Polymeric system for biological agent delivery

Abstract

This invention relates generally to polymeric delivery systems. The present polymeric delivery systems may be complexed with biological agents, including nucleic acids, peptides, proteins, or small molecules, for delivery to cells. In particular, the present polymeric delivery systems may be used in gene editing.

Description

[0001]PATENT ATTORNEY DOCKET NO.51618-005WO2 POLYMERIC SYSTEM FOR BIOLOGICAL AGENT DELIVERY BACKGROUND OF THE INVENTION Delivery of biological agents, such as nucleic acids, peptides, proteins, or small molecules, to cells both in vitro and in vivo has been performed using various recombinant viral vectors, lipid delivery systems, and electroporation. Such techniques have sought to treat various diseases and disorders by reducing or inhibiting gene expression, providing genetic constructs for gene therapy or to study various biological systems. Despite the vast curative potential of biological agents, widespread clinical deployment faces an uncertain outlook due to difficulties in delivery of the agent to the desired tissue. Novel and efficient polymer-based delivery vehicles are thus desired. SUMMARY OF THE INVENTION The present invention is related to polymeric delivery systems. The present polymeric delivery systems may be complexed with biological agents, including nucleic acids, peptides, proteins, or small molecules, for delivery to cells. In one aspect, the invention features a polymer including a backbone and first and second pendant groups or an ion or salt thereof. The first pendant group comprises an aminoalkyl group or amino aryl group, and the second pendant group comprises an aminoalkyl group, a dialkyl group, a neutral substituted alkyl group, an aryl group or a neutral heteroalkyl group. The first pendant group comprises 5- 95%, and the second pendant group comprises 5-95% of the total number of first and second pendant groups and the first and second pendant groups are different. In some embodiments, the backbone includes acrylate or methacrylate. In some embodiments, the backbone includes acrylamide or methacrylamide. In some embodiments, the backbone includes styrene. In some embodiments, the back includes a combination of any of acrylate, methacrylate, acrylamide, methacrylamide, or styrene. In some embodiments, the polymer includes a third pendant groups or an ion or salt thereof. The first pendant group includes an aminoalkyl group, e.g., a primary aminoalkyl group, the second pendant group includes a quaternary aminoalkyl group, and the third pendant group includes a dialkyl group, a neutral substituted alkyl group, e.g., being hydrophilic or hydrophilic and lipophilic, or a neutral heteroalkyl group, e.g., being hydrophilic or hydrophilic and lipophilic. The first pendant group includes 5-30%, the second pendant group includes 5-15%, and the third pendant group includes 55-90% of the total number of first, second, and third pendant groups. In some embodiments, the polymer includes formula (I) PATENT ATTORNEY DOCKET NO.51618-005WO2 , wherein R1 is -NRARBor ORA, wherein RA and RB are each independently H, aminoalkyl, or aminoheteroalkyl, provided that at least one of RAand RBis aminoalkyl or aminoheteroalkyl, R1’is amino, aminoalkyl, or aminoheteroalkyl, and u is 1 to 5, e.g., 1, and wherein R2is H or CH3; ORC, wherein RCand RDare each independently H or quaternary aminoalkyl or quaternary aminoheteroalkyl, provided that at least one of RCand RDis quaternary aminoalkyl or quaternary aminoheteroalkyl, R3’is quaternary amino, quaternary aminoalkyl, or quaternary aminoheteroalkyl, and v is 1 to 5, e.g., 1, and wherein R4is H or CH3; - , are or provided that at least one of REand RFis neutral substituted alkyl or neutral heteroalkyl, R5’is a neutral hydrophilic group, neutral substituted alkyl or neutral heteroalkyl, and w is 0 to 5, e.g., 1, and wherein R6is H or CH3; each W is independently wherein n is 1 to 8 and each R7is independently H, alkyl, alkenyl, alkynyl, or heteroalkyl; m is from 10 to 1000; a is 0 – 0.8, b is 0 – 0.5, c is 0 – 0.9, and d is 0 - 0.15 ; and each represents a fraction of m, and the sum of a, b, c, and d is 1, provided that at least two of a, b, c, and d are not 0; or an ion or salt thereof. In some embodiments, each X is independently , wherein R1is -NRARBor ORA, wherein RA and RB are each independently H or aminoalkyl, provided that at least one of RAand RBis aminoalkyl, and wherein R2is H or CH3; PATENT ATTORNEY DOCKET NO.51618-005WO2 each Y is independently , wherein R3is -NRCRDor -ORC, wherein RCand RDare each independently H or quaternary aminoalkyl, provided that at least one of RCand RDis quaternary aminoalkyl, and wherein each Z is independently , wherein R5is -NRERFor -ORE, wherein REand RFare each independently H or neutral substituted alkyl or neutral heteroalkyl, provided that at least one of REand RFis neutral substituted alkyl or neutral heteroalkyl and wherein R6is H or CH3; each W is independently , wherein n is 1 to 8 and each R7is independently H, alkyl, alkenyl, alkynyl, or heteroalkyl; m is from 10 to 1000; a is 0.04 – 0.3, b is 0.04 – 0.15, c is 0.4 – 0.9, and d is 0 – 0.15; and each represents a fraction of m, and the sum of a, b, c, and d is 1; or an ion or salt thereof. In some embodiments, a is 0.04 – 0.3, b is 0.04 – 0.15, and c is 0.4 – 0.9. wherein each R8is independently aminoalkyl and q is 1 to 4, or an ion or salt thereof. In some embodiments, q is 3. In some embodiments, , wherein q is 1 to 4, or an ion or salt thereof. In some embodiments, q is 3. wherein each R8is independently alkyl, each R8’is independently quaternary aminoalkyl, and r is 1 to 4, or a salt thereof. In some embodiments, r is 2. In some embodiments, R8is methyl. In some PATENT ATTORNEY DOCKET NO.51618-005WO2 embodiments, , wherein each R8is independently alkyl, and r is 1 to 4, or a salt thereof. In some embodiments, r is 2. In some embodiments, R8is methyl. wherein each R8is independently neutral substituted alkyl or neutral heteroalkyl, each R9is independently H, OH, or SH, and R10is aryl or heteroaryl, wherein s is 2 to 5. In some embodiments, R9is OH. In some embodiments, R10 is phenyl. In some embodiments, s is 3. In some embodiments, Z is , wherein each R9is H, OH, or SH, and R10is aryl or heteroaryl, wherein s is 2 to 5. In some embodiments, R10 is phenyl. In some embodiments, s is 3. In some embodiments, , wherein R7is H, alkyl, alkenyl, alkynyl, or heteroalkyl, e.g., substituted with amino, imino, amido, azido, cyano, cyanato, isocyanato, isothiocyanto, hydrazide, nitro, nitroso, nitrosooxy, hydroxyl, alkoxy, carboxyl, ester, acyl, halo, phosphino, phospho, sulfide, thiol, sulfonyl, sulfo, sulfinyl, or silyl, and n is 1 to 6. In some embodiments, the polymer includes formula (II): wherein R8is CH3 or C2H5, R9is OH or SH, R10is aryl or heteroaryl, or a salt thereof. In some embodiments, R8is methyl. In some embodiments, R9is OH. In some embodiments, R10is phenyl. PATENT ATTORNEY DOCKET NO.51618-005WO2 In some embodiments, the polymer including formula (III): wherein n is 1 to 6, each R7is independently H or alkyl, R8is CH3 or C2H5, R9is OH or SH, R10is aryl or heteroaryl, or a salt thereof. In 7 some embodiments, n is 3. In some embodiments, R is H or or a C1-3 ester or di C0-C3amide thereof, e.g., where one and only one R7is . In some embodiments, R8is methyl. In some embodiments, R9is OH. In some embodiments, R10is phenyl. In some embodiments, the polymer including formula (IV): In some embodiments, the polymer is terminated by a residue of a RAFT, SFRP, or ATRP agent or derivative thereof. In some embodiments, the polymer is terminated at one end with PATENT ATTORNEY DOCKET NO.51618-005WO2 PATENT ATTORNEY DOCKET NO.51618-005WO2 In some embodiments, the polymer further includes polyethylene glycol, e.g., 2000 to 10000 Da, a polynucleic acid, or a polypeptide. In some embodiments, the polymer further includes polyethylene glycol, e.g., 2000 to 10000 Da. In some embodiments, m is from 20 to 500. In some embodiments, m is from 50 to 300. In some embodiments, the first pendant group comprises a primary aminoalkyl group. In another aspect, the invention provides a complex including a polymer as described herein and a negatively charged biological