Polycarbonate-nucleic acid complex and use thereof

Abstract

Provided herein is a polycarbonate-nucleic acid complex. Also provided herein are a preparation method for the complex and use of the complex for delivering a nucleic acid.

Description

Polycarbonate nucleic acid complexes and uses thereof TECHNICAL FIELD

[0001] The present invention relates to the field of nucleic acid delivery; in particular, to a novel polycarbonate nanocarrier nucleic acid complex, a method for preparing the same, and uses thereof for delivering nucleic acids. BACKGROUND

[0002] Due to poor stability and difficulty in penetrating cell membranes, nucleic acid molecules such as DNA and RNA need to be delivered to the biological target site with the aid of delivery systems including nucleic acid carriers in current medical applications such as gene therapy and vaccine development.

[0003] Existing nucleic acid delivery systems still face many challenges. Known nucleic acid carriers such as viral carriers, cationic lipid carriers and polymer carriers have problems such as low loading and delivery efficiency, immune response and biological toxicity, which pose potential risks to the health of patients. In addition, some carrier materials such as PEI can accumulate in the body, causing long-term side effects. These problems seriously limit the large-scale production and industrial application of nucleic acid carriers.

[0004] Therefore, how to improve the delivery efficiency and safety of the delivery system is an important problem to be solved in the art. SUMMARY

[0005] After long-term extensive exploration and research, the inventors found that a complex nanoparticle formed by a new type of polymer carrier and nucleic acid can overcome the long-standing problems of low delivery efficiency and high biological toxicity that are prevalent in existing nucleic acid delivery systems, providing a safe, effective and reliable means for nucleic acid delivery, suitable for large-scale production and industrial application.

[0006] In one aspect, provided herein is a polycarbonate nucleic acid complex comprising:

[0007] (a) a polycarbonate having a repeating unit of Formula I or a salt thereof:

[0008] wherein,

[0009] A comprises at least one nitrogen-containing group;

[0010] R 1 each independently at each occurrence is selected from the group consisting of hydrogen, deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, substituted or unsubstituted ester, substituted or unsubstituted C 1-12 alkyl, substituted or unsubstituted C 1-12 alkoxy, substituted or unsubstituted C 1-12 alkylamino, substituted or unsubstituted C 1-12Alkyl ester group, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 Aryl, substituted or unsubstituted 3- to 10-membered heterocyclic alkyl and substituted or unsubstituted 5- to 14-membered heteroaryl;

[0011] a1 and a2 are each independent integers from 1 to 16;

[0012] b1 and b2 are each independent integers from 0 to 2; and

[0013] (b) At least one nucleic acid.

[0014] On the other hand, this article provides a method for preparing polycarbonate, comprising a polymerization reaction using at least one monomer having formula II and at least one monomer having formula III:

[0015] In Equations II and III,

[0016] A 1 and A 2 Each independently comprises one or more groups selected from the group consisting of formula IV:

[0017] in,

[0018] M is selected from: nitrogen-containing groups, redox-sensitive groups, hydrocarbon groups, oxygen-containing groups, and combinations thereof;

[0019] x1 and x2 are each an independent integer from 0 to 2;

[0020] y1 and y2 are each independent integers from 0 to 16;

[0021] R 1 Each group is independently selected from the following groups when it appears: hydrogen, deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, substituted or unsubstituted ester group, substituted or unsubstituted C group. 1-12 Alkyl, substituted or unsubstituted C 1-12 Alkoxy, substituted or unsubstituted C 1-12 Alkylamino, substituted or unsubstituted C 1-12 Alkyl ester group, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 Aryl, substituted or unsubstituted 3- to 10-membered heterocyclic alkyl and substituted or unsubstituted 5- to 14-membered heteroaryl;

[0022] Among them, A 1 and A 2 At least one of them has a nitrogen-containing group;

[0023] wherein, for A 1 and A 2 when comprising two or more groups independently of Formula IV, the two or more groups independently of Formula IV are connected to each other via L M wherein L M is a bond, a carbonate linkage, or a C 1-16 alkylene; and

[0024] wherein, the wavy line represents a bonding position to an adjacent atom.

[0025] In another aspect, provided herein is a method of making a polycarbonate nucleic acid complex, comprising:

[0026] - providing a polycarbonate or a salt thereof, and at least one nucleic acid; and

[0027] - mixing the polycarbonate or the salt thereof with the at least one nucleic acid. In another aspect, provided herein is a composition comprising the polycarbonate nucleic acid complex.

[0028] In another aspect, provided herein is a product comprising:

[0029] - a polycarbonate nucleic acid complex described herein;

[0030] - a composition described herein; or

[0031] - a polycarbonate or a salt thereof described herein and at least one nucleic acid.

[0032] In another aspect, provided herein is a method of delivering at least one nucleic acid to a cell or a subject, the method comprising:

[0033] - providing the complex or a composition comprising the complex, the complex being formed from a polycarbonate or a salt thereof complexed with the at least one nucleic acid; and

[0034] - contacting the cell or the subject with the complex or the composition.

[0035] In another aspect, provided herein is a method of expressing at least one nucleic acid in a cell or a subject, the method comprising:

[0036] - providing the complex or a composition comprising the complex, the complex being formed from a polycarbonate or a salt thereof complexed with the at least one nucleic acid; and

[0037] - contacting the cell or the subject with the complex or the composition.

[0038] - contacting the cell or the subject with the complex or the composition.

[0039] In another aspect, provided herein is use of the polycarbonate nucleic acid complex or a composition comprising the same for delivering at least one nucleic acid to a cell or a subject.

[0040] In another aspect, provided herein is use of the polycarbonate nucleic acid complex or a composition comprising the same for expressing at least one nucleic acid in a cell or a subject.

[0041] In another aspect, provided herein is use of the polycarbonate nucleic acid complex or a composition comprising the same for preparing a product for delivering at least one nucleic acid to a cell or a subject.

[0042] In another aspect, provided herein is use of the polycarbonate nucleic acid complex or a composition comprising the same for preparing a product for expressing at least one nucleic acid in a cell or a subject. BRIEF DESCRIPTION OF DRAWINGS

[0043] The present application is further described below by reference to the accompanying drawings, in which these show only embodiments of the present application and are not intended to limit the scope of the present application.

[0044] FIG. 1 shows a schematic diagram of polycarbonate forming a polycarbonate nucleic acid complex with nucleic acid according to the present disclosure.

[0045] FIG. 2 shows the effect of cell transfection using an exemplary polycarbonate / Luc-mRNA complex.

[0046] FIG. 3 shows the safety test results of treating cells using an exemplary polycarbonate / Luc-mRNA complex.

[0047] FIG. 4 shows the effect of cell transfection using an exemplary polycarbonate / pCMV-luc complex.

[0048] FIG. 5 shows the safety test results of treating cells using an exemplary polycarbonate / pCMV-luc complex.

[0049] FIG. 6 shows the CDS nucleotide sequence of Luc-mRNA used for preparing polycarbonate / Luc-mRNA complex (SEQ ID No.: 1).

[0050] FIG. 7 shows the full-length nucleotide sequence of Luc-mRNA used for preparing polycarbonate / Luc-mRNA complex (SEQ ID No.: 2). DETAILED DESCRIPTION

[0051] Nucleic acid molecules, as the material that carries and transmits the genetic information of organisms, are one of the most critical biological macromolecules in cells. DNA is responsible for storing genetic codes and guiding cell activities, while RNA is involved in various biological processes, including acting as a messenger of genetic information (mRNA), participating in protein synthesis (tRNA and rRNA), and regulating gene expression (small RNA). In modern medicine, by delivering specific nucleic acid drugs to the target site, antigen proteins can be expressed to stimulate immune responses, or genetic information that causes adverse consequences can be regulated or corrected to prevent or treat various diseases or conditions, including inflammatory conditions, cancer, genetic diseases, etc.

[0052] However, despite the huge market potential of nucleic acid drugs, their actual application faces many obstacles. Nucleic acid molecules are extremely susceptible to degradation before reaching the target site, and often cannot effectively cross biological barriers (such as cell membranes) to enter the interior of cells, because their large molecular weight and negative charge limit their ability to pass through hydrophobic cell membranes.

