Targeted drug polymer conjugates

Abstract

Disclosed herein are copolymers with a first repeating units having the structure of Formula (I), and a second repeating units having the structure of Formula (II). Disclosed herein are conjugates with a first repeating units having the structure of Formula (I), and a second repeating units having the structure of Formula (II) and compounds. Disclosed herein is a method of making the conjugate by reacting the copolymer with the compound, where Z1 of copolymer reacts with Z2 of the compound via click reaction in the absence of a metal. Disclosed herein is a method of treating a disease or condition by administering to a subject in need thereof the conjugate.

Description

TARGETED DRUG POLYMER CONJUGATES CROSS REFERENCE TO RELATED APPLICATIONS

[0001] The present application claims priority to U.S. Provisional Application No. 63 / 697408, filed on September 20, 2024, which is incorporated by reference in its entirety. BACKGROUND Field

[0002] The present invention relates to conjugates and methods thereof for bioactive payloads attached to a zwitterionic based polymer scaffold, and release of the bioactive payloads for therapeutic effects. Description of the Related Art

[0003] Antibody-drug conjugates (ADCs) are designed to deliver therapeutic or diagnostic payloads with precision, but existing approaches continue to face limitations in achieving high loading while maintaining solubility, stability, and favorable pharmacokinetics. Polymer-based scaffolds have been explored as a method to improve these characteristics. Incorporation of zwitterionic monomers has been shown to enhance hydrophilicity, reduce aggregation, and improve biocompatibility, while functional co-monomers can allow diverse conjugation chemistries. Continued development in this area is expected to provide more versatile and tunable conjugates with potential impact in therapeutic and diagnostic applications. SUMMARY

[0004] Disclosed herein is a copolymer with a first repeating units having the structure of Formula (I):

[0005] a second repeating units having the structure of Formula (II):,

[0006] where m > 0, n > 0, and m + n ≤ 8000, X1 is H or methyl; X2 is H, F, or methyl; Y is -O- or -NH-; R is absent, optionally substituted C1-12 alkylene, optionally substituted; q is 1, 2, 3, or 4; and Z1 comprises N3 or a cyclic olefin. In some embodiments, R is unsubstituted C1-3 alkylene, for example methylene, ethylene, or propylene.

[0007] Disclosed herein is a compound having the structure: , where, , bicyclic cycloalkyne, 7- to 9- membered monocyclic or bicyclic heterocycloalkyne, 10- to 16- membered tricyclic heterocycloalkyne, or optionally substituted tetrazine; and R7 is,, , , wherein z is 1, 2, 3, or 4, and is aryl; R’ is absent or is selected from the group consisting of, ,-2-integer between 1 and 15; X3 is H, –(CH2)b–phosphorylcholine, or C1-3 alkyl optionally substituted by COOH, OH, or C(O)NH2; X4is C1-3 alkyl; X5is C1-3 alkyl substituted by OH, COOH, C(O)NH2, or –(CH2)b–phosphorylcholine; b is 0, 1, 2, 3, or 4; L is absent or selected ,, or 4; M is O or N-alkyl; X6is optionally substituted C1-3 alkyl, optionally substituted phenyl, or optionally substituted pyridinyl; X7 and X8 are each H or methyl; and PL comprises a bioactive agent, tracer, or reactive functional group.

[0008] Disclosed herein is a conjugate with a first monomer repeating units having the structure of Formula (I):,

[0009] a second monomer repeating units having the structure of Formula (IIa):-3-

[0010] where m + n ≤ 8000; X1 is H or methyl; X2 is H, F, or methyl; Y is -O- or - NH-; R is optionally substituted C1-12 alkylene, optionally substituted, q is an integer between 1 and 4; and Z compriseswhere Z’ is a reaction productof (1) N3and cycloalkyne derivative or heterocycloalkyne derivative or (2) olefin and optionally substituted tetrazine derivative; R’ is absent or is selected from the group consisting , , ,, , , , , –(CH2)b–phosphorylcholine, or C1-3 alkyl optionally substituted by COOH, OH, or C(O)NH2; X4is C1-3 alkyl; X5is C1-3 alkyl substituted by OH, COOH, C(O)NH2, or –(CH2)b– phosphorylcholine; b is 0, 1, 2, 3, or 4; L is absent or selected from the group consisting of-4-optionally substituted phenyl, or optionally substituted pyridinyl; X7 and X8 are each H or methyl; and PL comprises a bioactive agent, tracer, or reactive functional group.

[0011] Disclosed herein is a method of making the conjugate by reacting the copolymer with the compound, where Z1 of copolymer reacts with Z2 of the compound via click reaction in the absence of a metal.

[0012] Disclosed herein is a method of treating a disease or condition by administering to a subject in need thereof the conjugate. DETAILED DESCRIPTION

[0013] Disclosed herein is a powerful and highly adaptable platform for payload conjugation through click chemistry. In some embodiments, a payload is modified such that the modified payload can be attached to a copolymer to form a polymer-drug conjugate. In some embodiments, the payload can be any suitable compound, including, but not limited to: small-molecule drugs, peptides, oligonucleotides, proteins, aptamers, tracers, and any combination thereof. In some embodiments, the copolymer comprises a phosphorylcholine (PC) polymer backbone. PC-based polymers provide a solid foundation for the copolymers and conjugates disclosed herein, as PC-based polymers are already well-recognized for their high water solubility, excellent biocompatibility, and favorable safety profile. Importantly, this architecture can support conjugation of more than 300 payloads per polymer, representing a major breakthrough compared to the drug-to-antibody ratio (DAR) limitations faced in antibody-drug-conjugate chemistry. Moreover, by employing linkers to regulate release half- life, the system allows precise control over payload pharmacokinetics in vivo, enabling extended circulation times, reduced dosing frequency, and improved therapeutic benefit— addressing key clinical challenges in drug delivery. Disclosed herein are a range of diverse copolymer backbones, multiple payload formats, and sequential conjugations, thereby enabling broad applicability across therapeutic, diagnostic, imaging, and material science contexts.

[0014] Further, the copolymers and conjugates disclosed herein have the capacity to support multiple modified payload (Z₂–R’–L–PL) conjugations on the same copolymer backbone, while still preserving independent release behavior of each payload. Even when two or more payload-linker constructs are conjugated sequentially, each payload demonstrates its-5-own predictable release kinetics, closely matching that of the corresponding single-payload conjugates. Beyond release half-life control, the copolymers and conjugates disclosed herein also allow for rational combination of different linker chemistries—including stable linkers, enzyme-cleavable linkers, and pH-sensitive linkers—thereby enabling fine-tuned pharmacokinetic and pharmacodynamic outcomes in clinical applications. In some embodiments, the modified payloads are attached to the copolymers via Strain-Promoted Azide-Alkyne Cycloaddition (SPAAC). SPAAC is a type of Click Chemistry that is well understood, entirely metal-free, requires no additional reagents, and proceeds with high efficiency and minimal by-products. This greatly simplifies pharmaceutical manufacturing by reducing purification burdens and eliminating concerns over residual metal contaminants. This high level of efficiency and cleanliness is not a generic property of all click chemistry; it arises specifically from the unique pairing of the Z1 group of the copolymers disclosed herein with the Z2–R’–L–PL construct in thie present disclosure, which together enable a balance of selectivity, stability, and scalability. Together, these design elements create a platform for synthesizing conjugates that are not only structurally stable and versatile but also highly translatable to clinical and commercial applications, positioning the conjugates disclosed herein as a true breakthrough in next-generation conjugate technology.

[0015] Disclosed herein is a copolymer with a first repeating units having the structure of Formula (I):,

[0016] a second repeating units having the structure of Formula (II):-6-

[0017] where m > 0, n > 0, and m + n ≤ 8000, X1is H or methyl; X2is H, F, or methyl; Y is -O- or -NH-; R is absent, optionally substituted C1-12 alkylene, optionally substituted; q is 1, 2, 3, or 4; and Z1 comprises N3 or a cyclic olefin.

[0018] Disclosed herein is a compound having the structure wheremembered monocyclic or bicyclic cycloalkyne, 7- to 9- membered monocyclic or bicyclic heterocycloalkyne, 10- to 16- membered tricyclic heterocycloalkyne, or optionally substituted tetrazine; and R7 is,, , , , , , , aryl; R’ is absent or is selected from the group consisting of, , , , ,, , , , , , n integer between 1 and 15; X3 is H, –(CH2)b–phosphorylcholine, or C1-3 alkyl optionally substituted by COOH, OH, or C(O)NH2; X4is C1-3 alkyl; X5is C1-3 alkyl substituted by OH, COOH, C(O)NH2, or –(CH2)b–phosphorylcholine; b is 0, 1, 2, 3, or 4; L is absent or selected-7-, or 4; M is O or N-alkyl; X6is optionally substituted C1-3 alkyl, optionally substituted phenyl, or optionally substituted pyridinyl; X7and X8are each H or methyl; and PL comprises a bioactive agent, tracer, or reactive functional group. In some embodiments, the reactive functional group may comprise maleimide, iodoacetamide, and / or bromoacetamide.

[0019] Disclosed herein is a conjugate with a first monomer repeating units having the structure of Formula (I):,

[0020] a second monomer repeating units having the structure of Formula (IIa):,

[0021] where m + n ≤ 8000; X1 is H or methyl; X2 is H, F, or methyl; Y is -O- or - NH-; R is optionally substituted C1-12 alkylene optionally substitutedq, q isan integer between 1 and 4; and Z comprises; where Z’ is a reaction productof (1) N3and cycloalkyne derivative or heterocycloalkyne derivative or (2) olefin and optionally substituted tetrazine derivative; R’ is absent or is selected from the group consisting-8-, , ,, –(CH2)b–phosphorylcholine, or C1-3 alkyl optionally substituted by COOH, OH, or C(O)NH2; X4 is C1-3 alkyl; X5 is C1-3 alkyl substituted by OH, COOH, C(O)NH2, or –(CH2)b– phosphorylcholine; b is 0, 1, 2, 3, or 4; L is absent or selected from the group consisting of, , , , ,7; p is 2, 3, or 4; r is 1, 2, 3, or 4; X6is optionally substituted C1-3 alkyl, optionally substituted phenyl, or optionally substituted pyridinyl; X7 and X8 are each H or methyl; and PL comprises a bioactive agent, tracer, or reactive functional group. In some embodiments, R is unsubstituted C1-3 alkylene, for example methylene, ethylene, or propylene. In some embodiments, R is substituted C1-3 alkylene. In some embodiments, R is substituted C3 alkylene; in some embodiments, the C3 alkylene is substituted with a hydroxy group. In some embodiments, R is optionally substituted; in some embodiments, q is 3. In some embodiments, R is absent. In some embodiments, the reactive functional group may comprise maleimide, iodoacetamide, and / or bromoacetamide.-9-

[0022] In some embodiments, the conjugate may further include monomer units that are unreacted, suchwherein X2, Y, R, Z1, and n are as disclosed herein.

[0023] Disclosed herein is a method of making the conjugate by reacting the copolymer with the compound, where Z1 of copolymer reacts with Z2 of the compound via click reaction in the absence of a metal.

[0024] Disclosed herein is a method of treating a disease or condition by administering to a subject in need thereof the conjugate. Definitions

[0025] For the purpose of the present disclosure the following terminology will be used in accordance with the definitions set forth below.

[0026] A “polymer” is a molecule composed of many repeating subunits. The subunits, also sometimes referred to as “monomers,” can be the same or different. A polymer composed of repeating units of a single monomer is called a homopolymer. A polymer composed of two or more monomers is called a copolymer or sometimes a heteropolymer. A copolymer in which certain monomer types are clustered together are sometimes called block copolymers. Polymers can be linear or branched. When the polymer is branched, polymer chains having a common origin (sometime called a core) are sometimes referred to as a polymer arm(s).

[0027] A “copolymer” refers to a polymer having at least two different monomer groups. The monomers of the copolymer are the chemical moieties that are bonded together to form the polymer. Each distinct chemical moiety is termed a monomer. The copolymers are prepared from monomers that include, but are not limited to, acrylates, methacrylates, acrylamides, methacrylamides, styrenes, vinyl-pyridine and vinyl-pyrrolidone. Additional monomers are useful in the copolymers of the present disclosure. When two different monomers are used, such as in the copolymers of the present disclosure, the two monomers are called “comonomers,” meaning that the different monomers are copolymerized to form a single polymer.-10-

[0028] “Block copolymer” refers to a type of copolymer in which certain monomer types are clustered together.

[0029] “Random copolymer” refers to a type of copolymer having at least two different monomer groups that are distributed randomly throughout the polymer backbone.

[0030] “Gradient copolymer” refers to a type of copolymer in which composition of the monomer units varies gradually along the polymer chain.

[0031] “Star copolymer” refers to a type of branched polymer with a general structure consisting of several linear chains connected to a central core.

[0032] “Initiator” refers to a compound capable of initiating a polymerization using the comonomers of the present disclosure. The polymerization can be a reversible-deactivation radical polymerization / controlled reversible-deactivation radical polymerization, such as Atom Transfer Radical Polymerization (ATRP), Reversible Addition-Fragmentation- Termination (RAFT) polymerization, or Nitroxide_Mediated Polymerization (NMP). The initiator (R-X) contains a labile bond that is cleaved to form two initiator fragments, R• and X•. When the initiator is suitable for ATRP, the labile bond can homolytically cleave to form two initiator fragments, the first (i.e., R•) being a radical capable of initiating a radical polymerization, and the second (i.e., X•) being a radical stabilized by a metal ion and a ligand. The first initiator fragment reacts with a monomer to begin growing the polymer chain. The radical of the first initiator fragment adds to the monomer, creating a new radical that propagates the polymer chain. In doing so, the first initiator fragment is attached to one end of the polymer. The first initiator fragment attached to one end of the polymer is referred to herein as “core.” The second initiator fragment reacts with the radical of the growing polymer chain to reversibly terminate the polymerization. The second initiator fragment is typically a halogen radical or pseudo halogen radical.

[0033] “Linker” refers to a chemical moiety that links two groups together. The linker can be cleavable or non-cleavable. Cleavable linkers can be hydrolyzable, enzymatically cleavable, pH sensitive, photolabile, or disulfide linkers, among others. Other linkers include homobifunctional and heterobifunctional linkers. A “linking group” is a functional group capable of forming a covalent linkage consisting of one or more bonds to a bioactive agent.-11-

[0034] “Hydrolyzable linker” refers to a chemical linkage or bond, such as a covalent bond, that undergoes hydrolysis under physiological conditions. The tendency of a bond to hydrolyze may depend not only on the general type of linkage connecting two central atoms between which the bond is severed, but also on the substituents attached to these central atoms. Non-limiting examples of hydrolytically susceptible linkages include esters of carboxylic acids, phosphate esters, acetals, ketals, acyloxyalkyl ether, imines, orthoesters, and some amide linkages.

[0035] “Enzymatically cleavable linker” refers to a linkage that is subject to degradation by one or more enzymes. Some hydrolytically susceptible linkages may also be enzymatically degradable. For example esterases may act on esters of carboxylic acid or phosphate esters, and proteases may act on peptide bonds and some amide linkages.

[0036] “pH sensitive linker” refers to a linkage that is stable at one pH and subject to degradation at another pH. For example, the pH sensitive linker can be stable at neutral or basic conditions, but labile at mildly acidic conditions.

[0037] “Photolabile linker” refers to a linkage, such as a covalent bond, that cleaves upon exposure to light. The photolabile linker includes an aromatic moiety in order to absorb the incoming light, which then triggers a rearrangement of the bonds in order to cleave the two groups linked by the photolabile linker.

[0038] “Functional agent” is defined to include a bioactive agent or a diagnostic agent. A “bioactive agent” is defined to include any agent, drug, compound, or mixture thereof that targets a specific biological location (targeting agent) and / or provides some local or systemic physiological or pharmacologic effect that can be demonstrated in vivo or in vitro.

[0039] Non-limiting examples include drugs, vaccines, antibodies, antibody fragments, vitamins and cofactors, polysaccharides, carbohydrates, steroids, lipids, fats, proteins, peptides, polypeptides, nucleotides, oligonucleotides, polynucleotides, and nucleic acids (e.g., mRNA, tRNA, snRNA, RNAi, DNA, cDNA, antisense constructs, ribozymes, etc).

[0040] A “diagnostic agent” is defined to include any agent that enables the detection or imaging of a tissue or disease. Examples of diagnostic agents include, but are not limited to, radiolabels, fluorophores and dyes.

[0041] “Therapeutic protein” refers to peptides or proteins that include an amino acid sequence which in whole or in part makes up a drug and can be used in human or animal-12-pharmaceutical applications. Numerous therapeutic proteins are known to practitioners of skill in the art including, without limitation, those disclosed herein.

[0042] “Phosphorylcholine,” also denoted as “PC,” refers to the following:

[0043] where * denotes the point of attachment. The phosphorylcholine is a zwitterionic group and includes salts (such as inner salts), and protonated and deprotonated forms thereof.

[0044] “Phosphorylcholine containing polymer” is a polymer that contains phosphorylcholine. It is specifically contemplated that in each instance where a phosphorylcholine containing polymer is specified in this application for a particular use, a single phosphorylcholine can also be employed in such use.

[0045] “Poly(acryloyloxyethyl phosphorylcholine) containing polymer” refers to a polymer of acrylic acid containing at least one acryloyloxyethyl phosphorylcholine monomer such as 2-methacryloyloxyethyl phosphorylcholine (i.e., 2-methacryloyl-2 '- trimethylammonium ethyl phosphate).

[0046] “Contacting” refers to the process of bringing into contact at least two distinct species such that they can react. It should be appreciated, however, that the resulting reaction product can be produced directly from a reaction between the added reagents or from an intermediate from one or more of the added reagents which can be produced in the reaction mixture.

[0047] “Water-soluble polymer” refers to a polymer that is soluble in water. A solution of a water-soluble polymer may transmit at least about 75%, more preferably at least about 95% of light, transmitted by the same solution after filtering. On a weight basis, a water- soluble polymer or segment thereof may be at least about 35%, at least about 50%, about 70%, about 85%, about 95% or 100% (by weight of dry polymer) soluble in water.

[0048] “Molecular weight” in the context of the polymer can be expressed as either a number average molecular weight or a weight average molecular weight. Unless otherwise indicated, all references to molecular weight herein refer to the weight average molecular weight. Both molecular weight determinations, number average and weight average, can be-13-measured using gel permeation chromatography or other liquid chromatography techniques. Other methods for measuring molecular weight values can also be used, such as the use of end- group analysis or the measurement of colligative properties (e.g., freezing-point depression, boiling-point elevation, or osmotic pressure) to determine number average molecular weight, or the use of light scattering techniques, ultracentrifugation or viscometry to determine weight average molecular weight. The polymeric reagents of the present disclosure are typically polydisperse (i.e., number average molecular weight and weight average molecular weight of the polymers are not equal), possessing low polydispersity values of preferably less than about 2, as judged by gel permeation chromatography.

[0049] “Dispersity,” “polydispersity,” or “polydispersity index” are used interchangeably herein to refer to the ratios Mw / Mn.

[0050] “Mw” refers to weight-average molecular weight.

[0051] “MN” refers to number-average molecular weight.

[0052] The phrase “a” or “an” entity as used herein refers to one or more of that entity; for example, a compound refers to one or more compounds or at least one compound. As such, the terms “a” (or “an”), “one or more”, and “at least one” can be used interchangeably herein.

[0053] “About” as used herein means variation one might see in measurements taken among different instruments, samples, and sample preparations.

[0054] “Protected,”, “protected form'', “protecting group” and “protective group” refer to the presence of a group (i.e., the protecting group) that prevents or blocks reaction of a particular chemically reactive functional group in a molecule under certain reaction conditions. Protecting group will vary depending upon the type of chemically reactive group being protected as well as the reaction conditions to be employed and the presence of additional reactive or protecting groups in the molecule, if any. The skilled artisan will recognize protecting groups known in the art, such as those found in the treatise by Greene et al., “Protective Groups In Organic Synthesis,” 3rd Edition, John Wiley and Sons, Inc., New York, 1999.

[0055] “Spacer,” and “spacer group” are used interchangeably herein to refer to an atom or a collection of atoms optionally used to link interconnecting moieties such as a terminus of a water-soluble polymer and a reactive group of a functional agent and a reactive-14-group. A spacer may be hydrolytically stable or may include a hydrolytically susceptible or enzymatically degradable linkage.

[0056] “Alkyl” refers to a straight or branched, saturated, aliphatic radical having the number of carbon atoms indicated. For example, C1-C6 alkyl includes, but is not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, hexyl, etc. Other alkyl groups include, but are not limited to heptyl, octyl, nonyl, decyl, etc. Alkyl can include any number of carbons, such as 1-2, 1-3, 1-4, 1-5, 1-6, 1-7, 1-8, 1-9, 1-10, 2-3, 2-4, 2-5, 2-6, 3-4, 3-5, 3-6, 4-5, 4-6 and 5-6. The alkyl group is typically monovalent, but can be divalent, such as when the alkyl group links two moieties together.

[0057] The term “lower” referred to above and hereinafter in connection with organic radicals or compounds respectively defines a compound or radical which can be branched or unbranched with up to and including 7, preferably up to and including 4 and (as unbranched) one or two carbon atoms.

[0058] “Alkylene” refers to an alkyl group, as defined above, linking at least two other groups, i.e., a divalent hydrocarbon radical. The two moieties linked to the alkylene can be linked to the same atom or different atoms of the alkylene. For instance, a straight chain alkylene can be the bivalent radical of -(CH2)n, where n is 1, 2, 3, 4, 5 or 6. Alkylene groups include, but are not limited to, methylene, ethylene, propylene, isopropylene, butylene, isobutylene, sec-butylene, pentylene and hexylene.

[0059] Substituents for the alkyl and heteroalkyl radicals (including those groups often referred to as alkylene, alkenyl, heteroalkylene, heteroalkenyl, alkynyl, cycloalkyl, heterocycloalkyl, cycloalkenyl, and heterocycloalkenyl) can be a variety of groups selected from: -OR', =O, =NR', =N-OR', -NR'R'', -SR', -halogen, -SiR'R”R”', -OC(O)R', -C(O)R', - CO2R', -CONR'R”, -OC(O)NR'R”, -NR”C(O)R', -NR'-C(O)NR”R”', -NR”C(O)2R', -NH- C(NH2)=NH, -NR'C(NH2)=NH, -NH-C(NH2)=NR', -S(O)R', -S(O)2R', -S(O)2NR'R”, -CN and -NO2in a number ranging from zero to (2m'+1), where m' is the total number of carbon atoms in such radical. R', R” and R”' each independently refer to hydrogen, unsubstituted (C1- C8)alkyl and heteroalkyl, unsubstituted aryl, aryl substituted with 1-3 halogens, unsubstituted alkyl, alkoxy or thioalkoxy groups, or aryl-(C1-C4)alkyl groups. When R' and R” are attached to the same nitrogen atom, they can be combined with the nitrogen atom to form a 5-, 6-, or 7- membered ring. For example, -NR'R” is meant to include 1-pyrrolidinyl and 4-morpholinyl.-15-From the above discussion of substituents, one of skill in the art will understand that the term “alkyl” is meant to include groups such as haloalkyl (e.g., -CF3 and -CH2CF3) and acyl (e.g., - C(O)CH3, -C(O)CF3, -C(O)CH2OCH3, and the like). Preferably, the substituted alkyl and heteroalkyl groups have from 1 to 4 substituents, more preferably 1, 2 or 3 substituents. Exceptions are those perhalo alkyl groups (e.g., pentafluoroethyl and the like) which are also preferred and contemplated by the present disclosure.

[0060] Substituents for the alkyl and heteroalkyl radicals (including those groups often referred to as alkylene, alkenyl, heteroalkylene, heteroalkenyl, alkynyl, cycloalkyl, heterocycloalkyl, cycloalkenyl, and heterocycloalkenyl) can be one or more of a variety of groups selected from, but not limited to: -OR', =O, =NR', =N-OR', -NR'R”, -SR', -halogen, - SiR'R”R”', -OC(O)R', -C(O)R', -CO2R', -CONR'R'', -OC(O)NR'R”, -NR”C(O)R', -NR'- C(O)NR”R”', -NR”C(0)2R', -NR-C(NR'R”R”')=NR”'', -NR-C(NR'R”)=NR”', -S(O)R', - S(O)2R', -S(O)2NR'R”, -NRSO2R', -CN and -NO2in a number ranging from zero to (2m'+ 1), where m' is the total number of carbon atoms in such radical. R', R”, R'“ and R”“ each preferably independently refer to hydrogen, substituted or unsubstituted heteroalkyl, substituted or unsubstituted aryl, e.g., aryl substituted with 1-3 halogens, substituted or unsubstituted alkyl, alkoxy or thioalkoxy groups, or arylalkyl groups. When a compound of the present disclosure includes more than one R group, for example, each of the R groups is independently selected as are each R', R'', R”' and R”“ groups when more than one of these groups is present. When R' and R” are attached to the same nitrogen atom, they can be combined with the nitrogen atom to form a 5-, 6-, or 7-membered ring. For example, -NR'R” is meant to include, but not be limited to, 1-pyrrolidinyl and 4-morpholinyl. From the above discussion of substituents, one of skill in the art will understand that the term “alkyl” is meant to include groups including carbon atoms bound to groups other than hydrogen groups, such as haloalkyl (e.g., -CF3 and -CH2CF3) and acyl (e.g., -C(O)CH3, -C(O)CF3, -C(O)CH2OCH3, and the like).

[0061] “Alkoxy” refers to alkyl group having an oxygen atom that either connects the alkoxy group to the point of attachment or is linked to two carbons of the alkoxy group. Alkoxy groups include, for example, methoxy, ethoxy, propoxy, iso-propoxy, butoxy, 2- butoxy, iso-butoxy, sec-butoxy, tert-butoxy, pentoxy, hexoxy, etc. The alkoxy groups can be-16-further substituted with a variety of substituents described within. For example, the alkoxy groups can be substituted with halogens to form a “halo-alkoxy'' group.

[0062] “Carboxyalkyl” means an alkyl group (as defined herein) substituted with a carboxy group. The term “carboxycycloalkyl” means a cycloalkyl group (as defined herein) substituted with a carboxy group. The term alkoxyalkyl means an alkyl group (as defined herein) substituted with an alkoxy group. The term “carboxy'' employed herein refers to carboxylic acids and their esters.

[0063] “Haloalkyl” refers to alkyl as defined above where some or all of the hydrogen atoms are substituted with halogen atoms. Halogen (halo) preferably represents chloro or fluoro, but may also be bromo or iodo. For example, haloalkyl includes trifluoromethyl, fluoromethyl, 1,2,3,4,5-pentafluoro-phenyl, etc. The term “perfluoro” defines a compound or radical which has all available hydrogens that are replaced with fluorine. For example, perfluorophenyl refers to 1,2,3,4,5-pentafluorophenyl, perfluoromethyl refers to 1,1,1-trifluoromethyl, and perfluoromethoxy refers to 1,1,1-trifluoromethoxy.

[0064] “Fluoro-substituted alkyl” refers to an alkyl group where one, some, or all hydrogen atoms have been replaced by fluorine.

[0065] “Cytokine” in the context of this disclosure is a member of a group of protein signaling molecules that may participate in cell-cell communication in immune and inflammatory responses. Cytokines are typically small, water-soluble glycoproteins that have a mass of about 8-35 kDa.

[0066] “Cycloalkyl” refers to a cyclic hydrocarbon group that contains from about 3 to 12, from 3 to 10, or from 3 to 7 endocyclic carbon atoms. Cycloalkyl groups include fused, bridged and spiro ring structures.

[0067] “Endocyclic” refers to an atom or group of atoms which comprise part of a cyclic ring structure.

[0068] “Exocyclic” refers to an atom or group of atoms which are attached but do not define the cyclic ring structure.

[0069] “Cyclic alkyl ether” refers to a 4 or 5 member cyclic alkyl group having 3 or 4 endocyclic carbon atoms and 1 endocyclic oxygen or sulfur atom (e.g., oxetane, thietane, tetrahydrofuran, tetrahydrothiophene); or a 6 to 7 member cyclic alkyl group having 1 or 2-17-endocyclic oxygen or sulfur atoms (e.g., tetrahydropyran, 1,3-dioxane, 1,4-dioxane, tetrahydrothiopyran, 1,3-dithiane, 1,4-dithiane, 1,4-oxathiane).

[0070] “Alkenyl” refers to either a straight chain or branched hydrocarbon of 2 to 6 carbon atoms, having at least one double bond. Examples of alkenyl groups include, but are not limited to, vinyl, propenyl, isopropenyl, 1-butenyl, 2-butenyl, isobutenyl, butadienyl, 1- pentenyl, 2-pentenyl, isopentenyl, 1,3-pentadienyl, 1,4-pentadienyl, 1-hexenyl, 2-hexenyl, 3- hexenyl, 1,3-hexadienyl, 1,4-hexadienyl, 1,5-hexadienyl, 2,4-hexadienyl, or 1,3,5-hexatrienyl. Alkenyl groups can also have from 2 to 3, 2 to 4, 2 to 5, 3 to 4, 3 to 5, 3 to 6, 4 to 5, 4 to 6 and 5 to 6 carbons. The alkenyl group is typically monovalent, but can be divalent, such as when the alkenyl group links two moieties together.

[0071] “Alkenylene” refers to an alkenyl group, as defined above, linking at least two other groups, i.e., a divalent hydrocarbon radical. The two moieties linked to the alkenylene can be linked to the same atom or different atoms of the alkenylene. Alkenylene groups include, but are not limited to, ethenylene, propenylene, isopropenylene, butenylene, isobutenylene, sec-butenylene, pentenylene and hexenylene.