agent. In some embodiments, the complex further includes a second polymer as described herein. In some embodiments, the negatively charged biological agent includes a nucleic acid. In some embodiments, the nucleic acid includes DNA or RNA. In some embodiments, the nucleic acid includes gRNA, mRNA, tmRNA, tRNA, rRNA, siRNA, shRNA, PNA, ssRNA, dsRNA, pDNA, ssDNA, dsDNA, a DNA:RNA hybrid molecule, a plasmid, an artificial chromosome, cDNA, a PCR product, a restriction fragment, a ribozyme, an antisense construct, or a combination thereof. In some embodiments, the negatively charged biological agent includes a protein. In some embodiments, the negatively charged biological agent includes a ribonucleoprotein. In some embodiments, the ribonucleoprotein includes a virus, a ribosome, telomerase, Ribonuclease P (RNase P), a heterogeneous ribonucleoprotein particle (hnRNP), or a small nuclear ribonucleoprotein particle (snRNP). PATENT ATTORNEY DOCKET NO.51618-005WO2 In some embodiments, the protein includes a nuclease. In some embodiments, the nuclease includes a zinc finger nuclease (ZFNs), a transcription-activator like effector nucleases (TALEN), or a Cas protein. In some embodiments, the Cas protein includes Cas2, Cas3, Cas4, Cas5, Cas5e (or CasD), Cas6, Cas6e, Cas6f, Cas7, Cas8a1, Cas8a2, Cas8b, Cas8c, Cas9, Cas10, Cas10d, CasF, CasG, CasH, CjCas9, SpCas9, Cas12, Cas13, Cas14, Cfpl, Casl, CaslB, Cpf1, Csy1, Csy2, Csy3, Cse1 (or CasA), Cse2 (or CasB), Cse3 (or CasE), Cse4 (or CasC), Csc1, Csc2, Csa5, Csn2, Csm2, Csm3, Csm4, Csm5, Csm6, Cmr1, Cmr3, Cmr4, Cmr5, Cmr6, Csb1, Csb2, Csb3, Csx17, Csx14, Csx10, Csx16, CsaX, Csx3, Csz1, Csx15, Csf1, Csf2, Csf3, Csf4, Cu1966, modified versions thereof, or combinations thereof. In some embodiments, the Cas protein is Cas9. In some embodiments, the negatively charged biological agent includes a nucleic acid and a nuclease. In some embodiments, the negatively charged biological agent includes gRNA and a Cas protein. In another aspect, the invention provides a composition including a complex as described herein and a liquid carrier. In another aspect, the invention provides a method including contacting a cell with the complex, wherein the biological agent is delivered into the cell. In some embodiments, the cell is a lung cell. Definitions To facilitate the understanding of this invention, a number of terms are defined below and throughout the disclosure. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology herein is used to describe specific embodiments of the invention, but their usage does not limit the invention, except as outlined in the claims. Terms such as “a”, “an,” and “the” are not intended to refer to only a singular entity but include the general class of which a specific example may be used for illustration. The term “about,” as used herein, refers to a value that is within 10% above or below the value being described. The term “effective amount,” as used herein refers to the amount that is necessary to result in a physiological change in the cell, organism, or tissue to which it is administered. The term “individual” or “subject” is an animal, such as a mammal, bird, amphibian, or reptile. Mammals, as used herein, include, but are not limited to, domesticated animals (e.g. cows, sheep, cats, dogs, and horses), primates (e.g. humans and non-human primates such as monkeys), rabbits, and rodents (e.g. mice and rats). Particularly, the individual or subject is a human. The term “pharmaceutical composition,” as used herein, refers to a preparation which is in such form as to permit the biological activity of an active ingredient contained therein to be effective, and which contains no additional components which are unacceptably toxic to a subject to which the formulation would be administered. The term “pharmaceutically acceptable carrier,” as used herein, refers to an ingredient in a pharmaceutical composition, other than an active ingredient, which is nontoxic to a subject. A PATENT ATTORNEY DOCKET NO.51618-005WO2 pharmaceutically acceptable carrier includes, but is not limited to, a buffer, excipient, stabilizer, or preservative. The term “therapeutically effective amount,” as used herein, e.g., of a pharmaceutical composition, refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired therapeutic or prophylactic result. A therapeutically effective amount of an agent for example eliminates, decreases, delays, minimizes, reduces, or prevents adverse effects of a disease. The term “alkenyl,” as used herein, refers to an acyclic straight or branched chain monovalent hydrocarbon group containing one or more double bonds, no triple bonds, and from 2 to 12 (e.g., 2 to 6) carbons, unless otherwise specified. Alkenyl groups may be substituted or unsubstituted. Exemplary substituents include alkoxy, alkylthio, alkynyl, amido, amino, carbonate, carboxyl, cyano, cycloalkyl, epoxy, halo, heterocyclyl, hydroxyl, oxo, and thiol. The term “alkyl,” as used herein, refers to an acyclic straight or branched chain, saturated, monovalent hydrocarbon group having from 1 to 12 carbons (e.g., 1 to 6), unless otherwise specified. Alkyl groups may be substituted or unsubstituted. Exemplary substituents include alkoxy, alkylthio, amido, amino, carbonate, carboxyl, cyano, cycloalkyl, epoxy, halo, heterocyclyl, hydroxyl, oxo, and thiol. An alkyl may be substituted with an oxo to form an aldehyde or ketone. The term “alkynyl,” as used herein, refers a straight or branched monovalent hydrocarbon group containing one or more triple bonds and from 2 to 12 (e.g., 2 to 6) carbons, unless otherwise specified. Alkynyl groups may be unsubstituted or substituted as alkenyl groups. Exemplary substituents include alkoxy, alkylthio, amido, amino, carbonate, carboxyl, cyano, cycloalkyl, epoxy, halo, heterocyclyl, hydroxyl, oxo, and thiol. The term “alkoxy,” as used herein, refers to a group of the formula RO-, wherein R is an alkyl group as defined herein. Alkoxy groups may be unsubstituted or substituted as alkyl groups. An alkoxy may be substituted with an oxo group to form an ester. Three alkoxy groups may be bound to the same carbon to form an orthoester. The term “alkylthio,” as used herein, refers to a group of the formula RS-, wherein R is an alkyl group as defined herein. Alkylthio groups may be unsubstituted or substituted as alkyl groups. The term “alkylene,” as used herein, refers to a divalent group obtained by removing a hydrogen from a carbon atom of an alkyl group. Alkylene groups may be unsubstituted or substituted as alkyl groups. The term “amido,” as used herein, refers to a group of the formula —C(=O)NRGRH, where each of RGand RHare independently H, alkyl, alkenyl, alkynyl, cycloalkyl, heteroalkyl, heterocyclyl, or aryl. The term “amino,” as used herein, refers to a group of formula —NRGRHor —NRGRHRI, where each of RG, RH, and RIis independently H, alkyl, alkenyl, alkynyl, cycloalkyl, heteroalkyl, heterocyclyl, or aryl. The term “aminoalkyl,” as used herein, refers to an alkyl group substituted with an amino group. Aminoalkyl groups may be primary, secondary, tertiary, or quaternary. The term “aminoheteroalkyl,” as used herein, refers to a heteroalkyl group substituted with an amino group. Aminoheteroalkyl groups may be primary, secondary, tertiary, or quaternary. PATENT ATTORNEY DOCKET NO.51618-005WO2 The term “aryl,” as used herein, refers to any monocyclic or fused ring bicyclic or multicyclic system containing only carbon atoms in the ring(s), which has the characteristics of aromaticity in terms of electron distribution throughout the ring system, e.g., phenyl, naphthyl, or phenanthryl. An aryl group may have, e.g., six to sixteen carbons (e.g., six carbons, ten carbons, thirteen carbons, fourteen carbons, or sixteen carbons). Aryl groups may be unsubstituted or substituted. Exemplary substituents include alkyl, alkenyl, alkynyl, alkoxy, alkylthio, amido, amino, aryl, carbonate, carboxyl, cyano, cycloalkyl, epoxy, halo, heteroalkyl, heterocyclyl, hydroxyl, and thiol. The term "acyl,” as used herein, refers to a group having the general formula -C(=O)RJ, wherein RJis hydrogen, alkyl, heteroalkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, or aryl. The term “azido,” as used herein, refers to —N3. The term “carbonate,” as used herein, refers