[0053] Taking messenger RNA (mRNA) as an example, as a transient, encodable genetic information carrier transcribed from a DNA strand and carrying protein synthesis coding information, mRNA can guide the translation and expression of functional proteins in cells. mRNA-based therapy can produce almost any functional protein by delivering mRNA encoding different proteins to the body for translation and expression, and thus has the potential to be used for the treatment of various (intractable) diseases, including infectious diseases, metabolic genetic diseases, cancer, cardiovascular and cerebrovascular diseases, etc. Recently, two mRNA vaccines (mRNA-1273 and BNT162b2) produced by Moderna and Pfizer-BioNTech have been successfully used to prevent COVID-19, highlighting the huge potential of mRNA technology in revolutionizing life sciences and medical research. In addition to COVID-19 vaccines, in recent years, there have been many mRNA vaccines targeting other infectious diseases (such as respiratory syncytial virus, seasonal influenza, rabies virus), cancer (such as melanoma), as well as mRNA drugs for protein therapy and gene editing in different clinical trial stages. With the breakthrough development of nucleic acid chemistry and RNA biology technology, such as the modification of traditional linear mRNA untranslated regions (UTRs) and the emergence of circular mRNA, the most concerning problems of immunogenicity and protein expression in mRNA have been largely solved. Therefore, the challenge of clinical translation of the therapeutic potential of mRNA drugs mainly comes from the development of mRNA delivery carriers.

[0054] However, mRNA is a polyanionic compound with large molecular weight, which is difficult to cross the non-polar cell membrane and tissue barrier. In addition, it is easily inactivated by rapid destruction by nucleases. Therefore, the delivery of a functional mRNA into the cytoplasm requires the protection of a delivery carrier and endosome escape to exert a therapeutic effect. Due to poor degradation performance, the existing mRNA delivery carriers generally have problems of toxicity and delayed nucleic acid release (low transfection efficiency). Specifically, if the delivery carrier is not degradable or the degradation product has high toxicity, it will cause safety problems; if the carrier degrades too slowly, it will cause the mRNA to be unable to be released quickly, which restricts the transfection efficiency.

[0055] Therefore, the development of safe and effective nucleic acid delivery carriers has become the biggest bottleneck restricting the release of the potential application of current nucleic acid drugs. The successful development of such carriers not only greatly improves the delivery efficiency and thus improves the efficacy of nucleic acids, but also enables the delivery process to be sufficiently safe for widespread application and promotion in multiple fields. This breakthrough will greatly promote the development of personalized and precision medicine, drive the transformation of the pharmaceutical industry, and significantly improve the efficacy and quality of life of patients.

[0056] After long-term extensive exploration and research, the inventors have developed a new type of carbonate polymer, which can be used as a polymeric nanocarrier for delivering nucleic acids. The polymeric nanocarrier is stable in vitro and can be rapidly and completely self-degraded after entering the target location (such as cells) for testing or pharmacology, and the degradation products are all small molecules with high safety (such as CO2, thiol, etc.). Due to its unique physicochemical properties, this material can form a complex with a wide range of molecular weights and lengths and different types of nucleic acids, effectively protecting them from enzymatic degradation, enhancing their stability, and promoting the transport across the cell membrane, thereby significantly improving the delivery efficiency of target nucleic acid molecules. Moreover, the nanomaterial has a simple and environmentally friendly preparation process, which is suitable for large-scale production to meet the needs of extensive testing and industrial applications, and provides a new choice for the field of nucleic acid delivery.

[0057] Definitions

[0058] To facilitate better understanding of the present disclosure, some key terms are first defined. Unless otherwise explicitly stated, each of the terms listed below should be understood based on the appended definitions. Unless otherwise stated, the meaning of other terms herein is consistent with the general understanding of those skilled in the art.

[0059] As used herein, "a" or a combination of "a" and various articles of

[0060] As used herein, each "embodiment" refers to and encompasses the embodiments of the methods and systems described herein. One or more features of any embodiment can be combined with one or more features of any one or more other embodiments, resulting in an embodiment that is within the scope of the disclosure.

[0061] All percentages and ratios used herein are by weight, unless otherwise specified. All temperatures are in degrees Celsius (°C) unless otherwise specified. Room temperature or ambient temperature means a temperature of 20 °C to 28 °C (e.g., 25 °C). All measurements are understood to be made at ambient conditions, i.e., at room temperature, at about one atmosphere of pressure, and at about 50% relative humidity, unless otherwise designated.

[0062] All numerical ranges are inclusive of narrower ranges unless otherwise indicated. The endpoints of the ranges described are to be included within the range unless otherwise indicated.

[0063] As used herein, the term "polycarbonate" refers to a class of polymers in which at least two repeating units are connected by carbonate linkages -OC(O)O-. The terms "polycarbonate", "carbonate polymer" are used interchangeably herein and mean the same thing. It is understood that when "polycarbonate" is referred to herein, it means polycarbonates and salt forms thereof. The salt forms of the polycarbonates are formed by salification with anions such as chloride, acetate, citrate, tetrafluoroacetate, and the like.

[0064] As used herein, the term "polymer" refers to a high molecular weight compound formed by the linking of repeating monomeric units through covalent bonds. The repeating monomeric units can be the same, forming a "homopolymer", or different, forming a "copolymer".

[0065] As used herein, the term "repeating unit" refers to a polymerized unit that occurs at least twice in the backbone of a polymer. Repeating units are typically derived from monomers. The term "monomer" refers to a small molecule compound that is capable of linking into a long chain molecule through polymerization. During polymerization, multiple chemical bonds of the monomer molecules are broken and reformed, resulting in the repeating linkage of monomeric units to form the backbone of a polymer.

[0066] Herein, the term "linear polymer" refers to a polymer having a linear structure of molecular chains. The linear polymer has a small number of side groups that do not belong to a branched structure. The term "crosslinked polymer" refers to a polymer having a three-dimensional network structure (e.g., by a crosslinking reaction).

[0067] Herein, the term "random copolymer" refers to a product obtained by random copolymerization, in which two or more monomer units are randomly distributed on the main chain, and no monomer unit forms a separate long segment on the polymer chain. The term "block copolymer" refers to a copolymer in which two or more monomer units each form a long sequence segment and are covalently bonded to each other.

[0068] Herein, the term "main chain" refers to the longest linear molecular chain constituting a polymer. All other molecular chains can be considered side groups or side chains with respect to the main chain. For example, the main chain of a polymer can be mainly composed of carbon atoms connected by covalent bonds, and can be interrupted by nitrogen, oxygen, sulfur, and the like. In general, end groups possessed by the ends of a polymer are not included in the main chain. When referring to "having" a certain group or atom in the main chain, or a certain group or atom "located in the main chain", it means that the group or atom constitutes a part of the main chain of the polymer (not a side group or an end). It can be considered that cleavage at the position of a certain group or atom "located in the main chain" will result in cleavage of the molecular chain of the polymer.

[0069] Herein, the term "amino" refers to the functional group of an amine, and can include primary amino, secondary amino, tertiary amino, quaternary amino.

[0070] Herein, the term "alkyl" refers to a saturated hydrocarbon group, which can be linear or branched (containing branched) structure, and is generally a chain group without a cyclic structure. When referring to "C 1-22 alkyl", it means an alkyl group having a number of carbon atoms of 1 to 22. Examples of such alkyl groups can include methyl, ethyl, propyl (such as n-propyl), butyl (such as isopropyl, n-butyl, isobutyl, t-butyl), pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, and the like, and branched isomers thereof.

[0071] Herein, the term "alkenyl" refers to an unsaturated hydrocarbon group containing at least one carbon-carbon double bond (C=C), which can be linear or branched (containing branched) structure, and is generally a chain group without a cyclic structure. When referring to "C 2-22 alkenyl", it means an alkenyl group having a number of carbon atoms of 2 to 22. Examples of such alkenyl groups can include ethenyl, 1-propenyl, 2-propenyl, 1-butenyl, 2-butenyl, 3-butenyl, 2-methylbut-2-enyl.

[0072] Herein, the term "alkynyl" refers to a hydrocarbyl group containing at least one carbon-carbon triple bond (C≡C), which can be straight-chain or branched (containing branching) in structure, and is typically a chain group without cyclic structure. When referring to "C 2-22 alkynyl," it is meant to refer to an alkynyl group having a number of carbon atoms from 2 to 22. Examples of such alkynyl groups can include ethynyl, 1-propynyl, 2-propynyl, 1-butynyl, 2-butynyl, 3-butynyl, 1-methylpent-2-ynyl.