[0072] “Alkynyl” refers to either a straight chain or branched hydrocarbon of 2 to 6 carbon atoms, having at least one triple bond. Examples of alkynyl groups include, but are not limited to, acetylenyl, propynyl, 1-butynyl, 2-butynyl, isobutynyl, sec-butynyl, butadiynyl, 1-pentynyl, 2-pentynyl, isopentynyl, 1,3-pentadiynyl, 1,4-pentadiynyl, 1-hexynyl, 2-hexynyl, 3-hexynyl, 1,3-hexadiynyl, 1,4-hexadiynyl, 1,5-hexadiynyl, 2,4-hexadiynyl, or 1,3,5- hexatriynyl. Alkynyl groups can also have from 2 to 3, 2 to 4, 2 to 5, 3 to 4, 3 to 5, 3 to 6, 4 to 5, 4 to 6 and 5 to 6 carbons. The alkynyl group is typically monovalent, but can be divalent, such as when the alkynyl group links two moieties together.

[0073] “Alkynylene” refers to an alkynyl group, as defined above, linking at least two other groups, i.e., a divalent hydrocarbon radical. The two moieties linked to the alkynylene can be linked to the same atom or different atoms of the alkynylene. Alkynylene groups include, but are not limited to, ethynylene, propynylene, butynylene, sec-butynylene, pentynylene and hexynylene.

[0074] “Cycloalkyl” refers to a saturated or partially unsaturated, monocyclic, fused bicyclic or bridged polycyclic ring assembly containing from 3 to 12 ring atoms, or the number of atoms indicated. Monocyclic rings include, for example, cyclopropyl, cyclobutyl,-18-cyclopentyl, cyclohexyl, and cyclooctyl. Bicyclic and polycyclic rings include, for example, norbornane, decahydronaphthalene and adamantane. For example, C3-8cycloalkyl includes cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclooctyl, and norbomane.

[0075] “Cycloalkylene” refers to a cycloalkyl group, as defined above, linking at least two other groups, i.e., a divalent hydrocarbon radical. The two moieties linked to the cycloalkylene can be linked to the same atom or different atoms of the cycloalkylene. Cycloalkylene groups include, but are not limited to, cyclopropylene, cyclobutylene, cyclopentylene, cyclohexylene, and cyclooctylene.

[0076] “Heterocycloalkyl” refers to a ring system having from 3 ring members to about 20 ring members and from 1 to about 5 heteroatoms such as N, 0 and S. Additional heteroatoms can also be useful, including, but not limited to, B, Al, Si and P. The heteroatoms can also be oxidized, such as, but not limited to, -S(O)- and -S(O)2-. For example, heterocycle includes, but is not limited to, tetrahydrofuranyl, tetrahydrothiophenyl, morpholino, pyrrolidinyl, pyrrolinyl, imidazolidinyl, imidazolinyl, pyrazolidinyl, pyrazolinyl, piperazinyl, piperidinyl, indolinyl, quinuclidinyl and l,4-dioxa-8-aza-spiro[ 4.5]dec-8-yl.

[0077] “Heterocycloalkylene” refers to a heterocyclalkyl group, as defined above, linking at least two other groups. The two moieties linked to the heterocycloalkylene can be linked to the same atom or different atoms of the heterocycloalkylene.

[0078] “Aryl” refers to a monocyclic or fused bicyclic, tricyclic or greater, aromatic ring assembly containing 6 to 16 ring carbon atoms. For example, aryl may be phenyl, benzyl or naphthyl, preferably phenyl. “Arylene” means a divalent radical derived from an aryl group. Aryl groups can be mono-, di- or tri-substituted by one, two or three radicals selected from alkyl, alkoxy, aryl, hydroxy, halogen, cyano, amino, amino-alkyl, trifluoromethyl, alkylenedioxy and oxy-C2-C3-alkylene; all of which are optionally further substituted, for instance as hereinbefore defined; or 1- or 2-naphthyl; or 1- or 2-phenanthrenyl. Alkylenedioxy is a divalent substitute attached to two adjacent carbon atoms of phenyl, e.g. methylenedioxy or ethylenedioxy. Oxy-C2-C3-alkylene is also a divalent substituent attached to two adjacent carbon atoms of phenyl, e.g. oxyethylene or oxypropylene. An example for oxy- C2-C3- alkylene-phenyl is 2,3-dihydrobenzofuran-5-yl.-19-

[0079] Preferred as aryl is naphthyl, phenyl or phenyl mono- or disubstituted by alkoxy, phenyl, halogen, alkyl or trifluoromethyl, especially phenyl or phenyl-mono- or disubstituted by alkoxy, halogen or trifluoromethyl, and in particular phenyl.

[0080] Examples of substituted phenyl groups as R are, e.g.4-chlorophen-1-yl, 3,4- dichlorophen-1-yl, 4-methoxyphen-1-yl, 4-methylphen-1-yl, 4-aminomethylphen-1-yl, 4- methoxyethylaminomethylphen-1-yl, 4-hydroxyethylaminomethylphen-1-yl, 4-hydroxyethyl- (methyl)-aminomethylphen-1-yl, 3-arninomethylphen-1-yl, 4-N-acetylaminomethylphen-1- yl, 4-arninophen-1-yl, 3-arninophen-1-yl, 2-aminophen-1-yl, 4-phenyl-phen-1-yl, 4- (imidazol-1-yl)-phen-yl, 4-(imidazol-1-ylmethyl)-phen-1-yl, 4-(morpholin-1-yl)-phen-1-yl, 4- (morpholin-1-ylrnethyl)-phen-1-yl, 4-(2-methoxyethylaminomethyl)-phen-1-yl and 4- (pyrrolidin-1-ylmethyl)-phen-1-yl, 4-(thiophenyl)-phen-1-yl, 4-(3-thiophenyl)-phen-1-yl, 4- (4-methylpiperazin-l-yl)-phen-1-yl, and 4-(piperidinyl)-phenyl and 4-(pyridinyl)-phenyl optionally substituted in the heterocyclic ring.

[0081] Similarly, substituents for the aryl and heteroaryl groups are varied and are selected from: -halogen, -OR', -OC(O)R', -NR'R”, -SR', -R', -CN, -NO2, -CO2R', -CONR'R”, -C(O)R', -OC(O)NR'R'', -NR”C(O)R', -NR”C(O)2R', ,-NR'-C(O)NR”R'“, -NH-C(NH2)=NH, - NR'C(NH2)=NH, -NH-C(NH2)=NR', -S(O)R', -S(O)2R', -S(O)2NR'R”, -N3, -CH(Ph)2, perfluoro(C1-C4)alkoxy, and perfluoro(C1-C4)alkyl, in a number ranging from zero to the total number of open valences on the aromatic ring system; and where R', R” and R”' are independently selected from hydrogen, (C1-C4)alkyl and heteroalkyl, unsubstituted aryl and heteroaryl, (unsubstituted aryl)-(C1-C4)alkyl, and (unsubstituted aryl)oxy-(C1-C4)alkyl.

[0082] Two of the substituents on adjacent atoms of the aryl or heteroaryl ring may optionally be replaced with a substituent of the formula -T-C(O)-(CH2)q-U-, wherein T and U are independently -NH-, -O-, -CHz- or a single bond, and q is an integer of from 0 to 2. Alternatively, two of the substituents on adjacent atoms of the aryl or heteroaryl ring may optionally be replaced with a substituent of the formula -A-(CH2)r-B-, wherein A and B are independently -CH2-, -O-, -NH-, -S-, -S(O)-, -S(O)-, -S(O)2NR'- or a single bond, and r is an integer of from 1 to 3. One of the single bonds of the new ring so formed may optionally be replaced with a double bond. Alternatively, two of the substituents on adjacent atoms of the aryl or heteroaryl ring may optionally be replaced with a substituent of the formula -(CH2)s-X- (CH2)t-, where s and t are independently integers of from 0 to 3, and X is -O-, -NR'-, -S-, --20-S(O)-, -S(O)2-, or -S(0)2NR'-. The substituent R' in -NR'- and -S(O)2NR'- is selected from hydrogen or unsubstituted (C1-C6)alkyl.

[0083] “Heteroaryl” refers to a monocyclic or fused bicyclic or tricyclic aromatic ring assembly containing 5 to 16 ring atoms, where from 1 to 4 of the ring atoms are a heteroatom each N, O or S. For example, heteroaryl includes pyridyl, indolyl, indazolyl, quinoxalinyl, quinolinyl, isoquinolinyl, benzothienyl, benzofuranyl, furanyl, pyrrolyl, thiazolyl, benzothiazolyl, oxazolyl, isoxazolyl, triazolyl, tetrazolyl, pyrazolyl, imidazolyl, thienyl, or any other radicals substituted, especially mono- or di-substituted, by e.g. alkyl, nitro or halogen. Pyridyl represents 2-, 3- or 4-pyridyl, advantageously 2- or 3-pyridyl. Thienyl represents 2- or 3-thienyl. Quinolinyl represents preferably 2-, 3- or 4-quinolinyl. Isoquinolinyl represents preferably 1-, 3- or 4-isoquinolinyl. Benzopyranyl, benzothiopyranyl represents preferably 3-benzopyranyl or 3-benzothiopyranyl, respectively. Thiazolyl represents preferably 2- or 4-thiazolyl, and most preferred, 4-thiazolyl. Triazolyl is preferably 1-, 2- or 5-(1,2,4-triazolyl). Tetrazolyl is preferably 5-tetrazolyl.

[0084] Preferably, heteroaryl is pyridyl, indolyl, quinolinyl, pyrrolyl, thiazolyl, isoxazolyl, triazolyl, tetrazolyl, pyrazolyl, imidazolyl, thienyl, furanyl, benzothiazolyl, benzofuranyl, isoquinolinyl, benzothienyl, oxazolyl, indazolyl, or any of the radicals substituted, especially mono- or di-substituted.

[0085] As used herein, the term “heteroalkyl” refers to an alkyl group having from 1 to 3 heteroatoms such as N, O and S. Additional heteroatoms can also be useful, including, but not limited to, B, Al, Si and P. The heteroatoms can also be oxidized, such as, but not limited to, -S(O)- and -S(O)2-. For example, heteroalkyl can include ethers, thioethers, alkyl- amines and alkyl-thiols.

[0086] As used herein, the term “heteroalkylene” refers to a heteroalkyl group, as defined above, linking at least two other groups. The two moieties linked to the heteroalkylene can be linked to the same atom or different atoms of the heteroalkylene.

[0087] “Electrophile” refers to an ion or atom or collection of atoms, which may be ionic, having an electrophilic center, i.e., a center that is electron seeking, capable of reacting with a nucleophile. An electrophile (or electrophilic reagent) is a reagent that forms a bond to its reaction partner (the nucleophile) by accepting both bonding electrons from that reaction partner.-21-

[0088] “Nucleophile” refers to an ion or atom or collection of atoms, which may be ionic, having a nucleophilic center, i.e., a center that is seeking an electrophilic center or capable of reacting with an electrophile. A nucleophile (or nucleophilic reagent) is a reagent that forms a bond to its reaction partner (the electrophile) by donating both bonding electrons. A “nucleophilic group” refers to a nucleophile after it has reacted with a reactive group. Non limiting examples include amino, hydroxyl, alkoxy, haloalkoxy and the like.

[0089] “Maleimido” refers to a pyrrole-2,5-dione-1-yl group having the structure:

[0090] which upon reaction with a sulfhydryl (e.g., a thio alkyl) forms an -S- maleimido group having the structure

[0091] where “•” indicates the point of attachment formaleimido group and “ “indicates the point of attachment of the sulfur atom the thiol to the remainder of the original sulfhydryl bearing group.

[0092] For the purpose of this disclosure, “naturally occurring amino acids” found in proteins and polypeptides are L-alanine, L-arginine, L-asparagine, L-aspartic acid, L- cysteine, L-glutamine, L-glutamic acid, L-glycine, L-histidine, L-isoleucine, L-leucine, L- lysine, L-methionine, L-phenylalanine, L-proline, L-serine, L-threonine, L-tryptophan, tyrosine, and or L-valine. ''Non-naturally occurring amino acids” found in proteins are any amino acid other than those recited as naturally occurring amino acids. Non-naturally occurring amino acids include, without limitation, the D isomers of the naturally occurring amino acids, and mixtures of D and L isomers of the naturally occurring amino acids. Other amino acids, such as 4-hydroxyproline, desmosine, isodesmosine, 5-hydroxylysine, epsilon- N-methyllysine, 3-methylhistidine, although found in naturally occurring proteins, are considered to be non-naturally occurring amino acids found in proteins for the purpose of this disclosure as they are generally introduced by means other than ribosomal translation of mRNA.-22-

[0093] “Linear” in reference to the geometry, architecture or overall structure of a polymer, refers to polymer having a single monomer derived backbone.

[0094] “Branched,” in reference to the geometry, architecture or overall structure of a polymer, refers to polymer having 2 or more polymer “arms” extending from a single group, such as an L group that may be derived from an initiator employed in an atom transfer radical polymerization reaction. A branched polymer may possess 2 polymer arms, 3 polymer arms, 4 polymer arms, 5 polymer arms, 6 polymer arms, 7 polymer arms, 8 polymer arms or more. For the purpose of this disclosure, compounds having three or more polymer arms extending from a single linear group are denoted as having a “comb” structure or “comb” architecture.

[0095] Branched can also be achieved through “statistical” structures to create broader dendrimer-like architectures.

[0096] “Pharmaceutically acceptable” composition or “pharmaceutical composition” refers to a composition comprising a compound of the present disclosure and a pharmaceutically acceptable excipient or pharmaceutically acceptable excipients.

[0097] “Pharmaceutically acceptable excipient” and “pharmaceutically acceptable carrier” refer to an excipient that can be included in the compositions of the present disclosure and that causes no significant adverse toxicological effect on the patient. Non-limiting examples of pharmaceutically acceptable excipients include water, NaCl, normal saline solutions, lactated Ringer's, normal sucrose, normal glucose and the like.

[0098] “Patient” or “subject in need thereof ' refers to a living organism suffering from or prone to a condition that can be prevented or treated by administration of a pharmaceutical composition as provided herein. Non-limiting examples include humans, other mammals and other non-mammalian animals.

[0099] “Therapeutically effective amount” refers to an amount of a conjugated functional agent or of a pharmaceutical composition useful for treating, ameliorating, or preventing an identified disease or condition, or for exhibiting a detectable therapeutic or inhibitory effect. The effect can be detected by any assay method known in the art.

[0100] The “biological half-life”of a substance is a pharmacokinetic parameter which specifies the time required for one half of the substance to be removed from an organism following introduction of the substance into the organism.-23-Copolymer

[0101] The present disclosure provides a copolymer with a first repeating units having the structure of Formula (I):a second repeating units having the structure of Formula (II):, where m > 0, n > 0, and m + n ≤ 8000, X1 is H or methyl; X2 is H, F, or methyl; Y is -O- or - NH-; R is optionally substituted C1-12 alkylene, optionally substituted; q is 1, 2, 3, or 4; and Z1 comprises N3 or a cyclic olefin.

[0102] In some embodiments of the copolymer, R is C1-12 alkylene, or C1-12 alkylene substituted by F, OH, or C1-6 alkyl.

[0103] In some embodiments of the copolymer, R is selected from the groupconsisting of C3 alkylene, and, each optionally substituted byF, OH, or C1-6 alkyl.

[0104] In some embodiments of the copolymer, R is C1 alkylene oroptionally substituted by F, OH or methyl. In some embodiments, R is unsubstituted C1-3 alkylene, for example methylene, ethylene, or propylene. In some embodiments, R is substituted C1-3 alkylene. In some embodiments, R is substituted C3 alkylene; in some embodiments, the C3 alkylene is substituted with a hydroxy group. In some-24-embodiments, R is optionally substitutedin some embodiments, q is 3. In some embodiments, R is absent.

[0105] In some embodiments of the copolymer, Y is O.

[0106] In some embodiments of the copolymer, Z1 is N3,,.

[0107] In some embodiments of the copolymer, Z1 is N3.

[0108] In some embodiments of the copolymer, n / (m+n) ≤ 0.4.

[0109] In some embodiments, the copolymer further comprises a core connecting to one or more polymer arms, each of the one or more polymer arms comprises the first repeating units and the second repeating units.

[0110] In some embodiments of the copolymer, the core is, consisting of –N3, –NH3+, and ; R2 is selected from the group consisting of alkylene, oxy, amido, and combination thereof;, , rR4, R5, and R6 are the same or different and are selected from the group consisting-25-connection to the polymer arm.

[0111] In some embodiments of the copolymer, the copolymer has the structure of:integer selected from 1-9.

[0112] In some embodiments of the copolymer, R2 is, and t is an integer selected from 1 to 8.

[0113] In some embodiments of the copolymer, R2 is, v is an integer selected from 1 to 6, and w is an integer selectedfrom 1 to 6.

[0114] In some embodiments of the copolymer, the copolymer is a star copolymer having 2 to 9 polymer arms where each polymer arm comprises the first repeating units and the second repeating units.

[0115] In some embodiments of the copolymer, the copolymer has a dispersity less than 2. In some embodiments of the copolymer, the copolymer has a dispersity of about 1.07 to about 1.9.

[0116] In some embodiments of the copolymer, the copolymer has a molecular weight of about 19 kDa to about 1,800 kDa.-26-

[0117] In some embodiments of the copolymer, the copolymer is a random copolymer.

[0118] In some embodiments of the copolymer, the copolymer is a gradient copolymer.

[0119] In some embodiments of the copolymer, the copolymer is a block copolymer.

[0120] In some embodiments of the copolymer, the copolymer has a refractive index increment of about 1.26 to about 1.47.

[0121] In some embodiments of the copolymer, the copolymer has an azide loading of about 0.5% to about 30%. In some embodiments of the copolymer, the copolymer has an azide loading of about 0.5% to about 14%.

[0122] In some embodiments of the copolymer, the copolymer is selected from the-27--28-. Modified Payload

[0123] The present disclosure provides a compound having the structure:,, , membered monocyclic or bicyclic cycloalkyne, 7- to 9- membered monocyclic or bicyclic heterocycloalkyne, 10- to 16-membered tricyclic heterocycloalkyne, or optionally substituted-29-is 1 or 2, and u is 1, 2, or 3, and w is an integer between 1 and 15; X3is H, –(CH2)b– phosphorylcholine, or C1-3 alkyl optionally substituted by COOH, OH, or C(O)NH2; X4is C1- 3 alkyl; X5 is C1-3 alkyl substituted by OH, COOH, C(O)NH2, or –(CH2)b–phosphorylcholine; b is 0, 1, 2, 3, or 4; L is absent or selected from the group consisting of,is 2, 3, or 4; r is 1, 2, 3, or 4; X6is optionally substituted C1-3 alkyl, optionally substituted phenyl, or optionally substituted pyridinyl; X7and X8are each H or methyl; and PL comprises a bioactive agent, tracer, or reactive functional group. In some embodiments, the reactive functional group may comprise maleimide, iodoacetamide, and / or bromoacetamide.

[0124] In some embodiments, the heterocycloalkyne, including monocyclic or bicyclic cycloalkyne, monocyclic or bicyclic heterocycloalkyne, and tricyclic heterocycloalkyne, may contain at least one O and / or at least one N. In some embodiments, is a 10- to 16-membered tricyclic heterocycloalkyne containing at least one O and / or at least one N.

[0125] In some embodiments of the compound, Z2 is selected from a group-30-,C1-3 alkyl, or optionally substituted phenyl. In some embodiments, Z2issome embodiments, Z2 is an Oxo, Aza-CycloNonyne (OACN), defined as a group containing the structure:. In some embodiments of the compound, the bioactive agent comprises a small molecule drug, molecular tracer, toxin, peptide, macrocycles, oligonucleotide, or aptamer.

[0126] In some embodiments of the compound, the bioactive agent is selected from the group consisting of NLRP3 inhibitors, Prostaglandins, Rho-kinase inhibitors, beta- adrenergic antagonists, carbonic anhydrase inhibitor, C3 inhibitor, C5 inhibitors, puromycin, and tirzepatide.

[0127] In some embodiments of the compound, the bioactive agent is selected from the group consisting of,-31-.

[0128] In some embodiments of the compound, L allows for the controlled or stimuli-responsive release of the bioactive agent.

[0129] In some embodiments of the compound, L is a stable linkage and allows for minimal release of the bioactive agent.

[0130] In some embodiments of the compound, L remains attached to R’ when the bioactive agent is released.

[0131] In some embodiments of the compound, R’ is absent or selected from the , ,-32-

[0132] In some embodiments, L is absent or selected from the group consisting of , , , ,,. Conjugates

[0133] The present disclosure provides a conjugate with a first monomer repeating units having the structure of Formula (I):-33-a second monomer repeating units having the structure of Formula (IIa):, where m + n ≤ 8000; X1is H or methyl; X2is H, F, or methyl; Y is -O- or -NH-; R is optionally substituted C1-12 alkylene, optionally substituted, q is an integer between 1and 4; and Z compriseswhere Z’ is a reaction product of (1) N3 andcycloalkyne derivative or heterocycloalkyne derivative or (2) olefin and optionally substituted, , , , a is 1 or 2, and u is 1, 2, or 3, and w is an integer between 1 and 15; X3is H, –(CH2)b– phosphorylcholine, or C1-3 alkyl optionally substituted by COOH, OH, or C(O)NH2; X4 is C1- 3 alkyl; X5 is C1-3 alkyl substituted by OH, COOH, C(O)NH2, or –(CH2)b–phosphorylcholine;-34-b is 0, 1, 2, 3, or 4; L is absent or selected from the group consisting of,optionally substituted C1-3 alkyl, optionally substituted phenyl, or optionally substituted pyridinyl; X7and X8are each H or methyl; and PL comprises a bioactive agent, tracer, or reactive functional group. In some embodiments, R is unsubstituted C1-3 alkylene, for example methylene, ethylene, or propylene. In some embodiments, R is substituted C1-3 alkylene. In some embodiments, R is substituted C3 alkylene; in some embodiments, the C3 alkylene is substituted with a hydroxy group. In some embodiments, R is optionally substituted; in some embodiments, q is 3. In some embodiments, R is absent. In some embodiments, b is 2. In some embodiments, the reactive functional group may comprise maleimide, iodoacetamide, and / or bromoacetamide.

[0134] In some embodiments of the conjugate, the bioactive agent comprises a small molecule drug, molecular tracer, toxin, peptide, macrocycles, oligonucleotide, or aptamer.

[0135] The present disclosure provides a method of making the conjugate by reacting the copolymer with the compound, where Z1of copolymer reacts with Z2of the compound via click reaction in the absence of a metal.

[0136] In some embodiments, the conjugate may further comprise monomer units that were unreacted.

[0137] The present disclosure provides a method of treating a disease or condition by administering to a subject in need thereof the conjugate.

[0138] In some embodiments of the method of treating a disease or condition, the disease or condition comprises dry age-related macular degeneration (AMD), wet AMD,-35-retinal vein occlusion (RVO), glaucoma, dry eyes, Duane-Radial Ray Syndrome (DR), neuroinflammation, neuroprotection, obesity, Parkinson's, Alzheimer's, cardiac, metabolic disease, diabetic macular edema (DME), cancer, non-infectious uveitis (NIU), inherited retinal disorder (IRD), hypercholesterolemia, inflammatory diseases, cystic fibrosis, b-thalassaemia, mucopolysacchariodosis type I, or hemophilia A.

[0139] In some embodiments, the conjugation process comprises coupling an antisense oligonucleotide (ASO) or small interfering RNA (siRNA), which is modified with a BCN-terminated protease-labile linker, to a copolymer by strain-promoted azide–alkyne cycloaddition (SPAAC) click chemistry, thereby producing a copolymer–oligonucleotide conjugate.

[0140] In some embodiments, the conjugation process comprises coupling an aptamer that is functionalized with a DBCO moiety to a copolymer by strain-promoted azide– alkyne cycloaddition (SPAAC) click chemistry, thereby producing an aptamer-copolymer conjugate.

[0141] In some embodiments, the modified payload / OACN is coupled to a cysteine residue of an antibody through maleimide–thiol conjugation, thereby forming an OACN– antibody intermediate. The resulting intermediate is subsequently conjugated to the copolymer by strain-promoted azide–alkyne cycloaddition (SPAAC) click chemistry to produce an antibody–copolymer conjugate.

[0142] In some embodiments, the modified payload / OACN is first conjugated to the copolymer via SPAAC click chemistry to yield a maleimide-functionalized copolymer intermediate. The maleimide-functionalized copolymer intermediate is then conjugated to a cysteine residue of an antibody via maleimide-thiol chemistry to generate an antibody- copolymer conjugate.

[0143] In some embodiments, the copolymer is first conjugated with a therapeutic agent via SPAAC click chemistry to yield a copolymer-drug intermediate. The modified payload / OACN is then conjugated to a cysteine residue of an antibody via maleimide-thiol chemistry to produce an OACN-antibody intermediate. The OACN-antibody intermediate is subsequently conjugated to the copolymer-drug intermediate via SPAAC click chemistry, thereby forming an antibody-copolymer-drug conjugate.-36-

[0144] In some embodiments, the copolymer is first conjugated with a therapeutic agent via SPAAC click chemistry to yield a copolymer-drug intermediate. The modified payload / OACN is then conjugated to the copolymer-drug intermediate via SPAAC click chemistry to produce a maleimide-functionalized copolymer-drug intermediate. The maleimide-functionalized copolymer-drug intermediate is subsequently conjugated to a cysteine residue of an antibody via maleimide-thiol chemistry to yield an antibody-copolymer- drug conjugate. Method of Preparation

[0145] The compounds disclosed herein may be synthesized by methods described below, or by modification of these methods. Ways of modifying the methodology include, among others, temperature, solvent, reagents etc., known to those skilled in the art and are part of routine reaction modification and optimization. In general, during any of the processes for preparation of the compounds disclosed herein, it may be necessary and / or desirable to protect sensitive or reactive groups on any of the molecules concerned. This may be achieved by means of conventional protecting groups, such as those described in Protective Groups in Organic Chemistry (ed. J.F.W. McOmie, Plenum Press, 1973); and P.G.M. Green, T.W. Wutts, Protecting Groups in Organic Synthesis (3rd ed.) Wiley, New York (1999), which are both hereby incorporated herein by reference in their entirety. The protecting groups may be removed at a convenient subsequent stage using methods known from the art. Synthetic chemistry transformations useful in synthesizing applicable compounds are known in the art and include e.g. those described in R. Larock, Comprehensive Organic Transformations, VCH Publishers, 1989, or L. Paquette, ed., Encyclopedia of Reagents for Organic Synthesis, John Wiley and Sons, 1995, which are both hereby incorporated herein by reference in their entirety. The routes shown and described herein are illustrative only and are not intended, nor are they to be construed, to limit the scope of the claims in any manner whatsoever. Those skilled in the art will be able to recognize modifications of the disclosed syntheses and to devise alternate routes based on the disclosures herein; all such modifications and alternate routes are within the scope of the claims.

[0146] In the following schemes, protecting groups for oxygen atoms are selected for their compatibility with the requisite synthetic steps as well as compatibility of the-37-introduction and deprotection steps with the overall synthetic schemes (P.G.M. Green, T.W. Wutts, Protecting Groups in Organic Synthesis (3rd ed.) Wiley, New York (1999)).

[0147] If the compounds of the present technology contain one or more stereogenic centers, such compounds can be prepared or isolated as pure stereoisomers, i.e., as individual enantiomers or d(l) stereoisomers, or as stereoisomer-enriched mixtures. All such stereoisomers (and enriched mixtures) are included within the scope of the present technology, unless otherwise indicated. Pure stereoisomers (or enriched mixtures) may be prepared using, for example, optically active starting materials or stereoselective reagents or catalysts well- known in the art. Alternatively, racemic mixtures of such compounds can be separated using, for example, chiral column chromatography, chiral resolving agents and the like.

[0148] The starting materials for the following reactions are generally known or commercial compounds or can be prepared by known procedures or obvious modifications thereof. For example, many of the starting materials are available from commercial suppliers such as Aldrich Chemical Co. (Milwaukee, Wisconsin, USA), Bachem (Torrance, California, USA), Emka-Chemce or Sigma (St. Louis, Missouri, USA). Others may be prepared by procedures, or obvious modifications thereof, described in standard reference texts such as Fieser and Fieser's Reagents for Organic Synthesis, Volumes 1-15 (John Wiley, and Sons, 1991), Rodd's Chemistry of Carbon Compounds, Volumes 1-5, and Supplementals (Elsevier Science Publishers, 1989), Organic Reactions, Volumes 1-40 (John Wiley, and Sons, 1991), March's Advanced Organic Chemistry, (John Wiley, and Sons, 5th Edition, 2001), and Larock's Comprehensive Organic Transformations (VCH Publishers Inc., 1989).

[0149] The methods disclosed herein may include using standard organic synthesis techniques to construct copolymers, compounds, and conjugates of the present disclosure. Copolymers of the present disclosure can be prepared by any suitable polymerization method. Thepolymerization can be a reversible-deactivation radical polymerization / controlled reversible-deactivation radical polymerization, such as Atom Transfer Radical Polymerization (ATRP), Reversible Addition-Fragmentation-Termination (RAFT) polymerization, or Nitroxide_Mediated Polymerization (NMP). The polymerization methods described herein are well-known in the art and a person of ordinary skill in the art would readily understand how to carry out such polymerizations, including the selection of suitable reagents, catalysts, initiators, solvents, reaction conditions, and techniques, without undue experimentation.-38-

[0150] The methods disclosed herein may include using standard organic synthesis techniques to construct conjugates of the present disclosure as shown in General Scheme 1 below. General Scheme 1 shows a copolymer containing reactive group Z1, where reactive group Z1 reacts with reactive group Z2 of a modified payload via click chemistry, to form a conjugate containing the click reaction product Z’. Click chemistry reactions are well-known in the art and a person of ordinary skill in the art would readily understand how to carry out such reactions, including the selection of suitable reagents, catalysts, solvents, reaction conditions, and techniques, without undue experimentation. General Scheme 1:Methods of Use

[0151] Disclosed herein are methods of using the conjugates of the present disclosure to deliver payloads as shown in General Scheme 2. General Scheme 2:L’ may be the same as L or different from L depending on the linkage.-39-EXAMPLES

[0152] To further illustrate the embodiments disclosed herein, the following examples are included. The examples should not, of course, be construed as specifically limiting the present disclosure. Variations of these examples within the scope of the claims are within the purview of one skilled in the art and are considered to fall within the scope of the present disclosure as described and claimed herein. The reader will recognize that the skilled artisan, armed with the present disclosure, and skill in the art is able to prepare and use the present disclosure without exhaustive examples. The following examples will further describe the present disclosure, and are used for the purposes of illustration only, and should not be considered as limiting.