to a group of the formula —OC(=O)OR, wherein R is H, alkyl, heteroalkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, or aryl. The term “carboxyl,” as used herein, refers to a group of the formula —(C=O)OH. A carboxy group may be esterified or amidated. For esters, the H atom is replaced with alkyl, heteroalkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, or aryl, and for amides, the -OH moiety is replaced with amino. The term “cyano,” as used herein, refers to —C≡N. The term “cyanato,” as used herein, refers to —O-C≡N. The term “cycloalkyl,” as used herein, refers to a cyclic, saturated, monovalent hydrocarbon group having from 3 to 12 carbons (e.g., 3 to 6), unless otherwise specified. Cycloalkyl groups may be substituted or unsubstituted. Exemplary substituents include alkyl, alkenyl, alkynyl, alkoxy, alkylthio, amido, amino, carbonate, carboxyl, cyano, cycloalkyl, epoxy, halo, heterocyclyl, hydroxyl, oxo, and thiol. The term “epoxy,” as used herein, refers to >O, where the oxygen is bound to adjacent carbon atoms. The term “ester,” as used herein, refers to a group having the general formula -C(=O)ORJ, wherein RJis alkyl, heteroalkyl, alkenyl, alkynyl, heterocyclyl, or aryl. The term “halide,” as used herein, refers to a F, Cl, Br, or I anion. The term “halo,” as used herein, refers to a F, Cl, Br, or I radical. The term “heteroalkyl,” as used herein, refers to an acyclic straight or branched chain, saturated, monovalent hydrocarbon group having from 1 to 12 carbons (e.g., 1 to 6), unless otherwise specified, wherein one or more of the carbon atoms has been replaced by a heteroatom selected from S, O, and N. A neutral heteroalkyl group includes only S or O as heteroatoms replacing carbon atoms. Heteroalkyl groups may be substituted or unsubstituted. Exemplary substituents include alkenyl, alkynyl, alkoxy, alkylthio, amido, amino, aryl, carbonate, carboxyl, cyano, cycloalkyl, epoxy, halo, heterocyclyl, hydroxyl, oxo, phospho, and thiol. The term “heteroaryl,” as used herein, refers to an aromatic heterocyclyl group. For example, a single ring heteroaryl group includes pyridyl; fused ring heteroaryl groups include benzimidazolyl, quinolinyl, acridinyl; and a non-fused bi-heteroaryl group includes bipyridinyl. Further examples of heteroaryls include, but are not limited to, furanyl, thienyl, oxazolyl, acridinyl, phenazinyl, benzimidazolyl, benzofuranyl, benzoxazolyl, benzothiazolyl, benzothiadiazolyl, benzothiophenyl, benzoxadiazolyl, PATENT ATTORNEY DOCKET NO.51618-005WO2 benzotriazolyl, imidazolyl, indolyl, isoxazolyl, isoquinolinyl, indolizinyl, isothiazolyl, isoindolyloxadiazolyl, indazolyl, pyridyl, pyridazyl, pyrimidyl, pyrazinyl, pyrrolyl, pyrazinyl, pyrazolyl, purinyl, phthalazinyl, pteridinyl, quinolinyl, quinazolinyl, quinoxalinyl, triazolyl, tetrazolyl, thiazolyl, triazinyl, thiadiazolyl and the like. Heteroaryl groups may be substituted or unsubstituted. Exemplary substituents include alkyl, alkenyl, alkynyl, alkoxy, alkylthio, amido, amino, aryl, carbonate, carboxyl, cyano, cycloalkyl, epoxy, halo, heterocyclyl, hydroxyl, and thiol. The term “heterocyclyl,” as used herein, represents a monovalent, monocyclic or fused ring bicyclic or multicyclic system having at least one heteroatom as a ring atom. For example, a heterocyclyl group may have, e.g., one to fifteen carbon ring atoms (e.g., a C1-C2, C1-C3, C1-C4, C1-C5, C1-C6, C1-C7, C1-C8, C1-C9, C1-C10, C1-C11, C1-C12, C1-C13, C1-C14, or C1-C15 heterocyclyl) and one or more (e.g., one, two, three, four, or five) ring heteroatoms independently selected from the group consisting of nitrogen, oxygen, and sulfur. Heterocyclyl groups may or may not include a ring that is aromatic. Heterocyclyl groups may be unsubstituted or substituted. In preferred embodiments of the invention, a heterocyclyl group is a 3- to 8-membered ring, a 3- to 6-membered ring, a 4- to 6-membered ring, a 5-membered ring, or a 6-membered ring. Exemplary 5-membered heterocyclyl groups may have zero to two double bonds, and exemplary 6-membered heterocyclyl groups may have zero to three double bonds. The term “basic nitrogen containing heterocycle,” as used herein, refers to a heterocyclyl group having at least one ring nitrogen (e.g., 1 to 4 nitrogen atoms, e.g., 1 or 2) that can accept a proton from solution, e.g., imidazolyl or benzimidazolyl. Heterocyclyl groups may be substituted or unsubstituted. Exemplary substituents include alkyl, alkenyl, alkynyl, alkoxy, alkylthio, amido, amino, aryl, carbonate, carboxyl, cyano, cycloalkyl, epoxy, halo, heteroalkyl, heterocyclyl, hydroxyl, and thiol. The term “hydrazide,” as used herein, refers to . The term “imino,” as used herein refers to =N-Rk, where each RKis H, alkyl, or heteroalkyl. The term “isocyanato,” as used herein, refers to —N=C=O. The term “isothiocyanato,” as used herein, refers to —N=C=S. The term “oxo,” as used herein, refers to =O. The term “nitro,” as used herein, refers to —NO2. The term “nitroso,” as used herein, refers to —N=O. The term “nitrosooxy,” as used herein, refers to —O-N=O. The term “phosphino,” as used herein, refers to —PRk2, where each RKis H, alkyl, or heteroalkyl. The term “phospho,” as used herein refers to —P(=O)(ORK)2, wherein each RKis hydrogen, alkyl, alkenyl, alkynyl, heteroalkyl, cycloalkyl, aryl, or heterocyclyl, e.g., heteroaryl. The term “silyl,” as used herein, refers to —SiRL3, wherein each RLis independently H, alkyl, or heteroalkyl. The term “sulfide,” as used herein refers to —SRM, wherein RMis alkyl, alkenyl, alkynyl, heteroalkyl, cycloalkyl, aryl, or heterocyclyl, e.g., heteroaryl. The term “sulfinyl,” as used herein refers to —S(=O)—RM, wherein RMis alkyl, alkenyl, alkynyl, heteroalkyl, cycloalkyl, aryl, or heterocyclyl, e.g., heteroaryl. PATENT ATTORNEY DOCKET NO.51618-005WO2 The term “sulfonyl,” as used herein refers to —S(=O)2—RM, wherein RMis alkyl, alkenyl, alkynyl, heteroalkyl, cycloalkyl, aryl, or heterocyclyl, e.g., heteroaryl. The term “sulfo,” as used herein refers to —S(=O)2O—RM, wherein RMis alkyl, alkenyl, alkynyl, heteroalkyl, cycloalkyl, aryl, or heterocyclyl, e.g., heteroaryl. The term “thiol,” as used herein, refers to —SH. BRIEF DESCRIPTION OF THE DRAWINGS The following detailed description of the embodiments of the invention will be better understood when read in conjunction with the appended drawings. For the purpose of illustrating the invention, there are shown in the drawing embodiments, which are presently exemplified. It should be understood, however, that the invention is not limited to the precise arrangement and instrumentalities of the embodiments shown in the drawings. FIG.1A shows non-PEGylated polymers (APMAm 15% / MAETMA 10% / PhHPMA 75%) assembled into complexes with the biological agent, pDNA, which were evaluated for efficient delivery of pDNA. FIG.1B shows non-PEGylated polymers (APMAm 15% / MAETMA 10% / PhHPMA 75%) assembled into complexes with the biological agent, mRNA, which were evaluated for efficient delivery of mRNA. FIG.2A shows PEGylated polymers (APMAm 15% / MAETMA 10% / PhHPMA 75%) assembled into complexes with the biological agent, pDNA, which were evaluated for efficient delivery of pDNA. FIG.2B shows PEGylated polymers (APMAm 15% / MAETMA 10% / PhHPMA 75%) assembled into complexes with the biological agent, mRNA, which were evaluated for efficient delivery of mRNA. FIG.3A shows polymers (APMAm 65% / MAETMA 10% / PhHPMA 25%) assembled into complexes with the biological agent, pDNA, which were evaluated for efficient delivery of pDNA. FIG.3B shows polymers(APMAm 65% / MAETMA 10% / PhHPMA 25%) assembled into complexes with the biological agent, mRNA, which were evaluated for efficient delivery of mRNA. FIG.4A shows polymers (APMAm 40% / MAETMA 10% / PhHPMA 50%) assembled into complexes with the biological agent, pDNA, which were evaluated for efficient delivery of pDNA. FIG.4B shows polymers (APMAm 40% / MAETMA 10% / PhHPMA 50%) assembled into complexes with the biological agent, mRNA, which were evaluated for efficient delivery of mRNA. FIG.5A shows the biodistribution of polymers from A-2, wherein n = 4 and including one multiplex, across liver, lung, spleen, heart, and kidney. FIG. 5B shows the real quantitation of biodistribution of polymers from A-2, wherein n = 4 and including one