B2is B; orRais H, and Rbis -OH.

127. An antibody or antibody fragment comprising a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide has at least 90% identity with SEQ ID NO. 1, further wherein a sequence corresponding to PRM1 or PRM2 is either at a C-terminal end of the polypeptide or is located between positions 38 to 50 or positions 94-104 of SEQ ID NO. 1, either:a. in place of any single amino acid in positions 38 to 50 or positions 94-104 of SEQ ID NO. 1, orb. between any two consecutive amino acids in positions 38 to 50 or positions 94-104 of SEQ ID NO. 1, wherein:the sequence of PRM1 is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the sequence of PRM2 is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Om, HLys, P2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

128. The antibody or antibody fragment of claim 127, wherein the PRM is a PRM1.

129. The antibody or antibody fragment of claim 127, wherein PRM1 comprises one of: X- His-His, His-X-His, His-His-X, X-His-His-His, His-X-His-His, His-His-X-His, His-His--139- SG Docket No.: 14989-700.600His-X, X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, or His-His-His-His-X, wherein X is Lys, Orn, HLys, p2-Lys, or p3-Lys.

130. The antibody or antibody fragment of claim 127, wherein PRM1 comprises His-Lys- His-His or His-His-Lys-His.

131. The antibody or antibody fragment of claim 127, wherein the PRM is PRM2.

132. The antibody or antibody fragment of claim 127, wherein PRM2 comprises one of SEQ ID NO. 32 to SEQ ID NO. 51.

133. The antibody or antibody fragment of claim 127, wherein PRM2 comprises Arg-Lys- Arg.

134. The antibody or antibody fragment of claim 127, wherein the PRM is covalently attached at a C-terminus of the antibody or antibody fragment.

135. The antibody or antibody fragment of claim 127, wherein the PRM is attached to a C- terminus of the antibody or antibody fragment through a linker.

136. The antibody or antibody fragment of claim 135, wherein the linker comprises between 1 and 10 Pro- Ala dipeptide units.

137. The antibody or antibody fragment of claim 135, wherein the linker comprises between 1 and 10 Gly-Ser dipeptide units.

138. An antibody or antibody fragment comprising a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a first amino acid sequence that has at least 95% identity with SEQ ID NO. 2, and a second amino acid sequence that has at least 95% identity with SEQ ID NO. 3 and a PRM1 or PRM2 sequence between the first amino acid sequence and the second amino acid sequence, wherein:the sequence of PRM1 is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the sequence of PRM2 is: X3-XLY -X4-X2-Y, wherein X3or X4is Lys, Om, HLys,-140- SG Docket No.: 14989-700.600P2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

139. An antibody or antibody fragment comprising a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a first amino acid sequence that has at least 95% identity with SEQ ID NO. 2, linked to a PRM sequence by v linking amino acids, where v is between 0 and 13, further wherein the PRM sequence consists of either a PRM1 sequence or a PRM2 sequence, further wherein:the PRM1 sequence is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the PRM2 sequence is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Om, HLys, P2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

140. The antibody or antibody fragment of claim 139, wherein, when v is nonzero, the linking amino acids consist of positions 1 to v of SEQ. ID. NO. 613.

141. An antibody or antibody fragment comprising a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a first amino acid sequence that has at least 95% identity with SEQ ID NO. 4, linked to a PRM sequence by v linking amino acids, where v is between 0 and 13, wherein the PRM sequence consists of either a PRM1 sequence or a PRM2 sequence, further wherein:the PRM1 sequence is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or -141- SG Docket No.: 14989-700.6004, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the PRM2 sequence is: X3-XLY -X4-X2-Y, wherein X3or X4is Lys, Om, HLys, P2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

142. The antibody or antibody fragment of claim 141, wherein when v is nonzero, the linking amino acids consist of positions 1 to v of SEQ. ID. NO. 613.

143. An antibody or antibody fragment comprising a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a first amino acid sequence that has at least 95% identity with SEQ ID NO. 1, linked at a C-terminus to a PRM sequence either directly or by a linker, wherein the PRM sequence consists of either a PRM1 sequence or a PRM2 sequence, further wherein:the PRM1 sequence is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the PRM2 sequence is: X3-XLY -X4-X2-Y, wherein X3or X4is Lys, Om, HLys, P2-Lys, or P3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

144. The antibody or antibody fragment of claim 143, wherein the linker comprises between 1 and 10 Pro- Ala dipeptide units.

145. The antibody or antibody fragment of claim 143, wherein the linker comprises between 1 and 10 Gly-Ser dipeptide units.-142- SG Docket No.: 14989-700.600146. An antibody or antibody fragment comprising a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide has at least 90% identity with SEQ ID NO. 12, further wherein a sequence corresponding to PRM1 or PRM2 is at one or more of: a C-terminal end of the polypeptide, an N-terminal end, or is located between positions 5 to 22 or positions 126 to 136 of SEQ ID NO. 12, either:c. in place of any single amino acid in positions 5 to 22 or positions 126 to 136 of SEQ ID NO. 12, ord. between any two consecutive amino acids in positions 5 to 22 or positions 126 to 136 of SEQ ID NO. 12, wherein:the sequence of PRM1 is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the sequence of PRM2 is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Om, HLys, P2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

147. The antibody or antibody fragment of claim 146, wherein the PRM is a PRM1.

148. The antibody or antibody fragment of claim 146, wherein PRM1 comprises one of: X- His-His, His-X-His, His-His-X, X-His-His-His, His-X-His-His, His-His-X-His, His-His- His-X, X-His-His-His-His, His-X-His-His-His, His-His-X-His-His, His-His-His-X-His, or His-His-His-His-X, wherein X is Lys, Orn, HLys, p2-Lys, or p3-Lys.