[0153] Compounds may be identified by their chemical structure and / or their chemical name. Chemical names were generated using the PerkinElmer (Waltham, MA 02451, USA) ChemDraw®Professional 25.0.2.14 and MestReNova®v15.0.0 (MNova IUPAC Name) nomenclature program. When the chemical structure and chemical name conflict, the chemical structure is determinative of the identity of the compound. General Procedures

[0154] It will be apparent to the skilled artisan that methods for preparing precursors and functionality related to the compounds claimed herein are generally described in the literature. In these reactions, it is also possible to make use of variants which are themselves known to those of ordinary skill in this art but are not mentioned in greater detail. The skilled artisan given the literature and this disclosure is well equipped to prepare any of the compounds.

[0155] It is recognized that the skilled artisan in the art of organic chemistry can readily carry out manipulations without further direction, that is, it is well within the scope and practice of the skilled artisan to carry out these manipulations. These include reduction of carbonyl compounds to their corresponding alcohols, oxidations, acylations, aromatic substitutions, both electrophilic and nucleophilic, etherifications, esterification and saponification and the like. These manipulations are discussed in standard texts such as March Advanced Organic Chemistry (Wiley), Carey and Sundberg, Advanced Organic Chemistry (incorporated herein by reference in their entirety) and the like. All the intermediate-40-compounds of the present disclosure were used without further purification unless otherwise specified.

[0156] The skilled artisan will readily appreciate that certain reactions are best carried out when other functionality is masked or protected in the molecule, thus avoiding any undesirable side reactions and / or increasing the yield of the reaction. Often the skilled artisan utilizes protecting groups to accomplish such increased yields or to avoid the undesired reactions. These reactions are found in the literature and are also well within the scope of the skilled artisan. Examples of many of these manipulations can be found for example in T. Greene and P. Wuts Protecting Groups in Organic Synthesis, 4th Ed., John Wiley & Sons (2007), incorporated herein by reference in its entirety.

[0157] The following example schemes are provided for the guidance of the reader and represent preferred methods for making the compounds exemplified herein. These methods are not limiting, and it will be apparent that other routes may be employed to prepare these compounds. Such methods specifically include solid phase based chemistry, including combinatorial chemistry. The skilled artisan is thoroughly equipped to prepare these compounds by those methods given the literature and this disclosure. The compound numberings used in the synthetic schemes depicted below are meant for those specific schemes only and should not be construed as or confused with same numberings in other sections of the application.

[0158] Trademarks used herein are examples only and reflect illustrative materials used at the time of the present disclosure. The skilled artisan will recognize that variations in lot, manufacturing processes, and the like, are expected. Hence the examples, and the trademarks used in them are non-limiting, and they are not intended to be limiting, but are merely an illustration of how a skilled artisan may choose to perform one or more of the embodiments of the present disclosure.

[0159] The following abbreviations have the indicated meanings:

[0160] 2N = a 2 normal solution of a species^ °C = degree Celsius^

[0161] ACN = acetonitrile^

[0162] AcOH = acetic acid^

[0163] AIBN = azobisisobutyronitrile^-41-

[0164] aq. = aqueous^

[0165] BINAP = (2,2’-bis(diphenylphosphino)-1,1’-binaphthyl)^

[0166] Bn = benzyl^

[0167] Boc = tert-butoxycarbonyl^

[0168] brine = saturated aqueous solution of sodium chloride (NaCl)^^

[0169] Bu = butyl^

[0170] CD3OD = deuterated methanol^

[0171] CHCl3 = chloroform^

[0172] CDCl3= deuterochloroform^

[0173] CH2Cl2= methylene chloride, or dichloromethane or DCM^

[0174] Cs2CO3 = cesium carbonate^

[0175] DCC = dicyclohexylcarbodiimide^

[0176] DCM = dichloromethane^

[0177] DIEA = N,N-diisopropylethylamine^

[0178] DMAP = 4-dimethylaminopyridine^

[0179] DMF = dimethylformamide^

[0180] DMSO = dimethylsulfoxide^

[0181] dppf = 1,1’-bis(dipenylphosphino)ferrocene^^

[0182] EDCI = 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC, EDC·HCl, EDAC)^

[0183] Et = ethyl^

[0184] Et2O = diethyl ether^

[0185] EtOH = ethanol^

[0186] EA = ethyl acetate^

[0187] g = gram^

[0188] FA = formic acid^

[0189] h = hour^

[0190] 1H+= proton^

[0191] H = hydrogen^

[0192] H2O = water^

[0193] HATU = hexafluorophosphate azabenzotriazole tetramethyl uronium^- -

[0194] HCl = hydrogen chloride^

[0195] Hex = hexanes^

[0196] HOBt = hydroxybenzotriazole^

[0197] HPLC = high-performance liquid chromatography^

[0198] iPrOH = isopropyl alcohol^

[0199] LC / MS = liquid chromatographic mass spectroscopy^

[0200] Me = methyl^

[0201] M = molarity^

[0202] m = multiplet^

[0203] [M+H]+= molecular ion plus one proton^

[0204] mCPBA = 3-chloroperbenzoic acid^

[0205] MeOH = methanol^

[0206] min = minute^

[0207] mL = milliliter^

[0208] mmol = millimole^

[0209] MHz = megahertz^^

[0210] m / z = mass to charge ratio^

[0211] N2 = nitrogen^

[0212] NaCl = sodium chloride^

[0213] Na2SO4= sodium sulfate^

[0214] NH3 = ammonia^^

[0215] NBS = N-bromo succinimide^

[0216] NH4HCO3= ammonium hydrogen carbonate^

[0217] NMR = nuclear magnetic resonance^

[0218] NMP = N-methyl-2-pyrrolidone or 1-methyl-2-pyrrolidone^

[0219] o.n. = overnight^

[0220] PBr3 = phosphorous tribromide^^

[0221] PE = petroleum ether^

[0222] PCl5= phosphorous pentachloride^

[0223] PCC = pyridinium chlorochromate^

[0224] Pd(PPh3)2Cl2 = bis(triphenylphosphine)palladium(II) dichloride^- -

[0225] Pd(PPh3)4 = tetrakis(triphenylphosphine)palladium(0)^

[0226] Pd / C = palladium on carbon^

[0227] PEG = polyethylene glycol^

[0228] Ph = phenyl^

[0229] pH = negative logarithm of hydrogen ion concentration^

[0230] ppm = parts per million^

[0231] q = quartet^

[0232] RBF = round bottom flask^

[0233] rt = room temperature^

[0234] RP = reverse phase^

[0235] s = singlet^

[0236] sat. = saturated^

[0237] t = triplet^

[0238] tR = retention time^

[0239] SFC = supercritical fluid chromatography^

[0240] TBSCl = tert-butyldimethylsilyl chloride^

[0241] tBu = tert-butyl^

[0242] TEA = triethylamine^

[0243] TFA = trifluoroacetic acid^

[0244] THF = tetrahydrofuran^

[0245] TLC = thin layer chromatography^

[0246] TMA = trimethlyamine^

[0247] TMS = trimethylsilyl^

[0248] wt% = weight percentage^ Examples

[0249] The following example schemes are provided for the guidance of the reader, and collectively represent an example method for making the compounds provided herein. Furthermore, other methods for preparing compounds described herein will be readily apparent to the person of ordinary skill in the art in light of the following reaction schemes and examples. Unless otherwise indicated, all variables are as defined above. Monomer Example 1: Synthesis of 3-azido propyl methacrylate-44-

[0250] 4-Dimethylaminopyridine (2.4 g, 20 mmol, 0.4 eq) was dissolved in dichloromethane (DCM, 40 mL) in a 250 mL dry round bottom flask. Next, methacrylic acid (3.9 mL, 46 mmol, 1.0 eq) and 3-azidopropanol (4.7 mL, 51 mmol, 1.1 eq) were added to the flask. The solution was then cooled in an ice water bath, and 1-ethyl-3-(3- dimethylaminopropyl)carbodiimide (12 g, 60 mmol, 1.3 eq) was slowly added over half an hour. The reaction mixture was allowed to warm up to room temperature and stirred overnight. Reaction completion was verified by1HNMR to confirm that all the methacrylic acid had been consumed. After 20 hours, the mixture was placed in an ice water bath and quenched with 33 vol.% 0.5 M hydrochloric acid (HCl) in saturated sodium chloride (NaCl, 40 mL). The product was extracted into DCM (3 x 30 mL). The organic extracts were combined and washed with saturated sodium bicarbonate (40 mL). Subsequently, the DCM layer was washed with 33 vol.% 0.5 M HCl in saturated NaCl (30 mL) and then dried over sodium sulfate. The solution was concentrated under reduced pressure by rotary evaporation. It was then further purified by column chromatography (0-20% ethyl acetate in hexanes) to afford 3-azido propyl methacrylate as a colorless liquid (4.9 g, 62%).1H NMR (600 MHz, CDCl3) δ 6.11 (m, 1H), 5.58 (m, 1H), 4.25 (t, J = 6.2 Hz, 2H), 3.42 (t, J = 6.7 Hz, 2H), 2.03 – 1.88 (m, 5H) ppm. Monomer Example 2: Synthesis of 3-Azidopropyl 2-bromo-2-methylpropanoate

[0251] Added N,N-diisopropylethylamine (3.4 mL, 20 mmol, 2.5 eq.) and 2- bromoisobutyryl bromide (1.2 mL, 9.5 mmol, 1.2 eq.) to anhydrous tetrahydrofuran (29 mL). Cooled the solution in an ice water bath and then slowly added 3-azido-1-propanol (0.80 g, 7.9 mmol, 1 eq.) to the reaction mixture. Allowed the mixture to warm up to room temperature and let it stir overnight. Added cold water to the reaction mixture and then extracted the product into DCM (3 x 80 mL). The DCM layer was washed with saturated sodium bicarbonate (3 x 15 mL) and then dried over sodium sulfate. The solution was concentrated under reduced pressure by rotary evaporation and purified by column chromatography (0-10% ethyl acetate in hexanes) to afford 3-azidopropyl 2-bromo-2-methylpropanoate as a slightly yellow liquid (0.8 g, 40%).1H NMR (600 MHz, CDCl3) δ 4.27 (t, J = 6.1 Hz, 2H), 3.45 (t, J = 6.7 Hz, 2H), 2.00 – 1.95 (m, 2H), 1.94 (s, 6H) ppm. Copolymer Example 1: Synthesis of 9-arm 10 mol% azide loading copolymer

[0252] Synthetic Scheme:-45-

[0253] 2-Methacryloyloxyethyl phosphorylcholine (5.00 g, 16.9 mmol, 3186 eq.) were added to a dry round bottom flask equipped with a 3-way stopcock in a glovebox, followed by addition (under dry N2) of degassed anhydrous ethanol (19.0 mL), glycidyl methacrylate (250 µL, 1.90 mmol, 354 eq.), CuBr2(89.5 mM in ethanol, 0.1 mL, 0.17 eq.), tris(2-pyridylmethyl)amine (TPMA) and ascorbic acid (21.4 mM and 21.6 mM respectively in ethanol, 0.5 mL, 2 eq. for both). The reaction was started with the addition of the initiator (2.71 mM in ethanol, 1.95 mL, 1 eq.) at r.t. and samples were taken periodically under N2to monitor the monomer conversion via1H NMR spectroscopic analysis. The polymerization was terminated after 2.5 h (monomer conversion = 0.67) by exposing to air. The crude product solution was diluted with ethanol (50 mL), passed through a silica column (Biotage® Sfär Silica HC D, 50 g, ethanol as eluent), concentrated and precipitated into ethyl acetate (400 mL) twice. The polymer precipitate was then redissolved in DI water (50 mL), and to the polymer solution (~ 10 wt%), sodium azide (0.39 g, 6.0 mmol, 1118 eq.) and ammonia chloride (0.32 g, 6.0 mmol, 1118 eq.) were then added. The solution was then stirred at r.t. for 56 hr, dialyzed against water (Spectra / Por®7, MWCO 1.0 kDa) for 48 hr, and freeze dried to give a white powder (3.18 g, 88%). GPC-MALS: Mn = 851 kDa, Ð = 1.23. Copolymer Example 2: Synthesis of 9-arm 10 mol% azide loading copolymer

[0254] Synthetic Scheme:-46-

[0255] 2-Methacryloyloxyethyl phosphorylcholine (5.00 g, 16.9 mmol, 2390 eq.) were added to a dry round bottom flask equipped with a 3-way stopcock in a glovebox, followed by addition (under dry N2) of degassed anhydrous ethanol (15.4 mL), stock solution of 3-azido-2-hydroxypropyl methacrylate (0.49 M in ethanol, 3.80 mL, 266 eq.), CuBr2(13.4 mM in ethanol, 0.10 mL, 0.20 eq.), tris(2-pyridylmethyl)amine (TPMA) and ascorbic acid (28.2 mM and 28.4 mM respectively in ethanol, 0.50 mL, 2 eq. for both). The reaction was started with the addition of the initiator (3.6 mM in ethanol, 1.95 mL, 1.0 eq.) at r.t. and samples were taken periodically under N2to monitor the monomer conversion viaNMR spectroscopic analysis. The polymerization was terminated after 7.5 h (monomer conversion = 0.90) by exposing to air. The crude product solution was diluted with ethanol (25 mL), passed through a silica column (Biotage® Sfär Silica HC D, 50 g, ethanol as eluent), concentrated and precipitated into ethyl acetate (400 mL) twice. The polymer precipitate was then collected, dried under reduced pressure, re-dissolved in DI water and freeze-dried to give a white powder (3.02 g, 62.5%). GPC-MALS: Mn= 641.5 kDa, Ð = 1.08. Copolymer Example 3: Synthesis of 9-arm 10 mol% azide loading copolymer

[0256] Synthetic Scheme:-47-

[0257] 2-Methacryloyloxyethyl phosphorylcholine (5.00 g, 16.9 mmol, 3186 eq.) were added to a dry round bottom flask equipped with a 3-way stopcock in a glovebox, followed by addition (under dry N2) of degassed anhydrous ethanol (19.5 mL), 3-azido propyl methacrylate (0.30 mL, 1.88 mmol, 354 eq.), CuBr2(13.4 mM in ethanol, 0.075 mL, 0.2 eq.), tris(2-pyridylmethyl)amine (TPMA) and ascorbic acid (28.2 mM and 28.4 mM respectively in ethanol, 0.35 mL, 1.86 eq. for both). The reaction was started with the addition of the initiator (3.6 mM in ethanol, 1.50 mL, 1 eq.) at r.t. and samples were taken periodically under N2to monitor the monomer conversion via1H NMR spectroscopic analysis. The polymerization was terminated after 4.8 h (monomer conversion = 0.73) by exposing to air. The crude product solution was diluted with ethanol (25.0 mL), passed through a silica column (Biotage® Sfär Silica HC D, 50 g, ethanol as eluent), concentrated and precipitated into diethyl ether (400 mL) twice. The polymer precipitate was then collected, dried under reduced pressure, re-dissolved in DI water and freeze-dried to give a white powder (2.66 g, 68.2%). GPC-MALS: Mn = 774.7 kDa, Ð = 1.11. Copolymer Example 4: Synthesis of 9-arm 10 mol% azide loading copolymer

[0258] Synthetic Scheme:-48-

[0259] 2-Methacryloyloxyethyl phosphorylcholine (5.00 g, 16.9 mmol, 3186 eq.) were added to a dry round bottom flask equipped with a 3-way stopcock in a glovebox, followed by addition (under dry N2) of degassed anhydrous ethanol (19.0 mL), 2-(2-(2-(2- azidoethoxy)ethoxy)ethoxy)ethyl methacrylate (0.55 g, 1.90 mmol, 354 eq.), CuBr2(6.26 mM in ethanol, 150 µL, 0.18 eq.), tris(2-pyridylmethyl)amine (TPMA) and ascorbic acid (12.7 mM and 12.5 mM respectively in ethanol, 0.85 mL, 2 eq. for both). The reaction was started with the addition of the initiator (3.6 mM in ethanol, 1.45 mL, 1 eq.) at r.t. and samples were taken periodically under N2to monitor the monomer conversion via1H NMR spectroscopic analysis. The polymerization was terminated after 6.5 h (monomer conversion = 0.71) by exposing to air. The crude product solution was diluted with ethanol (25 mL), passed through a silica column (Biotage® Sfär Silica HC D, 50 g, ethanol as eluent), concentrated and precipitated into diethyl ether (400 mL) twice. The polymer precipitate was then collected, dried under reduced pressure, re-dissolved in DI water and freeze-dried to give a white powder (2.45 g, 61.8%). GPC-MALS: Mn= 480 kDa, Ð = 1.42. Copolymer Example 5: Synthesis of 9-arm 5 mol% azide loading copolymer

[0260] Synthetic Scheme:-49-

[0261] 2-Methacryloyloxyethyl phosphorylcholine (20.0 g, 67.7 mmol, 2522 eq.) were added to a dry round bottom flask equipped with a 3-way stopcock in a glovebox, followed by addition (under dry N2) of degassed anhydrous ethanol (68 mL), CuBr2 (89.5 mM in ethanol, 2.7 mL, 9.0 eq.), ascorbic acid (28.4 mM in ethanol, 3.4 mL, 3.6 eq.), 2,2′- bipyridene (160 mM in ethanol, 3.0 mL, 18 eq.), and 3-azido propyl methacrylate (0.55 mL, 3.50 mmol, 130 eq.). The reaction was started with the addition of the initiator (6.2 mM in ethanol, 4.4 mL, 1.0 eq.) at 16 ºC, and samples were taken periodically under N2 to monitor the monomer conversion via1H NMR spectroscopic analysis. The polymerization was terminated after 61.5 h (monomer conversion ~ 0.99) by exposing to air. The crude product solution was diluted with ethanol (50 mL), passed through a silica column (Biotage® Sfär Silica HC D, 50 g, ethanol as eluent), dialyzed against water for 72 hr. The polymer solution was then collected, concentrated under reduced pressure, to give DI water stock solutions of the target polymer (145 mg / mL,114 ml, 80.8%). GPC-MALS: Mn = 694.8 kDa, Ð = 1.09. Copolymer Example 6: Synthesis of 9-arm 10 mol% azide loading copolymer

[0262] Synthetic Scheme:-50-

[0263] 2-Methacryloyloxyethyl phosphorylcholine (19.0 g, 64.3 mmol, 2390 eq.) were added to a dry round bottom flask equipped with a 3-way stopcock in a glovebox, followed by addition (under dry N2) of degassed anhydrous ethanol (68 mL), CuBr2 (89.5 mM in ethanol, 2.7 mL, 9.0 eq.), ascorbic acid (28.4 mM in ethanol, 3.4 mL, 3.6 eq.), 2,2′- bipyridene (160 mM in ethanol, 3.0 mL, 18 eq.), and 3-azido propyl methacrylate (1.15 mL, 7.27 mmol, 270 eq.). The reaction was started with the addition of the initiator (6.2 mM in ethanol, 4.9 mL, 1.12 eq.) at 16 ºC, and samples were taken periodically under N2to monitor the monomer conversion via1H NMR spectroscopic analysis. The polymerization was terminated after 48 h (monomer conversion = 0.98) by exposing to air. The crude product solution was diluted with ethanol (50 mL), passed through a silica column (Biotage® Sfär Silica HC D, 50 g, ethanol as eluent). The crude was sample dialyzed against water for 72 hr. The dialyzed copolymer solution was then filtered and concentrated under reduced pressure to give DI water stock solutions of the target polymer (89.2 mg / ml, 188 ml, 84.4%). GPC-MALS: Mn= 655 kDa, Ð = 1.14. Copolymer Example 7: Synthesis of 9-arm 2.5 mol% azide loading copolymer

[0264] Synthetic Scheme:-51-

[0265] 2-Methacryloyloxyethyl phosphorylcholine (20.0 g, 67.7 mmol, 2589 eq.) were added to a dry round bottom flask equipped with a 3-way stopcock in a glovebox, followed by addition (under dry N2) of degassed anhydrous ethanol (66 mL), CuBr2 (89.5 mM in ethanol, 2.6 mL, 9.0 eq.), ascorbic acid (28.4 mM in ethanol, 3.3 mL, 3.6 eq.), 2,2′- bipyridene (160 mM in ethanol, 2.9 mL, 18 eq.), and 3-azido propyl methacrylate (275 ul, 1.74 mmol, 66 eq.). The reaction was started with the addition of the initiator (6.2 mM in ethanol, 4.2 mL, 1 eq.) at 16 ºC, and samples were taken periodically under N2to monitor the monomer conversion via1H NMR spectroscopic analysis. The polymerization was terminated after 67 h (monomer conversion = 0.99) by exposing to air. The crude product solution was diluted with ethanol (100 mL), passed through a silica column (Biotage® Sfär Silica HC D, 50 g, ethanol as eluent). The crude polymer solution was then dialyzed against water for 72 hr. The crude reaction was concentrated under reduced vacuum to142 mg / ml. The final product was then stored frozen at -20 ºC (14.7 g, 73%). GPC-MALS: Mn = 791.4 kDa, Ð = 1.11 Copolymer Example 8: Synthesis of 9-arm 5 mol% azide loading copolymer

[0266] Synthetic Scheme:-52-

[0267] 2-Methacryloyloxyethyl phosphorylcholine (20.0 g, 67.7 mmol, 2522 eq.) were added to a dry round bottom flask equipped with a 3-way stopcock in a glovebox, followed by addition (under dry N2) of degassed anhydrous ethanol (68 mL), CuBr2 (89.5 mM in ethanol, 2.7 mL, 9.0 eq.), ascorbic acid (28.4 mM in ethanol, 3.4 mL, 3.6 eq.), 2,2′-bipyridine (160 mM in ethanol, 3.0 mL, 18 eq.), and 3-azidopropyl 2-fluoroacrylate (580 µl, 3.56 mmol, 133 eq.). The reaction was started with the addition of the initiator (6.2 mM in ethanol, 4.3 mL, 1 eq.) at 16 ºC, and samples were taken periodically under N2 to monitor the monomer conversion via1H NMR spectroscopic analysis. The polymerization was terminated after 67.5 h (monomer conversion = 0.99) by exposing to air. The crude product solution was diluted with ethanol (150 mL), passed through a silica column (Biotage® Sfär Silica HC D, 50 g, ethanol as eluent). The crude polymer solution was then dialyzed against water for 72 hr. The crude reaction was concentrated under reduced vacuum to 158.9 mg / ml. The final product was then stored frozen at -20 ºC (11.4 g, 55.3%). GPC-MALS: Mn = 738 kDa, Ð = 1.18 Copolymer Example 9: Synthesis of 9-arm 5 mol% azide loading copolymer

[0268] Synthetic Scheme:-53-

[0269] 2-Methacryloyloxyethyl phosphorylcholine (5.0 g, 16.9 mmol, 3027 eq.) were added to a dry 2-neck round bottom flask equipped with a 3-way stopcock in a glovebox, followed by addition (under dry N2) of degassed anhydrous ethanol (16.5 mL), CuBr2 (89.5 mM in ethanol, 0.70 mL, 9.0 eq.), ascorbic acid (28.4 mM in ethanol, 0.8 mL, 3.4 eq.), 2,2′- bipyridine (160 mM in ethanol, 0.70 mL, 18 eq.), and 3-azido propyl methacrylate (150 µl, 0.95 mmol, 170 eq.). The reaction was started with the addition of the initiator (3.6 mM in ethanol, 1.5 mL, 1 eq.) at r.t,, and samples were taken periodically under N2to monitor the monomer conversion via1H NMR spectroscopic analysis.5 mL of the crude polymerization solution were taken at t = 4 hr, 45% monomer conversion (1H NMR spectroscopic analysis), dialyzed against water and freeze-dried to give the final product as a white powder. GPC- MALS: Mn = 396 kDa, Ð = 1.09 Copolymer Example 10: Synthesis of 9-arm 5 mol% azide loading copolymer

[0270] Synthetic Scheme:-54-

[0271] The rest of the polymerization of Copolymer Example 9 was terminated after 10 h (monomer conversion = 0.90) by exposing to air. The crude product solution was diluted with ethanol (~ 30 mL), passed through a silica column (Biotage® Sfär Silica HC D, 25g, ethanol as eluent). The crude polymer solution was then concentrated and dialyzed against water for 72 hr. The dialyzed polymer solution was then concentrated under reduced vacuum to 76 mg / ml and stored at -20 ºC. GPC-MALS: Mn = 851 kDa, Ð = 1.13 Copolymer Example 11: Synthesis of linear homopolymer with azide end group

[0272] Synthetic Scheme:-55-2-methacryloyloxyethyl phosphorylcholine (2.0 g, 6.8 mmol, 50 eq.) was added to a 25 mL round bottom flask equipped with a 3-way stopcock in a dry box. Anhydrous ethanol (5.5 mL) that had been sparged with nitrogen was added to the flask and dissolved the 2- methacryloyloxyethyl phosphorylcholine. Stock solutions were prepared by dissolving Initiator 3 (71 mg) in ethanol (3.6 mL), copper(II) bromide (200 mg) in ethanol (5.0 mL), ascorbic acid (38 mg) in ethanol (5.0 mL), and 2,2'-bipyridine (500 mg) in ethanol (5.0 mL). The copper(II) bromide stock solution (0.76 mL, 0.14 mmol, 1 eq.) was added to the reaction flask, followed by the ascorbic acid stock solution (1.3 mL, 0.05 mmol, 0.4 eq.) and the 2,2'- bipyridyl stock solution (0.42 mL, 0.27 mmol, 2 eq.). The polymerization was started by addition of the initiator stock solution (1.7 mL, 0.14 mmol, 1 eq.). The polymerization was terminated after 26 hours by exposing the reaction to air. The final monomer conversion was 68%. The reaction solution was diluted with deionized water and then dialyzed against water for 2 days with Spectra / Por® 1 kDa MWCO dialysis tubing. Following purification by dialysis, the polymer solution was dried by lyophilization and stored in the freezer at -20 °C. GPC- MALS: Mn = 19.3 kDa, Ð = 1.11 Copolymer Example 12: Synthesis of 3-arm 3 mol% azide loading coplymer

[0273] Synthetic Scheme:-56-

[0274] 2-methacryloyloxyethyl phosphorylcholine (4.0 g, 14 mmol, 666 eq.) was added to a 2-neck 50 mL round bottom flask equipped with a 3-way stopcock in a dry box. Anhydrous ethanol (6.4 mL) that had been sparged with nitrogen was added to the flask and dissolved the 2-methacryloyloxyethyl phosphorylcholine. Stock solutions were prepared by dissolving initiator (43 mg) in ethanol (2.5 mL), copper(II) bromide (50 mg) in ethanol (2.5 mL), ascorbic acid (35 mg) in ethanol (5.0 mL), and 2,2'-bipyridine (63 mg) in ethanol (2.5 mL). The copper(II) bromide stock solution (0.68 mL, 0.06 mmol, 3 eq.) was added to the reaction flask, followed by the ascorbic acid stock solution (0.61 mL, 0.02 mmol, 1.2 eq.). Then, the 2,2'-bipyridyl stock solution (0.76 mL, 0.12 mmol, 6 eq.) was added. A vial containing 3-azidopropyl methacrylate was degassed with nitrogen for 2 minutes and then 0.17 mL was added to degassed ethanol (9.0 mL) in a vial, forming a stock solution. This 3- azidopropyl methacrylate stock solution (3.4 mL, 0.41 mmol, 20 eq.) was added to the reaction flask. The polymerization was started by addition of the initiator stock solution (1.4 mL, 0.02 mmol.1 eq.). The polymerization was allowed to continue overnight and was terminated after 19 hours by exposing the reaction to air. The final monomer conversion was 77%. The reaction solution was diluted with ethanol (50 mL) and deionized water (30 mL) and then dialyzed against water for 2 days with Spectra / Por® 12 - 14 kDa MWCO dialysis tubing. After dialysis, the aqueous solution was filtered and concentrated under reduced pressure. The solution was stored in the freezer at -20 °C at a concentration of 123 mg / mL. GPC-MALS: Mn = 157 kDa, Ð = 1.14. Copolymer Example 13: Synthesis of 3-arm 10 mol% azide loading copolymer

[0275] Synthetic Scheme:-57-

[0276] 2-methacryloyloxyethyl phosphorylcholine (4.7 g, 16 mmol, 733 eq.) was added to a 2-neck 50 mL round bottom flask equipped with a 3-way stopcock in a dry box. Anhydrous ethanol (5.3 mL) that had been sparged with nitrogen was added to the flask and dissolved the 2-methacryloyloxyethyl phosphorylcholine. Stock solutions were prepared by dissolving initiator (43 mg) in ethanol (2.5 mL), copper(II) bromide (50 mg) in ethanol (2.5 mL), ascorbic acid (50 mg) in ethanol (5.0 mL), and 2,2'-bipyridine (200 mg) in ethanol (2.5 mL). The copper(II) bromide stock solution (0.73 mL, 0.07 mmol, 3 eq.) was added to the reaction flask, followed by the ascorbic acid stock solution (0.46 mL, 0.03 mmol, 1.2 eq.). Then, the 2,2'-bipyridyl stock solution (0.25 mL, 0.13 mmol, 6 eq.) was added. A vial containing 3-azidopropyl methacrylate was degassed with nitrogen for 2 minutes and then 0.28 mL (1.8 mmol, 81 eq) was added to the reaction flask. The polymerization was started by addition of the initiator stock solution (1.4 mL, 0.02 mmol, 1 eq.). The polymerization was allowed to continue overnight and was terminated after 19 hours by exposing the reaction to air. The final monomer conversion was 100%. The reaction solution was diluted with ethanol (40 mL) and deionized water (40 mL) and then dialyzed against water for 2 days with Spectra / Por® 12 - 14 kDa MWCO dialysis tubing. After dialysis, the aqueous solution was filtered and concentrated under reduced pressure. The solution was stored in the freezer at -20 °C at a concentration of 158 mg / mL. GPC-MALS: Mn = 229 kDa, Ð = 1.25. Copolymer Example 14: Synthesis of 9-arm 10 mol% azide loading copolymer