[0073] Herein, the terms "alkyl," "alkenyl," and "alkynyl" can be collectively referred to as "hydrocarbyl." When referring to "C 1-22 hydrocarbyl," it is meant to refer to a straight-chain or branched hydrocarbyl group having a number of carbon atoms from 1 to 22, i.e., an alkane, alkene, or alkyne. "Heterohydrocarbyl" refers to a hydrocarbyl group having at least one heteroatom in the backbone, e.g., O, N, S, P, Si, etc.

[0074] Herein, the term "cyclohydrocarbyl" refers to a saturated or unsaturated cyclic hydrocarbyl group, including "cycloalkyl" (cyclic alkyl), "cycloalkenyl" (cyclic alkenyl), and "cycloalkynyl" (cyclic alkynyl). When referring to "C 1-20 cyclohydrocarbyl," it is meant to refer to a cyclohydrocarbyl group having a number of carbon atoms from 1 to 20. Examples of such cyclohydrocarbyl groups can include monocyclic hydrocarbyl groups, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, cyclodecyl, cyclopentenyl, cyclohexenyl, cycloheptenyl, cyclopentynyl, cyclohexynyl, cycloheptynyl; bridged cyclic hydrocarbyl groups, such as spirocyclic, norbornyl; fused cyclic hydrocarbyl groups, such as decalinyl, adamantyl.

[0075] Herein, the term "heterocyclohydrocarbyl" refers to an organic cyclic group including at least one (e.g., 1, 2, or 3) ring heteroatom selected from the group consisting of N, O, S, Si, and P (e.g., selected from N, O, and S). Examples of such heterocycloalkyl groups can include morpholinyl, piperazinyl, piperidinyl, furanyl, pyrrolyl, thienyl, pyranyl, and the like.

[0076] Herein, the term "heteroatom" refers to a non-carbon atom selected from oxygen (O), nitrogen (N), sulfur (S), phosphorus (P), and silicon (Si), among others.

[0077] Herein, "aryl" refers to a cyclic group having aromaticity, i.e., having a planar or near-planar ring structure. Most aryl groups have a benzene ring structure. Aryl groups can be broadly classified into monocyclic aryl groups (e.g., phenyl), polycyclic aryl groups (e.g., biphenyl, benzhydryl, trityl), and fused aryl groups (e.g., naphthyl, anthryl, phenanthryl, pyrenyl, azulenyl, tetraphenyl).

[0078] Herein, the term "5- to 14-membered heteroaryl" refers to an aryl group having 5 to 14 (i.e., 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14) ring skeletal atoms and including at least one (e.g., 1, 2, 3, or 4) ring heteroatom selected from the group consisting of N, O, S, Si, and P (e.g., selected from N, O, and S). Such heteroaryl groups can be monocyclic or fused rings condensed with at least one benzene ring, and can be a heteroaryl formed by connecting at least one heteroaryl or aryl group to a heteroaryl group via one or more single bonds. Such heteroaryl groups can include monocyclic-type heteroaryl groups and fused ring-type heteroaryl groups. Examples of monocyclic-type heteroaryl groups can include furanyl, thienyl, pyrrolyl, imidazolyl, pyrazolyl, thiazolyl, thiadiazolyl, isothiazolyl, isoxazolyl, oxazolyl, oxadiazolyl, carbazolyl, triazinyl, tetrazinyl, triazolyl, tetrazolyl, furazanyl, pyridyl, pyrazinyl, pyrimidinyl, and pyridazinyl. Examples of fused ring-type heteroaryl groups can include benzofuranyl, benzothienyl, isobenzofuranyl, dibenzofuranyl, dibenzothienyl, benzoimidazolyl, benzothiazolyl, benzoisothiazolyl, benzoisoxazolyl, benzoaxazolyl, isoindolyl, indolyl, benzoindolyl, indazolyl, benzothiadiazolyl, quinolyl, isoquinolyl, cinnolinyl, quinazolinyl, quinoxalinyl, benzoquinoxalinyl, naphthidinyl, carbazolyl, benzocarbazolyl, phenoxazinyl, phenothiazinyl, phenanthridinyl, benzodioxolyl, and dihydroacridinyl.

[0079] Herein, the term "3- to 10-membered heterocycloalkyl" refers to an organic cyclic group having 3 to 10 (i.e., 3, 4, 5, 6, 7, 8, 9, 10, or a range formed by any two of the above values as end values) ring skeletal atoms and including at least one (e.g., 1, 2, or 3) ring heteroatom selected from the group consisting of N, O, S, Si, and P (e.g., selected from N, O, and S). Examples of such heterocycloalkyl groups can include morpholinyl, piperazinyl, piperidinyl, furanyl, pyrrolyl, thienyl, pyranyl, and the like. The heterocycle can be an alicyclic (such as tetrahydrofuran) or an aromatic ring (such as pyridine). Complex heterocyclic systems can be formed by the fusion of two or more simple rings (such as indole).

[0080] Herein, "C x-y " means that the corresponding group or moiety has x to y (inclusive) carbon atoms. For example, C 1-22 represents the case where the number of carbon atoms is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, or a range formed by any two of the above values as end values. "C 3-20 ", "C 1-16 ", "C 1-12 ", "C 1-8 ", "C 1-6 ", "C1-4 “C” 3-14 “C” 6-14 "etc." have similar meanings.

[0081] In this article, the term "halogen" includes fluorine (F), chlorine (Cl), bromine (Br), and iodine (I).

[0082] In this article, the term "ester group" refers to a carboxylic acid ester group, i.e., a molecular chain containing -C(O)OR. x The structural group, wherein R x C 1- 12 Alkyl groups, such as C 1-8 Alkyl, C 1-6 Alkyl or C 1-4 Alkyl groups. Examples of such ester groups may include methyl ester, ethyl ester, propyl ester, butyl ester, etc.

[0083] In this article, the term "amide group" refers to a group represented by any of the following chemical formulas: -CONH2, -CONH-, -CON<.

[0084] In this text, the term "substituted or unsubstituted" refers to the substitution of a hydrogen atom in a functional group by another atom or functional group (i.e., a substituent). For example, a substituted methyl group refers to a methyl group (-CH) in which one or more hydrogen atoms are replaced by other atoms or functional groups (i.e., substituents). In this text, the substituents of a "substituted" group can be independently selected from: deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, ester, C... 1-22 Alkyl, Halogenated C 1-22 Alkyl, C 2-22 alkenyl, C 2-22 alkynyl group, C 1-22 Alkoxy, C 1-22 Alkylthio, C 1-22 Alkyl ester group, C 3-20 Cycloalkyl, 3- to 10-membered heterocycloalkyl, C 6-20 Aryl, C 6-20 aryloxy group, C 6-20 Arylthioyl, 5- to 14-membered heteroaryl, C 1-22 Alkyl C 3-20 cycloalkyl, C 1-22 Alkyl 3- to 10-membered heterocyclic alkyl, C 1-22 Alkyl C 6-20 Aryl, C 1-22 Alkylsilyl, C 6-20 Arylsilane, C 1-22 Alkyl C 6-20 Arylsilane, C 1-22 Alkylamino, C 6-20 arylamino, C 1-22alkyl C 6-20 aryl C 1-22 alkylcarbonyl C 1-22 alkoxycarbonyl C 6-20 arylcarbonyl C 6-20 arylboronyl C 1-22 alkylboronyl C 1-22 alkyl C 6-20 arylboronyl and C 6-20 aryl C 1-22 alkyl.

[0085] Herein, the term "redox-sensitive group" refers to a group whose molecular structure is responsive to one or both of an oxidative or a reductive environment; that is, a group that is susceptible to chemical reaction under oxidative or reductive conditions. An exemplary oxidative environment includes a high reactive oxygen species (ROS) environment. An exemplary reductive environment includes a high glutathione (GSH) environment.

[0086] Herein, the term "complex" means the product resulting from the complexation of a polycarbonate described herein with at least one nucleic acid through non-covalent interactions (e.g., electrostatic interactions, ionic interactions, hydrogen bonding, or van der Waals forces, etc.). "Polycarbonate complex", "carbonate polymer complex", "PC complex" mean the same and are used interchangeably herein.