multiplex, across liver, lung, spleen, heart, and kidney. FIG.5C shows the biodistribution of polymers from A-2, wherein n = 2 and including one multiplex, across liver, lung, spleen, heart, and kidney. FIG. 5D shows the real quantitation of biodistribution of polymers from A-2, wherein n = 2 and including one multiplex, across liver, lung, spleen, heart, and kidney. FIG.5E shows the biodistribution of polymers from A-2, wherein n = 3 with all as singleplex, across liver, lung, spleen, heart, and kidney. PATENT ATTORNEY DOCKET NO.51618-005WO2 FIG.5F shows the real quantitation of biodistribution of polymers from A-2, wherein n = 3 with all as singleplex, across liver, lung, spleen, heart, and kidney. FIG.5G shows a spider plot of the biodistribution of polymers from the three A-2 complexes. FIG.5H shows a spider plot of the real quantitation of the biodistribution of polymers from the three A-2 complexes. DETAILED DESCRIPTION OF THE INVENTION The disclosure provides polymeric delivery vehicles for the delivery of biological agents. Polymers described herein may be advantageous in their ability to deliver biological agents to a desired tissue, e.g., the lungs. The present polymers may be synthesized from at least two monomers to form a backbone that includes first and second pendant groups. The polymer includes a mixture of charged and / or neutral pendant groups. For example, the first pendant group includes an aminoalkyl group or amino aryl group (e.g., primary, secondary, or tertiary), and the second pendant group includes an aminoalkyl group, a dialkyl group, a neutral substituted alkyl group, an aryl group or a neutral heteroalkyl group , e.g., being hydrophilic or hydrophilic and lipophilic. Some polymers further include poly(ethylene glycol) (e.g., PEG2k), polynucleic acids, polypeptides, and / or disulfide groups in the backbone, or both PEG and disulfide groups. In general, the present disclosure is directed to polymers that may be associated with at least one biological agent payload such as pDNA, ribonucleoproteins (RNP), mRNA, and the like. The complexes may be internalized by a cell via various endocytic routes, the biological agent may be released inside the cell, and it subsequently may enter the cell nucleus to alter gene expression. The polymers disclosed herein thus provide a polymeric scaffold that provides a well-defined host configured to bind with biological macromolecular agents and facilitate intracellular delivery thereof. The polymers have physiochemical properties such as, for example, composition, molecular weight, ζ-potential, pKa, complex diameter, nucleic acid condensation, and combinations thereof selected for efficient nucleic acid payload delivery using, for example, a CRISPR / Cas9 delivery process. The polymers and related complexes may also have good gene editing efficiency, cellular internalization, cytotoxicity, and combinations thereof. The present polymeric delivery systems harness the use of cationic polymers as delivery systems to take advantage of biological agents having a negative charge, e.g., nucleic acids due to the phosphate groups along each nucleotide. The present polymeric delivery vehicles may condense biological agents, such as CRISPR payloads (for example, mRNA, pDNA or RNP), which can vary widely in their lengths, topologies, physical characteristics and biological mechanisms, into discrete nanosized polyelectrolyte complexes. Upon administration, the complexes may navigate both extracellular barriers such as serum DNAases (or RNAases) and reticuloendothelial system clearance, as well as intracellular barriers such as endosomal interrogation and lysosomal degradation. Finally, the biological agent may be released within the spatiotemporal window that is optimal for payload translocation to the nucleus, where the biological agent can undergo further processing and realization of targeted edits. In addition to meeting high PATENT ATTORNEY DOCKET NO.51618-005WO2 standards for safety, efficiency and cost-effectiveness, synthetic delivery systems may minimize immune activation and cellular toxicity. Polymers The present polymers include a backbone and first and second pendant groups. The first pendant group includes an aminoalkyl group or amino aryl group (e.g., primary, secondary, or tertiary), and the second pendant group includes an aminoalkyl group, a dialkyl group, a neutral substituted alkyl group an aryl group or a neutral heteroalkyl group, e.g., being hydrophilic or hydrophilic and lipophilic. The polymer may include the first pendant group and the second pendant group comprises , where each is 5-95% of the total number of first and second pendant groups and the first and second pendant groups are different. Exemplary backbones include styrene, acrylate / acrylamide, methacrylate / methacrylamide, and mixtures thereof. The backbone may also include disulfide groups, e.g., separated by alkylene, polyethylene glycol, polynucleic acids, and / or polypeptides. In some embodiments, the polymer includes a third pendant group or an ion or salt thereof, wherein the first pendant group includes an aminoalkyl group, the second pendant group includes a quaternary aminoalkyl group, and the third pendant group includes a dialkyl group, a neutral substituted alkyl group, or a neutral heteroalkyl group, wherein the first pendant group is 5-30%, the second pendant group is 5-15%, and the third pendant group is 55-90% of the total number of first, second, and third pendant groups. Use of disulfide monomers in the synthesis of the polymers may result in the incorporation of sulfur atoms and disulfide bonds in the backbone. Incorporation of sulfur atoms and disulfide bonds in the backbone introduces sites for cleavage or reduction, e.g., by thiols or antioxidants in vivo. Reduction of disulfides may be pH dependent and, e.g., allow for delivery of the intact polymer inside a cell prior to degradation. Degradation of the polymer results in smaller fragments that may be more able to be excreted or eliminated. The polymer may include formula (I): wherein R1is -NRARBor ORA, wherein RAand RBare each independently H, aminoalkyl, or aminoheteroalkyl, provided that at least one of RAand RBis aminoalkyl or aminoheteroalkyl, R1’is amino, aminoalkyl, or aminoheteroalkyl, and u is 1 to 5, and wherein R2is H or CH3; PATENT ATTORNEY DOCKET NO.51618-005WO2 each Y is independently , wherein R3is -NRCRDor -ORC, wherein RC and RD are each independently H or quaternary aminoalkyl or quaternary aminoheteroalkyl, provided that at least one of RCand RDis quaternary aminoalkyl or quaternary aminoheteroalkyl, R3’is quaternary amino, quaternary aminoalkyl, or quaternary aminoheteroalkyl, and v is 1 to 5, and wherein R4is H or each Z is independently , wherein R5is -NRERFor -ORE, wherein REand RFare each independently H or neutral substituted alkyl or neutral heteroalkyl, provided that at least one of REand RFis neutral substituted alkyl or neutral heteroalkyl, R5’is a neutral hydrophilic group, neutral substituted alkyl or neutral heteroalkyl, and w is 0 to 5, and wherein R6is H or CH3; each W is independently , wherein n is from 1 to 8 and each R7is independently H, alkyl, alkenyl, alkynyl, or heteroalkyl; m is from 10 to 1000; a is 0 – 0.8, b is 0 – 0.5, c is 0 – 0.9, and d is 0 - 0.15 and each represents a fraction of m, and the sum of a, b, c, and d is 1, provided that at least two of a, b, c, and d are not 0; e.g., a is 0 – 0.3, b is 0 – 0.15, c is 0 – 0.9, and d is 0 - 0.15, or an ion or salt thereof. In some embodiments, a is 0.04 – 0.3, b is 0.04 – 0.15, and c is 0.4 – 0.9. In some embodiments, X is wherein each R8is independently aminoalkyl, and q is 1 to 4, or an ion or salt thereof. In some embodiments, Y is PATENT ATTORNEY DOCKET NO.51618-005WO2 independently alkyl, each R8’is independently quaternary aminoalkyl, and r is 1 to 4, or a salt thereof. In some embodiments, Z is neutral substituted alkyl or neutral heteroalkyl, each R9is independently H, OH, or SH, and R10is aryl or heteroaryl, wherein s is 2 to 5. In some embodiments, the monomer employed resulting in the first pendant group or X is 3- aminopropyl methacrylamide. In some embodiments, the monomer resulting in the second pendant group or Y is 3-trimethylaminoethyl methacrylate. In some