[0277] Synthetic Scheme:-58-

[0278] 2-methacryloyloxyethyl phosphorylcholine (4.7 g, 16 mmol, 127 eq.) was added to a 2-neck 50 mL round bottom flask equipped with a 3-way stopcock in a dry box. Anhydrous ethanol (4.9 mL) that had been sparged with nitrogen was added to the flask and dissolved the 2-methacryloyloxyethyl phosphorylcholine. Stock solutions were prepared by dissolving initiator (420 mg) in ethanol (6.0 mL), copper(II) bromide (320 mg) in ethanol (8.0 mL), ascorbic acid (110 mg) in ethanol (8.0 mL), and 2,2'-bipyridine (550 mg) in ethanol (5.0 mL). The copper(II) bromide stock solution (6.3 mL, 1.1 mmol, 9 eq.) was added to the reaction flask. Subsequently, the ascorbic acid stock solution (5.7 mL, 0.45 mmol, 3.6 eq.) and the 2,2'- bipyridyl stock solution (3.2 mL, 2.3 mmol, 18 eq.) were added. A vial containing 3- azidopropyl methacrylate was degassed with nitrogen for 10 minutes and then 0.28 mL (1.8 mmol, 14 eq.) was added to the reaction flask. The polymerization was started by addition of the initiator stock solution (4.9 mL, 0.12 mmol, 1 eq.). The polymerization was allowed to continue overnight and was terminated after 20 hours by exposing the reaction to air. The final monomer conversion was 93%. The reaction solution was diluted with ethanol (50 mL) and deionized water (50 mL) and then dialyzed against water for 2 days with Spectra / Por® 1 kDa MWCO dialysis tubing. After dialysis, the aqueous solution was filtered and concentrated under reduced pressure. The solution was stored in the freezer at -20 °C at a concentration of 86.5 mg / mL. GPC-MALS: Mn= 41.2 kDa, Ð = 1.12. Copolymer Example 15: Synthesis of 9-arm star statistical copolymer, 10 mol% azide loading

[0279] Synthetic Scheme:-59-Br

[0280] 2-methacryloyloxyethyl phosphorylcholine (4.7 g, 16 mmol, 317 eq.) was added to a 2-neck 50 mL round bottom flask equipped with a 3-way stopcock in a dry box. Anhydrous ethanol (9.4 mL) that had been sparged with nitrogen was added to the flask and dissolved the 2-methacryloyloxyethyl phosphorylcholine. Stock solutions were prepared by dissolving initiator (170 mg) in ethanol (10 mL), copper(II) bromide (200 mg) in ethanol (5.0 mL), ascorbic acid (60 mg) in ethanol (6.0 mL), and 2,2'-bipyridine (400 mg) in ethanol (5.0 mL). The copper(II) bromide stock solution (2.5 mL, 0.45 mmol, 9 eq.) was added to the reaction flask, followed by the ascorbic acid stock solution (3.2 mL, 0.18 mmol, 3.6 eq.) and the 2,2'-bipyridyl stock solution (1.8 mL, 0.9 mmol, 18 eq.). A vial containing 3-azidopropyl methacrylate was degassed with nitrogen for 2 minutes and then 0.28 mL (1.8 mmol, 35 eq.) was added to the reaction flask. The polymerization was started by addition of the initiator stock solution (8.1 mL, 0.05 mmol, 1 eq.). The polymerization was allowed to continue overnight and was terminated after 15.5 hours by exposing the reaction to air. The final monomer conversion was 99%. The reaction solution was diluted with ethanol (50 mL) and deionized water (50 mL) and then dialyzed against water for 2 days with Spectra / Por® 12 - 14 kDa MWCO dialysis tubing. After dialysis, the aqueous solution was filtered and concentrated under reduced pressure. The solution was stored in the freezer at -20 °C at a concentration of 167 mg / mL. GPC-MALS: Mn = 103 kDa, Ð = 1.08. Copolymer Example 16: Synthesis of 9-arm 10 mol% azide loading copolymer

[0281] Synthetic Scheme:-60-

[0282] 2-methacryloyloxyethyl phosphorylcholine (4.7 g, 16 mmol, 733 eq.) was added to a 2-neck 50 mL round bottom flask equipped with a 3-way stopcock in a dry box. Anhydrous ethanol (18 mL) that had been sparged with nitrogen was added to the flask and dissolved the 2-methacryloyloxyethyl phosphorylcholine. Stock solutions were prepared by dissolving Initiator (102 mg) in ethanol (6.0 mL), copper(II) bromide (200 mg) in ethanol (5.0 mL), ascorbic acid (50 mg) in ethanol (5.0 mL), and 2,2'-bipyridine (200 mg) in ethanol (5.0 mL). The copper(II) bromide stock solution (1.1 mL, 0.20 mmol, 9 eq.) was added to the reaction flask, followed by the ascorbic acid stock solution (1.4 mL, 0.08 mmol, 3.6 eq.) and the 2,2'-bipyridyl stock solution (1.5 mL, 0.39 mmol, 18 eq.). A vial containing 3-azidopropyl methacrylate was degassed with nitrogen for 10 minutes and then 0.28 mL (1.8 mmol, 81 eq.) was added to the reaction flask. The polymerization was started by addition of the initiator stock solution (3.5 mL, 0.02 mmol, 1 eq.). The polymerization was allowed to continue overnight and was terminated after 16 hours by exposing the reaction to air. The final monomer conversion was 88%. The reaction solution was diluted with ethanol (50 mL) and deionized water (50 mL) and then dialyzed against water for 4 days with Spectra / Por® 12 - 14 kDa MWCO dialysis tubing. After dialysis, the aqueous solution was filtered and concentrated under reduced pressure. The solution was stored in the freezer at -20 °C at a concentration of 104 mg / mL. GPC-MALS: Mn = 199 kDa, Ð = 1.07. Copolymer Example 17: Synthesis of 9-arm 5 mol% azide loading copolymer

[0283] Synthetic Scheme:-61-

[0284] To a solution of a mixture of 2-methacryloyloxyethyl phosphorylcholine (HEMA-PC) (2.588g ) and 3-azidopropyl methacrylate (78.12mg) in 5.517 g of degassed ethanol, is added under inert atmosphere (nitrogen): a solution of CuBr2 (7.637mg) already dissolved in 309uL of degassed ethanol, a solution of ascorbic acid (2.667mg) already dissolved in 410uLof degassed ethanol, and a solution of 2,2′-bipyridine (10.675mg ) already dissolved in 309uL of ethanol) under stirring (400rpm). To the brownish solution formed is added a solution of initiator (10mg in 100uL of ethanol). Aliquots are taken over regular times (1h30, 3h30, 5h30, 8h, 20h, 32h, 46h) to check on the conversion profiles of both monomers and molecular weight evolution. The reaction mixture is finally quenched after 46 hours by exposing the reaction to air to give a blue-greenish solution (Conversions were estimated by1H NMR). A sampler of aliquot is passed through a short column of silica gel, eluting with methanol to give a colorless solution. The solution was dried under vacuum, dissolved in DI water and purified by dialysis (MWCO is 3,500g / mol). Characterization of the final polymer by SEC-MALS gave Mn: 750kDa, Mp: 735kDa, Mw: 785kDa and Ð:1.04 (dn / dc=0.136±0.001mL / g). Copolymer Example 18: Synthesis of 4-arm 5 mol% azide loading copolymer

[0285] Synthetic Scheme:-62-

[0286] To a solution of a mixture of 2-acryloyloxyethyl phosphorylcholine (HEA- PC) (0.475g) and 3-azidopropyl methacrylate (15mg) in 1.5mL of degassed methanol is added under inert atmosphere (nitrogen): a solution of CuBr2 (4.46mg) already dissolved in 50uL of degassed methanol, a solution of ascorbic acid (4mg) already dissolved in 200uL of degassed methanol, and 5.5uL of Tris [2-(dimethylamino) ethyl]amine (6-methyl TREN) under stirring. To the light blueish solution formed is added a solution of initiator 4fBiB (3.66mg in 36.6uL of N,N-Dimethylformamide (DMF)). Aliquots are taken over regular times (2h, 12h, 19h, 25h, 39h and 113h) to check on the conversion profiles for both monomers. The reaction mixture is finally quenched after 113h hours by exposing the reaction to air to give a blue-greenish solution. Conversions from the samplings were estimated by1H NMR analysis. Crude reaction was purified by dialysis (MWCO 14kDa) before drying by lyophilization and characterized by SEC MALS. Final polymer was characterized by SEC-MALS after purification (dialysis, MWCO 14kDa). Mn: 66kDa, Mp: 75kDa, Mw: 85kDa, Ð: 1.28 (dn / dc=0.158±0.002mL / g). Copolymer Example 19: Synthesis of 9-arm 5 mol% azide loading copolymer

[0287] Synthetic Scheme:-63-

[0288] To a solution of a mixture of 2-acryloyloxyethyl phosphorylcholine (HEA- PC) (1.26g) in 1.32mL of methanol and 3-azidopropyl methacrylate (39.9mg) in 62.5mL of degassed methanol is added under inert atmosphere (nitrogen): a solution of CuBr2(3.73mg) already dissolved in 41.1uL of degassed methanol, a solution of ascorbic acid (3.32mg ) already dissolved in 166uLof degassed methanol, and 3.84mg of Tris [2- (dimethylamino)ethyl]amine (6-methyl TREN) under stirring. To the light blueish solution formed is added a solution of initiator (5.1mg in 25uL of N,N-Dimethylformamide (DMF)). Aliquots are taken over regular times (2h, 5h, 17h, 27h, 41h) to check on the conversion profiles for both monomers. The reaction mixture is finally quenched after 41 hours by exposing the reaction to air to give a blue-greenish solution. Final polymer was characterized by SEC-MALS after purification (dialysis, MWCO 14kDa). Mn: 1,050kDa, Mp: 2,500kDa, Mw: 1,880kDa, Ð: 1.85 (dn / dc=0.143±0.000mL / g). Copolymer Example 20: Synthesis of 9-arm 5 mol% azide loading copolymer

[0289] Synthetic Scheme:-64-

[0290] To a solution of a mixture of 2-acryloyloxyethyl phosphorylcholine (HEA- PC) (1.26g) in 2.28mL of methanol and 3-azidopropyl methacrylate (39.9mg) in 3mL of degassed methanol is added under inert atmosphere ( nitrogen): a solution of CuBr2 (3.73mg) already dissolved in 41.1uL of degassed methanol, a solution of ascorbic acid (3.32mg ) already dissolved in 166uLof degassed methanol, and 3.84mg of Tris [2- (dimethylamino)ethyl]amine (6-methyl TREN) under stirring. To the light blueish solution formed is added a solution of initiator (5.1mg in 100uL of N,N-Dimethylformamide (DMF)). Aliquots are taken over regular times (2h, 4h, 16h, 28h, 44h, 120h, 164h, 212h, 283h) to check on the conversion profiles for both monomers. The reaction mixture is finally quenched after 283h hours by exposing the reaction to air to give a blue-greenish solution. Final polymer was characterized by SEC-MALS after purification (dialysis, MWCO 14kDa). Mn: 95 kDa, Mp: 200 kDa, Mw: 180 kDa, Ð: 1.90 (dn / dc=0.162±0.001mL / g). Copolymer Example 21: Synthesis of 9-arm 5 mol% azide loading copolymer

[0291] Synthetic Scheme:-65-

[0292] To a solution of a mixture of 2-acryloyloxyethyl phosphorylcholine (HEA- PC) (1.26g) and 3-azidopropyl acrylate (37.7mg) in 1.5mL of degassed methanol is added under inert atmosphere (nitrogen): a solution of CuBr2 (3.73mg) already dissolved in 41.1uL of degassed methanol, a solution of ascorbic acid (3.32mg) already dissolved in 166uL of degassed methanol, and 3.84mg of Tris [2-(dimethylamino) ethyl]amine (6-methyl TREN) under stirring. To the light blueish solution formed is added a solution of initiator (5.1mg in 25uL of N, N-Dimethylformamide (DMF)). Aliquots are taken over regular times (2h, 5h, 17h, 29h) to check on the conversion profiles for both monomers. The reaction mixture is finally quenched after 29 hours by exposing the reaction to air to give a blue-greenish solution. Conversions from the samplings were estimated by1H NMR analysis. Crude reaction was purified by dialysis (MWCO: 14kDa) before drying by lyophilization and characterized by SEC-MALS. Mn: 385 kDa, Mp: 500 kDa, Mw: 700 kDa, Ð: 1.80 (dn / dc=0.140±0.003mL / g). Copolymer Example 22: Synthesis of 9-arm 10 mol% azide loading copolymer

[0293] Synthetic Scheme:-66-

[0294] To a solution of a mixture of 2-acryloyloxyethyl phosphorylcholine (HEA- PC) (1.26g) and 3-azidopropyl acrylate (77.2mg) in 1.5mL of degassed methanol is added under inert atmosphere ( nitrogen): a solution of CuBr2 (3.73mg) already dissolved in 41.1uL of degassed methanol, a solution of ascorbic acid (3.32mg ) already dissolved in 166uLof degassed methanol, and 3.84mg of Tris [2-(dimethylamino)ethyl]amine (6-methyl TREN) under stirring. To the light blueish solution formed is added a solution of initiator (5.1mg in 25uL of N, N-Dimethylformamide (DMF)). Aliquots are taken over regular times (2h, 5h, 17h, 29h) to check on the conversion profiles for both monomers. The reaction mixture is finally quenched after 29 hours by exposing the reaction to air to give a blue-greenish solution. Conversions from the samplings were estimated by1H NMR analysis. Crude reaction was purified by dialysis (MWCO 14kDa) before drying by lyophilization and characterized by SEC-MALS. Mn: 175 kDa, Mp: 265 kDa, Mw: 300 kDa, Ð: 1.75 (dn / dc=0.145±0.002mL / g). Copolymer Example 23: Synthesis of 9-arm 10 mol% comonomer loading

[0295] Synthetic Scheme:

[0296] In a scintillation vial was added a mixture of 1.25g of 2- methacryloyloxyethyl phosphorylcholine (HEMA-PC) and 179mg of comonomer in degassed ethanol (4g of ethanol) is added under inert atmosphere (nitrogen): CuBr2 (3.6) , ascorbic acid (1.1mg) and bpy (5.1mg). To the brownish solution form is added a solution of initiator (5mg in 50uL of ethanol). Aliquots are taken over regular times (2h, 4h, 7h, 23h, 30h and 45h) to check on the conversion profiles and molecular weight evolution. The reaction mixture is quenched after 48 hours by exposing the reaction to air to give a blue-greenish solution.-67-Samplings were performed at 12h, 24h, 36h and 46h. Conversions were estimated by SEC- UV. Final polymer R14008 was characterized by SEC-MALS after purification (dialysis, MWCO 1kDa). Mn: 300 kDa, Mp: 460 kDa, Mw: 480 kDa, Ð: 1.60 (dn / dc used is 0.142mL / g.). Incorporation estimated by1H NMR is 9 % Copolymer Example 24: Synthesis of 9-arm 10 mol% comonomer loading copolymer

[0297] Synthetic Scheme:

[0298] In a scintillation vial was added a mixture of 1g of 2-methacryloyloxyethyl phosphorylcholine (HEMA-PC) and 140mg of comonomer in degassed ethanol (3g of ethanol) is added under inert atmosphere ( nitrogen): CuBr2(2.68mg) , ascorbic acid (0.8mg) and bpy (3.7mg). To the brownish solution form is added a solution of initiator (3.6mg in 40mg of ethanol). Aliquots are taken over regular times (4h, 16h, 24h, 40h) to check on the conversion profiles and molecular weight evolution. The reaction mixture is quenched after 48 hours by exposing the reaction to air to give a blue-greenish solution. Samplings were performed at 12h, 24h, 36h and 46h. Conversions were estimated by SEC UV. Final polymer was characterized by SEC-MALS after purification (dialysis, MWCO 1kDa). Mn: 1,400 kDa, Mp: 1,390 kDa, Mw: 1785 kDa, Ð: 1.27 (dn / dc used is 0.142mL / g). Incorporation estimated by1H NMR is 9 . Copolymer Example 25: Synthesis of 9-arm 10 mol% comonomer loading copolymer

[0299] Synthetic Scheme:-68-

[0300] To a 40mL flask equipped with a stir bar under an inert atmosphere (glovebox) was added a mixture of 18.408g of 2-methacryloyloxyethyl phosphorylcholine (HEMA-PC) and 1.3525g of comonomer already dissolved in 58.21g of ethanol.

[0301] To the solution were added 52.95mg of CuBr2already dissolved in 1.185 g of ethanol, followed, after 5 minutes, by the addition of a stock solution of ascorbic acid (18.5mg of ascorbic acid already dissolved in 1.58g of ethanol.) After 5 minutes, to the reaction mixture was added a final stock solution of 1.185g of bpy already dissolved in 1.185g of ethanol. A brown coloration is seen instantaneously. After 5mn of stirring, the initiator is added as a solution (72.32mg dissolved in 570.21mg of ethanol). Conversions were estimated by SEC-UV using a reference. Samplings for both conversion and molecular analysis were performed after 2h for 1st data point), 13 h, 24h and 37h when quenching took place. Final polymer was characterized by SEC-MALS after purification (dialysis, MWCO 1kDa). Mn: 1,000 kDa, Mp: 880 kDa, Mw: 1,300 kDa, Ð: 1.3 (dn / dc=0.134±0.013mL / g). Copolymer Example 26: Synthesis of 9-arm 10 mol% comonomer loading copolymer-69-

[0303] To a solution of a mixture of 47.3g of 2-methacryloyloxyethyl phosphorylcholine (HEMA-PC) and 3.3g of comonomer in degassed ethanol (146.3g of ethanol) is added under inert atmosphere (nitrogen): a solution of CuBr2(134mg) already dissolved in 1g of degassed ethanol and immediately rinsed with 2g of degassed ethanol, a solution of ascorbic acid (46.8mg) already dissolved in 3g of degassed ethanol and immediately rinsed with 1g of dry ethanol, and a solution of bpy (187mg) already dissolved in 1g of ethanol) and immediately rinsed with 2g of dry ethanol under stirring (400rpm). To the brownish solution form is added a solution of initiator (183mg in 1.443g of ethanol). Aliquots are taken over regular times (4h, 16h, 24h, 40h) to check on the conversion profiles and molecular weight evolution. The reaction mixture is quenched after 48 hours by exposing the reaction to air to give a blue-greenish solution (Conversion on a aliquot was estimated by 1H NMR to be about 97%). A sampler of aliquot is passed through a short column of silica gel, eluting with methanol to give a colorless solution. The solution was dried under vacuum, dissolved in DI water and purified by dialysis (MWCO 1,000g / mol). Characterization of the final polymer by SEC-MALS gave Mn: 840kDa, Mp: 790kDa, Mw: 1,080kDa and Ð: 1.3 (dn / dc=0.146±0.002mL / g). Copolymer Example 27: Synthesis of 9-arm 5 mol% comonomer loading copolymer

[0304] Synthetic Scheme:

[0305] To a solution of a mixture of 47.5 of 2-methacryloyloxyethyl phosphorylcholine (HEMA-PC) and 1.653g of comonomer in degassed ethanol (146.3g of ethanol) is added under inert atmosphere ( nitrogen): a solution of CuBr2(134mg) already dissolved in 1g of degassed ethanol and immediately rinsed with 2g of degassed ethanol, a-70-solution of ascorbic acid (46.8mg) already dissolved in 3g of degassed ethanol and immediately rinsed with 1g of dry ethanol, and a solution of bpy (187mg) already dissolved in 1g of ethanol) and immediately rinsed with 2g of dry ethanol under stirring (400rpm). To the brownish solution form is added a solution of initiator (183mg in 1.443g of ethanol). Aliquots are taken over regular times (2h, 4h, 7h, 19h, 31h, 55h) to check on the conversion profiles and molecular weight evolution. The reaction mixture is quenched after 55 hours by exposing the reaction to air to give a blue-greenish solution. A sampler of aliquot is passed through a short column of silica gel, eluting with methanol to give a colorless solution. The solution was dried under vacuum, dissolved in DI water and purified by dialysis (MWCO 1,000g / mol). Characterization of the final polymer by SEC-MALS after purification gave Mn: 600kDa, Mp: 590 kDa, Mw: 610 Da and Ð: 1.03 (dn / dc=0.135±0.001mL / g). Copolymer Example 28: Synthesis of 9-arm 10 mol% comonomer loading copolymer

[0306] Synthetic Scheme:

[0307] To a solution of a mixture of 47.3g of 2-methacryloyloxyethyl phosphorylcholine (HEMA-PC) and 3.3g of comonomer in degassed ethanol (146.3g of ethanol) is added under inert atmosphere ( nitrogen): a solution of CuBr2(134mg) already dissolved in 1g of degassed ethanol and immediately rinsed with 2g of degassed ethanol, a solution of ascorbic acid (46.8mg) already dissolved in 3g of degassed ethanol and immediately rinsed with 1g of dry ethanol, and a solution of bpy (187mg) already dissolved in 1g of ethanol) and immediately rinsed with 2g of dry ethanol under stirring (400rpm). To the brownish solution form is added a solution of INITIATOR (183mg in 1.443g of ethanol). Reaction mixture was allowed to react for 2 hours at room temperature before being cooled down at 5C via a jacketed reactor. Aliquots are taken over regular times (2h, 14h, 26h, 69h,-71-137h) to check on the conversion profiles and molecular weight evolution. A sampler of aliquot is passed through a short column of silica gel, eluting with methanol to give a colorless solution. The solution was dried under vacuum, dissolved in DI water and purified by dialysis (MWCO 1000g / mol). Final polymer was characterized by SEC-MALS after purification (dialysis, MWCO 1kDa). Mn: 510kDa, Mp:505 kDa, Mw:535 Da and Ð: 1.05 (dn / dc=0.148±0.001mL / g). Copolymer Example 29: Synthesis of 9-arm 10 mol% comonomer loading copolymer

[0308] Synthetic Scheme:

[0309] To a solution of a mixture of 2-methacryloyloxyethyl phosphorylcholine (HEMA-PC) (5.13g ) and comonomer (375mg) in 15.725 g of degassed ethanol is added under inert atmosphere ( nitrogen): a solution of CuBr2(15.2mg) already dissolved in 618uL of degassed ethanol, a solution of ascorbic acid (5.3mg) already dissolved in 820uLof degassed ethanol, and a solution of bpy (21.3mg ) already dissolved in 618uL of ethanol) under stirring (400rpm). To the brownish solution formed is added a solution of initiator (20mg in 200uL of ethanol). Aliquots are taken over regular times (2h30, 7h, 19h, 24h, 31h, 48h, 115h and 336h) to check on the conversion profiles of both monomers and molecular weight evolution. The reaction mixture is finally quenched after 336 hours by exposing the reaction to air to give a blue-greenish solution (Conversions were estimated by1H NMR). A sampler of aliquot is passed through a short column of silica gel, eluting with methanol to give a colorless solution. The solution was dried under vacuum, dissolved in DI water and purified by dialysis (MWCO is 1,000g / mol). Characterization of the final polymer by SEC-MALS gave Mn: 840kDa, Mp: 835kDa, Mw: 990kDa and Ð: 1.18 (dn / dc=0.144±0.003mL / g). Copolymer Example 30: Synthesis of 3-arm 14 mol% azide loading copolymer-72-

[0310] Synthetic Scheme:

[0311] 2-Methacryloyloxyethyl phosphorylcholine (5.0 g, 16.9 mmol, 60 eq.) were added to a dry 2-neck round bottom flask equipped with a 3-way stopcock in a glovebox, followed by addition (under dry N2) of degassed anhydrous ethanol (6.3 mL), CuBr2 (179 mM in ethanol, 4.7 mL, 3.0 eq.), ascorbic acid (56.7 mM in ethanol, 6.0 mL, 1.2 eq.), 2,2′-bipyridine (0.64 M in ethanol, 2.6 mL, 6.0 eq.), and 3-azido propyl methacrylate (3-azidopropyl methacrylate, 450 µl, 2.85 mmol, 10 eq.). The reaction was started with the addition of the initiator (2-(((2-bromo-2-methylpropanoyl)oxy)methyl)-2-methylpropane-1,3-diyl bis(2- bromo-2-methylpropanoate), 113 mM in ethanol, 2.5 mL, 1 eq.) at r.t., and samples were taken periodically under N2to monitor the monomer conversion via1H NMR spectroscopic analysis. The polymerization was terminated after 24 h (monomer conversion = 99%) by exposing to air. The crude product solution was diluted with ethanol (~ 10 mL), and water (50 mL). The crude polymer solution was then concentrated and dialyzed (MWCO = 1000) against water for 72 hr. The dialyzed polymer solution was then concentrated under reduced vacuum to 186 mg / ml. The final product was aliquoted, stored frozen at -20oC (5.21g, 92.4%). GPC-MALS: Mn= 25.7 kDa, Ð =1.11. Copolymer Example 31: Synthesis of 9-arm 10 mol% azide loading copolymer

[0312] Synthetic Scheme:-73-

[0313] 2-Methacryloyloxyethyl phosphorylcholine (65.0 g, 220 mmol, 1600 eq.) was first dissolved with 82 mL anhydrous ethanol, and then added to 300 mL reactor under nitrogen atmosphere. To the reactor, ethanol (135 mL anhydrous) was then added followed by the addition of CuBr2 solution (90 mM in ethanol, 14.0 mL, 9.0 eq.), ascorbic acid solution (28 mM in ethanol, 17.4 mL, 3.6 eq.), 2,2′-bipyridine solution (160 mM in ethanol,15.5 mL, 18 eq.), and 3-azido propyl methacrylate (3.9 ml, 24.8 mmol, 180 eq.). The reaction was started with the addition of the initiator (7.7 mM in ethanol, 18 mL, 1 eq.) at r.t., and samples were taken periodically under N2to monitor the monomer conversion via1H NMR spectroscopic analysis. The polymerization was terminated after 54 h (monomer conversion = 98%) by exposing to air. The crude product solution was diluted with ethanol (~ 150 mL). The crude polymer solution was then dialyzed (Spectra / Por®4, MWCO = 15-20 kDa) against water for 72 hr. The dialyzed polymer solution was then concentrated under reduced vacuum to 176 mg / ml. The final product was then stored frozen at -20 ºC (66.8 g, 98.1%). GPC-MALS: Mn = 472 kDa, Ð = 1.03. Copolymer Example 32: Synthesis of linear 10 mol% azide loading copolymer

[0314] Synthetic Scheme:-74-

[0315] 2-methacryloyloxyethyl phosphorylcholine (3.7 g, 13 mmol, 318 eq.) was added to a 2-neck 50 mL round bottom flask equipped with a 3-way stopcock in a dry box. Anhydrous ethanol (11 mL) that had been sparged with nitrogen was added to the flask and dissolved the 2-methacryloyloxyethyl phosphorylcholine. Stock solutions were prepared by dissolving the initiator ETHYL Α-BROMOISOBUTYRATE (136 mg) in ethanol (8.0 mL), copper(II) bromide (50 mg) in ethanol (2.5 mL), ascorbic acid (25 mg) in ethanol (5.0 mL), and 2,2'-bipyridine (63 mg) in ethanol (2.5 mL). The copper(II) bromide stock solution (0.44 mL, 0.04 mmol, 1 eq.) was added to the reaction flask, followed by the ascorbic acid stock solution (0.55 mL, 0.02 mmol, 0.4 eq.). Then, the 2,2'-bipyridyl stock solution (0.49 mL, 0.08 mmol, 2 eq.) was added. A vial containing 3-azidopropyl methacrylate was degassed with nitrogen for 2 minutes and then 0.22 mL (1.4 mmol, 35 eq.) was added to the reaction flask. The polymerization was started by addition of the initiator stock solution (0.45 mL, 0.04 mmol, 1 eq.). The polymerization was allowed to continue overnight and was terminated after 24 hours by exposing the reaction to air. The final monomer conversion was 84%. The reaction solution was diluted with ethanol (50 mL) and deionized water (30 mL) and then dialyzed against water for 4 days with Spectra / Por® 1 kDa MWCO dialysis tubing. After dialysis, the aqueous solution was filtered and concentrated under reduced pressure. The solution was stored in the freezer at -20 °C at a concentration of 130 mg / mL. GPC-MALS: Mn = 104 kDa, Ð = 1.37.-75-Copolymer Example 33: Synthesis of 3-arm 3 mol% azide loading copolymer

[0316] Synthetic Scheme:

[0317] 2-methacryloyloxyethyl phosphorylcholine (16 g, 54 mmol, 863 eq.) was added to a 2-neck 100 mL round bottom flask equipped with a 3-way stopcock in a dry box. Anhydrous ethanol (43 mL) that had been sparged with nitrogen was added to the flask and dissolved the 2-methacryloyloxyethyl phosphorylcholine. Stock solutions were prepared by dissolving the initiator (85 mg) in ethanol (5.0 mL), copper(II) bromide (100 mg) in ethanol (5.0 mL), ascorbic acid (40 mg) in ethanol (8.0 mL), and 2,2'-bipyridine (130 mg) in ethanol (5.0 mL). The copper(II) bromide stock solution (2.1 mL, 0.19 mmol, 3 eq.) was added to the reaction flask, followed by the ascorbic acid stock solution (2.7 mL, 0.08 mmol, 1.2 eq.). Then, the 2,2'-bipyridyl stock solution (2.4 mL, 0.38 mmol, 6 eq.) was added. 3-azidopropyl methacrylate was degassed with nitrogen for 2 minutes and then 0.26 mL (1.7 mmol, 27 eq.) was added to the reaction flask. The polymerization was started by addition of the initiator stock solution (3.3 mL, 0.06 mmol, 1 eq.). The polymerization was allowed to continue overnight and was terminated after 23 hours by exposing the reaction to air. The final monomer conversion was 96%. The reaction solution was diluted with ethanol (30 mL) and deionized water (40 mL) and then dialyzed against water for 4 days with Spectra / Por® 12 - 14 kDa MWCO dialysis tubing. After dialysis, the aqueous solution was filtered and concentrated under reduced pressure. The solution was stored in the freezer at -20 °C at a concentration of 186 mg / mL. GPC-MALS: Mn= 231 kDa, Ð = 1.11. Copolymer Example 34: Synthesis of 9-arm 30 mol% azide loading copolymer

[0318] Synthetic Scheme:-76-

[0319] 2-methacryloyloxyethyl phosphorylcholine (6.5 g, 22 mmol, 2113 eq.) was added to a 2-neck 50 mL round bottom flask equipped with a 3-way stopcock in a dry box. Anhydrous ethanol (23 mL) that had been sparged with nitrogen was added to the flask and dissolved the 2-methacryloyloxyethyl phosphorylcholine. Stock solutions were prepared by dissolving Initiator 1 (85 mg) in ethanol (5.0 mL), copper(II) bromide (100 mg) in ethanol (5.0 mL), ascorbic acid (40 mg) in ethanol (8.0 mL), and 2,2'-bipyridine (130 mg) in ethanol (5.0 mL). The copper(II) bromide stock solution (1.0 mL, 0.09 mmol, 9 eq.) was added to the reaction flask, followed by the ascorbic acid stock solution (1.3 mL, 0.04 mmol, 3.6 eq.), and the 2,2'-bipyridyl stock solution (1.2 mL, 0.19 mmol, 18 eq.). A vial containing 3-azidopropyl methacrylate was degassed with nitrogen for 2 minutes and then 1.5 mL (9.4 mmol, 905 eq.) was added to the reaction flask. The polymerization was started by addition of the initiator stock solution (1.7 mL, 0.01 mmol, 1 eq.). The polymerization was allowed to continue overnight and was terminated after 48 hours by exposing the reaction to air. The final monomer conversion was 89%. The reaction solution was diluted with ethanol (15 mL) and deionized water (35 mL) and then dialyzed against water for 4 days with Spectra / Por® 12 - 14 kDa MWCO dialysis tubing. After dialysis, the aqueous solution was filtered and concentrated under reduced pressure. The solution was stored in the freezer at -20 °C at a concentration of 124 mg / mL. GPC-MALS: Mn= 629 kDa, Ð = 1.05. Copolymer Example 35: Synthesis of 9-arm 10 mol% azide loading copolymer

[0320] Synthetic Scheme:-77-

[0321] 2-methacryloyloxyethyl phosphorylcholine (8.0 g, 27 mmol, 2646 eq.) was added to a 2-neck 50 mL round bottom flask equipped with a 3-way stopcock in a dry box. Anhydrous ethanol (37 mL) that had been sparged with nitrogen was added to the flask and dissolved the 2-methacryloyloxyethyl phosphorylcholine. Stock solutions were prepared by dissolving initiator (85 mg) in ethanol (5.0 mL), copper(II) bromide (100 mg) in ethanol (5.0 mL), ascorbic acid (25 mg) in ethanol (5.0 mL), and 2,2'-bipyridine (130 mg) in ethanol (5.0 mL). The copper(II) bromide stock solution (1.0 mL, 0.09 mmol, 9 eq.) was added to the reaction flask, followed by the ascorbic acid stock solution (1.3 mL, 0.04 mmol, 3.6 eq.) and the 2,2'-bipyridyl stock solution (1.2 mL, 0.18 mmol, 18 eq.). The polymerization was started by addition of the initiator stock solution (1.7 mL, 0.01 mmol, 1 eq.). A vial containing 3- azidopropyl methacrylate was degassed with nitrogen for 10 minutes and then 0.48 mL (3.0 mmol, 294 eq.) was added to the reaction flask 2.5 hours after starting the polymerization. The polymerization was allowed to continue overnight and was terminated after 25 hours by exposing the reaction to air. The final monomer conversion was 86%. The reaction solution was diluted to 75 mL with deionized water and then dialyzed against water for 5 days with Spectra / Por® 12 - 14 kDa MWCO dialysis tubing. After dialysis, the aqueous solution was filtered and concentrated under reduced pressure. The solution was stored in the freezer at -20 °C at a concentration of 130 mg / mL. GPC-MALS: Mn = 556 kDa, Ð = 1.03.

[0322] Copolymer Example 36: Synthesis of 3-arm 10 mol% azide loading copolymer

[0323] Synthetic Scheme:-78-

[0324] 2-methacryloyloxyethyl phosphorylcholine (7.0 g, 24 mmol, 77 eq.) was added to a 2-neck 50 mL round bottom flask equipped with a 3-way stopcock in a dry box. Anhydrous ethanol (13 mL) that had been sparged with nitrogen was added to the flask and dissolved the 2-methacryloyloxyethyl phosphorylcholine. Stock solutions were prepared by dissolving initiator (320 mg) in ethanol (8.0 mL), copper(II) bromide (250 mg) in ethanol (10 mL), ascorbic acid (100 mg) in ethanol (10 mL), and 2,2'-bipyridine (400 mg) in ethanol (4.0 mL). The copper(II) bromide stock solution (8.3 mL, 0.93 mmol, 3 eq.) was added to the reaction flask, followed by the ascorbic acid stock solution (7.3 mL, 0.42 mmol, 1.4 eq.) and the 2,2'-bipyridyl stock solution (2.9 mL, 1.9 mmol, 6 eq.). The polymerization was started by addition of the initiator stock solution (6.8 mL, 0.31 mmol, 1 eq.). A vial containing 3- azidopropyl methacrylate was degassed with nitrogen for 2 minutes and then 0.42 mL (2.6 mmol, 9 eq.) was added to the reaction flask 18.5 hours after starting the polymerization. The polymerization was terminated after 42.5 hours by exposing the reaction to air. The final monomer conversion was 99%. The reaction solution was diluted to 70 mL with deionized water and then dialyzed against water for 5 days with Spectra / Por® 1 kDa MWCO dialysis tubing. After dialysis, the aqueous solution was filtered and concentrated under reduced pressure. The solution was stored in the freezer at -20 °C at a concentration of 199 mg / mL. GPC-MALS: Mn= 32.9 kDa, Ð = 1.14. Intermediate Example 1: Synthesis of 7,8-Didehydro-2,3,4,5,6,9-hexahydro-1,5-oxazonine

[0325] Synthetic Scheme:-79-

[0326] Step 1: Synthesis of but-2-yne-1,4-diol cobalt complex

[0327] To a stirred mixture of but-2-yne-1,4-diol (10.0 g, 116.0 mmol) in CH2Cl2 (4.0 L) was added dicobalt octacarbonyl (41.7 g, 122.1 mmol) in portions at rt. The resulting mixture was stirred at rt overnight. The reaction mixture was then concentrated by rotary evaporation under reduced pressure. The residue was purified by silica gel column chromatography eluting with EA / CH2Cl2 gradients (0-30%) to afford but-2-yne-1,4-diol cobalt complex as a brownish solid (36.0 g, 83.1% yield).1H NMR (400 MHz, CDCl3) δ 4.86 (s, 4H), 2.54 (s, 2H) ppm.

[0328] Step 2: Synthesis of N-(3-hydroxypropyl)-2-nitrobenzenesulfonamide

[0329] To a solution of 3-aminopropan-1-ol (10.0 g, 133.1 mmol) in anhydrous CH2Cl2(500 mL) was added pyridine (21.0 g, 266.2 mmol) and 2-nitrobenzenesulfonyl chloride (35.4 g, 159.7 mmol) in portions at 0 °C under an atmosphere of N2. The resulting mixture was stirred at 0-5 °C for 1 hour. The reaction mixture was then concentrated by rotary evaporation under reduced pressure and the residue was subjected to silica gel column chromatography purification eluting with MeOH / CH2Cl2(0-1%) to afford N-(3- hydroxypropyl)-2-nitrobenzenesulfonamide as a pale-yellow oil (10.6 g, 30.2% yield).1H NMR (400 MHz, CD3OD) δ 8.12 – 8.04 (m, 1H), 7.89 – 7.84 (m, 1H), 7.83 – 7.78 (m, 2H), 3.58 (t, J = 6.1 Hz, 2H), 3.14 (t, J = 6.9 Hz, 2H), 1.71 (p, J = 6.5 Hz, 2H) ppm. LC-MS: mass calcd for C9H12N2O5S: 260.05, found: m / z = 260.90 [M+H]+.-80-

[0330] Step 3: Synthesis of 5-[(2-Nitrophenyl)sulfonyl]-7,8-didehydro-2,3,4,5,6,9- hexahydro-1,5-oxazonine cobalt complex

[0331] To a mixture of N-(3-hydroxypropyl)-2-nitrobenzenesulfonamide (10.6 g, 40.7 mmol) and but-2-yne-1,4-diol cobalt complex (11.5 g, 30.9 mmol) in anhydrous CH2Cl2 (2.0 L) was added boron trifluoride diethyl etherate (10.5 g, 74.0 mmol) in portions at 0 °C under an atmosphere of N2. The resulting mixture was allowed to warm to rt and stirred for 1 hour. The reaction was quenched by sat. NaHCO3(aq., 500 mL) and the resulting mixture was extracted with CH2Cl2 (3 x 500 mL). The combined organic layers were dried over Na2SO4, filtered, and concentrated by rotary evaporation under reduced pressure. The residue was purified by silica gel column chromatography eluting with CH2Cl2to afford 5-[(2- nitrophenyl)sulfonyl]-7,8-didehydro-2,3,4,5,6,9-hexahydro-1,5-oxazonine cobalt complex as a dark-brown solid (14.0 g, 76% yield).1H NMR (400 MHz, CD3OD) δ 8.08 – 8.04 (m, 1H), 7.88 – 7.79 (m, 3H), 5.04 (s, 2H), 4.85 (s, 2H), 3.94 (t, J = 5.4 Hz, 2H), 3.57 (t, J = 5.9 Hz, 2H), 2.05 – 1.96 (m, 2H) ppm.

[0332] Step 4: Synthesis of 5-[(2-nitrophenyl)sulfonyl]-7,8-didehydro-2,3,4,5,6,9- hexahydro-1,5-oxazonine

[0333] To a stirred mixture of 5-[(2-nitrophenyl)sulfonyl]-7,8-didehydro- 2,3,4,5,6,9-hexahydro-1,5-oxazonine cobalt complex (5.0 g, 8.4 mmol) in anhydrous CH2Cl2 (1.0 L) was added ceric ammonium nitrate (25.6 g, 46.7 mmol) and silica gel (100.0 g, 100~200 mesh) in portions at rt. The resulting mixture was allowed to stir at rt for 16 h. The mixture was then filtered through silica gel, washed with 25% EA in CH2Cl2. The filtrate was concentrated by rotary evaporation under reduced pressure. The residue was purified by silica gel column chromatography eluting with EA / Hex gradients (10~30%) to afford 5-[(2- nitrophenyl)sulfonyl]-7,8-didehydro-2,3,4,5,6,9-hexahydro-1,5-oxazonine as a white solid (1.2 g, 46.1% yield).1H NMR (400 MHz, CDCl3) δ 8.03 – 7.96 (m, 1H), 7.78 – 7.67 (m, 2H), 7.66 – 7.61 (m, 1H), 4.15 – 4.08 (m, 4H), 3.87 (t, J = 5.3 Hz, 2H), 3.53 (t, J = 5.6 Hz, 2H), 1.96 (p, J = 5.4 Hz, 2H) ppm. LC-MS: mass calcd for C13H14N2O5S: 310.06, found: m / z = 310.85 [M+H]+.

[0334] Step 5: Synthesis of 7,8-didehydro-2,3,4,5,6,9-hexahydro-1,5-oxazonine.

[0335] To a stirred mixture of 5-[(2-nitrophenyl)sulfonyl]-7,8-didehydro- 2,3,4,5,6,9-hexahydro-1,5-oxazonine (1.0 g, 3.2 mmol) in anhydrous ACN (20 mL) was added-81-Cs2CO3 (3.2 g, 9.8 mmol) and p-toluenethiol (1.2 g, 9.7 mmol) in portions at rt. The reaction mixture was stirred at rt for 16 h under an atmosphere of N2. The mixture was diluted with diethyl ether (30 mL). Then the pH of the mixture was adjusted to 2~3 with 1M HCl (aq., ~30 mL). The resulting mixture was extracted with Et2O (3 x 30 mL). The aqueous layer was basified to pH 13~14 and extracted with CH2Cl2 (3 x 30 mL). All organic layers were combined, dried over Na2SO4, and concentrated by rotary evaporation under reduced pressure to afford 7,8-didehydro-2,3,4,5,6,9-hexahydro-1,5-oxazonine as a dark-brownish solid (300.8 mg, 74.2% yield).1H NMR (400 MHz, CDCl3) δ 4.18 – 4.11 (m, 2H), 3.90 – 3.82 (m, 2H), 3.47 – 3.40 (m, 2H), 3.05 – 2.97 (m, 2H), 1.84 – 1.67 (m, 2H) ppm. Intermediate Example 2: Synthesis of 2,5-dioxopyrrolidin-1-yl 4-[7,8-didehydro-3,4,6,9- tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutanoate

[0336] Scheme:

[0337] Step 1: Synthesis of 4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin- 5(2H)-yl]-4-oxobutanoic acid

[0338] In a 100 mL round bottom flask, 7,8-didehydro-2,3,4,5,6,9-hexahydro-1,5- oxazonine (260.0 mg, 2.1 mmol), succinic anhydride (416.0 mg, 4.2 mmol) were dissolved in CH2Cl2(10 mL), before the addition of TEA (1.2 mL, 8.4 mmol). The resulting mixture was allowed to stir at rt for 2 hours. The mixture was concentrated by rotary evaporation under reduced pressure, and re-dissolved in 1 M HCl (aq., 30 mL). The resulting mixture was extracted with EA (3 x 20 mL). The combined organic layer was dried over anhydrous Na2SO4, filtered, and concentrated by rotary evaporation under reduced pressure. The crude was further purified with silica gel column chromatography purification eluting with MeOH / CH2Cl2 gradients (0-7%) to afford 4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-4- oxobutanoic acid as a pale yellowish solid (382.3 mg, 82.0% yield).1H NMR (600 MHz, CD3OD) δ 4.24 (dt, J = 8.9, 2.5 Hz, 2H), 4.16 – 4.10 (m, 2H), 3.86 – 3.80 (m, 1H), 3.79 – 3.75-82-(m, 1H), 3.74 – 3.69 (m, 1H), 3.62 – 3.57 (m, 1H), 2.69 – 2.62 (m, 3H), 2.61 – 2.57 (m, 1H), 2.07 – 2.01 (m, 1H), 1.95 – 1.88 (m, 1H) ppm.

[0339] Step 2: Synthesis of 2,5-dioxopyrrolidin-1-yl 4-[7,8-didehydro-3,4,6,9- tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutanoate

[0340] In a 100 mL round bottom flask, 4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]-4-oxobutanoic acid (200.0 mg, 0.89 mmol), N, N′-disuccinimidyl carbonate (341.1 mg, 1.3 mmol) were dissolved in CH2Cl2(10 mL), before the addition of DIEA (0.23 ml, 1.3 mmol). The resulting mixture was allowed to stir at rt for 1 hour. The mixture was concentrated by rotary evaporation under reduced pressure, then further purified with silica gel column chromatography purification eluting with EA / Hexane gradients (50- 100%) to afford 2,5-dioxopyrrolidin-1-yl 4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin- 5(2H)-yl]-4-oxobutanoate as a white solid (260.1 mg, 90.2% yield).1NMR (600 MHz, CDCl3) δ 4.21 (t, J = 2.6 Hz, 1H), 4.11 – 3.99 (m, 3H), 3.78 – 3.64 (m, 2H), 3.61 – 3.47 (m, 2H), 2.97 – 2.85 (m, 2H), 2.76 (s, 4H), 2.68 (t, J = 6.7 Hz, 2H), 1.94 – 1.85 (m, 2H) ppm. Intermediate Example 3: Synthesis of (S)-3-((tert-butoxycarbonyl)amino)-3-carboxypropyl (2-(trimethylammonio)ethyl) phosphate-83-

[0342] Step 1: Synthesis of benzyl N-(tert-butoxycarbonyl)-O-(2-oxido-1,3,2- dioxaphospholan-2-yl)-L-homoserinate.

[0343] To a dried and nitrogen-purged flask was added benzyl (tert- butoxycarbonyl)-L-homoserinate (2.0 g, 6.5 mmol) in anhydrous THF (20 mL). The flask was evacuated and refilled with N2 three times. TEA (2.6 g, 25.9 mmol) was added via a syringe. The reaction mixture was cooled to -78 °C using a dry ice / acetone bath, and 2-chloro-1,3,2- dioxaphospholane 2-oxide (3.7 g, 25.9 mmol) was added dropwise. The mixture was stirred at -78 °C for 2 h, during which TLC indicated completion of the reaction. The mixture was filtered through a Celite pad and eluted with EA (20 mL). The filtrate was directly purified on a silica gel chromatography column, eluting with 100% EA to afford benzyl N-(tert- butoxycarbonyl)-O-(2-oxido-1,3,2-dioxaphospholan-2-yl)-L-homoserinate as a pale-yellow oil (2.6 g, 96.8% yield).1H NMR (400 MHz, CDCl3) δ 7.36 (s, 4H), 5.17 (s, 2H), 4.50 – 4.31 (m, 5H), 4.27 – 4.17 (m, 2H), 2.33 – 2.21 (m, 1H), 2.17 – 2.07 (m, 1H), 1.43 (s, 9H) ppm.

[0344] Step 2: Synthesis of (S)-4-(benzyloxy)-3-((tert-butoxycarbonyl)amino)-4- oxobutyl (2-(trimethylammonio)ethyl) phosphate.

[0345] Benzyl N-(tert-butoxycarbonyl)-O-(2-oxido-1,3,2-dioxaphospholan-2-yl)- L-homoserinatev (2.6 g, 6.3 mmol) was dissolved in anhydrous ACN (5.2 mL), then TMA (38 mL, 76.0 mmol, 2.0 N in ACN) was added. The reactor was purged with N2, sealed, and then heated in an oil bath at 70 °C with stirring for 16 h. The progress of the reaction was monitored by LC-MS. Upon completion of the reaction, the mixture was filtered through a Celite pad and washed with ACN (50 mL). The filtrate was evaporated to dryness under reduced pressure. The resulting solid residue was triturated with PE (10 mL) at rt for 1 h, filtered, and the solid was collected and dried to afford (S)-4-(benzyloxy)-3-((tert-butoxycarbonyl)amino)-4- oxobutyl (2-(trimethylammonio)ethyl) phosphate as a pale-yellow solid (2.1 g, 74.1% yield). LC / MS: mass calcd for C21H35N2O8P: 474.21, found: m / z = 475.05 [M+H]+.1H NMR (400 MHz, D2O) δ 7.46 (s, 5H), 5.25 (s, 2H), 4.34 – 4.21 (m, 3H), 4.06 – 3.86 (m, 3H), 3.55 (s, 2H), 3.16 (s, 9H), 2.26 – 2.16 (m, 1H), 2.06 – 1.97 (m, 1H), 1.49 – 1.28 (m, 9H) ppm.

[0346] Step 3: Synthesis of (S)-3-((tert-butoxycarbonyl)amino)-3-carboxypropyl (2-(trimethylammonio)ethyl) phosphate

[0347] To a solution of (S)-4-(benzyloxy)-3-((tert-butoxycarbonyl)amino)-4- oxobutyl (2-(trimethylammonio)ethyl) phosphate (600.2 mg, 1.3 mmol) in MeOH (6 mL) was-84-added Pd / C (60 mg, 10%) under a nitrogen atmosphere. The mixture was purged with H2 and stirred at 35 °C for 2 h under an atmosphere of H2 (1 atm). The mixture was cooled to rt, filtered through a Celite cake, and washed with MeOH (20 mL). The filtrate was concentrated under reduced pressure by rotary evaporation to afford (S)-3-((tert-butoxycarbonyl)amino)-3- carboxypropyl (2-(trimethylammonio)ethyl) phosphate as a grey solid (480.0 mg, 99.3% yield).1H NMR (400 MHz, D2O) δ 4.19 – 4.13 (m, 2H), 3.91 – 3.78 (m, 3H), 3.56 – 3.49 (m, 2H), 3.08 (s, 9H), 2.07 – 1.91 (m, 1H), 1.83 – 1.70 (m, 1H), 1.29 (s, 9H) ppm. Intermediate Example 4: Synthesis of (3S)-3-amino-4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate hydrochloride

[0349] Step 1: Synthesis of (3S)-3-[(tert-butoxycarbonyl)amino]-4-[7,8- didehydro-3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate.

[0350] To a solution of (S)-3-((tert-butoxycarbonyl)amino)-3-carboxypropyl (2- (trimethylammonio)ethyl) phosphate (1.6 g, 4.1 mmol) in DMF (16 mL), DIEA (1.5 g, 11.2 mmol), 7,8-didehydro-2,3,4,5,6,9-hexahydro-1,5-oxazonine (470.0 mg, 3.8 mmol), HOBt monohydrate (1.2 g, 7.5 mmol) and EDCI (1.4 g, 7.5 mmol) were added sequentially at rt, and the mixture was stirred at rt overnight. LC-MS monitoring indicated completion of the reaction. The mixture was concentrated under reduced pressure. The crude reaction mixture was purified using preparative RP-LC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–50% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford (3S)-3-[(tert-butoxycarbonyl)amino]-4-[7,8-didehydro-3,4,6,9- tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate as a-85-white solid (1.3 g, 70.4% yield). LC / MS: mass calcd for C21H38N3O8P: 491.24, found: m / z = 492.35 [M+H]+.1H NMR (400 MHz, CD3OD) δ 4.70 – 4.60 (m, 1H), 4.44 – 4.25 (m, 4H), 4.24 – 4.04 (m, 3H), 3.99 (q, J = 6.5 Hz, 2H), 3.90 – 3.73 (m, 3H), 3.70 – 3.59 (m, 3H), 3.25 (s, 9H), 2.22 – 1.85 (m, 4H), 1.46 (d, J = 3.9 Hz, 9H) ppm.

[0351] Step 2: Synthesis of (3S)-3-amino-4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate hydrochloride.

[0352] To a solution of (3S)-3-[(tert-butoxycarbonyl)amino]-4-[7,8-didehydro- 3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate (400.0 mg, 0.8 mmol) in MeOH (4 mL) was added HCl (4 mL, 8.0 mmol, 2.0 N in methanol) at 0 °C. The mixture was stirred at rt for 2 h. Reaction progress was monitored by LC-MS, after which the mixture was concentrated under reduced pressure and lyophilized to afford (3S)-3-amino-4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutyl 2- (trimethylazaniumyl)ethyl phosphate hydrochloride as a yellow solid (317.1 mg, 91.2% yield). LC / MS: mass calcd for C16H30N3O6P: 391.19, found: m / z = 392.27 [M+H]+. “Z2-R’-L-PL” Modified Payload Example 1: Synthesis of DBCO-Tirzepatide

[0354] Step 1: Synthesis of DBCO-Tirzepatide “Z2-R’-L-PL” Modified Payloads

[0355] Tirzepatide (50.0 mg, 10.4 μmol) was dissolved in DMSO (2 mL). 2,5- Dioxopyrrolidin-1-yl 6-[11,12-didehydrodibenzo[b,f]azocin-5(6H)-yl]-6-oxohexanoate (5.4 mg, 12.5 μmol) was added, followed by DIEA (2.7 mg, 20.8 μmol). The mixture was stirred-86-at rt with reaction progress monitored by LC-MS. The crude was diluted with CH3OH (2 mL) and purified using preparative RP-LC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (30–100% ACN in 35 min). Fractions corresponding to the mono- conjugated product (tR = 14.6 min) and the di-conjugated product (tR = 22.7 min) were collected and lyophilized to give mono-DBCO-Tirzepatide as a white solid (30.8 mg, 57.8% yield), and di-DBCO-Tirzepatide as a white solid (2.9 mg, 5.1% yield). Mono-DBCO– Tirzepatide LC-MS: mass calcd for C246H365N49O70: 5125.65, found: m / z = 1709.33 [M+3H]3+Di-DBCO–Tirzepatide LC-MS: mass calcd for C267H382N50O72: 5440.78, found: m / z = 1814.47 [M+3H]3+“Z2-R’-L-PL” Modified Payload Example 2: Synthesis of Synthesis of (43R)-12-hydroxy-24- methyl-6,9,12,15-tetraoxo-14-(trifluoromethyl)-167,168-didehydro-163,164,166,169- tetrahydro-162H-8-oxa-3,13-diaza-16(5)-1,5-oxazonina-2(3,6)-pyridazina-4(3,1),7(1,4)- dipiperidina-1(1)-benzenahexadecaphane

[0356] Scheme:-87-

[0357] Step 1: Synthesis of benzyl (1-(tert-butoxycarbonyl)piperidin-4-yl) succinate.

[0358] To a solution of tert-butyl 4-hydroxypiperidine-1-carboxylate (3.0 g, 14.9 mmol) in CH2Cl2 (30 mL) was added 4-(benzyloxy)-4-oxobutanoic acid (3.1 g, 14.9 mmol) and DMAP (181.0 mg, 1.5 mmol) under an atmosphere of N2. The reaction mixture was cooled to 0 °C (ice bath), and EDCI (5.7 g, 29.8 mmol) was added portion wise. The resulting mixture was stirred at rt overnight. Reaction progress was monitored by LC-MS. The mixture was concentrated by rotary evaporation under reduced pressure and the residue was subjected to silica gel column chromatography purification eluting with EA / Hex gradients (0-10%) to afford the title compound benzyl (1-(tert-butoxycarbonyl)piperidin-4-yl) succinate as a pale- yellow oil (5.5 g, 93.8% yield). LC-MS: mass calcd for C21H29NO6: 391.20, found: m / z = 414.23 [M+Na]+.1H NMR (400 MHz, CDCl3) δ 7.42 – 7.32 (m, 5H), 5.15 (s, 2H), 4.98 – 4.90-88-(m, 1H), 3.75 – 3.63 (m, 2H), 3.27 – 3.18 (m, 2H), 2.73 – 2.64 (m, 4H), 1.86 – 1.77 (m, 2H), 1.62 – 1.52 (m, 2H), 1.48 (s, 9H) ppm.

[0359] Step 2: Synthesis of benzyl piperidin-4-yl succinate hydrochloride.

[0360] Benzyl (1-(tert-butoxycarbonyl)piperidin-4-yl) succinate (5.5 g, 14.1 mmol) was dissolved in HCl (50 mL, 200.0 mmol, 4 M in 1,4-dioxane) in a single-neck flask and stirred at rt for 1 h. The reaction progress was monitored by TLC. The reaction mixture was concentrated by rotary evaporation under reduced pressure to afford the crude product benzyl piperidin-4-yl succinate hydrochloride as a white solid (4.5 g, 96.6%). LC-MS: mass calcd for C16H21NO4: 291.15, found: m / z = 292.23 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.41 – 7.28 (m, 5H), 5.15 (s, 2H), 5.06 – 4.99 (m, 1H), 3.32 – 3.26 (m, 2H), 3.23 – 3.15 (m, 2H), 2.77 – 2.63 (m, 4H), 2.12 – 2.00 (m, 2H), 1.95 – 1.85 (m, 2H) ppm.

[0361] Step 3: Synthesis of (R)-benzyl (1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl) succinate.

[0362] Benzyl piperidin-4-yl succinate hydrochloride (780.1 mg, 2.4 mmol) was dissolved in DMF (10 mL) in a single-neck flask. To this solution were added (R)-2-(3-((6-(2- hydroxy-4-(trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetic acid (998.0 mg, 2.4 mmol), DIEA (1.2 g, 9.6 mmol), HOBt monohydrate (648.0 mg, 4.2 mmol), and EDCI (920.0 mg, 4.8 mmol). The reaction mixture was purged with N2and stirred at rt overnight. The mixture was diluted with water (50 mL) and extracted with EA (3 x 50 mL). The combined organic layers were washed with sat. brine (3 x 50 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was subjected to silica gel column chromatography purification eluting with MeOH / CH2Cl2gradients (0-5%) to afford the target compound (R)-benzyl (1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)-5- methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl) succinate as a pale-yellow solid (1.0 g, 61.3% yield). LC-MS: mass calcd for C35H40F3N5O6: 683.73, found: m / z = 684.63 [M+H]+.1H NMR (400 MHz, CDCl3) δ 7.54 (d, J = 8.2 Hz, 1H), 7.37 (s, 6H), 7.16 (d, J = 8.0 Hz, 1H), 6.72 (s, 1H), 5.15 (s, 2H), 5.04 (s, 1H), 4.28 (s, 1H), 3.92 – 3.62 (m, 2H), 3.55 – 3.23 (m, 4H), 2.73 – 2.62 (m, 6H), 2.46 (s, 3H), 1.96 – 1.59 (m, 12H) ppm.-89-

[0363] Step 4: Synthesis of (R)-4-((1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4- yl)oxy)-4-oxobutanoic acid

[0364] (R)-benzyl (1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)-5- methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl) succinate (1.0 g, 1.4 mmol) was dissolved in MeOH (10 mL) in a single-neck flask and purged with N2. Pd / C (10 wt% dry, 50.0 mg) was added, and the mixture was again purged with H2. The reaction mixture was stirred at rt for 45 min under 1 atm H2 atmosphere. The reaction progress was monitored by TLC. The mixture was filtered through Celite and concentrated under reduced pressure to crude (R)-4-((1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)-5-methylpyridazin-3- yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl)oxy)-4-oxobutanoic acid as a yellow solid (0.8 g, 93.0% yield). LC-MS: mass calcd for C28H34F3N5O6: 593.25, found: m / z = 594.42 [M+H]+.