[0087] Herein, the term "nucleotide", also referred to as "mononucleotide", is formed by the linkage of a base, a five-carbon sugar (deoxyribose or ribose), and a phosphate in a specific way. It is the building block of nucleic acids (e.g., DNA, RNA). Nucleotides can be polymerized into dinucleotides, trinucleotides, and oligonucleotides (fewer than 25 nucleotides), polynucleotides (more than 25 nucleotides), etc., through phosphodiester bonds.

[0088] Herein, the term "nucleic acid" refers to a class of widely existing biological macromolecules. Depending on the composition, structure, and function, it can be mainly divided into ribonucleic acid (RNA) and deoxyribonucleic acid (DNA). RNA is formed by the linkage of nucleotides through 3,5'-phosphodiester bonds, and DNA is formed by the linkage of deoxynucleotides through 3',5'-phosphodiester bonds. Herein, the term "endogenous nucleic acid" refers to a nucleic acid molecule that naturally exists in a cell or an organism, is produced or inherited by the delivery target cell or organism itself; the term "exogenous nucleic acid" refers to a nucleic acid molecule that cannot be produced or inherited by the delivery target cell or organism itself, and is from the outside.

[0089] In this document, "pharmaceutically acceptable" means a substance that, within reasonable medical judgment, is suitable for contact with human and animal tissues without excessive toxicity, irritation, allergic reactions, or other problems or complications, and is commensurate with a reasonable benefit / risk ratio. The term "pharmaceutically acceptable carrier" includes pharmaceutically acceptable substances, compositions, or carriers suitable for administration to mammals of the polycarbonates or polycarbonate complexes described herein.

[0090] In this article, the term "unit dose" refers to the amount of substance expected to produce a single effect (e.g., prevention or treatment). Unit doses are typically prepared for a single use or for a single subject.

[0091] Polycarbonate nucleic acid complex

[0092] On the one hand, this article provides a polycarbonate nucleic acid complex comprising:

[0093] (a) a polycarbonate or a salt thereof having nitrogen-containing repeating units; and

[0094] (b) At least one nucleic acid.

[0095] Specifically, the polycarbonate or its salt having nitrogen-containing repeating units has repeating units of Formula I:

[0096] in,

[0097] A contains at least one nitrogen-containing group;

[0098] R 1 Each group is independently selected from the following groups when it appears: hydrogen, deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, substituted or unsubstituted ester group, substituted or unsubstituted C group. 1-12 Alkyl, substituted or unsubstituted C 1-12 Alkoxy, substituted or unsubstituted C 1-12 Alkylamino, substituted or unsubstituted C 1-12 Alkyl ester group, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 Aryl, substituted or unsubstituted 3- to 7-membered heterocyclic alkyl and substituted or unsubstituted 5- to 14-membered heteroaryl;

[0099] a1 and a2 are each an integer from 1 to 16, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or an integer within a range formed by taking any two of the above values ​​as endpoints.

[0100] b1 and b2 are each an integer from 0 to 2, for example, 0, 1 or 2.

[0101] The polycarbonates described herein have one or more nitrogen-containing groups in their repeat units (e.g., repeat units of Formula I). The one or more nitrogen-containing groups can be located in the backbone and / or pendant groups of the polycarbonate. Preferably, at least one nitrogen-containing group is located in the backbone of the polycarbonate. For example, at least one (e.g., at least 1 or 2) nitrogen atom of the at least one nitrogen-containing group is located in the backbone of the polycarbonate. In some embodiments, the number of nitrogen-containing groups or nitrogen atoms located in the backbone can be greater than the number of nitrogen-containing groups or nitrogen atoms located in the pendant groups. In some embodiments, the repeat units of Formula I contain no more than 5, 4, 3, 2, 1 nitrogen-containing groups or nitrogen atoms in the pendant groups. In some embodiments, some or all of the nitrogen-containing groups or nitrogen atoms in the repeat units of Formula I are located in the backbone of the polycarbonate. In some embodiments, greater than 20%, greater than 30%, greater than 40%, greater than 50%, greater than 60%, greater than 70%, greater than 80%, greater than 90%, or 100% of the nitrogen-containing groups or nitrogen atoms in the repeat units of Formula I are located in the backbone of the polycarbonate rather than in the pendant groups.

[0102] Advantageously, the polycarbonates are protonatable. Specifically, the polycarbonates are protonatable due to the incorporation of at least one protonatable nitrogen-containing group in their repeat units (e.g., repeat units of Formula I). In some embodiments, the at least one protonatable nitrogen-containing group has at least one protonatable nitrogen atom. In some embodiments, the protonatable atom of the protonatable nitrogen-containing group is nitrogen. In some embodiments, for example in the repeat units of Formula I, at least one protonatable nitrogen-containing group or at least one protonatable nitrogen atom is located in the backbone of the polycarbonate.

[0103] In some embodiments, the protonatable nitrogen-containing group is a divalent group.

[0104] In some exemplary embodiments, the protonatable nitrogen-containing group can include a group selected from the group consisting of:

[0105] a group having a protonatable nitrogen atom located in the backbone and no pendant groups;

[0106] a group having a protonatable nitrogen atom located in the backbone and pendant hydrocarbon groups (e.g., saturated or unsaturated hydrocarbon groups);

[0107] a group having a protonatable nitrogen atom located in the backbone and pendant heteroatom-containing (e.g., one or more of O, N, S, P, Si) groups.

[0108] The protonatable groups on the polymer are capable of absorbing protons and becoming positively charged, thereby imparting or increasing the positive charge of the polymer. Without wishing to be bound by a particular theory, it is found that this change can advantageously affect the physicochemical properties of the polymer, helping it to effectively load nucleic acids.

[0109] In some embodiments, the protonatable nitrogen-containing groups contained in the polycarbonate can have a group selected from the group consisting of an amino group, a nitrogen-containing cyclic group, and any combination thereof.

[0110] In the case where at least one of the protonatable groups comprises at least one amino group, the amino group can be selected from the group consisting of a secondary amino group, a tertiary amino group, a quaternary amino group, and combinations thereof.

[0111] In some embodiments, the amino group can be a monovalent amino group (having the structure -N(R 3 )2or -N(R 3 )3 + ) or a divalent amino group (having the structure -N(R 3 )- or -N(R 3 )2 + ), wherein R 3 is selected from hydrogen, deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, amide, carbamate, urea, substituted or unsubstituted ester, substituted or unsubstituted C 1-22 alkyl, substituted or unsubstituted C 2-22 alkenyl, substituted or unsubstituted C 2-22 alkynyl, substituted or unsubstituted C 1-22 alkoxy, substituted or unsubstituted C 1-22 alkylamino, substituted or unsubstituted C 1-22 alkyl ester, substituted or unsubstituted C 3-20 cycloalkyl, substituted or unsubstituted C 6-14 aryl, substituted or unsubstituted 3- to 10-membered heterocycloalkyl, substituted or unsubstituted 5- to 14-membered heteroaryl, substituted or unsubstituted C 1-22 alkyl 3- to 10-membered heterocycloalkyl, and substituted or unsubstituted C 1-22 alkyl 5- to 14-membered heteroaryl.

[0112] In various specific embodiments, non-limiting examples of R 3 may include:

[0113] wherein,

[0114] n1and n2are each independently at each occurrence an integer from 0 to 21, for example, 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, or an integer within a range formed by any two of the above recited numerical values as endpoints;

[0115] n3 is at each occurrence independently an integer from 2 to 21, for example, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, or an integer in a range formed between any two of these values;

[0116] n4 and n5 are at each occurrence independently an integer from 0 to 8, for example, 0, 1, 2, 3, 4, 5, 6, 7, 8, or an integer in a range formed between any two of these values;

[0117] n6 is at each occurrence independently an integer from 0 to 8, for example, 0, 1, 2, 3, 4, 5, 6, 7, 8, or an integer in a range formed between any two of these values;

[0118] n7 is at each occurrence independently an integer from 1 to 8, for example, 1, 2, 3, 4, 5, 6, 7, 8, or an integer in a range formed between any two of these values;

[0119] wherein the wavy line represents the position of bonding with the adjacent atom; and

[0120] Optionally, the above groups can be substituted with one or more substituents selected from the group consisting of deuterium, tritium, hydroxyl, carboxyl, nitro, amino, halogen, cyano, ester, C 1-12 alkyl, C 2-12 alkenyl, C 2-12 alkynyl, C 1-12 alkoxy, C 1-12 alkylamino, C 1- 12 alkyl ester, C 3-14 cycloalkyl, C 6-14 aryl, 3- to 10-membered heterocycloalkyl, 5- to 14-membered heteroaryl, C 1-12 alkyl 5- to 14-membered heteroaryl, C 1-12 alkyl 3- to 10-membered heterocycloalkyl, and any combination thereof.