149. The antibody or antibody fragment of claim 146, wherein PRM1 comprises His-Lys- His-His or His-His-Lys-His.

150. The antibody or antibody fragment of claim 146, wherein the PRM is PRM2.

151. The antibody or antibody fragment of claim 146, wherein PRM2 comprises one of SEQ ID NO. 32 to SEQ ID NO. 51.-143- SG Docket No.: 14989-700.600152. The antibody or antibody fragment of claim 146, wherein PRM2 comprises Arg-Lys- Arg.

153. The antibody or antibody fragment of claim 146, wherein the PRM is covalently attached at a C-terminus of the antibody or antibody fragment.

154. The antibody or antibody fragment of claim 146, wherein the PRM is attached to a C- terminus of the antibody or antibody fragment through a linker.

155. The antibody or antibody fragment of claim 154, wherein the linker comprises between 1 and 10 Pro- Ala dipeptide units.

156. The antibody or antibody fragment of claim 154, wherein the linker comprises between 1 and 10 Gly-Ser dipeptide units.

157. An antibody or antibody fragment comprising a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a first amino acid sequence that has at least 95% identity with SEQ ID NO. 15, and a second amino acid sequence that has at least 95% identity with SEQ ID NO. 16 and a PRM1 or PRM2 sequence between the first amino acid sequence and the second amino acid sequence, wherein:the sequence of PRM1 is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the sequence of PRM2 is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Om, HLys, P2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

158. An antibody or antibody fragment comprising a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a PRM sequence linked to a second-144- SG Docket No.: 14989-700.600amino acid sequence that has at least 95% identity with SEQ ID NO. 14, by v linking amino acids, where v is between 0 and 18, further wherein the PRM sequence consists of either a PRM1 sequence or a PRM2 sequence, further wherein:the PRM1 sequence is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the PRM2 sequence is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Om, HLys, P2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

159. The antibody or antibody fragment of claim 158, wherein, when v is nonzero, the linking amino acids consist of positions (19-v) to 18 of SEQ. ID. NO. 614.

160. An antibody or antibody fragment comprising a polypeptide including a precision conjugation site, the antibody or antibody fragment including a peptidic recognition motif (PRM) consisting of peptidic recognition motif 1 (PRM1) or peptidic recognition motif 2 (PRM2), wherein the polypeptide consist of: a first amino acid sequence that has at least 95% identity with SEQ ID NO. 12, linked at a C-terminus to a PRM sequence either directly or by a linker, wherein the PRM sequence consists of either a PRM1 sequence or a PRM2 sequence, further wherein:the PRM1 sequence is (His)m-X-(His)n, wherein X is lysine (Lys), ornithine (Om), homolysine (HLys), p2-lysine (P2-Lys), or p3-lysine (P3-Lys), m is 0, 1, 2, 3, or 4, and n is 0, 1, 2, 3, or 4, provided that m + n is between 2 and 4; and the PRM2 sequence is: X3-X1-Y -X4-X2-Y, wherein X3or X4is Lys, Om, HLys, P2-Lys, or p3-Lys, if X3is present, then X4is not present, if X4is present, then X3is not present, each Y is independently selected from Arg, HArg, Cit, AGBA, P2-Arg, and p3-Arg, X1comprises 0 to 10 amino acid residues other than X3, X4, and Y, and X2comprises 0 to 10 amino acids other than X3, X4, and Y.

161. The antibody or antibody fragment of claim 160, wherein the linker comprises between 1 and 10 Pro- Ala dipeptide units.-145- SG Docket No.: 14989-700.600162. The antibody or antibody fragment of claim 160, wherein the linker comprises between 1 and 10 Gly-Ser dipeptide units.

163. A method of selectively modifying a lysine residue within an antibody or antibody fragment comprising:contacting a compound of any one of claims 1-19, with the antibody or antibody fragment of any one of claims 127-162, in the presence of a nickel (Ni2+) salt, cobalt (Co2+) salt, copper (Cu2+) salt, or zinc (Zn2+) salt, in a suitable solvent; wherein the PRM comprises at least one PRM1.

164. The method of claim 163, wherein the lysine residue that is selectively modified is the lysine in the at least one PRM1.

165. A method of selectively modifying a lysine residue within a polypeptide comprising:contacting a compound of any one of claims 1-19, with an antibody or antibody fragment of any one of claims 127-162, in a suitable solvent in the absence of a transition metal salt;wherein the PRM comprises at least one PRM2.

166. The method of claim 165, wherein the lysine residue that is selectively modified is the lysine in the at least one PRM2.-146- SG Docket No.: 14989-700.600

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