[0365] Step 5: Synthesis of tert-butyl {2-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]-2-oxoethyl}carbamate.

[0366] 7,8-Didehydro-2,3,4,5,6,9-hexahydro-1,5-oxazonine (250.0 mg, 2.0 mmol) was dissolved in DMF (10 mL) in a single-neck flask. To this solution were added HOBt monohydrate (540.2 mg, 3.5 mmol), EDCI (766.0 mg, 4.0 mmol), DIEA (1.0 g, 8.0 mmol), and (tert-butoxycarbonyl)glycine (385.0 mg, 2.2 mmol). The reaction mixture was purged with N2(3 cycles) and stirred at rt for 16 h. Reaction progress was monitored by TLC until complete consumption of the starting material. The reaction mixture was concentrated under reduced pressure and the residue was subjected to silica gel column chromatography purification eluting with 100% CH2Cl2to afford tert-butyl {2-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]-2-oxoethyl}carbamate as a colorless oil (500.0 mg, 88.8% yield).1H NMR (400 MHz, CDCl3) δ 5.44 (s, 1H), 4.30 (t, J = 2.5 Hz, 1H), 4.14 – 4.10 (m, 2H), 4.02 – 3.91 (m, 3H), 3.83 – 3.75 (m, 2H), 3.66 – 3.57 (m, 2H), 2.01 – 1.94 (m, 2H), 1.44 (s, 9H) ppm.

[0367] Step 6: Synthesis of 2-amino-1-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]ethan-1-one hydrochloride.

[0368] tert-Butyl {2-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-2- oxoethyl}carbamate (600.0 mg, 2.1 mmol) was dissolved in HCl (15 mL, 60.0 mmol, 4 M in 1,4-dioxane) in a single-neck flask and stirred at rt for 2 h. The reaction progress was monitored by TLC. The reaction mixture was concentrated by rotary evaporation under reduced pressure-90-and the residue was subjected to silica gel column chromatography purification eluting with MeOH / CH2Cl2 gradients (0-10%) to afford 2-amino-1-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]ethan-1-one hydrochloride as a white solid (430.0 mg, 92.3% yield).1H NMR (400 MHz, CD3OD) δ 4.32 – 4.27 (m, 1H), 4.15 – 4.10 (m, 3H), 3.88 (d, J = 7.4 Hz, 2H), 3.82 – 3.75 (m, 2H), 3.63 (q, J = 6.4 Hz, 2H), 2.02 – 1.90 (m, 2H) ppm.

[0369] Step 7: Synthesis of (43R)-12-hydroxy-24-methyl-6,9,12,15-tetraoxo-14- (trifluoromethyl)-167,168-didehydro-163,164,166,169-tetrahydro-162H-8-oxa-3,13-diaza- 16(5)-1,5-oxazonina-2(3,6)-pyridazina-4(3,1),7(1,4)-dipiperidina-1(1)- benzenahexadecaphane.

[0370] 2-amino-1-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)- yl]ethan-1-one hydrochloride (46.0 mg, 0.2 mmol) was dissolved in DMF (4 mL) in a single- neck flask. To this solution were added HOBt monohydrate (68.0 mg, 0.4 mmol), EDCI (97.0 mg, 0.5 mmol), DIEA (130.0 mg, 1.0 mmol), and (R)-4-((1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4- yl)oxy)-4-oxobutanoic acid (150.0 mg, 0.2 mmol). The mixture was purged with N2 (3 cycles) and stirred at rt for 16 h. The reaction progress was monitored by LC-MS until complete consumption of the starting material. The crude reaction mixture was purified using preparative RP-HPLC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford (43R)- 12-hydroxy-24-methyl-6,9,12,15-tetraoxo-14-(trifluoromethyl)-167,168-didehydro- 163,164,166,169-tetrahydro-162H-8-oxa-3,13-diaza-16(5)-1,5-oxazonina-2(3,6)-pyridazina- 4(3,1),7(1,4)-dipiperidina-1(1)-benzenahexadecaphane as a white solid (46.0 mg, 30.0% yield). LC-MS: mass calcd for C37H46F3N7O7: 757.34, found: m / z = 758.15 [M+H]+.1H NMR (400 MHz, CD3OD) δ 8.33 (s, 1H), 7.42 (d, J = 7.8 Hz, 1H), 7.25 (d, J = 8.8 Hz, 1H), 7.19 (s, 1H), 6.83 (s, 1H), 5.09 – 4.98 (m, 2H), 4.27 (s, 2H), 4.19 – 4.13 (m, 3H), 4.07 (s, 2H), 3.85 – 3.77 (m, 2H), 3.76 – 3.64 (m, 5H), 3.62 – 3.49 (m, 3H), 2.98 (s, 1H), 2.76 – 2.55 (m, 6H), 2.17 (s, 3H), 2.10 – 1.88 (m, 6H), 1.88 – 1.72 (m, 3H), 1.71 – 1.58 (m, 2H) ppm.19F NMR (376 MHz, CD3OD) δ -64.31 (s, 3F) ppm. “Z2-R’-L-PL” Modified Payload Example 3: Synthesis of 1-(2-((R)-3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4--91-methylpiperidin-4-yl 1-((1R,8S,9s)-bicyclo[6.1.0]non-4-yn-9-yl)-3,11-dioxo-2,7-dioxa-4,10- diazatetradecan-14-oate

[0372] Step 1: Synthesis of benzyl 4-chloro-4-oxobutanoate.

[0373] To a solution of 4-(benzyloxy)-4-oxobutanoic acid (1.0 g, 4.8 mmol) in CH2Cl2(10 mL), oxalyl chloride (3.0 g, 24.0 mmol) was added dropwise under an ice-water bath. The mixture was then stirred at rt for 1 h. The reaction mixture was concentrated under-92-reduced pressure to afford benzyl 4-chloro-4-oxobutanoate as a colorless oil (660.1 mg, 60.6% yield).

[0374] Step 2: Synthesis of benzyl (1-(tert-butoxycarbonyl)-4-methylpiperidin-4- yl) succinate.

[0375] To a solution of benzyl 4-chloro-4-oxobutanoate (1.0 g, 4.4 mmol) and tert- butyl 4-hydroxy-4-methylpiperidine-1-carboxylate (900.2 mg, 3.7 mmol) in CHCl3(20 mL), sodium tripolyphosphate (1.4 g, 3.7 mmol) was added, and the reaction was heated at 65 °C for 48 h. Completion of the reaction was confirmed by TLC. The mixture was filtered and concentrated under reduced pressure. The residue was purified by flash chromatography eluting with 100% CH2Cl2to afford benzyl (1-(tert-butoxycarbonyl)-4-methylpiperidin-4-yl) succinate as a colorless oil (660.0 mg, 33.8% yield). LC-MS: mass calcd for C22H31NO6: 405.22, found: m / z = 306.00 [M-Boc+H]+.1H NMR (400 MHz, CDCl3) δ 7.40 – 7.29 (m, 5H), 5.13 (s, 2H), 3.76 (d, J = 13.6 Hz, 2H), 3.02 (t, J = 13.6 Hz, 2H), 2.68 – 2.59 (m, 4H), 2.14 (d, J = 13.5 Hz, 2H), 1.49 (s, 3H), 1.45 (s, 9H) ppm.

[0376] Step 3: Synthesis of benzyl (4-methylpiperidin-4-yl) succinate hydrochloride.

[0377] To a solution of benzyl (1-(tert-butoxycarbonyl)-4-methylpiperidin-4-yl) succinate (200.1 mg, 0.5 mmol) in 1,4-dioxane (1 mL), HCl (2 mL, 8 mmol, 4 M in 1,4- dioxane) was added. The mixture was stirred at rt for 1 h. The reaction mixture was concentrated under reduced pressure to afford benzyl (4-methylpiperidin-4-yl) succinate hydrochloride as a white solid (200.2 mg, 118.5% yield).1H NMR (400 MHz, CD3OD) δ 7.35 – 7.28 (m, 8H), 5.11 (s, 3H), 3.22 – 3.11 (m, 6H), 2.69 – 2.65 (m, 3H), 2.63 – 2.58 (m, 3H), 2.45 – 2.35 (m, 2H), 1.80 – 1.70 (m, 3H), 1.51 (s, 4H) ppm.

[0378] Step 4: Synthesis of (R)-benzyl (1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4- methylpiperidin-4-yl) succinate.

[0379] To a solution of benzyl (4-methylpiperidin-4-yl) succinate hydrochloride (150.3 mg, 0.5 mmol) in DMF (2 mL), DIEA (251.1 mg, 2.0 mmol), (R)-2-(3-((6-(2-hydroxy- 4-(trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetic acid (158.2 mg, 0.5 mmol), HOBt monohydrate (153.5 mg, 1.0 mmol), and EDCI (187.4 mg, 1.0 mmol) were added, and the mixture was stirred at rt for 16 h. The reaction mixture was concentrated-93-under reduced pressure. The residue was purified by flash column chromatography eluting with 0–40% MeOH in CH2Cl2 to afford (R)-benzyl (1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4- methylpiperidin-4-yl) succinate as a yellow solid (157.3 mg, 45.8% yield).NMR (400 MHz, CD3OD) δ 7.40–7.34 (m, 1H), 7.34–7.22 (m, 4H), 7.20 (d, J = 6.8 Hz, 1H), 7.14 (d, J = 1.5 Hz, 1H), 6.75 (s, 1H), 5.10 (s, 2H), 4.18–4.04 (m, 2H), 3.87–3.77 (m, 1H), 3.32 (s, 1H), 3.27–3.16 (m, 2H), 3.08–2.82 (m, 3H), 2.68–2.54 (m, 4H), 2.47–2.32 (m, 1H), 2.31–2.13 (m, 2H), 2.11 (d, J = 4.0 Hz, 3H), 1.99–1.83 (m, 2H), 1.77–1.50 (m, 3H), 1.49–1.43 (m, 3H), 1.42– 1.27 (m, 1H) ppm.19F NMR (376 MHz, CD3OD) δ -64.31 ppm.

[0380] Step 5: Synthesis of (R)-4-((1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4- methylpiperidin-4-yl)oxy)-4-oxobutanoic acid.

[0381] To a single-neck flask were added (R)-benzyl (1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4- methylpiperidin-4-yl) succinate (157.3 mg, 0.2 mmol) and iPrOH (2 mL). The sealed flask was evacuated / backfilled with N2(3 cycles).10 wt% dry Pd / C (20.1 mg) was added, the flask was purged with N2(3 cycles) and then with H2(3 cycles). The mixture was stirred at rt for 1 h under H2 (1 atm). The reaction mixture was filtered through a pad of Celite, the pad was washed with 2 mL of MeOH, and the combined filtrates were concentrated under reduced pressure to afford (R)-4-((1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)-5-methylpyridazin-3- yl)amino)piperidin-1-yl)acetyl)-4-methylpiperidin-4-yl)oxy)-4-oxobutanoic acid as a yellow solid (150.4 mg, 110.3% yield). LC-MS: mass calcd for C29H36F3N5O6: 607.26, found: m / z = 608.15 [M+H]+.1H NMR (400 MHz, CD3OD) δ 8.29 (s, 1H), 7.38 (d, J = 9.9 Hz, 1H), 7.21 (d, J = 9.9 Hz, 1H), 7.15 (s, 1H), 6.80 (s, 1H), 4.29 (s, 1H), 4.23 – 4.14 (m, 1H), 3.98 – 3.83 (m, 9H), 3.85 – 3.61 (m, 3H), 3.36 (d, J = 10.3 Hz, 1H), 3.13 – 2.98 (m, 2H), 2.81 – 2.64 (m, 2H), 2.34 – 2.19 (m, 2H), 2.13 (s, 3H), 2.07 – 1.96 (m, 2H), 1.90 – 1.78 (m, 1H), 1.70 – 1.56 (m, 2H), 1.50 (d, J = 9.2 Hz, 3H), 1.31 (d, J = 9.8 Hz, 1H) ppm.19F NMR (376 MHz, CD3OD) δ -64.30 ppm.

[0382] Step 6: Synthesis of (R)-1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)- 5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4-methylpiperidin-4-yl 2,2-dimethyl- 4,12-dioxo-3,8-dioxa-5,11-diazapentadecan-15-oate.-94-

[0383] To a solution of (R)-4-((1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)- 5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4-methylpiperidin-4-yl)oxy)-4- oxobutanoic acid (145.6 mg, 0.2 mmol) in DMF (2 mL), DIEA (62.1 mg, 0.5 mmol), tert-butyl (2-(2-aminoethoxy)ethyl)carbamate (97.2 mg, 0.5 mmol), and HATU (136.0 mg, 0.4 mmol) were added, and the mixture was stirred at rt for 2 h. LC-MS monitoring indicated complete consumption of the starting material. The reaction was quenched with water and extracted with EA (3 × 10 mL). The combined organic layers were dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by flash column chromatography eluting with 0–4% MeOH in CH2Cl2to afford (R)-1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4- methylpiperidin-4-yl 2,2-dimethyl-4,12-dioxo-3,8-dioxa-5,11-diazapentadecan-15-oate as a yellow solid (127.2 mg, 67.0% yield). LC-MS: mass calcd for C38H54F3N7O8: 793.40, found: m / z = 794.15 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.43 – 7.37 (m, 1H), 7.22 (d, J = 8.6 Hz, 1H), 7.16 (s, 1H), 6.79 (d, J = 5.3 Hz, 1H), 4.25 – 4.11 (m, 2H), 3.94 – 3.84 (m, 1H), 3.53 – 3.41 (m, 5H), 3.40 – 3.32 (m, 3H), 3.20 (t, J = 6.4 Hz, 3H), 3.06 – 2.95 (m, 2H), 2.60 (t, J = 7.3 Hz, 2H), 2.50 (d, J = 6.8 Hz, 2H), 2.41 (s, 1H), 2.31 (d, J = 17.0 Hz, 1H), 2.21 (d, J = 16.4 Hz, 1H), 2.14 (d, J = 2.6 Hz, 3H), 2.00 – 1.80 (m, 2H), 1.75 – 1.58 (m, 2H), 1.52 (d, J = 15.4 Hz, 4H), 1.43 (s, 12H) ppm.19F NMR (376 MHz, CD3OD) δ -64.31 ppm.

[0384] Step 7: Synthesis of (R)-1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)- 5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4-methylpiperidin-4-yl 4-((2-(2- aminoethoxy)ethyl)amino)-4-oxobutanoate hydrochloride.

[0385] To a solution of (R)-1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)-5- methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4-methylpiperidin-4-yl 2,2-dimethyl- 4,12-dioxo-3,8-dioxa-5,11-diazapentadecan-15-oate (125.1 mg, 0.2 mmol) in 1,4-dioxane (1 mL), HCl (1 mL, 4.0 mmol, 4 M in 1,4-dioxane) was added. The mixture was stirred at rt for 1 h. The reaction mixture was concentrated under reduced pressure to afford (R)-1-(2-(3-((6- (2-hydroxy-4-(trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)- 4-methylpiperidin-4-yl 4-((2-(2-aminoethoxy)ethyl)amino)-4-oxobutanoate hydrochloride as a yellow solid (150.0 mg, 130.4 % yield). LC-MS: mass calcd for C33H46F3N7O6: 693.35, found: m / z = 694.15 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.67 – 7.48 (m, 2H), 7.34 (d, J = 9.3 Hz, 1H), 7.27 (s, 1H), 4.42 – 4.33 (m, 2H), 4.22 (s, 1H), 3.95 (s, 1H), 3.81 (d, J = 4.7 Hz,-95-1H), 3.60 – 3.42 (m, 5H), 3.37 (s, 4H), 3.17 – 2.92 (m, 5H), 2.59 (s, 2H), 2.48 (s, 2H), 2.34 (s, 1H), 2.30 (s, 3H), 2.25 (s, 2H), 2.14 – 1.99 (m, 2H), 1.72 (s, 2H), 1.50 (s, 4H) ppm.19F NMR (376 MHz, CD3OD) δ -64.81 ppm.

[0386] Step 8: Synthesis of 1-(2-((R)-3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)- 5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4-methylpiperidin-4-yl 1-((1R,8S,9s)- bicyclo[6.1.0]non-4-yn-9-yl)-3,11-dioxo-2,7-dioxa-4,10-diazatetradecan-14-oate.

[0387] To a solution of (R)-1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)-5- methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)-4-methylpiperidin-4-yl 4-((2-(2- aminoethoxy)ethyl)amino)-4-oxobutanoate hydrochloric acid salt (150.0 mg, 0.2 mmol) in DMF (2 mL), TEA (41.1 mg, 0.4 mmol) and ((1R,8S,9s)-bicyclo[6.1.0]non-4-yn-9-yl)methyl (4-nitrophenyl) carbonate (57.2 mg, 0.2 mmol) were added at 0oC, and the resulting mixture was stirred at rt for 1 h. The crude reaction mixture was purified using preparative RP-HPLC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford 1-(2-((R)-3-((6- (2-hydroxy-4-(trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)- 4-methylpiperidin-4-yl 1-((1R,8S,9s)-bicyclo[6.1.0]non-4-yn-9-yl)-3,11-dioxo-2,7-dioxa- 4,10-diazatetradecan-14-oate as a white solid (63.8 mg, 36.3% yield). LC-MS: mass calcd for C44H58F3N7O8: 869.43, found: m / z = 870.40 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.38 (dd, J = 7.3, 3.8 Hz, 1H), 7.20 (d, J = 7.8 Hz, 1H), 7.14 (s, 1H), 6.77 (s, 1H), 4.18 (s, 1H), 4.12 (d, J = 8.4 Hz, 2H), 3.86 – 3.76 (m, 1H), 3.52 (s, 1H), 3.46 (s, 4H), 3.39 (s, 1H), 3.31 (d, J = 5.5 Hz, 1H), 3.23 (d, J = 6.5 Hz, 2H), 3.12 (s, 2H), 3.03 – 2.95 (m, 1H), 2.79 (s, 1H), 2.57 (d, J = 6.7 Hz, 2H), 2.47 (t, J = 6.5 Hz, 3H), 2.30 (d, J = 13.6 Hz, 1H), 2.20 (d, J = 15.6 Hz, 6H), 2.13 – 2.08 (m, 4H), 2.02 – 1.81 (m, 3H), 1.79 – 1.63 (m, 2H), 1.63 – 1.54 (m, 3H), 1.50 (d, J = 13.7 Hz, 4H), 1.45 – 1.25 (m, 3H), 0.95 – 0.86 (m, 2H) ppm.19F NMR (376 MHz, CD3OD) δ -64.31 ppm. “Z2-R’-L-PL” Modified Payload Example 4: Synthesis of (3S)-4-[7,8-didehydro-3,4,6,9- tetrahydro-1,5-oxazonin-5(2H)-yl]-3-({4-[(1-{[(3R)-3-({6-[2-hydroxy-4- (trifluoromethyl)phenyl]-5-methylpyridazin-3-yl}amino)piperidin-1-yl]acetyl}piperidin-4- yl)oxy]-4-oxobutanoyl}amino)-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate-96-

[0389] Step 1: Synthesis of (R)-2,5-dioxopyrrolidin-1-yl (1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl) succinate.

[0390] To a solution of (R)-4-((1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)- 5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl)oxy)-4-oxobutanoic acid (200.0 mg, 0.3 mmol) in THF (3 mL), DCC (123.1 mg, 0.6 mmol), DMAP (4.0 mg, 0.03 mmol), and HOSu (35.6 mg, 0.3 mmol) were added sequentially under an atmosphere of N2 after 3 purge cycles. The mixture was stirred at rt for 3 h. Reaction progress was monitored by TLC to completion. The mixture was then filtered, the filtrate was concentrated under reduced pressure, and the residue was dissolved in CH2Cl2 and purified on a silica gel column, elution with 1% MeOH in CH2Cl2to provide (R)-2,5-dioxopyrrolidin-1-yl (1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl) succinate as an off-white oil (200.5 mg, 85.1% yield). LC-MS: mass calcd for C32H37F3N6O8: 690.26, found: m / z = 691.22 [M+H]+.-97-

[0391] Step 2: Synthesis of (3S)-4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]-3-({4-[(1-{[(3R)-3-({6-[2-hydroxy-4-(trifluoromethyl)phenyl]-5- methylpyridazin-3-yl}amino)piperidin-1-yl]acetyl}piperidin-4-yl)oxy]-4- oxobutanoyl}amino)-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate.

[0392] To a single-neck flask were added (3S)-3-amino-4-[7,8-didehydro-3,4,6,9- tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate hydrochloride (114.1 mg, 0.3 mmol) and DMF (3 mL) to afford a homogeneous solution. TEA (58.0 mg, 0.6 mmol) was added, and the mixture was cooled in an ice / water bath. (R)-2,5- Dioxopyrrolidin-1-yl (1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)-5-methylpyridazin- 3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl) succinate (200.0 mg, 0.3 mmol) was added, the cooling bath was removed, and the reaction was stirred at rt for 2 h. The crude reaction mixture was purified using preparative RP-HPLC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford (3S)-4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]-3-({4-[(1-{[(3R)-3-({6-[2-hydroxy-4-(trifluoromethyl)phenyl]-5- methylpyridazin-3-yl}amino)piperidin-1-yl]acetyl}piperidin-4-yl)oxy]-4- oxobutanoyl}amino)-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate as a white solid (63.8 mg, 36.3% yield). LC-MS: mass calcd for C44H62F3N8O11P: 966.42, found: m / z = 967.37 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.42 (d, J = 7.9 Hz, 1H), 7.25 (dd, J = 8.0, 1.7 Hz, 1H), 7.20 (d, J = 1.7 Hz, 1H), 6.83 (d, J = 2.3 Hz, 1H), 5.09 – 4.98 (m, 1H), 4.91 (s, 2H), 4.41 – 4.17 (m, 5H), 4.14 (d, J = 3.0 Hz, 2H), 3.98 – 3.89 (m, 2H), 3.89 – 3.69 (m, 5H), 3.67 – 3.47 (m, 7H), 3.24 (s, 9H), 3.03 – 2.90 (m, 1H), 2.71 – 2.50 (m, 6H), 2.18 – 2.15 (m, 3H), 2.14 – 1.73 (m, 10H), 1.70 – 1.56 (m, 2H) ppm.31P NMR (162 MHz, CD3OD) δ -0.49 (d, J = 13.0 Hz) ppm. “Z2-R’-L-PL” Modified Payload Example 5: Synthesis of (R)-1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl 5-((4-(6-methyl-1,2,4,5-tetrazin-3-yl)benzyl)amino)-5-oxopentanoate-98-

[0394] Step 1: Synthesis of 1-((benzyloxy)carbonyl)piperidin-4-yl tert-butyl glutarate.

[0395] Benzyl 4-hydroxypiperidine-1-carboxylate (5.0 g, 21.2 mmol) was dissolved in DCM (100 mL) in a single-neck flask. To this solution were added 5-(tert-butoxy)- 5-oxopentanoic acid (6.0 g, 31.8 mmol), DCC (13.2 g, 63.5 mmol), and DMAP (259.1 mg, 2.2 mmol). The reaction mixture was stirred at 40 °C overnight. Reaction progress was monitored by TLC. The mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was subjected to silica gel column chromatography purification eluting with EA / Hex gradients (0-25%) to afford 1-((benzyloxy)carbonyl)piperidin-4-yl tert-butyl glutarate as a colorless oil (10.2 g).1H NMR (400 MHz, CDCl3) δ 7.39 – 7.29 (m, 5H), 5.13 (s, 2H), 4.99 – 4.91 (m, 1H), 3.80 – 3.71 (m, 2H), 3.39 – 3.29 (m, 2H), 2.47 (t, J = 7.2 Hz, 1H), 2.36 (t, J = 7.5 Hz, 2H), 2.32 – 2.23 (m, 3H), 1.96 – 1.88 (m, 4H), 1.44 (s, 9H) ppm.

[0396] Step 2: Synthesis of tert-butyl piperidin-4-yl glutarate.-99-

[0397] 1-((Benzyloxy)carbonyl)piperidin-4-yl tert-butyl glutarate (10.0 g, 53.1 mmol) was dissolved in MeOH (100 mL) in a single-neck flask and purged with N2 (3 cycles). Pd / C (1.0 g, 10 wt% dry) was added under N2The mixture was evacuated and backfilled with H2 (3 cycles) and stirred at 50 °C overnight. Reaction progress was monitored by TLC until complete consumption of the starting material. The reaction mixture was filtered through a pad of Celite, and the filtrate was concentrated under reduced pressure. The residue was subjected to silica gel column chromatography purification eluting with EA / Hex gradients (0-50%) to afford tert-butyl piperidin-4-yl glutarate as a pale-yellow oil (3.0 g, 52.7% yield over two steps).1H NMR (400 MHz, CDCl3) δ 4.88 – 4.80 (m, 1H), 3.09 – 3.00 (m, 2H), 2.75 – 2.65 (m, 2H), 2.33 (t, J = 7.5 Hz, 2H), 2.25 (t, J = 7.3 Hz, 2H), 1.92 – 1.84 (m, 4H), 1.59 – 1.49 (m, 2H), 1.43 (s, 9H) ppm.

[0398] Step 3: Synthesis of (R)-tert-butyl (1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl) glutarate

[0399] (R)-2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)-5-methylpyridazin-3- yl)amino)piperidin-1-yl)acetic acid (300.0 mg, 0.7 mmol) was dissolved in pyridine (10 mL) in a single-neck flask. To this solution were added tert-butyl piperidin-4-yl glutarate (298.0 mg, 1.1 mmol) and EDCI (280.2 mg, 1.5 mmol). The mixture was stirred at 50 °C for 16 h, and the reaction was monitored by LC-MS until complete consumption of the starting material. The mixture was concentrated under reduced pressure, and the residue was subjected to silica gel column chromatography purification eluting with CH3OH / CH2Cl2 gradients (0-3%) to afford (R)-tert-butyl (1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)-5-methylpyridazin- 3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl) glutarate as a yellow oil (330.0 mg, 67.9% yield). LC-MS: mass calcd for C33H44F3N5O6: 663.32, found: m / z = 664.30 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.38 (d, J = 9.6 Hz, 1H), 7.21 (d, J = 6.3 Hz, 1H), 7.15 (s, 1H), 6.76 (d, J = 6.6 Hz, 1H), 5.01 - 4.96 (m, 1H), 4.12 (s, 1H), 4.05 - 4.01 (m, 1H), 3.86 – 3.78 (m, 2H), 3.76 - 3.69 (m, 1H), 3.55 (s, 1H), 3.44 (s, 1H), 3.27 – 3.21 (m, 2H), 3.19 – 3.06 (m, 1H), 2.45 – 2.34 (m, 4H), 2.29 - 2.23 (m, 4H), 2.12 (d, J = 4.0 Hz, 2H), 1.86 (s, 2H), 1.84 - 1.80 (m, 4H), 1.43 (s, 9H) ppm.-100-

[0400] Step 4: Synthesis of (R)-5-((1-(2-(3-((6-(2-hydroxy-4- (trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4- yl)oxy)-5-oxopentanoic acid hydrochloride.

[0401] (R)-tert-butyl (1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)-5- methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl) glutarate (330.1 mg, 0.5 mmol) was dissolved in HCl (10 mL, 40.0 mmol, 4 M in 1,4-dioxane) in a single-neck flask and stirred at rt for 16 h. The reaction progress was monitored by TLC. The reaction mixture was concentrated by rotary evaporation under reduced pressure to afford (R)-5-((1-(2-(3-((6- (2-hydroxy-4-(trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1- yl)acetyl)piperidin-4-yl)oxy)-5-oxopentanoic acid hydrochloride as a white solid (330.1 mg, 102.5% yield). LC-MS: mass calcd for C29H36F3N5O6: 607.26, found: m / z = 608.25 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.59 – 7.50 (m, 2H), 7.34 (d, J = 8.3 Hz, 1H), 7.27 (s, 1H), 5.01 (s, 1H), 4.09 – 4.01 (m, 1H), 4.00 – 3.93 (m, 1H), 3.85 – 3.79 (m, 3H), 3.56 – 3.40 (m, 2H), 3.40 – 3.32 (m, 1H), 3.28 – 3.21 (m, 1H), 3.15 – 3.06 (m, 2H), 2.38 (s, 6H), 2.32 – 2.29 (m, 3H), 1.88 – 1.84 (m, 4H), 1.74 – 1.57 (m, 2H), 1.51 – 1.30 (m, 2H) ppm.19F NMR (376 MHz, CD3OD) δ -64.79 (s, 3F) ppm.

[0402] Step 5: Synthesis of (R)-1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)- 5-methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl 5-((4-(6-methyl-1,2,4,5- tetrazin-3-yl)benzyl)amino)-5-oxopentanoate.