[0121] In some embodiments, the polycarbonate comprises, in the backbone, an amino group, or specifically, one or more of a secondary amino group, a tertiary amino group, a quaternary amino group, or more specifically, one or more of a nitrogen atom of a secondary amino group, a tertiary amino group, a quaternary amino group. In some embodiments, the polycarbonate (e.g., in the backbone) can be free of a secondary amino group as the only protonatable nitrogen-containing group. In some embodiments, the polycarbonate (e.g., in the backbone) can be free of a secondary amino group as a protonatable nitrogen-containing group. In some embodiments, the polycarbonate (e.g., in the backbone) can be free of an amino group other than a tertiary amino group as the only protonatable nitrogen-containing group. In some embodiments, the polycarbonate (e.g., in the backbone) can be free of an amino group other than a tertiary amino group as a protonatable nitrogen-containing group.

[0122] In the case where the at least one protonatable group comprises at least one nitrogen-containing ring group, the nitrogen-containing ring group can be selected from the group consisting of: a 3- to 10-membered saturated or unsaturated aliphatic or aromatic heterocyclic group having at least one ring nitrogen atom, a 5- to 20-membered heterocyclic group fused from the 3- to 10-membered saturated or unsaturated aliphatic or aromatic heterocyclic group having at least one ring nitrogen atom, each other, or with an aliphatic or aromatic carbocyclic ring, and combinations thereof. Examples of such nitrogen-containing ring groups can include, but are not limited to: (substituted or unsubstituted) aziridinyl, (substituted or unsubstituted) azetidinyl, (substituted or unsubstituted) pyrrolidinyl, (substituted or unsubstituted) thiazolidinyl, (substituted or unsubstituted) oxazolidinyl, (substituted or unsubstituted) pyrazolidinyl, (substituted or unsubstituted) imidazolidinyl, (substituted or unsubstituted) triazolidinyl, (substituted or unsubstituted) piperidinyl, (substituted or unsubstituted) morpholinyl, (substituted or unsubstituted) thiomorpholinyl, (substituted or unsubstituted) piperazinyl, (substituted or unsubstituted) hexahydropyrimidinyl, (substituted or unsubstituted) triazinanyl, (substituted or unsubstituted) azepinyl, (substituted or unsubstituted) oxazepinyl, (substituted or unsubstituted) homopiperazinyl, (substituted or unsubstituted) triazepinyl, (substituted or unsubstituted) aziridinyl, (substituted or unsubstituted) azetidinyl, (substituted or unsubstituted) pyrrolyl, (substituted or unsubstituted) pyrazolyl, (substituted or unsubstituted) imidazolyl, (substituted or unsubstituted) triazolyl, (substituted or unsubstituted) pyridinyl, (substituted or unsubstituted) dihydropyridinyl, (substituted or unsubstituted) tetrahydropyridinyl, (substituted or unsubstituted) pyrazinyl, (substituted or unsubstituted) pyrimidinyl, (substituted or unsubstituted) pyridazinyl, (substituted or unsubstituted) azepinyl, (substituted or unsubstituted) diazepinyl, (substituted or unsubstituted) triazepinyl, (substituted or unsubstituted) thiazolyl, (substituted or unsubstituted) thiazolinyl, (substituted or unsubstituted) thiazinyl, (substituted or unsubstituted) oxazinyl, (substituted or unsubstituted) azocinyl, (substituted or unsubstituted) oxazocinyl, (substituted or unsubstituted) azacyclononanyl, (substituted or unsubstituted) oxazacyclononanyl, (substituted or unsubstituted) azacyclodecanyl, (substituted or unsubstituted) oxazacyclodecanyl, (substituted or unsubstituted) benzothiazolyl, (substituted or unsubstituted) benzothiazinyl, and 5- to 20-membered heterocyclic groups fused from each other or with an aliphatic or aromatic carbocyclic ring, and combinations thereof.

[0123] In some embodiments, the nitrogen-containing ring group can have a structure of Formula la:

[0124] wherein,

[0125] Y 1 is N or C; and

[0126] Ring R A is a 3- to 20-membered aliphatic heterocycle having 1-6 heteroatoms selected from N, O, S, and combinations thereof;

[0127] Optionally, one or more ring carbon atoms and / or one or more ring nitrogen atoms of Ring R A are each independently substituted with one or two R 2 ; wherein R 2 is each independently at each occurrence selected from the group consisting of deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, amido, carboxylic ester, urea, substituted or unsubstituted C 1-12 alkyl, substituted or unsubstituted C 1-12 alkoxy, substituted or unsubstituted C 1-12 alkylamino, substituted or unsubstituted C 1-12 alkyl ester, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 aryl, substituted or unsubstituted 3- to 10-membered heterocycloalkyl, and substituted or unsubstituted 5- to 14-membered heteroaryl; and, two adjacent R 2 may form a ring together with the ring atom to which they are attached (e.g., forming a monocyclic or bicyclic ring);

[0128] wherein the wavy line represents a bonding position to an adjacent atom.

[0129] In some embodiments, Ring R A may be a 3- to 20-membered aliphatic heterocycle (e.g., a 3- to 10-membered aliphatic heterocycle, such as a 3- to 7-membered aliphatic heterocycle) having 1-4 (e.g., 1, 2, 3, or 4) heteroatoms selected from N, O, S, and combinations thereof.

[0130] In some embodiments, the nitrogen-containing ring group can have the structure of Formula Ib:

[0131] wherein,

[0132] Y 2 and Y 4 are each independently N or C, and Y 2 and Y 4 at least one of which is N;

[0133] Y 3 is selected from the group consisting of: a bond (e.g., a single bond), C, N, O, and S;

[0134] Y 5 is selected from the group consisting of: a bond (e.g., a single bond), C, N, O, and S;

[0135] n is, at each occurrence, independently an integer from 0 to 2;

[0136] Optionally, one or more ring carbon atoms and / or one or more ring nitrogen atoms of the above-mentioned nitrogen-containing ring group are each independently substituted with one or two R B 2 wherein each R 2 is independently defined above with respect to Formula la;

[0137] wherein the wavy line indicates a position of bonding to an adjacent atom.

[0138] In some embodiments, the ring R B may be a 3- to 20-membered aliphatic heterocyclic ring (e.g., 3- to 10-membered aliphatic heterocyclic ring, such as 3- to 7-membered aliphatic heterocyclic ring) having 1 to 4 (e.g., 1, 2, 3, or 4) heteroatoms selected from N, O, S, and combinations thereof.

[0139] In some example embodiments, the nitrogen-containing ring group can include the following groups:

[0140] Optionally, one or more ring carbon atoms and / or one or more ring nitrogen atoms of the above-mentioned nitrogen-containing ring group are each independently substituted with one or two R 2 2 wherein each R

[0141] Optionally, one or more (e.g., 1, 2, or more) ring carbon atoms of the above-mentioned nitrogen-containing ring group are each independently replaced with a heteroatom selected from N, O, and S;

[0142] wherein the wavy line indicates a position of bonding to an adjacent atom.

[0143] In some embodiments, the polycarbonate includes a nitrogen-containing ring group in the main chain. In some embodiments, the nitrogen-containing ring group is connected to the main chain via one or both ring nitrogen atoms thereof. In some embodiments, the polycarbonate (e.g., in the main chain) can be free of other nitrogen-containing groups as the only protonatable nitrogen-containing group other than the nitrogen-containing ring group. In some embodiments, the polycarbonate (e.g., in the main chain) can be free of other nitrogen-containing groups as the protonatable nitrogen-containing group other than the nitrogen-containing ring group.