embodiments, the monomer employed resulting in the third pendant group or Z is selected from PhHPMA (2-hydroxy-3-phenoxypropyl methacrylate), HEMA (hydroxy ethylmethacrylate), PEGMA (poly(ethylene glycol) methacrylate), e.g., PEG300 or PEG500, EGPhEMA (ethylene glycol phenyl ether methacrylate), THFMA (tetrahydrofurfuryl methacrylate), GlyMA (glycidyl methacrylate), MPC (2-methacryloyloxyethyl phosphorylcholine), DMMAm (N,N-dimethylmethacrylamide), and HPMAm (2-hydroxypropyl methacrylate). In some embodiments, each W is independently , wherein n is from 1 to 8, e.g., 1 to 6, e.g., 3, and each R7is independently H, alkyl, alkenyl, alkynyl, or heteroalkyl, e.g., substituted with amino, imino, amido, azido, cyano, cyanato, isocyanato, isothiocyanto, hydrazide, nitro, nitroso, nitrosooxy, hydroxyl, alkoxy, carboxyl, ester, acyl, halo, phosphino, phospho, sulfide, thiol, sulfonyl, sulfo, sulfinyl, or silyl, e.g., as described in WO2023 / 122807. In some embodiments, the monomer employed resulting in group W is lipoic acid, which can be further modified via esterification or amidation. The polymers described herein may be synthesized be any suitable method, e.g., controlled polymerization mechanisms such as atom transfer radical polymerization (ATRP), stable free radical polymerization (SFRP), or reversible addition fragmentation chain transfer (RAFT) polymerization. The present invention may employ RAFT polymerization or ATRP. In particular embodiments, the end groups of formula (I), formula (II), or formula (III) may be from any RAFT or ATRP agent or a chemical derivative thereof. Specific examples include, but are not limited to, RAFT agents that incorporate dithiobenzoate or trithiocarbonate, such as 2-(dodecylthiocarbonothioylthio)-2-methylpropionic acid, 4-cyano-4- PATENT ATTORNEY DOCKET NO.51618-005WO2 [(dodecylsulfanylthiocarbonyl)sulfanyl]pentanoic acid, 2-cyano-2-propyl dodecyl trithiocarbonate, 4-cyano- 4-(phenylcarbonothioylthio)pentanoic acid, and 2-cyano-2-propylbenzodithioate. Specific examples include, but are not limited to, ATRP agents that incorporate halogens, such as methyl-2- bromopropionate, methyl-2-chloropropionate, ethyl-2-bromoisobutyrate, and 2-hydroxyethyl 2-bromo-2- methylpropanoate. End groups may also include modifications made to the reacted RAFT or ATRP agent, e.g. displacement or thioesterification. In some embodiments, the polymer is terminated at one end with PATENT ATTORNEY DOCKET NO.51618-005WO2 ATRP and RAFT agents also include macromolecular agents, such as poly(ethylene)glycol. These also include branched structures that are formed from branched to dendrimer RAFT reagents: PATENT ATTORNEY DOCKET NO.51618-005WO2 , or . Other groups may also be included in the polymers. Such groups include polyethylene glycol, a polynucleic acid (e.g., sequence specific, such as an aptamer), and / or a polypeptide (e.g., sequence specific such as an antibody). Such polymers may be attached by standard conjugation methods known in the art. In some embodiments, the polymer includes formula (II): wherein each R8is independently CH3 or C2H5, R9is OH or SH, R10is aryl or heteroaryl, or a salt thereof. In some embodiments, R8is methyl. In some embodiments, R9is OH. In some embodiments, R10is phenyl. PATENT ATTORNEY DOCKET NO.51618-005WO2 In particular embodiments, the compound of formula (II) is selected from: wherein m is from 10 to 1000; a is 0 – 0.8, b is 0 – 0.5, and c is 0 – 0.9; and each represents a fraction of m, and the sum of a, b, and c is 1; or an ion or salt thereof. In some embodiments, the polymer including formula (III): wherein n is 1 to 6, each R7is independently H or alkyl, each R8is independently CH3 or C2H5, R9is OH or SH, R10is aryl or heteroaryl,or a salt thereof. In some embodiments, n is 3. In some embodiments, C3 ester or di-C0-C3 amide thereof, e.g., where one and only one ester or di-C0-C3 amide thereof. In some embodiments, R8is methyl. In some embodiments, R9is OH. In some embodiments, R10is phenyl. In particular embodiments, the compound of formula (III) is selected from: PATENT ATTORNEY DOCKET NO.51618-005WO2 wherein m is from 10 to 1000; a is 0 – 0.8, b is 0 – 0.5, c is 0 – 0.9, and d is 0 – 0.15; and each represents a fraction of m, and the sum of a, b, c, and d is 1, provided that at least two of a, b, c, and d are not 0; or an ion or salt thereof. In some embodiments, the polymer includes formula (IV): wherein t is from 10 to 100, and formula (IV) is bound to X. In formulas (I), (II), (IIa), (IIb), (III), (IIIa), and (IIIb), m may be from about 10 to about 1000, e.g., from 10 to 20, 10 to 25, 10 to 30, 10 to 40, 10 to 50, 10 to 60, 10 to 70, 10 to 75, 10 to 80, 10 to 90, 10 to 100, 10 to 125, 10 to 150, 10 to 175, 10 to 200, 10 to 250, 10 to 300, 10 to 400, 10 to 500, 10 to 600, 10 to 700, 10 to 800, 10 to 900, 15 to 20, 15 to 25, 15 to 30, 15 to 35, 15 to 40, 15 to 45, 15 to 50, 15 to 55, 15 to 60, 15 to 70, 15 to 75, 15 to 80, 15 to 90, 15 to 100, 16 to 24, 17 to 23, 18 to 22, 19 to 21, 20 to 30, 20 to 40, 20 to 50, 20 to 60, 20 to 70, 20 to 75, 20 to 80, 20 to 90, 20 to 100, 25 to 30, 25 to 35, 25 to 40, 25 to 50, 25 to 75, 25 to 100, 25 to 125, 26 to 34, 27 to 33, 28 to 32, 29 to 31, 30 to 35, 30 to 40, 30 to 45, 30 to 50, 30 to 60, 30 to 70, 30 to 80, 30 to 90, 30 to 100, 35 to 65, 40 to 50, 40 to 60, 40 to 70, 40 to 80, 40 to 90, 40 to 100, 50 to 60, 50 to 70, 50 to 80, 50 to 90, 50 to 100, 50 to 125, 50 to 150, 50 to 175, 50 to 200, 50 to 250, 50 to 300, 50 to 350, 50 to 400, 50 to 450, 50 to 500, 50 to 550, 50 to 600, 50 to 700, 50 to 800, 50 to 900, 50 to 1000, 60 to 100, 60 to 140, 70 to 100, 70 to 130, 80 to 100, 80 to 120, 90 to 100, 100 to 125, 100 to 150, 100 to 200, 100 to 300, 100 to 500, 100 to 750, 100 to 1000, 150 to 200, 150 to PATENT ATTORNEY DOCKET NO.51618-005WO2 250, 200 to 300, 200 to 500, 200 to 700, 200 to 1000, 250 to 500, 250 to 750, 250 to 1000, 300 to 500, 300 to 700, 400 to 500, 400 to 600, 500 to 600, 500 to 750, 500 to 800, 500 to 1000, 750 to 1000, 800 to 900, 800 to 1000, or 900 to 1000, e.g., is about 10, 25, 50, 75, 90, or 100. In formulas (I), (II), (IIa), (IIb), (III), (IIIa), and (IIIb), a may be from about 0 to about 0.8, e.g., from 0 to 0.05, 0 to 0.1, 0 to 0.15, 0.15 to 0.2, 0.15 to 0.25, 0.15 to 0.8, 0.2 to 0.8, 0.25 to 0.8, 0.3 to 0.8, 0.35 to 0.8, 0.4 to 0.8, 0.45 to 0.8, 0.5 to 0.8, 0.55 to 0.8, 0.6 to 0.8, 0.65 to 0.8, 0.7 to 0.8, 0.75 to 0.8, e.g., is about 0, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.55, 0.6, 0.65, 0.7, 0.75 or 0.8. In formulas (I), (II), (IIa), (IIb), (III), (IIIa), and (IIIb), b may be from about 0 to about 0.5, e.g., from 0 to 0.05, 0 to 0.1, 0 to 0.15, 0.15 to 0.2, 0.15 to 0.25, 0.15 to 0.5, 0.2 to 0.5, 0.25 to 0.5, 0.3 to 0.5, 0.35 to 0.5, 0.4 to 0.5, 0.45 to 0.5, e.g., is about 0, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, or 0.5. In formulas (I), (II), (IIa), (IIb), (III), (IIIa), and (IIIb), c may be from about 0 to about 0.9, e.g., from 0 to 0.05, 0 to 0.1, 0 to 0.15, 0.15 to 0.2, 0.15 to 0.25, 0.15 to 0.9, 0.2 to 0.9, 0.25 to 0.9, 0.3 to 0.9, 0.35 to 0.9, 0.4 to 0.9, 0.45 to 0.9, 0.5 to 0.9, 0.55 to 0.9, 0.6 to 0.9, 0.65 to 0.9, 0.7 to 0.9, 0.75 to 0.9, 0.8 to 0.9, 0.85 to 0.9, e.g., is about 0, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.55, 0.6, 0.65, 0.7, 0.75, 0.8, 0.85, or 0.9. In formulas (I), (II), (IIa), (IIb), (III), (IIIa), and (IIIb), d may be from about 0 to about 0.15, e.g., from 0 to 0.05, 0 to 0.1, 0 to 0.15, 0.05 to 0.1, 0.05 to 0.15, 0.1 to 0.15 e.g., is about 0, 0.05, 0.1, 0.15. Pendant groups may be present in the monomers during polymerization or may be added after polymerization. For example, groups substituted with thiols may be reacted with pendant haloalkyl or alkenyl groups on the polymer. Exemplary thiolated groups are shown in Table 1. Neutral and positively charged side chains may be formed. In some embodiments, this is accomplished via conjugation reactions with various thiols as shown in Tables 1 and 2. For other conjugation reactions, the thiol in Tables 1 and 2 may be replaced with another group, e.g., amino, hydroxy, carboxy (or activated derivative), azido, alkyne (linear or cyclic), diene, or alkene. Table 1: Neutral thiols PATENT ATTORNEY DOCKET NO.51618-005WO2 Table 2: Positively charged thiols