[0403] (R)-5-((1-(2-(3-((6-(2-hydroxy-4-(trifluoromethyl)phenyl)-5- methylpyridazin-3-yl)amino)piperidin-1-yl)acetyl)piperidin-4-yl)oxy)-5-oxopentanoic acid hydrochloride (320.0 mg, 0.5 mmol) was dissolved in pyridine (10 mL) in a single-neck flask. To this solution were added (4-(6-methyl-1,2,4,5-tetrazin-3-yl)phenyl)methanamine (159.1 mg, 0.8 mmol) and EDCI (202.2 mg, 1.1 mmol). The mixture was stirred at 50 °C for 16 h, and the reaction progress was monitored by LC-MS until complete consumption of the starting material. The crude reaction mixture was purified using preparative RP-HPLC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford (R)-1-(2-(3-((6-(2- hydroxy-4-(trifluoromethyl)phenyl)-5-methylpyridazin-3-yl)amino)piperidin-1- yl)acetyl)piperidin-4-yl 5-((4-(6-methyl-1,2,4,5-tetrazin-3-yl)benzyl)amino)-5-oxopentanoate as a red solid (94.1 mg, 23.8% yield). LC-MS: mass calcd for C39H45F3N10O5: 790.35, found:-101-m / z = 791.30 [M+H]+.1H NMR (400 MHz, CD3OD) δ 8.48 (d, J = 8.2 Hz, 2H), 8.35 (s, 1H), 7.53 (d, J = 8.0 Hz, 2H), 7.38 (d, J = 7.9 Hz, 1H), 7.22 (d, J = 8.0 Hz, 1H), 7.16 (s, 1H), 6.81 (s, 1H), 4.47 (s, 2H), 4.27 (s, 1H), 3.76 – 3.61 (m, 4H), 3.53 – 3.42 (m, 2H), 3.36 – 3.31 (m, 1H), 3.07 (s, 1H), 3.01 (d, J = 3.5 Hz, 3H), 2.80 – 2.59 (m, 2H), 2.44 – 2.28 (m, 5H), 2.13 (s, 3H), 2.08 – 1.90 (m, 5H), 1.88 – 1.78 (m, 2H), 1.75 – 1.67 (m, 1H), 1.65 – 1.55 (m, 2H) ppm.19F NMR (376 MHz, CD3OD) δ -64.27 (s, 3F) ppm. “Z2-R’-L-PL” Modified Payload Example 6: Synthesis of N-[(4-{[({[5-({4-[7,8-didehydro- 3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutanoyl}amino)-3,3-dimethyl-1- (phenylsulfonyl)pentan-2-yl]oxy}carbonyl)amino]methyl}-5-methylfuran-2-yl)(oyl)-λ6- sulfananimidoyl]-N'-(1,2,3,5,6,7-hexahydros-indacen-4-yl)urea

[0404] Scheme:-102-

[0405] Step 1: Synthesis of methyl 4-chloro-2,2-dimethylbutanoate.

[0406] To a 100 mL RBF was added LDA (18 mL, 18.0 mmol, 1.0 M in THF) at - 78 °C, followed by the slow addition of methyl isobutyrate (1.5 g, 15.0 mmol). The mixture was stirred at -78 °C for 20 min and 1-bromo-2-chloroethane (2.4 g, 16.5 mmol) was added. The mixture was slowly warmed to rt and stirred for 16 h. The reaction mixture was slowly quenched at 0 °C (ice bath) with 1M HCl (aq., 50 mL) and extracted with EA (3 x 100 mL). The combined organic phases were dried over Na2SO4, filtered, and concentrated by rotary evaporation under reduced pressure. The residue was subjected to silica gel column-103-chromatography purification eluting with EA / Hex gradients (0-5%) to afford methyl 4-chloro- 2,2-dimethylbutanoate as a light-yellow liquid (1.0 g, 41.1% yield).1H NMR (600 MHz, CDCl3) δ 3.68 (s, 3H), 3.52 – 3.46 (m, 2H), 2.08 – 2.02 (m, 2H), 1.22 (s, 6H) ppm.

[0407] Step 2: Synthesis of methyl 4-azido-2,2-dimethylbutanoate.

[0408] To a 75 mL sealed tube were added methyl 4-chloro-2,2-dimethylbutanoate (1.0 g, 6.1 mmol), sodium azide (520.1 mg, 8.0 mmol) and DMSO (15 mL). The mixture was stirred at 70 °C for 16 h. The mixture was cooled rt and diluted with EA (100 mL) and washed with water (3 x 50 mL) and dried over Na2SO4. After filtration, the filtrate was concentrated by rotary evaporation under reduced pressure and the residue was subjected to silica gel column chromatography purification eluting with EA / Hex gradients (0-5%) to afford methyl 4-azido-2,2-dimethylbutanoate as a light-yellow liquid (875.3 mg, 84.0% yield).1H NMR (600 MHz, CDCl3) δ 3.68 (s, 3H), 3.29 – 3.23 (m, 2H), 1.88 – 1.81 (m, 2H), 1.21 (s, 6H) ppm.

[0409] Step 3: Synthesis of 5-azido-3,3-dimethyl-1-(phenylsulfonyl)pentan-2-one.

[0410] To a 50 mL RBF was added methyl phenyl sulfone (531.0 mg, 3.4 mmol) in THF (18 mL) at -78 °C followed by the slow addition of LDA (4.4 mL, 4.4 mmol, 1.0 M in THF). The mixture was stirred at -78 °C for 20 min and methyl 4-azido-2,2-dimethylbutanoate (580.0 mg, 3.4 mmol) in THF (2 mL) was added. The mixture was stirred at -78 °C for 2 h and rt for 3 h. The mixture was slowly quenched at 0 °C (ice bath) with 1M HCl (aq., 20 mL) and extracted with EA (3 x 50 mL). The combined organic phases were dried over Na2SO4, filtered, and concentrated by rotary evaporation under reduced pressure and the residue was subjected to silica gel column chromatography purification eluting with EA / Hex gradients (5- 30%) to afford a mixture of 5-azido-3,3-dimethyl-1-(phenylsulfonyl)pentan-2-one (70 wt%) and methyl phenyl sulfone (30 wt%) as a white solid (572.2 mg).1H NMR (600 MHz, CDCl3) δ 7.95 – 7.88 (m, 2H), 7.66 – 7.60 (m, 1H), 7.57 – 7.50 (m, 2H), 4.36 (s, 2H), 3.18 (t, J = 7.2 Hz, 2H), 1.74 (t, J = 7.2 Hz, 2H), 1.11 (s, 6H) ppm.

[0411] Step 4: Synthesis of 5-azido-3,3-dimethyl-1-(phenylsulfonyl)pentan-2-ol.

[0412] To a 50 mL RBF was added 5-azido-3,3-dimethyl-1- (phenylsulfonyl)pentan-2-one (70 wt% mixture, 110.0 mg, 0.4 mmol) in MeOH (3 mL) at 0 °C (ice bath) followed by the addition of NaBH4(28.0 mg, 0.7 mmol). The mixture was stirred at 0 °C for 1 h. The mixture was quenched with water (30 mL) and extracted with CH2Cl2(3 x 50 mL). The combined organic phases were dried over Na2SO4, filtered, and concentrated-104-by rotary evaporation under reduced pressure and the residue was purified using preparative RP-HPLC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford 5- azido-3,3-dimethyl-1-(phenylsulfonyl)pentan-2-ol as a colorless liquid (50.0 mg, 64.1% yield). NMR (600 MHz, CDCl3) δ 7.96 – 7.91 (m, 2H), 7.71 – 7.67 (m, 1H), 7.63 – 7.57 (m, 2H), 3.86 (dd, J = 9.9, 1.1 Hz, 1H), 3.56 – 3.35 (m, 1H), 3.34 – 3.27 (m, 1H), 3.26 – 3.19 (m, 2H), 3.14 (dd, J = 14.1, 9.9 Hz, 1H), 1.73 – 1.65 (m, 1H), 1.48 – 1.38 (m, 1H), 0.86 (d, J = 1.8 Hz, 6H) ppm.

[0413] Step 5: Synthesis of 5-amino-3,3-dimethyl-1-(phenylsulfonyl)pentan-2-ol.

[0414] To a 50 mL RBF were added 5-azido-3,3-dimethyl-1- (phenylsulfonyl)pentan-2-ol (50.0 mg, 0.2 mmol), Pd / C (10 wt% dry, 20.2 mg) and MeOH (2 mL). The mixture was degassed under vacuum and filled with H2. The mixture was stirred at rt under H2for 1 h. The mixture was filtered through Celite and concentrated by rotary evaporation under reduced pressure to afford crude 5-amino-3,3-dimethyl-1- (phenylsulfonyl)pentan-2-ol as a colorless liquid (45.0 mg, 97.6% yield).1H NMR (600 MHz, CDCl3) δ 7.99 – 7.95 (m, 2H), 7.64 – 7.59 (m, 1H), 7.56 – 7.51 (m, 2H), 4.02 – 3.94 (m, 1H), 3.51 – 3.46 (m, 1H), 3.24 (d, J = 14.2 Hz, 1H), 3.20 – 3.12 (m, 1H), 3.11 – 3.02 (m, 1H), 1.95 – 1.86 (m, 1H), 1.69 (s, 1H), 0.87 (s, 3H), 0.80 (s, 3H) ppm.

[0415] Step 6: Synthesis of 4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin- 5(2H)-yl]-N-[4-hydroxy-3,3-dimethyl-5-(phenylsulfonyl)pentyl]-4-oxobutanamide.

[0416] To a 50 mL RBF were added 5-amino-3,3-dimethyl-1- (phenylsulfonyl)pentan-2-ol (45.0 mg, 0.2 mmol), 2,5-dioxopyrrolidin-1-yl 4-[7,8-didehydro- 3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutanoate (77.0 mg, 0.2 mmol), DIEA (349 uL, 2.0 mmol) and CH2Cl2 (2 mL). The mixture was stirred at rt for 1 h. The mixture was concentrated by rotary evaporation under reduced pressure and the residue was subjected to silica gel column chromatography purification eluting with MeOH / CH2Cl2gradients (0-10%) to afford 4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-N-[4-hydroxy-3,3- dimethyl-5-(phenylsulfonyl)pentyl]-4-oxobutanamide as an orange liquid (48.0 mg, 60.8% yield).1H NMR (600 MHz, CDCl3) δ 7.95 – 7.90 (m, 2H), 7.68 – 7.63 (m, 1H), 7.60 – 7.54 (m, 2H), 6.57 (s, 1H), 4.23 (t, J = 2.5 Hz, 1H), 4.14 – 4.08 (m, 3H), 3.91 (d, J = 9.6 Hz, 1H), 3.83 – 3.78 (m, 1H), 3.76 – 3.71 (m, 1H), 3.67 – 3.62 (m, 1H), 3.60 – 3.54 (m, 1H), 3.27 – 3.11-105-(m, 4H), 2.68 – 2.64 (m, 2H), 2.53 – 2.44 (m, 2H), 2.00 – 1.94 (m, 1H), 1.94 – 1.88 (m, 1H), 1.61 – 1.54 (m, 1H), 1.38 – 1.30 (m, 1H), 0.86 – 0.78 (m, 6H) ppm.

[0417] Step 7: Synthesis of 5-({4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin- 5(2H)-yl]-4-oxobutanoyl}amino)-3,3-dimethyl-1-(phenylsulfonyl)pentan-2-yl 2,5- dioxopyrrolidin-1-yl carbonate.

[0418] A 100 mL round bottom flask with 4-[7,8-didehydro-3,4,6,9-tetrahydro- 1,5-oxazonin-5(2H)-yl]-N-[4-hydroxy-3,3-dimethyl-5-(phenylsulfonyl)pentyl]-4- oxobutanamide (323.0 mg, 0.7 mmol) was purged with N2 / vacuum cycle 3 times, before the addition of triphosgene (502.1 mg, 1.7 mmol) and 10 mL of dry THF. The mixture was stirred at 0 °C (ice bath) for 5 min, followed by the dropwise addition of dry pyridine (164 μL, 2.0 mmol) in dry THF (1 mL). The mixture was stirred at 0 °C for 10 min and slowly warmed to rt for 1 hour. The mixture was filtered and washed with THF (3 x 5 mL). The filtrate was concentrated by rotary evaporation under reduced pressure. HOSu (393.0 mg, 3.4 mmol) was added to the residue, and the flask was purged with N2 / vacuum cycle 3 times, before the addition of dry THF (10 mL) and dropwise addition of dry pyridine (164 μL, 2.0 mmol) in dry THF (1 mL) at 0 °C (ice bath). The mixture was stirred at 0 °C for 10 min and slowly warmed to rt for 1.5 hour. The mixture was filtered and washed with THF (3 x 5 mL). The filtrate was concentrated by rotary evaporation under reduced pressure and the residue was purified using preparative RP-HPLC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford 5-({4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-4- oxobutanoyl}amino)-3,3-dimethyl-1-(phenylsulfonyl)pentan-2-yl 2,5-dioxopyrrolidin-1-yl carbonate as a white solid (128.0 mg, 30.8% yield). LC-MS: mass calcd for C29H37N3O10S: 619.22, found: m / z = 620.32 [M+H]+.1H NMR (600 MHz, CDCl3) δ 7.94 (dd, J = 8.2, 1.0 Hz, 2H), 7.71 – 7.65 (m, 1H), 7.64 – 7.57 (m, 2H), 6.09 (br s, 1H), 5.19 (dd, J = 9.4, 1.4 Hz, 1H), 4.29 – 4.09 (m, 4H), 3.84 – 3.74 (m, 2H), 3.68 – 3.57 (m, 2H), 3.52 – 3.46 (m, 1H), 3.41 – 3.35 (m, 1H), 3.32 – 3.25 (m, 1H), 3.14 – 3.05 (m, 1H), 2.85 (s, 4H), 2.71 – 2.61 (m, 2H), 2.57 – 2.46 (m, 2H), 2.02 – 1.92 (m, 2H), 1.51 – 1.42 (m, 2H), 0.92 (d, J = 9.5 Hz, 6H) ppm.

[0419] Step 8: Synthesis of 4-(aminomethyl)-N-((1,2,3,5,6,7-hexahydro-s-indacen- 4- yl)carbamoyl)furan-2-sulfonimidamide-106-

[0420] To a solution of 1-[[4-(bromomethyl)furan-2-yl](imino)oxo-lambda6- sulfanyl]-3-(1,2,3,5,6,7-hexahydro-s-indacen-4-yl)urea 1j (150 mg, 0.342 mmol) in MeOH (10 ml) was added NH3(g) (7 M / in MeOH) (10 ml). The reaction mixture was stirred at rt for 2 h and concentrated. The residue was purified by Prep-HPLC using the following conditions Column: YMC-Actus Triart C18 ExRS, 30 *150 mm, 5Pm; Mobile Phase A: Water (10mmol / L NH4HCO3), Mobile Phase B: ACN; Flow rate:60 mL / min; Gradient:21%B to 33%B in 7 min; 254 / 220 nm; RT1: 6.18 min. After lyophilization, the titled compound 4- (aminomethyl)-N-((1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)furan-2- sulfonimidamide Ex.22 was obtained as a white solid (22.1 mg, 16.83%).1HNMR (300 MHz, CD3OD) δ (ppm): 7.64 - 7.71 (m, 1H), 7.04 - 7.11 (m, 1H), 6.90 (s, 1H), 3.71 - 3.78 (m, 2H), 2.80 - 2.88 (m, 4H), 2.72 - 2.79 (m, 4H), 1.94 - 2.09 (m, 4H); LC-MS: m / z [M+H]+= 375.1.

[0421] Step 9: Synthesis of N-[(4-{[({[5-({4-[7,8-didehydro-3,4,6,9-tetrahydro- 1,5-oxazonin-5(2H)-yl]-4-oxobutanoyl}amino)-3,3-dimethyl-1-(phenylsulfonyl)pentan-2- yl]oxy}carbonyl)amino]methyl}-5-methylfuran-2-yl)(oyl)-λ6-sulfananimidoyl]-N'- (1,2,3,5,6,7-hexahydros-indacen-4-yl)urea.

[0422] To a solution of 5-({4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin- 5(2H)-yl]-4-oxobutanoyl}amino)-3,3-dimethyl-1-(phenylsulfonyl)pentan-2-yl 2,5- dioxopyrrolidin-1-yl carbonate (400.0 mg, 0.6 mmol) in DMF (5 mL) were added TEA (140.0 mg, 3.5 mmol) and 4-(aminomethyl)-N-((1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)-5- methylfuran-2-sulfonimidamide (180.0 mg, 0.5 mmol) at 0 °C (ice bath). The reaction progress was monitored by LC-MS until complete consumption of the starting material. The crude reaction mixture was purified using preparative RP-HPLC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford N-[(4-{[({[5-({4-[7,8-didehydro-3,4,6,9- tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutanoyl}amino)-3,3-dimethyl-1- (phenylsulfonyl)pentan-2-yl]oxy}carbonyl)amino]methyl}-5-methylfuran-2-yl)(oyl)-λ6- sulfananimidoyl]-N'-(1,2,3,5,6,7-hexahydros-indacen-4-yl)urea as a pale-yellow solid (54.7 mg, 12.7% yield).

[0423] LC-MS: mass calcd for C44H56N6O10S2: 892.35, found: m / z = 893.36 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.95 (d, J = 6.3 Hz, 2H), 7.83 – 7.66 (m, 2H), 7.66 – 7.52 (m, 2H), 7.13 (s, 1H), 7.02 (s, 1H), 6.92 (s, 1H), 5.13 – 5.01 (m, 1H), 4.29 – 4.19 (m, 2H),-107-4.15 (s, 2H), 4.10 – 3.91 (m, 2H), 3.89 – 3.75 (m, 2H), 3.74 – 3.64 (m, 1H), 3.63 – 3.44 (m, 3H), 3.24 – 2.99 (m, 2H), 2.92 – 2.82 (m, 5H), 2.79 (s, 4H), 2.67 (s, 2H), 2.53 – 2.42 (m, 2H), 2.38 (s, 3H), 2.11 – 1.98 (m, 6H), 1.96 – 1.84 (m, 1H), 1.50 – 1.26 (m, 3H), 0.88 – 0.73 (m, 6H) ppm. “Z2-R’-L-PL” Modified Payload Example 7: Synthesis o didehydro- 3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutanoyl}amino)-3,3-dimethyl-1- (phenylsulfonyl)pentan-2-yl]oxy}carbonyl)(methyl)amino]methyl}furan-2-yl)(oyl)- sulfananimidoyl]-N'-(1,2,3,5,6,7-hexahydros-indacen-4-yl)urea

[0425] N-((1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)-4- ((methylamino)methyl)furan-2-sulfonimidamide was prepared as described in WO 2024 / 010772, which is hereby incorporated by reference.

[0426] Step 1: Synthesis of N-[(4-{[({[5-({4-[7,8-didehydro-3,4,6,9-tetrahydro- 1,5-oxazonin-5(2H)-yl]-4-oxobutanoyl}amino)-3,3-dimethyl-1-(phenylsulfonyl)pentan-2- yl]oxy}carbonyl)(methyl)amino]methyl}furan-2-yl)(oyl)-λ6-sulfananimidoyl]-N'-(1,2,3,5,6,7- hexahydros-indacen-4-yl)urea.

[0427] To a solution of N-((1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)-4- ((methylamino)methyl)furan-2-sulfonimidamide (80 mg, 0.21 mmol) in DMF (1.5 mL) were added TEA (62.1 mg, 0.6 mmol) and 5-({4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin- 5(2H)-yl]-4-oxobutanoyl}amino)-3,3-dimethyl-1-(phenylsulfonyl)pentan-2-yl 2,5- dioxopyrrolidin-1-yl carbonate (260.2 mg, 0.4 mmol) in DMF (1.5 mL) at 0 °C (ice bath). The mixture was stirred at rt for 2 h. The reaction progress was monitored by LC-MS until complete consumption of the starting material. The crude reaction mixture was purified using preparative-108-RP-HPLC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford N-[(4- {[({[5-({4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutanoyl}amino)- 3,3-dimethyl-1-(phenylsulfonyl)pentan-2-yl]oxy}carbonyl)(methyl)amino]methyl}furan-2- yl)(oyl)-λ6-sulfananimidoyl]-N'-(1,2,3,5,6,7-hexahydros-indacen-4-yl)urea as a pale-yellow solid (32.5 mg, 17.9% yield). LC-MS: mass calcd for C44H56N6O10S2: 892.35, found: m / z = 893.21 [M+H]+.1H NMR (400 MHz, CD3OD) δ 8.02 – 7.94 (m, 2H), 7.80 – 7.68 (m, 2H), 7.67 – 7.55 (m, 2H), 7.09 (s, 1H), 6.92 (s, 1H), 5.13 (s, 1H), 4.37 – 4.18 (m, 4H), 4.16 – 4.12 (m, 2H), 3.86 – 3.76 (m, 2H), 3.74 – 3.62 (m, 2H), 3.61 – 3.55 (m, 2H), 3.19 – 2.97 (m, 2H), 2.91 – 2.80 (m, 6H), 2.76 (s, 4H), 2.72 – 2.61 (m, 4H), 2.52 – 2.37 (m, 2H), 2.09 – 1.97 (m, 6H), 1.90 (s, 1H), 1.37 – 1.29 (m, 5H), 0.89 – 0.82 (m, 6H). “Z2-R’-L-PL” Modified Payload Example 8: Synthesis of (3S)-3-({4-[7,8-didehydro-3,4,6,9- tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutanoyl}amino)-4-({5-({[(5-{[(1,2,3,5,6,7- hexahydros-indacen-4-ylcarbamoyl)amino](oyl)-λ6-sulfananimidoyl}furan-3- yl)methyl](methyl)carbamoyl}oxy)-4,4-dimethyl-6-[(4- methylphenyl)sulfonyl]hexyl}amino)-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate

[0428] Scheme:-109-

[0429] Step 1: Synthesis of methyl 5-chloro-2,2-dimethylpentanoate.

[0430] To a three-neck flask was added LDA (100 mL, 100 mmol, 1 M in THF) at -78 °C (dry ice / acetone bath). Methyl isobutyrate (10.0 g, 97.9 mmol) was added slowly and the mixture was stirred for 20 min.1-Bromo-3-chloropropane (16.9 g, 108.0 mmol) was added slowly. The reaction mixture was stirred at rt overnight. The mixture was quenched by slow-110-addition of conc. HCl (14 mL) followed by HCl (aq., 200 mL, 1 M) at 0 °C (ice bath). The aqueous phase was extracted with EA (3 × 150 mL). The combined organic layers were dried over Na2SO4, filtered, and concentrated by rotary evaporation under reduced pressure. The residue was subjected to silica gel column chromatography purification eluting with 100% PE to afford methyl 5-chloro-2,2-dimethylpentanoate as a yellow oil (5.8 g, 33.3% yield).1H NMR (400 MHz, CDCl3) δ 3.68 (s, 3H), 3.54 – 3.51 (m, 2H), 1.77 – 1.64 (m, 4H), 1.21 (s, 6H) ppm.

[0431] Step 2: Synthesis of methyl 5-azido-2,2-dimethylpentanoate.

[0432] Methyl 5-chloro-2,2-dimethylpentanoate (5.8 g, 32.5 mmol) was dissolved in DMSO (120 mL) in a single-neck flask. Sodium azide (3.2 g, 48.7 mmol, 1.5 eq) was added, and the mixture was heated at 70 °C overnight. After cooling to rt, the reaction mixture was diluted with water (150 mL) and extracted with EA (3 x 100 mL). The combined organic layers were washed with 5% LiCl solution (2 x 100 mL) and sat. NaCl (aq., 2 x 100 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was subjected to silica gel column chromatography purification eluting with EA / Hex gradients (0-5%) to afford methyl 5-azido-2,2-dimethylpentanoate as a yellow oil (3.8 g, 63.3% yield).1H NMR (400 MHz, CDCl3) δ 3.69 (s, 3H), 3.27 (t, J = 6.4 Hz, 2H), 1.62 – 1.51 (m, 4H), 1.21 (s, 6H) ppm.

[0433] Step 3: Synthesis of 6-azido-3,3-dimethyl-1-tosylhexan-2-one.

[0434] 1-Methyl-4-(methylsulfonyl)benzene (5.2 g, 30.5 mmol) was dissolved in THF (60 mL) in a three-neck flask. The solution was added LDA (20.5 mL, 41.0 mmol, 2 mol / L in THF) slowly at -78 °C (dry ice / acetone bath). The mixture was stirred at -78 °C for 20 min and methyl 5-azido-2,2-dimethylpentanoate (3.8 g, 20.5 mmol) was added. The reaction mixture was warmed to rt and stirred overnight. The reaction was quenched by slow addition of sat. NH4Cl solution (aq., 60 mL) at 0 °C (ice bath). The mixture was extracted with EA (3 × 60 mL). The combined organic layers were dried over Na2SO4 and concentrated under reduced pressure. The residue was subjected to silica gel column chromatography purification eluting with EA / Hex gradients (0-10%) to afford 6-azido-3,3-dimethyl-1-tosylhexan-2-one as a yellow oil (3.4 g, 51.3% yield). LC-MS: mass calcd for C15H21N3O3S: 323.13, found: m / z = 346.12 [M+Na]+.1H NMR (400 MHz, CDCl3) δ 7.84 (s, 2H), 7.38 (s, 2H), 4.30 (s, 2H), 3.26 (t, J = 6.5 Hz, 2H), 2.48 (s, 3H), 1.59 – 1.54 (m, 2H), 1.46 – 1.39 (m, 2H), 1.13 (s, 6H) ppm.

[0435] Step 4: Synthesis of 6-azido-3,3-dimethyl-1-tosylhexan-2-ol.-111-

[0436] 6-Azido-3,3-dimethyl-1-tosylhexan-2-one as a yellow oil (3.4 g, 10.5 mmol) was dissolved in CH3OH (40 mL) in a single-neck flask. The solution was cooled to 0 °C (ice bath) and NaBH4(199.0 mg, 5.2 mmol) was added. The reaction mixture was stirred at rt for 1 h. The reaction was quenched with water (40 mL) and extracted with EA (3 × 40 mL). The combined organic layers were dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by reverse phase column chromatography employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford 6-azido-3,3- dimethyl-1-tosylhexan-2-ol as a colorless oil (1.3 g, 38.2% yield).

[0437] LC-MS: mass calcd for C15H23N3O3S: 325.15, found: m / z = 326.11 [M+H]+.1H NMR (400 MHz, CDCl3) δ 7.84 (d, J = 8.3 Hz, 2H), 7.41 (d, J = 8.1 Hz, 2H), 3.87 (d, J = 9.8 Hz, 1H), 3.34 (s, 1H), 3.30 – 3.18 (m, 3H), 3.17 – 3.09 (m, 1H), 2.49 (s, 3H), 1.61 – 1.53 (m, 1H), 1.45 – 1.42 (m, 1H), 1.25 – 1.16 (m, 1H), 0.86 (d, J = 13.7 Hz, 6H) ppm.

[0438] Step 5: Synthesis of 6-azido-3,3-dimethyl-1-tosylhexan-2-yl (2,5- dioxopyrrolidin-1-yl) carbonate.

[0439] 6-Azido-3,3-dimethyl-1-tosylhexan-2-ol (1.0 g, 3.1 mmol) was dissolved in THF (40 mL) in a single-neck flask. Pyridine (729.2 mg, 9.2 mmol) and triphosgene (2.7 g, 9.2 mmol) were added at 0 °C (ice bath). The mixture was purged with N2 (3 cycles) and stirred at rt for 1 h. The reaction mixture was filtered through Celite and washed with THF (10 mL). The filtrate was concentrated under reduced pressure. The residue was dissolved in THF (20 mL) and pyridine (729.2 mg, 9.2 mmol) and HOSu (1.0 g, 9.2 mmol) were added at 0 °C (ice bath). The mixture was purged with N2(3 cycles) and stirred at rt for 1 h. The reaction mixture was filtered through Celite, washed with THF, and concentrated. The residue was purified by reverse phase column chromatography employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford 6-azido-3,3-dimethyl-1-tosylhexan-2-yl (2,5-dioxopyrrolidin-1-yl) carbonate as a white solid (780.0 mg, 54.2% yield). LC-MS: mass calcd for C20H26N4O7S: 466.15, found: m / z = 467.16 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.86 (d, J = 8.2 Hz, 2H), 7.48 (d, J = 8.1 Hz, 2H), 5.06 (dd, J = 7.7, 2.9 Hz, 1H), 3.66 – 3.62 (m, 2H), 3.26 – 3.15 (m, 2H), 2.85 (s, 4H), 2.49 (s, 3H), 1.55 – 1.45 (m, 1H), 1.36 – 1.23 (m, 3H), 0.91 (d, J = 7.1 Hz, 6H) ppm.-112-

[0440] Step 6: Synthesis of 6-azido-3,3-dimethyl-1-tosylhexan-2-yl ((5-(N- ((1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)sulfamidimidoyl)furan-3- yl)methyl)(methyl)carbamate.

[0441] N-((1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)-4- ((methylamino)methyl)furan-2-sulfonimidamide (195.1 mg, 0.5 mmol) was dissolved in DMF (5 mL) in a single-neck flask. TEA (152.2 mg, 1.5 mmol) and 6-azido-3,3-dimethyl-1- tosylhexan-2-yl (2,5-dioxopyrrolidin-1-yl) carbonate (257.3 mg, 1.1 mmol) were added at 0 °C (ice bath). The mixture was purged with N2 (3 cycles) and stirred at rt overnight. Reaction progress was monitored by LC-MS until complete consumption of starting materials The crude reaction mixture was purified using preparative RP-HPLC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford 6-azido-3,3-dimethyl-1-tosylhexan-2-yl ((5- (N-((1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)sulfamidimidoyl)furan-3- yl)methyl)(methyl)carbamate as a white solid (150.1 mg, 40.5% yield). LC-MS: mass calcd for C35H45N7O7S2: 739.28, found: m / z = 740.36 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.93 – 7.82 (m, 2H), 7.80 – 7.71 (m, 1H), 7.44 (dd, J = 18.2, 7.4 Hz, 2H), 7.24 – 7.06 (m, 1H), 6.92 (s, 1H), 5.21 – 5.08 (m, 1H), 4.47 – 4.15 (m, 2H), 3.52 (d, J = 5.7 Hz, 2H), 3.17 (s, 2H), 2.94 – 2.69 (m, 11H), 2.53 – 2.41 (m, 3H), 2.03 (p, J = 7.2 Hz, 4H), 1.61 – 1.46 (m, 1H), 1.33 – 1.14 (m, 3H), 0.84 (d, J = 6.6 Hz, 6H) ppm.

[0442] Step 7: Synthesis of 6-amino-3,3-dimethyl-1-tosylhexan-2-yl ((5-(N- ((1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)sulfamidimidoyl)furan-3- yl)methyl)(methyl)carbamate.