[0144] ​​The protonable nitrogen-containing group may be unsubstituted or substituted with one or more substituents. Exemplary substituents may include: deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, amide, urethane, urea, substituted or unsubstituted ester groups, substituted or unsubstituted C groups. 1-12 Alkyl, substituted or unsubstituted C 1-12 Alkoxy, substituted or unsubstituted C 1-12 Alkylamino, substituted or unsubstituted C 1-12 Alkyl ester group, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 Aryl, substituted or unsubstituted 3- to 10-membered heterocyclic alkyl and substituted or unsubstituted 5- to 14-membered heteroaryl; and, the two adjacent substituents may form a ring together with the ring atom to which they are attached (e.g., forming a monocyclic or bicyclic ring).

[0145] In some embodiments, in addition to the protonable nitrogen-containing groups described above, the polycarbonate may also contain other nitrogen-containing groups in its repeating units (e.g., repeating units of Formula I), such as, but not limited to, nitrile, isonitrile, azide, hydrazone, nitro, nitroso, azaaryl, amide, urethane, urea, etc. In some embodiments, the other nitrogen-containing groups may be located in the main chain and / or side groups of the polycarbonate.

[0146] In some cases, the polycarbonate described herein may also contain one or more additional groups selected from the group consisting of redox-sensitive groups, hydrocarbon groups (e.g., saturated or unsaturated, substituted or unsubstituted chain hydrocarbon groups or cyclic hydrocarbon groups), oxygen-containing groups (e.g., ether groups, epoxy groups, ketone groups, etc.), and combinations thereof. The one or more additional groups may be monovalent or divalent groups.

[0147] In some embodiments, the one or more additional groups may be located on the main chain and / or side groups of the polycarbonate.

[0148] In some embodiments, the one or more additional groups may be located in the nitrogen-containing repeating unit of the polycarbonate (e.g., the repeating unit of Formula I) or other repeating units.

[0149] The polycarbonate described herein may have at least one (e.g., 1, 2, 3, 4, or 5) redox-sensitive groups in its nitrogen-containing repeating unit (e.g., a repeating unit of Formula I, such as structure A) or other repeating units. In some embodiments, the redox-sensitive groups include oxidation-sensitive and / or reduction-sensitive groups. In some embodiments, the redox-sensitive groups include reduction-sensitive groups.

[0150] Such redox-sensitive groups can include, for example, a single sulfur bond (-S-), a single selenium bond (-Se-), a disulfide bond (-S-S-), a diselenide bond (-Se-Se-), a trisulfide bond (-S-S-S-), a triselenide bond (-Se-Se-Se-), a tetrasulfide bond (-S-S-S-S-), a ketosulfide bond where R a and R b each independently is a C 1-3 alkyl group, a wavy line represents a bonding position to an adjacent atom.

[0151] The one or more redox-sensitive groups can be located in the main chain and / or side groups of the polycarbonate. For example, at least one redox-sensitive group is located in the main chain of the polycarbonate. In some embodiments, the number of redox-sensitive groups located in the main chain of the polycarbonate can be greater than the number of redox-sensitive groups located in the side groups in the repeating unit of Formula I. In some embodiments, the repeating unit of Formula I contains no more than 5, 4, 3, 2, 1, or no redox-sensitive groups in the side groups. In some embodiments, some or all of the redox-sensitive groups in the repeating unit of Formula I are located in the main chain of the polycarbonate. In some embodiments, greater than 50%, greater than 60%, greater than 70%, greater than 80%, greater than 90%, or 100% of the redox-sensitive groups in the repeating unit of Formula I are located in the main chain of the polycarbonate rather than in the side groups.

[0152] The polycarbonates described herein can also have one or more (e.g., 1, 2, 3, 4, 5, or 6) hydrocarbyl groups in its nitrogen-containing repeating unit (e.g., the repeating unit of Formula I, e.g., the A structure) or other repeating unit.

[0153] Such hydrocarbyl groups can include, for example, saturated or unsaturated, substituted or unsubstituted, chain or cyclic hydrocarbyl groups, etc. In some embodiments, the hydrocarbyl groups can include unsubstituted or substituted C 1-24 (e.g., C 1-20 , C 1-16 , or C 1-12 hydrocarbyl groups (e.g., saturated or unsaturated, straight chain or cyclic aliphatic hydrocarbyl groups or aromatic hydrocarbyl groups), where the substituents can be selected from hydrogen, deuterium, tritium, hydroxyl, halogen (e.g., fluorine, chlorine, bromine, iodine), carboxyl, nitro, amino, cyano, ether, substituted or unsubstituted C 1-12 ester groups, substituted or unsubstituted C 1-12 alkyl groups, substituted or unsubstituted C 1-12 alkoxy groups, substituted or unsubstituted C 1-12 alkylamino groups, substituted or unsubstituted C1-12 alkyl ester group, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 aryl, substituted or unsubstituted 3- to 10-membered heterocycloalkyl, and substituted or unsubstituted 5- to 14-membered heteroaryl.

[0154] The one or more hydrocarbyl groups can be located in the main chain and / or pendant groups of the polycarbonate. For example, at least one of the hydrocarbyl groups is located in the main chain of the polycarbonate. In some embodiments, the number of the hydrocarbyl groups located in the main chain of the polycarbonate can be greater than the number of the hydrocarbyl groups located in the pendant groups, for example, in the repeating unit of Formula I.

[0155] The polycarbonates described herein can also have one or more (e.g., 1, 2, 3, 4, 5, or 6) oxygen-containing groups in their nitrogen-containing repeating units (e.g., the repeating unit of Formula I, e.g., the A structure) or other repeating units.

[0156] Such oxygen-containing groups can include, for example, ether groups, epoxy groups, ketone groups, and the like.

[0157] The one or more oxygen-containing groups can be located in the main chain and / or pendant groups of the polycarbonate. For example, at least one of the oxygen-containing groups is located in the main chain of the polycarbonate. In some embodiments, the number of the oxygen-containing groups located in the main chain of the polycarbonate can be greater than the number of the oxygen-containing groups located in the pendant groups, for example, in the repeating unit of Formula I.

[0158] Any two of the at least one nitrogen-containing group and the at least one additional group contained in the repeating unit of Formula I are connected to each other via L I is a bond (such as a single bond), a carbonate bond, or a C I alkylene group. 1-16 alkylene group.

[0159] The degree of polymerization of the repeating units (e.g., the repeating unit of Formula I) in the polycarbonate is not less than 2, for example, can be 2-2000, e.g., 2, 3, 5, 8, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 250, 300, 400, 500, 600, 800, 1000, 1200, 1500, 1800, 2000, or a range formed with any two of the above as the terminal values. In some embodiments, the degree of polymerization of the repeating unit of Formula I can be 2-1500, 2-1000, 3-800, 3-500, 3-200, 3-150, or 3-100. In some embodiments, the repeating units can be directly bonded.

[0160] In some example cases, the polycarbonate or salt thereof can have repeating units of Formula VI:

[0161] wherein,

[0162] X and Y each independently comprise at least one nitrogen-containing group and / or at least one redox-sensitive group, with the proviso that at least one of X and Y comprises at least one protonatable nitrogen-containing group, and X is different from Y;

[0163] L is a bond (such as a single bond), C 1-16 (e.g., C 1-12 , C 1-10 , C 1-8 , C 1-6 , C 1-4 or C 1-3 )alkylene (e.g., alkylene or alkenylene) or a carbonate bond;

[0164] R 1 each occurrence is independently selected from the group consisting of hydrogen, deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, substituted or unsubstituted ester, substituted or unsubstituted C 1-12 alkyl, substituted or unsubstituted C 1-12 alkoxy, substituted or unsubstituted C 1-12 alkylamino, substituted or unsubstituted C 1-12 alkyl ester, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 aryl, substituted or unsubstituted 3- to 10-membered heterocycloalkyl, and substituted or unsubstituted 5- to 14-membered heteroaryl;

[0165] each of p1 to p4 is independently an integer from 0 to 2, for example, 0, 1, or 2;

[0166] each of q1 and q4 is independently an integer from 1 to 16, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or a range formed between any two of the above values;

[0167] each of q2and q3is independently an integer from 0 to 16, e.g., 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or a range formed by any two of the foregoing values as endpoints. In such polycarbonates, the degree of polymerization of the repeating unit of Formula VI is not less than 2, e.g., can be from 2 to 2000, e.g., 2, 3, 5, 8, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 250, 300, 400, 500, 600, 800, 1000, 1200, 1500, 1800, 2000, or a range formed by any two of the foregoing values as endpoints. In some embodiments, the degree of polymerization of the repeating unit of Formula VI can be from 2 to 1500, 2 to 1000, 3 to 800, 3 to 500, 3 to 200, 3 to 150, or 3 to 100. In some embodiments, the repeating units (e.g., of Formula VI) are directly bonded.