PATENT ATTORNEY DOCKET NO.51618-005WO2 Complexes The polymers described herein may bind with a biological agent. The biological agent may be bound to the polymer in a variety of methods. In some embodiments, the biological agent is bound noncovalently to the polymer, e.g., electrostatically to the polymer. In some embodiments, the polymer may be complexed with the biological agent. In some embodiments, the polymer may be condensed with the biological agent. Without wishing to be bound to theory, the polymers described herein, including those bound, complexed, or condensed to negatively charged biological agents, may be able to evade the immune system by mimicking bacteria-like, less-foreign morphologies, thereby showing great promise as a multiplexable system. The present disclosure provides a complex including a polymer described herein, e.g., of formula (I), (II), or (III), and a biological agent. In some embodiments, the biological agent is a negatively charged biological agent. The biological agent may include a therapeutic agent. The biological agent may include a nucleic acid, a peptide, a protein, or a small molecule. The biological agent may include a small molecule. Small molecules are compounds with low molecular weight that are capable of modulating biochemical processes to diagnose, treat, or prevent diseases. The present polymeric delivery systems may be used to transport small-molecule therapeutics to cells. The biological agent may include a nucleic acid. Without wishing to be bound to theory, when negatively charged biological agents (e.g., nucleic acids such as RNA or other small molecule therapeutics) are covalently attached to the bottlebrush architecture, the circulation time of the cargo may be increased in vivo. The nucleic acid may be DNA, RNA, or chimeric. In some embodiments, the nucleic acid includes gRNA, mRNA (e.g., that encodes for proteins (fluorescent or therapeutic), tmRNA, tRNA, rRNA, siRNA, shRNA, PNA, ssRNA, dsRNA, pDNA (e.g., that encodes for proteins (fluorescent or therapeutic), ssDNA, dsDNA, a DNA:RNA hybrid molecule, DNA editing templates, miRNA, an artificial chromosome, oligo nucleotide, a nucleic acid encoding a nuclease, cDNA, a PCR product, a restriction fragment, a ribozyme, an antisense construct, or a combination thereof. Nucleic acids may include modification to the sugar, backbone, and / or base and may include synthetic or non-canonical bases. A gRNA includes an engineered, non-naturally occurring Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) gRNA that hybridizes with a target nucleic acid sequence of interest. In various embodiments, DNA editing templates include an exogenous strand of DNA that bears homology arms to a section of genomic DNA that has been cut by a nuclease (for example, Cas9, TALEN or zinc finger) along with an intervening sequence between these homology arms that differs with the natural segment of genomic DNA that has been cut. This intervening segment serves as the template for repair of the cut genomic DNA and, in so doing, the cell corrects its own DNA to match that of the DNA PATENT ATTORNEY DOCKET NO.51618-005WO2 template. The DNA template may be included in a single DNA expression vector that also encodes the nuclease. siRNAs refer to a double-stranded interfering RNA. In addition to siRNA molecules, other interfering RNA molecules and RNA-like molecules may be used. Examples of other interfering RNA molecules that may to inhibit target biomolecules include, but are not limited to, short hairpin RNAs (shRNAs), single-stranded siRNAs, microRNAs (miRNAs), piwiRNA, Dicer-substrate 27-mer duplexes, and variants thereof containing one or more chemically modified nucleotides, one or more non- nucleotides, one or more deoxyribonucleotides, and / or one or more non-phosphodiester linkages. Typically, all RNA or RNA-like molecules that may interact with transcripts RISC complexes and participate in RISC-related changes in gene expression may be referred to as interfering RNAs or “interfering RNA molecules. Suitable interfering RNAs may readily be produced based on the well-known nucleotide sequences of target biomolecules. In various embodiments interfering RNAs that inhibit target biomolecules may include partially purified RNA, substantially pure RNA, synthetic RNA, recombinant produced RNA, as well as altered RNA that differs from naturally occurring RNA by the addition, deletion, substitution, and / or alteration of one or more nucleotides. Such alterations may include, for example, addition of non-nucleotide material, such as to the end(s) of the interfering RNAs or to one or more internal nucleotides of the interfering RNAs, including modifications that make the interfering RNAs resistant to nuclease digestion. Such alterations result in sequences that are generally at least about 80%, or more, or even 100% identical to the sequence of the target biomolecule. When the gene to be down regulated is in a family of highly conserved genes, the sequence of the duplex region may be chosen with the aid of sequence comparison to target only the desired gene. On the other hand, if there is sufficient identity among a family of homologous genes within an organism, a duplex region may be designed that would down regulate a plurality of genes simultaneously. The N / P ratio of a complex is the ratio of positively-chargeable polymer amine (N = nitrogen) groups to negatively-charged nucleic acid phosphate (P) groups. The N / P character of a polymer / nucleic acid complex may influence complex properties, such as its net surface charge, size, and stability of the complex. The N / P ratio of the present complexes may be from 0 to 10, e.g., from 0 to 1, 0 to 2, 0 to 3, 0 to 4, 0 to 5, 0 to 6, 0 to 7, 0 to 8, 0 to 9, 1 to 2, 1 to 3, 1 to 4, 1 to 5, 1 to 6, 1 to 7, 1 to 8, 1 to 9, 1 to 10, 2 to 3, 2 to 4, 2 to 5, 2 to 6, 2 to 7, 2 to 8, 2 to 9, 2 to 10, 3 to 4, 3 to 5, 3 to 6, 3 to 7, 3 to 8, 3 to 9, 3 to 10, 4 to 5, 4 to 6, 4 to 7, 4 to 8, 4 to 9, 4 to 10, 5 to 6, 5 to 7, 5 to 8, 5 to 9, 5 to 10, 6 to 7, 6 to 8, 6 to 9, 6 to 10, 7 to 8, 7 to 9, 7 to 10, 8 to 9, 8 to 10, 9 to 10, or about 0.5, about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, or about 10. Alternatively, or in addition, the biological agent may include a peptide. In various embodiments, peptide fragments include two or more amino acids covalently linked by at least one amide bond. For example, in some embodiments the peptide fragments may include pDNA encoded fluorescent or therapeutic peptides. Alternatively, or in addition, the biological agent may include a protein. In some embodiments, the protein may include an antibody. The antibody may be a monoclonal antibody. PATENT ATTORNEY DOCKET NO.51618-005WO2 In some embodiments, the protein includes a ribonucleoprotein. The ribonucleoprotein may include a ribosome, telomerase, ribonuclease P (RNase P), a heterogeneous ribonucleoprotein particle (hnRNP), or a small nuclear ribonucleoprotein particle (snRNP). sgRNA may be chemically modified to improve stability and prevent intracellular degradation. In some embodiments, the protein may include a nuclease. The nuclease may include a zinc finger nuclease (ZFNs), a transcription-activator like effector nucleases (TALEN), or a Cas protein. The Cas protein may be a Cas2, Cas3, Cas4, Cas5, Cas5e (or CasD), Cas6, Cas6e, Cas6f, Cas7, Cas8a1, Cas8a2, Cas8b, Cas8c, Cas9, Cas10, Cas10d, CasF, CasG, CasH, CjCas9, SpCas9, Cas12, Cas13, Cas14, Cfpl, Casl, CaslB, Cpf1, Csy1, Csy2, Csy3, Cse1 (or CasA), Cse2 (or CasB), Cse3 (or CasE), Cse4 (or CasC), Csc1, Csc2, Csa5, Csn2, Csm2, Csm3, Csm4, Csm5, Csm6, Cmr1, Cmr3, Cmr4, Cmr5, Cmr6, Csb1, Csb2, Csb3, Csx17, Csx14, Csx10, Csx16, CsaX, Csx3, Csz1, Csx15, Csf1, Csf2, Csf3, Csf4, Cu1966, modified versions thereof, or combinations thereof. In some embodiments, the Cas protein is Cas9. In some embodiments, the biological agent includes a nucleic acid and a nuclease. The negatively charged biological agent may be gRNA and a Cas protein, as in the CRISPR-Cas system. The CRISPR-Cas system is useful for precise editing of genomic nucleic acids (e.g., for creating null mutations). For example, a composition containing only the guide RNA can be administered to an animal or cells transgenic for the Cas9 enzyme. Similar