[0443] 6-Azido-3,3-dimethyl-1-tosylhexan-2-yl ((5-(N-((1,2,3,5,6,7-hexahydro-s- indacen-4-yl)carbamoyl)sulfamidimidoyl)furan-3-yl)methyl)(methyl)carbamate (150.0 mg, 0.2 mmol) was dissolved in iPrOH (3 mL) in a single-neck flask and purged with N2 (3 cycles). Pd(OH)2(30.3 mg, 20%) was added under N2, and the mixture was evacuated and backfilled with H2 (3 cycles). The reaction mixture was stirred at rt for 3 h under H2 atmosphere (1 atm). The mixture was filtered through a pad of Celite and concentrated under reduced pressure to afford crude 6-amino-3,3-dimethyl-1-tosylhexan-2-yl ((5-(N-((1,2,3,5,6,7-hexahydro-s- indacen-4-yl)carbamoyl)sulfamidimidoyl)furan-3-yl)methyl)(methyl)carbamate as a white-113-solid (135 mg, crude). LC-MS: mass calcd for C35H47N5O7S2: 713.29, found: m / z = 714.40 [M+H]+.

[0444] Step 8: Synthesis of (3S)-3-((tert-butoxycarbonyl)amino)-4-((5-((((5-(N- ((1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)sulfamidimidoyl)furan-3- yl)methyl)(methyl)carbamoyl)oxy)-4,4-dimethyl-6-tosylhexyl)amino)-4-oxobutyl (2- (trimethylammonio)ethyl) phosphate.

[0445] 6-Amino-3,3-dimethyl-1-tosylhexan-2-yl ((5-(N-((1,2,3,5,6,7-hexahydro- s-indacen-4-yl)carbamoyl)sulfamidimidoyl)furan-3-yl)methyl)(methyl)carbamate (135 mg, 0.2 mmol) was dissolved in DMF (3 mL) in a single-neck flask. DIEA (73.1 mg, 0.6 mmol), (S)-3-((tert-butoxycarbonyl)amino)-3-carboxypropyl (2-(trimethylammonio)ethyl) phosphate (72.6 mg, 0.3 mmol), HOBt monohydrate (38.2 mg, 0.3 mmol), and EDCI (54.3 mg, 0.3 mmol) were added. The mixture was stirred at rt for 16 h, and reaction completion was monitored by LC-MS. The crude reaction mixture was purified using preparative RP-HPLC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford (3S)-3-((tert- butoxycarbonyl)amino)-4-((5-((((5-(N-((1,2,3,5,6,7-hexahydro-s-indacen-4- yl)carbamoyl)sulfamidimidoyl)furan-3-yl)methyl)(methyl)carbamoyl)oxy)-4,4-dimethyl-6- tosylhexyl)amino)-4-oxobutyl (2-(trimethylammonio)ethyl) phosphate as a white solid (68.1 mg, 33.2% yield). LC-MS: mass calcd for C49H74N7O14PS2: 1079.45, found: m / z = 1080.43 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.91 – 7.81 (m, 2H), 7.80 – 7.71 (m, 1H), 7.49 – 7.34 (m, 2H), 7.20 – 7.06 (m, 1H), 6.92 (s, 1H), 5.18 (s, 1H), 4.37 – 4.22 (m, 3H), 4.16 (s, 1H), 4.07 – 3.87 (m, 2H), 3.71 – 3.59 (m, 4H), 3.53 (d, J = 6.5 Hz, 2H), 3.32 (s, 1H), 3.26 – 3.10 (m, 9H), 3.07 (s, 1H), 2.90 – 2.70 (m, 10H), 2.47 (s, 3H), 2.11 – 1.96 (m, 5H), 1.89 (s, 1H), 1.45 (d, J = 2.7 Hz, 9H), 1.40 – 1.08 (m, 4H), 0.93 – 0.71 (m, 6H) ppm.

[0446] Step 9: Synthesis of (3S)-3-amino-4-((5-((((5-(N-((1,2,3,5,6,7-hexahydro-s- indacen-4-yl)carbamoyl)sulfamidimidoyl)furan-3-yl)methyl)(methyl)carbamoyl)oxy)-4,4- dimethyl-6-tosylhexyl)amino)-4-oxobutyl (2-(trimethylammonio)ethyl) phosphate trifluoroacetate.

[0447] (3S)-3-((tert-Butoxycarbonyl)amino)-4-((5-((((5-(N-((1,2,3,5,6,7- hexahydro-s-indacen-4-yl)carbamoyl)sulfamidimidoyl)furan-3- yl)methyl)(methyl)carbamoyl)oxy)-4,4-dimethyl-6-tosylhexyl)amino)-4-oxobutyl (2--114-(trimethylammonio)ethyl) phosphate (65.0 mg, 0.06 mmol) was dissolved in CH2Cl2 (2 mL) in a single-neck flask. TFA (0.5 mL) was added, and the reaction mixture was stirred at rt for 1 h. The solvent was removed under reduced pressure to afford crude (3S)-3-amino-4-((5-((((5- (N-((1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)sulfamidimidoyl)furan-3- yl)methyl)(methyl)carbamoyl)oxy)-4,4-dimethyl-6-tosylhexyl)amino)-4-oxobutyl (2- (trimethylammonio)ethyl) phosphate trifluoroacetate as a pale-yellow oil (55.2 mg, 84.9% yield). LC-MS: mass calcd for C44H66N7O12PS2: 979.39, found: m / z = 980.42 [M+H]+.

[0448] Step 10: Synthesis of (3S)-3-({4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]-4-oxobutanoyl}amino)-4-({5-({[(5-{[(1,2,3,5,6,7-hexahydros-indacen-4- ylcarbamoyl)amino](oyl)-λ6-sulfananimidoyl}furan-3-yl)methyl](methyl)carbamoyl}oxy)-4,4- dimethyl-6-[(4-methylphenyl)sulfonyl]hexyl}amino)-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate.

[0449] (3S)-3-Amino-4-((5-((((5-(N-((1,2,3,5,6,7-hexahydro-s-indacen-4- yl)carbamoyl)sulfamidimidoyl)furan-3-yl)methyl)(methyl)carbamoyl)oxy)-4,4-dimethyl-6- tosylhexyl)amino)-4-oxobutyl (2-(trimethylammonio)ethyl) phosphate trifluoroacetate (44.0 mg, 0.04 mmol) was dissolved in DMF (1 mL) in a single-neck flask. TEA (13.6 mg, 0.134 mmol) and 2,5-dioxopyrrolidin-1-yl 4-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)- yl]-4-oxobutanoate (14.4 mg, 0.04 mmol) were added at 0 °C (ice bath). The reaction mixture was stirred at rt for 1 h. The crude reaction mixture was purified using preparative RP-HPLC employing ACN / water mobile phases acidified with 0.1 vol-% formic acid (5–95% ACN in 30 min). Product-containing fractions were combined and lyophilized to afford (3S)-3-({4-[7,8- didehydro-3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-4-oxobutanoyl}amino)-4-({5-({[(5- {[(1,2,3,5,6,7-hexahydros-indacen-4-ylcarbamoyl)amino](oyl)-λ6-sulfananimidoyl}furan-3- yl)methyl](methyl)carbamoyl}oxy)-4,4-dimethyl-6-[(4- methylphenyl)sulfonyl]hexyl}amino)-4-oxobutyl 2-(trimethylazaniumyl)ethyl phosphate as a white solid (9.4 mg, 17.1% yield). LC-MS: mass calcd for C55H79N8O15PS2: 1186.48, found: m / z = 1187.45 [M+H]+.1H NMR (400 MHz, CD3OD) δ 7.93 – 7.80 (m, 2H), 7.79 – 7.70 (m, 1H), 7.50 – 7.34 (m, 2H), 7.23 – 7.02 (m, 1H), 6.92 (s, 1H), 5.17 – 5.12 (m, 1H), 4.46 – 4.40 (m, 1H), 4.34 – 4.18 (m, 6H), 4.14 (s, 2H), 4.05 – 3.99 (m, 1H), 3.98 – 3.91 (m, 1H), 3.84 – 3.75 (m, 2H), 3.71 – 3.63 (m, 3H), 3.62 (s, 2H), 3.54 (s, 3H), 3.28 – 3.16 (m, 9H), 3.14 – 3.05 (m, 2H), 2.89 – 2.80 (m, 6H), 2.79 – 2.69 (m, 6H), 2.64 – 2.56 (m, 1H), 2.55 – 2.49 (m, 1H),-115-2.47 (d, J = 4.2 Hz, 3H), 2.28 (s, 1H), 2.09 – 1.97 (m, 5H), 1.94 – 1.89 (m, 1H), 1.52 (s, 1H), 1.39 – 1.31 (m, 1H), 1.28 – 1.11 (m, 2H), 0.93 – 0.74 (m, 6H) ppm. “Z2-R’-L-PL” Modified Payload Example 9: Synthesis of 5-[7,8-didehydro-3,4,6,9- tetrahydro-1,5-oxazonin-5(2H)-ylsulfonyl]-N,N-dimethylnaphthalen-1-amine

[0451] Step 1: Synthesis of 5-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin- 5(2H)-ylsulfonyl]-N,N-dimethylnaphthalen-1-amine.

[0452] To a solution of 7,8-didehydro-2,3,4,5,6,9-hexahydro-1,5-oxazonine (300.0 mg, 2.4 mmol) in CH2Cl2 (8 mL), 5-(dimethylamino)naphthalene-1-sulfonyl chloride (646.5 mg, 2.4 mmol) and TEA (485.7 mg, 4.8 mmol) were added. The mixture was stirred at rt for 1 h. Upon completion of the reaction, the mixture was concentrated by rotary evaporation under reduced pressure, and the residue was triturated with MeOH (3 mL) for 1 h. The resulting suspension was filtered, the solid was washed with cold MeOH (1 mL), and the filter cake was dried under reduced pressure to afford 5-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin- 5(2H)-ylsulfonyl]-N,N-dimethylnaphthalen-1-amineas a yellow solid (599.2 mg, 70.1% yield).1H NMR (400 MHz, CDCl3) δ 8.56 (d, J = 8.5 Hz, 1H), 8.39 (d, J = 8.7 Hz, 1H), 8.19 (dd, J = 7.3, 1.2 Hz, 1H), 7.59 – 7.50 (m, 2H), 7.19 (d, J = 7.5 Hz, 1H), 4.09 – 3.99 (m, 4H), 3.80 – 3.71 (m, 2H), 3.52 – 3.42 (m, 2H), 2.89 (s, 6H), 1.79 (t, J = 5.5 Hz, 2H) ppm. “Z2-R’-L-PL” Modified Payload Example 10: Synthesis of N-{3-[7,8-didehydro-3,4,6,9- tetrahydro-1,5-oxazonin-5(2H)-yl]-3-oxopropyl}-3-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1- yl)propenamide-116-

[0454] Step 1: Synthesis of N-{3-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin- 5(2H)-yl]-3-oxopropyl}-3-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)propenamide.

[0455] In a 100 mL RBF, 3-amino-1-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]propan-1-one (129.0 mg, 0.7 mmol), 2,5-dioxopyrrolidin-1-yl 3-(2,5- dioxo-2,5-dihydro-1H-pyrrol-1-yl)propanoate (175.0 mg, 0.7 mmol) were dissolved THF (15 mL), and then TEA (1.5 mL, 1.1 g, 10.8 mmol) was added. The mixture was allowed to stir at rt overnight while being monitored with TLC. The crude was concentrated by rotary evaporation under reduced pressure, and purified with silica gel column chromatography purification eluting with MeOH / CH2Cl2gradients (0-5%) to afford N-{3-[7,8-didehydro- 3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-3-oxopropyl}-3-(2,5-dioxo-2,5-dihydro-1H- pyrrol-1-yl)propenamide as a white solid (120.7 mg, 52.6% yield). MS: mass calcd for C17H21N3O5: 347.12, found: m / z = 348.10 [M+H]+.1H NMR (600 MHz, CDCl3) δ 6.66 (s, 2H), 6.52 (br s, 1H), 4.25 – 4.02 (m, 4H), 3.80 – 3.72 (m, 4H), 3.59 – 3.54 (m, 2H), 3.50 – 3.45 (m, 2H), 2.53 – 2.46 (m, 2H), 2.45 – 2.40 (m, 2H), 1.96 – 1.90 (m, 2H) ppm. “Z2-R’-L-PL” Example 11: Synthesis of N-{3-[7,8-didehydro-3,4,6,9-tetrahydro-1,5- oxazonin-5(2H)-yl]-3-oxopropyl}-3-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)propenamide.

[0456] Scheme:-117-

[0457] Step 1: Synthesis of 1-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin- 5(2H)-yl]-2-iodoethan-1-one:

[0458] In a 100 mL RBF, 7,8-didehydro-2,3,4,5,6,9-hexahydro-1,5-oxazonine (168.0 mg, 1.3 mmol) was dissolved in 10 mL of DCM, then 2,5-dioxopyrrolidin-1-yl 2- iodoacetate (760.0 mg, 2.6 mmol), DIEA (0.5 mL, 2.9 mmol) were added. The mixture was stirred at rt for 6 h, while being monitored by TLC. Once the reaction was completed, the mixture was concentrated by rotary evaporation under reduced pressure, the residue was subjected to silica gel column chromatography purification eluting with EA / Hex gradients (60- 100%) to afford 1-[7,8-didehydro-3,4,6,9-tetrahydro-1,5-oxazonin-5(2H)-yl]-2-iodoethan-1- one as a white solid (239.2 mg, 60.8% yield).1H NMR (600 MHz, CDCl3) δ 4.64 (d, J = 17.2 Hz, 2H), 4.32 – 4.16 (m, 2H), 4.07 (dt, J = 4.1, 2.4 Hz, 2H), 3.79 – 3.72 (m, 2H), 3.71 – 3.52 (m, 2H), 1.97 – 1.86 (m, 2H) ppm. Click Example 1: Click Conjugation and Payload Release Analysis (with one Z2-R’-L-PL)

[0459] Copolymer (stored in water) was used as the starting material. The polymer concentration was adjusted to 25 mg / mL in pH 5.0 sodium acetate buffer, with a total reaction volume of 2.5 mL. Z2-R’-L-PL was dissolved in DMSO (100 µL) and added dropwise to the polymer solution. The mixture was incubated at 37 °C for 3–12 h to complete the click reaction.

[0460] Progress of the click reaction was monitored by size-exclusion chromatography (SEC; Agilent 1100, Shodex OHpak SB-804 / 806M HQ columns, 0.1 M NaNO₃ with 0.02 wt% NaN₃, 1 mL / min, 35 °C), where the polymer peak (280 nm absorbance) area no longer increased, and by reverse-phase C4 HPLC (Thermo HyPURITY C4, 10 mM NH₄OAc / acetonitrile, 0.5 mL / min, 50 °C), where the Z2-R’-L-PL peak completely disappeared.

[0461] After reaction, pH was adjusted to 7.4 by the addition of NaOH (0.5 N, aq.) followed by PBS buffer (pH 7.4), until the concentration was adjusted to 10 mg / mL. The solution was filtered through a 0.22 µm syringe filter and stored at 37 °C.-118-

[0462] Release kinetics were monitored by size-exclusion chromatography (SEC; Agilent 1100, Shodex OHpak SB-804 / 806M HQ columns, 0.1 M NaNO₃ with 0.02 wt% NaN₃, 1 mL / min, 35 °C) and reverse-phase C4 HPLC (Thermo HyPURITY C4, 10 mM NH₄OAc / acetonitrile, 0.5 mL / min, 50 °C). Data were analyzed by measuring the AUC of conjugates (SEC) and released payload (C4) vs release time and fitted to first-order release kinetics to calculate half-life values. Click Example 2: Click Conjugation and Payload Release Analysis (with multiple Z2-R’-L- PL)

[0463] Copolymer (stored in water) was used as the starting material. The polymer concentration was adjusted to 25 mg / mL in pH 5.0 sodium acetate buffer, with a total reaction volume of 2.5 mL.

[0464] A solution of Z2a-R’a-La-PLa in DMSO (100 µL) was added dropwise to the polymer solution. The mixture was incubated at 37 °C for 3–12 h to complete the first click reaction. Progress of the click reaction was monitored by size-exclusion chromatography (SEC; Agilent 1100, Shodex OHpak SB-804 / 806M HQ columns, 0.1 M NaNO₃ with 0.02 wt% NaN₃, 1 mL / min, 35 °C), where the polymer peak (280 nm absorbance) area no longer increased, and by reverse-phase C4 HPLC (Thermo HyPURITY C4, 10 mM NH₄OAc / acetonitrile, 0.5 mL / min, 50 °C), where the Z2a-R’a-La-PLa peak completely disappeared.

[0465] After completion of the first conjugation, a second solution of Z2b-R’b-Lb- PLbin DMSO (100 µL) was added dropwise to the same reaction mixture, and the mixture was incubated at 37 °C for 3–12 h to complete the second click reaction, with progress monitored by SEC and C4 HPLC as described above

[0466] This sequential conjugation procedure can be repeated with further payload- linker constructs (e.g., Z2c-R’c-Lc-PLc, etc.), each addition followed by incubation and reaction monitoring. The combined molar equivalents of all added Z2a-R’a-La-PLa, Z2b-R’b-Lb-PLb, Z2c-R’c-Lc-PLc, and subsequent payloads did not exceed the total molar equivalents of Z1 units present in the copolymer.

[0467] After completion of conjugation steps, the pH was adjusted to 7.4 by the addition of NaOH (0.5 N, aq.) followed by PBS buffer (pH 7.4), until the concentration was adjusted to 10 mg / mL. The solution was filtered through a 0.22 µm syringe filter and stored at 37 °C.-119-

[0468] Step 4. Payload release analysis:

[0469] Release kinetics were monitored by reverse-phase C4 HPLC (Thermo HyPURITY C4, 10 mM NH₄OAc / acetonitrile, 0.5 mL / min, 50 °C). Data were analyzed by measuring the AUC of released payload (C4) vs release time and fitted to first-order release kinetics to calculate half-life values. Table 1: Conjugate Examples

[0470] Conjugate Examples 1-8 were performed following the procedure described for Click Example 1. Conjugate Example 9 was performed following the procedure described for Click Example 2.

[0471] Table 2 shows the half-life data of the various conjugates. Release half-life can be systematically tuned (1-100+ day) by the linker chemistry (steric and electronic modulation of pKa). Within a comparable copolymer size range, the copolymer and payload have relatively little effect on the half-life; the control comes primarily from the linker design. Table 2: Half-Life Data of Conjugates Physiological pH = 7.4 release half-life data of conjugates-120-Conjugate Example 10: Generation of an Aptamer Copolymer Conjugate

[0472] This non-limiting example describes the generation of aptamer copolymer conjugates.

[0473] Aptamers are single-stranded DNA or RNA molecules that bind to targets through the formation of specific three-dimensional structures, analogous to antibodies. In this example, a thiol-modified aptamer (synthesized by IDT DNA, Inc.) was conjugated to a copolymer backbone.

[0474] The aptamer was first structurally folded in 1× PBS (pH 7.4) supplemented with 2 mM MgCl₂. Folding was achieved by heating the aptamer to 95°C for 5 minutes in a PCR thermocycler, followed by gradual cooling to room temperature.

[0475] To prepare the aptamer-conjugated copolymer, a DBCO-PEG₁₂-maleimide linker (Cat. # BP-25730, BroadPharm) was used. The linker was conjugated to anti-TfR1 aptamer by reacting with a 3-fold molar excess of DBCO-PEG₁₂-maleimide in aptamer conjugation buffer (20 mM Tris-HCl pH 8.5, 50 mM NaCl, 2 mM MgCl2). for 24 hours at room temperature.

[0476] The resulting aptamer-DBCO conjugate was then reacted with copolymer (Mw 38 kDa; 10% azide) via a strain-promoted azide-alkyne cycloaddition (SPAAC) “click” reaction. 3-fold molar excess of the aptamer-DBCO was incubated with copolymer for 24 hours at room temperature.

[0477] The resulting product was purified by size-exclusion chromatography using a Superdex 200 Increase 10 / 30 GL column in a running buffer containing 20 mM HEPES (pH 7.5), 150 mM NaCl, and 2 mM MgCl₂. Fractions containing aptamer-copolymer conjugate were pooled and stored in conjugation buffer. Conjugate Example 11: Generation of antibody-copolymer-drug conjugate via strain-promoted azide–alkyne cycloaddition (SPAAC) click chemistry

[0478] This non-limiting example describes the generation of an antibody copolymer drug conjugate via strain-promoted azide–alkyne cycloaddition (SPAAC) click chemistry.-121-

[0479] A small-molecule cytotoxic drug was loaded onto copolymer (Mw 290 kDa; 10% azide) by mixing copolymer at a final concentration of 10 mg / mL with drug functionalized with a BCN functional group in 20 mM acetate buffer, 70 mM NaCl, pH 5.0 for 72 hours. The reaction employed a molar ratio of 1:93 copolymer:drug to target drug loading of 95% of the total azide sites (98) on each copolymer. The loaded copolymer was then buffer- exchanged into 0.5 M pH 7.8 Na Phosphate to remove the free drug reagent. A DBCO-PEG2- maleimide (Cat.# BP-25727, BroadPharm) linker was then reacted with the loaded copolymer via a strain-promoted azide-alkyne cycloaddition (SPAAC) “click” reaction in 20 mM Tris pH 8.5, 50 mM NaCl to yield a maleimide-copolymer-drug intermediate.

[0480] The antibody decapping was done by reducing antibody with 30x molar excess of Tris (2- carboxyethyl) phosphine hydrochloride (TCEP) for 45 minutes at room temperature followed by buffer exchange into 20 mM Tris-HCl pH 8.0, 50 mM NaCl to remove the cysteine cap and TCEP. Then, the reduced antibodies were oxidized with 15x molar excess of dehydro-ascorbic acid (DHAA) for 75 minutes at room temperature, followed by buffer exchange into 20 mM Tris, pH 8.5, 50 mM NaCl to remove the oxidizing reagent.

[0481] The conjugation process was done by mixing the decapped antibody intermediate at a final concentration of 2.0 mg / mL (or 14 µM) with 3.5x molar excess of maleimide-copolymer-drug intermediate in 20 mM acetate buffer, 70 mM NaCl, pH 5.0 for 72 hours. Once the reaction was complete, the solution contained antibody-copolymer-drug conjugate, free maleimide-copolymer-drug, and free antibody.

[0482] Antibody-copolymer-drug conjugate was purified from unconjugated material by Protein A chromatography with HiTrap MabSelect SuRe 1 mL column (Cytiva US LLC) using a step pH gradient. Step 1 is 50 mM Citrate pH 6.2 to elute free maleimide- copolymer-drug. Step 2 is 30% 50 mM Citrate pH 6.2 and 70% 50 mM Citrate pH 3.5 to elute antibody-copolymer-drug conjugate. Step 3 is 50 mM Citrate pH 3.5 to elute free antibodies. The fractions containing antibody-copolymer-drug conjugate were pooled and buffer- exchanged into 1x PBS, pH 7.4.-122-

Claims

WHAT IS CLAIMED IS:

1. A copolymer comprising: a first repeating units having the structure ofa second repeating units having the structure ofwherein m > 0, n > 0, and m + n ≤ 8000; X1 is H or methyl; X2is H, F, CF3or methyl; Y is -O- or -NH-; R is optionally substituted C1-12 alkylene, optionally substituted, wherein q is 1, 2, 3, or 4; and Z1 comprises N3 or a cyclic olefin.

2. The copolymer of Claim 1, wherein R is C1-12 alkylene, or C1-12 alkylene substituted by F, OH or C1-6 alkyl.

3. The copolymer of Claim 1 wherein R is selected from the group consisting ofC3 alkylene,and, each optionally substituted by F, OH, or C1-6 alkyl.-123-4. The copolymer of Claim 1, wherein R is C1 alkylene oroptionally substituted by F, OH or methyl.

5. The copolymer of Claim 1, wherein Y is O.

6. The copolymer of Claim 1, wherein Z1 is N3,, ,.

7. The copolymer of Claim 1, wherein Z1 is N3.

8. The copolymer of Claim 1 further comprising a core connecting to one or more polymer arms, each of the one or more polymer arms comprises the first repeating units and the second repeating units. Claim 8, wherein the core is,, , or , wherein: R1 is selected from the group consisting of –N3, –NH3+, and; R2 is selected from the group consisting of alkylene, oxy, amido, and combination thereof;, , , wherein R4, R5, and R6 are the same or different and are selected from the group consisting of:, , ; d * indicates a connection to the polymer arm.-124-10. The copolymer of Claim 9 having the following structure:; wherein s is an integer selected from 1-9.

11. The copolymer of Claim 9, wherein R2 comprises, wherein t is an integer selected from 1 to 8.

12. The copolymer of Claim 9, wherein R2 comprisesor, wherein v is an integer selected from 1 to 6 and w is an integerselected from 1 to 6.

13. The copolymer of Claim 1, wherein the copolymer is a star copolymer having 2 to 9 polymer arms each comprising the first repeating units and the second repeating units.

14. The copolymer of Claim 1, wherein the copolymer has a dispersity less than 2.

15. The copolymer of Claim 1, wherein the copolymer has a molecular weight of about 19 kDa to about 1,800 kDa.

16. The copolymer of Claim 1, wherein the copolymer is a random copolymer.

17. The copolymer of Claim 1, wherein the copolymer is a gradient copolymer.

18. The copolymer of Claim 1, wherein the copolymer is a block copolymer.

19. A compound having the following structure: wherein Z2comprises, , ;-125-wherein is a 7- to 9- membered monocyclic or bicyclic cycloalkyne, 7- to 9- membered monocyclic or bicyclic heterocycloalkyne, 10- to 16-membered tricyclic heterocycloalkyne, or optionally substituted tetrazine; and , , ,, , , , d; wherein a is 1 or 2, and u is 1, 2, or 3, and w is an integer between 1 and 15; wherein X3 is H, –(CH2)b–phosphorylcholine, or C1-3 alkyl optionally substituted by COOH, OH, or C(O)NH2; X4is C1-3 alkyl; X5 is C1-3 alkyl substituted by OH, COOH, C(O)NH2, or –(CH2)b– phosphorylcholine; wherein b is 0, 1, 2, 3, or 4;-126-L is absent or selected from the group consisting,; wherein p is 2, 3, or 4; r is 1, 2, 3, or 4; M is O or N-alkyl; X6 is optionally substituted C1-3 alkyl, optionally substituted phenyl, or optionally substituted pyridinyl; X7 and X8 are each H or methyl; and PL comprises a bioactive agent, tracer, or reactive functional group.

20. The compound of Claim 19, wherein Z2 is selected from a group consisting of, , ; , y , r optionally substituted phenyl.

21. The compound of Claim 19, wherein the bioactive agent comprises a small molecule drug, molecular tracer, toxin, peptide, macrocycles, oligonucleotide, or aptamer.-127-22. The compound of Claim 19, wherein the bioactive agent is selected from the group consisting of NLRP3 inhibitors, Prostaglandins, Rho-kinase inhibitors, beta-adrenergic antagonists, carbonic anhydrase inhibitor, C3 inhibitor, C5 inhibitors, puromycin, and tirzepatide.

23. The compound of Claim 19, wherein the bioactive agent is selected from the ,.

24. The compound of Claim 19, wherein L allows for the controlled or stimuli- responsive release of the bioactive agent.

25. The compound of Claim 19, wherein L is a stable linkage and allows for minimal release of the bioactive agent.

26. The compound of Claim 19, wherein L remains attached to R’ when the bioactive agent is released.

27. A conjugate comprising: a first monomer repeating units having the structure of-128-a second monomer repeating units having the structure of; wherein m + n ≤ 8000; X1 is H or methyl; X2 is H, F, CF3, or methyl; Y is -O- or -NH-; R is optionally substituted C1-12 alkylene, optionally substituted, wherein q is an integer between 1 and 4; and Zcomprises;wherein Z’ is a reaction product of (1) N3and cycloalkyne derivative or heterocycloalkyne derivative or (2) olefin and optionally substituted tetrazine derivative; R’ is absent or is selected from the group consisting of, , ,-129-; wherein a is 1 or 2, and u is 1, 2, or 3, and w is an integer between 1 and 15; wherein X3is H, –(CH2)b–phosphorylcholine, or C1-3 alkyl optionally substituted by COOH, OH, or C(O)NH2; X4is C1-3 alkyl; X5is C1-3 alkyl substituted by OH, COOH, C(O)NH2, or –(CH2)b–phosphorylcholine; wherein b is 0, 1, 2, 3, or 4; ,, , , ; wherein p is 2, 3, or 4; r is 1, 2, 3, or 4; X6is optionally substituted C1-3 alkyl, optionally substituted phenyl, or optionally substituted pyridinyl; X7 and X8 are each H or methyl; and PL comprises a bioactive agent, tracer, or reactive functional group.

28. The conjugate of Claim 27, wherein the bioactive agent comprises a small molecule drug, molecular tracer, toxin, peptide, macrocycles, oligonucleotide, or aptamer.

29. A method of making the conjugate of Claim 27 comprising reacting the copolymer of Claim 1 with the compound of Claim 19, wherein Z1 of copolymer react with Z2 of the compound via click reaction in the absence of a metal.

30. A method of treating a disease or condition comprising administer to a subject in need thereof a conjugate of Claim 27.

31. The method of Claim 29, wherein the disease or condition comprises dry age- related macular degeneration (AMD), wet AMD, retinal vein occlusion (RVO), glaucoma, dry eyes, Duane-Radial Ray Syndrome (DR), neuroinflammation, neuroprotection, obesity, Parkinson's, Alzheimer's, cardiac, metabolic disease, diabetic macular edema (DME), cancer, non-infectious uveitis (NIU), inherited retinal disorder (IRD), hypercholesterolemia,-130-inflammatory diseases, cystic fibrosis, b-thalassaemia, mucopolysacchariodosis type I, or hemophilia A.-131-

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