[0168] In Formula VI, X and Y can each independently comprise: at least one nitrogen-containing group and / or at least one redox-sensitive group, with the proviso that at least one of X and Y comprises at least one protonatable nitrogen-containing group and X is different from Y. The nitrogen-containing group can be a protonatable nitrogen-containing group or other nitrogen-containing group, e.g., as described herein. In some embodiments, each of X and Y comprises at least one (e.g., at least two, three, or more) nitrogen-containing group. In some embodiments, each of X and Y comprises at least one (e.g., at least two, three, or more) redox-sensitive group. In some embodiments, one of X and Y comprises at least one (e.g., at least two, three, or more) nitrogen-containing group and the other comprises at least one (e.g., at least two, three, or more) redox-sensitive group. In some embodiments, one or both of X and Y comprises a carbonate group on the backbone. In some embodiments, one or both of X and Y does not comprise a carbonate group on the backbone. In some embodiments, one or both of X and Y does not comprise a carbonate group.

[0169] In some embodiments, the repeating unit of Formula VI has one or more nitrogen-containing groups. The one or more nitrogen-containing groups can be located in the backbone and / or pendant groups of the polycarbonate. Preferably, at least one nitrogen-containing group is located in the backbone of the polycarbonate. For example, at least one (e.g., at least 1 or 2) of the at least one nitrogen atom in the nitrogen-containing group is located in the backbone of the polycarbonate. In some embodiments, the number of nitrogen-containing groups or nitrogen atoms located in the backbone of the polycarbonate can be greater than the number of nitrogen-containing groups or nitrogen atoms located in the pendant groups. In some embodiments, the repeating unit of Formula VI contains no more than 5, 4, 3, 2, 1, or no nitrogen-containing groups or nitrogen atoms in the pendant groups. In some embodiments, some or all of the nitrogen-containing groups or nitrogen atoms in the repeating unit of Formula VI are located in the backbone of the polycarbonate. In some embodiments, greater than 20%, greater than 30%, greater than 40%, greater than 50%, greater than 60%, greater than 70%, greater than 80%, greater than 90%, or 100% of the nitrogen-containing groups or nitrogen atoms in the repeating unit of Formula VI are located in the backbone of the polycarbonate rather than in the pendant groups.

[0170] In some embodiments, the polycarbonate is protonatable due to the incorporation of at least one protonatable nitrogen-containing group in the repeating unit of Formula VI as described herein.

[0171] In some embodiments, the one or more redox-sensitive groups can be located in the backbone and / or pendant groups of the polycarbonate. For example, at least one redox-sensitive group is located in the backbone of the polycarbonate. In some embodiments, the number of redox-sensitive groups located in the backbone of the polycarbonate in the repeating unit of Formula VI can be greater than the number of redox-sensitive groups located in the pendant groups. In some embodiments, the repeating unit of Formula VI contains no more than 5, 4, 3, 2, 1, or no redox-sensitive groups in the pendant groups. In some embodiments, some or all of the redox-sensitive groups in the repeating unit of Formula VI are located in the backbone of the polycarbonate. In some embodiments, greater than 50%, greater than 60%, greater than 70%, greater than 80%, greater than 90%, or 100% of the redox-sensitive groups in the repeating unit of Formula VI are located in the backbone of the polycarbonate rather than in the pendant groups.

[0172] The repeating unit of Formula VI can contain one or more nitrogen-containing groups and / or one or more redox-sensitive groups. In some embodiments, two adjacent ones of the one or more nitrogen-containing groups and / or one or more redox-sensitive groups can be separated by a bond (e.g., a single bond) or a C 1-20 alkylene (e.g., C 1-16 , C 1-12 , C 1-10 , C 1-8 , C 1-6 , C 1-4 , or C 1-3) alkylene (e.g., alkylene or alkenylene) group.

[0173] In some cases, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% of the repeating units in the polycarbonate are linked by carbonate linkages.

[0174] In some cases, the polycarbonate can be a linear polymer. In some embodiments, the polycarbonate is not a cross-linked polymer. In some particular embodiments, the polycarbonate can or can not be a random copolymer.

[0175] The polycarbonate can not require long polyethylene glycol (PEG) segments as part of its backbone or pendant groups to impart the necessary carrier properties to the polymer. Generally, PEG segments having a degree of polymerization greater than 40, greater than 30, or greater than 25 can be considered long PEG segments. In some embodiments, the polycarbonate can not include long PEG segments or PEG segments.

[0176] The polycarbonate can have a weight average molecular weight (Mw) of 0.5 to 100 kDa. w For example, the polycarbonate can have a Mw of 0.5, 0.8, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 15, 18, 20, 22, 25, 28, 30, 35, 40, 50, 60, 70, 80, 90, 100 kDa, or a range formed by any two of these values as end points. w In some embodiments, the polycarbonate can have a Mw of 1 to 100 kDa, 2 to 100 kDa, 2 to 90 kDa, 2 to 80 kDa, 2 to 70 kDa, 2 to 60 kDa, 2 to 50 kDa, 2 to 40 kDa, 2 to 30 kDa, or 2 to 20 kDa. w .

[0177] In some embodiments, the polycarbonate has a pKa of 3.5-9.5, for example, 5.0-8.5, 5.5-7.5, for example 5.5-7.0.

[0178] In some embodiments, the polycarbonate can also exist in its salt form. For example, the salt of the polycarbonate is formed by salification with anions such as chloride, acetate, citrate, tetrafluoroacetate, and the like.

[0179] In some exemplary embodiments, the polycarbonate can be prepared by a method comprising polymerizing at least one monomer having Formula II with at least one monomer having Formula III:

[0180] In Formulas II and III,

[0181] A 1 and A 2 Each independently comprises one or more groups selected from the group consisting of formula IV:

[0182] in,

[0183] M is selected from: nitrogen-containing groups, redox-sensitive groups, hydrocarbon groups, oxygen-containing groups, and combinations thereof;

[0184] x1 and x2 are each an integer from 0 to 2, for example, 0, 1 or 2;

[0185] y1 and y2 are each an integer from 0 to 16, for example, 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or an integer within a range formed by taking any two of the above values ​​as endpoints;

[0186] R 1 Each group is independently selected from the following groups when it appears: hydrogen, deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, substituted or unsubstituted ester group, substituted or unsubstituted C group. 1-12 Alkyl, substituted or unsubstituted C 1-12 Alkoxy, substituted or unsubstituted C 1-12 Alkylamino, substituted or unsubstituted C 1-12 Alkyl ester group, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 Aryl, substituted or unsubstituted 3- to 10-membered heterocyclic alkyl and substituted or unsubstituted 5- to 14-membered heteroaryl;

[0187] Among them, A 1 and A 2 At least one of them has a nitrogen-containing group;

[0188] Among them, for A 1 and A 2 In the case of two or more groups that independently have Formula IV, the two or more groups that independently have Formula IV are connected to each other via L M Connected, where L M It is a bond, carbonate bond or C 1-16 Hydroxyl group; and

[0189] Among them, wavy lines It indicates the bonding position with adjacent atoms.

[0190] For the sake of brevity, specific descriptions of the nitrogen-containing groups, redox-sensitive groups, hydrocarbon groups, oxygen-containing groups contained in the monomers used in the above methods can be found in the above descriptions related to the polycarbonates contained in the polycarbonate nucleic acid complexes.

[0191] In some embodiments, A 1 contains one group having Formula IV. In some embodiments, A 2 contains one group having Formula IV.