strategies may be used (e.g., zinc finger, transcription activator-like effectors (TALEs) or homing meganucleases). The CRISPR-Cas system is known in the art for deleting, modifying genome sequences or incorporating transgenes. Transgene refers to any nucleotide sequence, particularly a DNA sequence, that is integrated into one or more chromosomes of a host cell by human intervention, such as by the methods of the present invention. For example, a transgene can be an RNA coding region or a gene of interest, or a nucleotide sequence, preferably a DNA sequence, that is used to mark the chromosome where it has integrated or may indicate a position where nucleic acid editing, such as by the CRISPR- CAS system, may occur. In this situation, the transgene does not have to include a gene that encodes a protein that may be expressed. CRISPR-Cas genome editing has rapidly emerged as a multi-faceted technology to enable gene insertion, deletion, activation, suppression, and even single base editing of target genes within the nucleus of any cell. This highly efficient and facile technique has broad utility from white biotechnology and agriculture to biomedical research, pharmaceutics, and regenerative medicine. Currently, the CRISPR / Cas9 system can be delivered in vitro, ex vivo, and in vivo in three different payload forms: i) pDNA that encodes Cas9 protein and / or sgRNA ii) mRNA that encodes for Cas9 nuclease and a separate sgRNA, or iii) a ribonucleoprotein (RNP) that consists of recombinant Cas9 protein precomplexed directly with a sgRNA. CRISPR-Cas9 pDNA needs to enter the cellular nucleus to express, and consistent expression produces an overabundance of Cas9 protein, which can lead to increased off-target editing and mutagenesis. Researchers have utilized the CRISPR / Cas9 system in mRNA form to circumvent the barrier of nuclear entry, which has been reported with polymer-based nanoparticles. However, sgRNA often needs to be delivered separately, presenting challenges in trafficking kinetics of different payloads. PATENT ATTORNEY DOCKET NO.51618-005WO2 Direct delivery of CRISPR / Cas9 ribonucleoprotein (RNP) has several benefits, including precision in endonuclease dosing and potential to avoid uncontrolled integration of the transgene into the cellular genome. Formulations In another aspect, the present disclosure is directed to compositions including the polymer complexes described which have been dispersed in a solution. In some embodiments, the composition includes a plurality of polymers described herein which have been dispersed in a solution, e.g., a mixture of 2, 3, 4, 5, or more. When multiple polymers are employed, they may differ in the fraction of monomeric units, i.e., in o and n, or they may differ in the chemical structure of the monomeric units. In some embodiments, the complexes may be added to a liquid carrier and stored in liquid form until needed, or alternatively may be dried and introduced into and dispersed in the liquid carrier prior to use, e.g., administration to a subject. In some embodiments the liquid carrier is a pharmaceutically acceptable carrier. Some non- limiting examples of materials which can serve as pharmaceutically-acceptable carriers include: pyrogen- free water; isotonic saline; Ringer’s solution; ethyl alcohol; pH buffered solutions. The formulation may include solvents, dispersing medium (containing, e.g., water, cell culture medium, buffers (e.g., phosphate buffered saline), polyol (for example, glycerol, propylene glycol, or liquid polyethylene glycol), wetting agents, emulsifying agents, coloring agents, release agents, coating agents, sweetening agents, flavoring agents, perfuming agents, pH buffering agents, gelling or viscosity enhancing additives, preservatives, colors, preservatives, or antioxidants. The polymers, e.g., of formula (I), (II), or (III), may be ionized in dry or liquid formulation. For example, amine or basic nitrogen groups may be protonated. The polymers, e.g., of formula (I), (II), or (III) may be in salt form with one or more anions, e.g., acetate, ascorbate, benzoate, bicarbonate, bisulfate, carbonate, cholate, citrate, dihydrogen citrate, glycocholate, halide (such as F, Cl, Br, or I), hydrogen citrate, hydroxide, mandelate, methanesulfonate, nitrate, oxalate, p-toluenesulfonate, persulfate, phosphate, lactate, succinate, sulfate, sulfite, tartrate, taurocholate, or trifluoroacetate. Methods of Use The present disclosure provides methods of contacting a cell with the herein-described complexes, wherein the biological agent is delivered into the cell. For example, after a composition including the polymer and bound biological agent are contacted with a cell, the complexes internalize into the cell, and the biological agent disassociate partially or completely from the polymeric carriers. The formulations described herein can be delivered to a cell or an organism via any administration mode known to a skilled practitioner. For example, the formulations described herein can be delivered via administration routes such as, but not limited to, in vitro, oral, intravenous, intramuscular, intraperitoneal, intradermal, and subcutaneous. In some embodiments, the compositions described herein are in a form that is suitable for injection. In other embodiments, the formulations described herein are PATENT ATTORNEY DOCKET NO.51618-005WO2 formulated for oral administration. In various embodiments, for in vivo administration a delivery device can be used to facilitate the administration of any composition described herein to a subject, e.g., a syringe, a dry powder injector, a nasal spray, a nebulizer, or an implant such as a microchip, e.g., for sustained release or controlled release of any formulation described herein. In various embodiments, the compositions may be administered to a cell in vitro by removing a cell from a subject, culturing the cells, applying to the cells a composition including polymer vehicles and bonded biological agent to deliver a therapeutic amount of the biological agent into at least a portion of the cells, and optionally re-introducing the cell to the subject. In another embodiment, a tissue cell therapy technique may be used in which a tissue sample is removed from a subject, a composition including a polymer and a bonded biological agent is applied to the tissue to deliver a therapeutic amount of the biological agent to modify a selected cell or region of the tissue, and the modified tissue is transplanted into the subject. EXAMPLES The following examples are put forth so as to provide those of ordinary skill in the art with a complete disclosure and description of how the methods and compounds claimed herein are performed, made, and evaluated, and are intended to be purely exemplary of the invention and are not intended to limit the scope of what the inventors regard as their invention. Example Polymer Synthesis To a dried 4 mL glass vial, the reactants (e.g., APMAm / MAETMA / PhHPMA, chain transfer agent, thermal initiator; 800 to 10 to 1 eq., respectively at desired mmol for targeted monomer compositions) were dissolved in acetic acid / methanol (1:1 v / v). The solution was sealed, degassed with nitrogen for at least 30 min, and set at 60 °C under constant stirring for 18 h. The reaction was quenched by cooling to room temperature and exposure to air. Crude solutions were purified with dialysis against filtered Milli-Q water for 3 cycles of 8 h each, followed by lyophilization to afford free flowing yellow solids. Polyplex Preparation Polymer stock solutions were prepared in 4 mL glass vials by direct dissolution of polymer in filtered Milli-Q water at 150 mM total concentration. Samples were vortexed for at least 30 sec. Polyplex assemblies were prepared by mixing polymer stock solutions with nucleic acid stock solutions at specific N / P ratios (number of nitrogen to phosphate between polymer and nucleic acid, respectively). Solutions are allowed to equilibrate for at least 30 min before further bioanalysis. Results of various A-2 polyplexes are shown below in Table 3, and all four A-2 polyplexes were successful at delivering a biological agent. PATENT ATTORNEY DOCKET NO.51618-005WO2 Table 3: Components of A-2 (Y = yes, included; N = no, not included) Other embodiments are in the claims.

Claims

PATENT ATTORNEY DOCKET NO.51618-005WO2 What is claimed is: CLAIMS 1. A polymer comprising a backbone and first and second pendant groups or an ion or salt thereof, wherein the first pendant group comprises an aminoalkyl group or amino aryl group, the second pendant group comprises an aminoalkyl group, a dialkyl group, a neutral substituted alkyl group, an aryl group or a neutral heteroalkyl group, wherein the first pendant group comprises 5-95%, the second pendant group comprises 5-95% of the total number of first and second pendant groups and the first and second pendant groups are different.

2. The polymer of claim 1, further comprising a third pendant group or an ion or salt thereof, wherein the first pendant group comprises an aminoalkyl group, the second pendant group comprises a quaternary aminoalkyl group, and the third pendant group comprises a dialkyl group, a neutral substituted alkyl group, or a neutral heteroalkyl group, wherein the first pendant group comprises 5-30%, the second pendant group comprises 5-15%, and the third pendant group comprises 55-90% of the total number of first, second, and third pendant groups.

3. The polymer of claim 1, comprising formula (I)wherein each X is independentlywherein R1is -NRARBor ORA, wherein RAand RBare each independently H, aminoalkyl, or aminoheteroalkyl, provided that at least one of RAand RBis aminoalkyl or aminoheteroalkyl, R1’is amino, aminoalkyl, or aminoheteroalkyl, and u is 1 to 5, and wherein R2is each Y is independently, wherein R3is - NRCRDor -ORC, wherein RC and RD are each independently H or quaternary aminoalkyl or quaternary aminoheteroalkyl, provided that at least one of RCand RDis quaternary aminoalkyl or quaternary aminoheteroalkyl, R3’is quaternary amino, quaternary aminoalkyl, or quaternary aminoheteroalkyl, and v is 1 to 5, and wherein R4is H or CH3;PATENT ATTORNEY DOCKET NO.51618-005WO2 each Z is independently, wherein R5is - NRERFor -ORE, wherein REand RFare each independently H or neutral substituted alkyl or neutral heteroalkyl, wherein alkyl is substituted with a neutral hydrophilic group, provided that at least one of REand RFis neutral substituted alkyl or neutral heteroalkyl, R5’is a neutral hydrophilic group, neutral substituted alkyl or neutral heteroalkyl, and w is 0 to 5, and wherein R6is H or CH3; each W is independently, wherein n is from 1 to 8 and each R7is independently H, alkyl, alkenyl, alkynyl, or heteroalkyl; m is from 10 to 1000; a is 0 – 0.8, b is 0 – 0.5, c is 0 – 0.9, and d is 0 - 0.15 and each represents a fraction of m, and the sum of a, b, c, and d is 1, provided that at least two of a, b, c, and d are not 0; or an ion or salt thereof.

4. The polymer of claim 3, wherein a is 0.04 – 0.3, b is 0.04 – 0.15, and c is 0.4 – 0.

9.

5. The polymer of claim 3, wherein X iswherein each R8is independently aminoalkyl, and q is 1 to 4, or an ion or salt thereof. The polymer of claim 5, wherein q is 3. The polymer of any one of claims 3-6, wherein Y iswherein each R8is independently alkyl, each R8’is independently quaternary aminoalkyl, and r is 1 to 4, or a salt thereof.PATENT ATTORNEY DOCKET NO.51618-005WO2 8. The polymer of claim 7, wherein r is 2.

9. The polymer of claim 7 or 8, wherein each R8is methyl.

10. The polymer of claim 3, wherein Z isindependently neutral substituted alkyl or neutral heteroalkyl, each R9is independently H, OH, or SH, and each R10is independently aryl or heteroaryl, wherein s is 2 to 5.

11. The polymer of claim 10, wherein R9is OH.

12. The polymer of claim 10 or 11, wherein R10is phenyl.

13. The polymer of any one of claims 10-12, wherein s is 3. The polymer of claim 3, whereinalkyl, alkenyl, alkynyl, or heteroalkyl, and n is 1 to 6.

15. The polymer of claim 3, comprising formula (II):wherein R8is CH3 or C2H5; R9is OH or SH; R10is aryl or heteroaryl;PATENT ATTORNEY DOCKET NO.51618-005WO2 or a salt thereof.

16. The polymer of claim 15, wherein R8is methyl.

17. The polymer of claim 15 or 16, wherein R9is OH.

18. The polymer of any one of claims 15-17, wherein R10is phenyl.

19. The polymer of claim 3, comprising formula (III):wherein n is 1 to 6; each R7is independently H or alkyl; R8is CH3 or C2H5; R9is OH or SH; R10is aryl or heteroaryl; or a salt thereof.

20. The polymer of claim 19, wherein n is 3. The polymer of claim 19 or 20, whereinester or di-C0-C3 amide thereof.

22. The polymer of any one of claims 19-21, wherein R8is methyl.

23. The polymer of any one of claims 19-22, wherein R9is OH.

24. The polymer of any one of claims 19-23, wherein R10is phenyl.PATENT ATTORNEY DOCKET NO.51618-005WO2 25. The polymer of claim 3, further comprising formula (IV)wherein t is from 10 to 100, wherein formula (IV) is bound to X.

26. The polymer of any one of claims 1 to 25, wherein the polymer is terminated by a residue of a RAFT, SFRP, or ATRP agent or derivative thereof.

27. The polymer of claim 26, wherein the polymer is terminated at one end with, wherein U is H or alkyl, and V is alkylene, or.PATENT ATTORNEY DOCKET NO.51618-005WO229. The polymer of any one of claims 1 to 28, further comprising polyethylene glycol, a polynucleic acid, or a polypeptide.

30. The polymer of any one of claims 3 to 29, wherein m is from 20 to 500.

31. The polymer of claim 30, wherein m is from 50 to 300.PATENT ATTORNEY DOCKET NO.51618-005WO2 32. The polymer of any one of claims 1-30, wherein the first pendant group comprises a primary aminoalkyl group.

33. A complex comprising a first polymer of any one of claims 1 to 32 and a negatively charged biological agent.

34. The complex of claim 33, further comprising a second polymer of any one of claims 1 to 32.

35. The complex of claim 33 or 34, wherein the negatively charged biological agent comprises a nucleic acid.

36. The complex of claim 35, wherein the nucleic acid comprises DNA or RNA.

37. The complex of claim 36, wherein the nucleic acid comprises gRNA, mRNA, tmRNA, tRNA, rRNA, siRNA, shRNA, PNA, ssRNA, dsRNA, pDNA, ssDNA, dsDNA, a DNA:RNA hybrid molecule, a plasmid, an artificial chromosome, cDNA, a PCR product, a restriction fragment, a ribozyme, an antisense construct, or a combination thereof.

38. The complex of claim 33 or 34, wherein the negatively charged biological agent comprises a protein.

39. The complex of claim 33 or 34, wherein the negatively charged biological agent comprises a ribonucleoprotein.

40. The complex of claim 39, wherein the ribonucleoprotein comprises a virus, a ribosome, telomerase, Ribonuclease P (RNase P), a heterogeneous ribonucleoprotein particle (hnRNP), or a small nuclear ribonucleoprotein particle (snRNP).

41. The complex of claim 38, wherein the protein comprises a nuclease.

42. The complex of claim 41, wherein the nuclease comprises a zinc finger nuclease (ZFNs), a transcription-activator like effector nucleases (TALEN), or a Cas protein.

43. The complex of claim 42, wherein the Cas protein comprises Cas2, Cas3, Cas4, Cas5, Cas5e (or CasD), Cas6, Cas6e, Cas6f, Cas7, Cas8a1, Cas8a2, Cas8b, Cas8c, Cas9, Cas10, Cas10d, CasF, CasG, CasH, CjCas9, SpCas9, Cas12, Cas13, Cas14, Cfpl, Casl, CaslB, Cpf1, Csy1, Csy2, Csy3, Cse1 (or CasA), Cse2 (or CasB), Cse3 (or CasE), Cse4 (or CasC), Csc1, Csc2, Csa5, Csn2, Csm2, Csm3, Csm4, Csm5, Csm6, Cmr1, Cmr3, Cmr4, Cmr5, Cmr6, Csb1, Csb2, Csb3, Csx17, Csx14, Csx10,PATENT ATTORNEY DOCKET NO.51618-005WO2 Csx16, CsaX, Csx3, Csz1, Csx15, Csf1, Csf2, Csf3, Csf4, Cu1966, modified versions thereof, or combinations thereof.

44. The complex of claim 42, wherein the Cas protein is Cas9.

45. The complex of claim 33 or 34, wherein the negatively charged biological agent comprises a nucleic acid and a nuclease.

46. The complex of claim 45, wherein the negatively charged biological agent comprises gRNA and a Cas protein.

47. A composition comprising the complex of any one of claims 33 to 46 and a liquid carrier.

48. A method comprising contacting a cell with the complex of any one of claims 33 to 44, wherein the biological agent is delivered into the cell.

49. The method of claim 48, wherein the cell is a lung cell.

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