[0192] In other exemplary embodiments, the polycarbonate can be prepared by a method comprising polymerizing a monomer having Formula VII with a monomer having Formula VIII:

[0193] wherein,

[0194] X 1 and X 2 are each independently: a bond (such as a single bond) or

[0195] Y 1 and Y 2 are each independently: a bond (such as a single bond) or

[0196] X and Y each independently contain at least one nitrogen-containing group and / or at least one redox-sensitive group;

[0197] L is a bond (such as a single bond), C 1-16 (e.g., C 1-12 , C 1-10 , C 1-8 , C 1-6 , C 1-4 , or C 1-3 )alkylene (e.g., alkylene or alkenylene) or a carbonate bond;

[0198] R 1 are each independently at each occurrence selected from the group consisting of hydrogen, deuterium, tritium, hydroxy, halogen, carboxyl, nitro, amino, cyano, substituted or unsubstituted ester, substituted or unsubstituted C 1-12 alkyl, substituted or unsubstituted C 1-12 alkoxy, substituted or unsubstituted C 1-12 alkylamino, substituted or unsubstituted C 1-12 alkyl ester, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14substituted or unsubstituted 3- to 10-membered heterocycloalkyl, and substituted or unsubstituted 5- to 14-membered heteroaryl;

[0199] each p1 to p4 is independently an integer from 0 to 2, for example, 0, 1, or 2;

[0200] each q1 and q4 is independently an integer from 1 to 16, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or in a range formed between any two of the above recited values;

[0201] each q2 and q3 is independently an integer from 0 to 16, for example, 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or in a range formed between any two of the above recited values;

[0202] with the proviso that:

[0203] (1) Formula VII and Formula VIII together have at least one X and at least one Y, and at least one X and at least one Y are not both from Formula VII or Formula VIII;

[0204] (2) At least one of X and Y comprises at least one protonatable nitrogen-containing group, and X is different from Y;

[0205] wherein the wavy line represents a bonding position to an adjacent atom.

[0206] For brevity, specific descriptions of the (protonatable) nitrogen-containing groups contained in the monomers used in the above methods can be found in the above descriptions related to the polycarbonates contained in the polycarbonate-nucleic acid complexes.

[0207] In some embodiments, X 1 may be the same or different. In some embodiments, Y 2 may be the same or different. In some embodiments, -X 1 -Y 2 may be the same or different. In some embodiments, -X 1 -Y 1 may be the same or different. 2 2

[0208] In one exemplary embodiment, X 1 is and Y 1 is a single bond; X 2 is a single bond and Y 2 is L is a bond.

[0209] ​​In another exemplary embodiment, X 1 is and Y 1 is X 2 is or Y 2 is and L is a bond; wherein X 1 may be the same or different from X 2 , Y 1 may be the same or different from Y 2 . In another exemplary embodiment, X 1 is or Y 1 is X 2 is and Y 2 is and L is a bond; wherein X 1 may be the same or different from X 2 , Y 1 may be the same or different from Y 2 .

[0210] The above polymerization reaction can be carried out in a certain solvent system, typically an organic solvent is selected.

[0211] As an example, the organic solvent can include, but is not limited to: dimethyl sulfoxide; C 1-10 aliphatic hydrocarbon (including halogenated hydrocarbon) solvents, such as chloroform, dichloromethane, pentane (such as cyclopentane), hexane (such as n-hexane, cyclohexane), heptane (such as n-heptane, cycloheptane), octane, nonane, decane, and isomers and mixtures thereof; C 6-14 aromatic hydrocarbon (including halogenated aromatic hydrocarbon) solvents, such as benzene, toluene, xylene, chlorobenzene, bromobenzene, and isomers and mixtures thereof; ester solvents, such as methyl acetate, ethyl acetate, γ-butyrolactone, and ε-caprolactone; ether solvents, such as diethyl ether, tetrahydrofuran, dioxane, and isomers and mixtures thereof.

[0212] The temperature and time of the reaction can be determined according to the specific situation, for example, the type of the reactants (such as monomers) used, the desired degree of polymerization, and the desired properties of the target polymer (such as molecular weight and molecular weight distribution).

[0213] Generally, the process does not have stringent requirements on reaction temperature and can be suitable for a wide range of types and scale-up production applications. Illustratively, the reaction can be carried out at a temperature range of 20 °C to 200 °C. For example, the reaction can be carried out at 20 °C, 22 °C, 25 °C, 30 °C, 40 °C, 50 °C, 60 °C, 70 °C, 80 °C, 90 °C, 100 °C, 110 °C, 120 °C, 130 °C, 140 °C, 150 °C, 160 °C, 170 °C, 180 °C, 190 °C, 200 °C, or a temperature range defined by any two of the above values as the end values. In some embodiments, the reaction can be carried out at a temperature no higher than 120 °C, no higher than 110 °C, no higher than 100 °C, for example, no higher than 90 °C, no higher than 80 °C, no higher than 70 °C, no higher than 65 °C, no higher than 60 °C, no higher than 55 °C, or no higher than 50 °C. For example, when it is desired that the polymerization reaction be carried out under relatively mild reaction conditions, it can be contemplated to carry out the above reaction at a temperature range of 20 °C to 70 °C, 25 °C to 70 °C, 30 °C to 70 °C, or 35 °C to 70 °C.

[0214] There is no particular limitation on the reaction time. The reaction time can be determined based on the reaction temperature and the desired molecular properties of the polymer. Illustratively, the reaction can last, for example, 4 to 96 hours. For example, the polymerization reaction can last 4, 8, 12, 18, 24, 36, 48, 60, 72, 96 hours, or a time range defined by any two of the above values as the end values. In some embodiments, the reaction can be carried out for 4-72 hours, 4-60 hours, 4-48 hours, 8-36 hours, or 8-24 hours.

[0215] In some cases, a catalyst can also be added to the reaction system to accelerate the polymerization reaction. The amount of catalyst to be added is not particularly limited and can be determined based on the actual need. Examples of the catalyst can include, but are not limited to, cesium fluoride, potassium fluoride, sodium fluoride, 4-dimethylaminopyridine (DMAP), and combinations thereof.

[0216] Optionally, the reaction product can be subjected to post-treatment after the reaction is completed, such as precipitation, separation, drying, ultrafiltration, etc. In some illustrative embodiments, the desired reaction product can be precipitated by subjecting the reaction solution to cooling, evaporation of solvent, centrifugation, etc. In some illustrative embodiments, the reaction solution can be dropped into anhydrous cold diethyl ether to precipitate the reaction product. In some illustrative embodiments, the resulting precipitate can be further subjected to washing and / or drying (e.g., vacuum drying) treatment to obtain the desired reaction product.

[0217] For the purposes herein, the nucleic acid can be one that needs to be protected for delivery to a certain biological target site (e.g., a cell, a tissue, an organ, etc.) to exert its effect.

[0218] In some cases, the nucleic acid includes, but is not limited to, deoxyribonucleic acid (DNA), ribonucleic acid (RNA), peptide nucleic acid (PNA), and locked nucleic acid (LNA).

[0219] In some embodiments, the nucleic acid can be selected from the group consisting of cDNA, mtDNA, ssDNA, dsDNA, antisense DNA, plasmid DNA, linear DNA, supercoiled DNA, siRNA, dsRNA, ssRNA, shRNA, saRNA, miRNA, rRNA, hnRNA, mRNA, tRNA, snRNA, pre-messenger RNA, catalytic RNA, antisense RNA, long non-coding RNA (lncRNA), and any combination thereof.

[0220] There is no particular limitation on the length and form of the nucleic acid to be carried, and oligonucleotides of a relatively small length (e.g., 10 to 200 nt or bp), mRNA of a relatively large length, circular plasmid DNA, and the like can all form complexes with the polycarbonates described herein and be carried to a target site. For non-limiting purposes, in general, nucleic acids of a length of no more than 100,000 nt or bp, no more than 80,000 nt or bp, no more than 60,000 nt or bp, no more than 50,000 nt or bp, no more than 40,000 nt or bp, no more than 30,000 nt or bp, no more than 25,000 nt or bp, no more than 20,000 nt or bp, no more than 18,000 nt or bp, no more than 15,000 nt or bp, or no more than 12,000 nt or bp can be selected as the cargo. In some embodiments, nucleic acids of a length of 1 to 100,000 nt or bp, e.g., 1, 2, 5, 10, 20, 30, 40, 50, 80, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1,000, 1,500, 2,000, 2,500, 3,000, 3,500, 4,000, 4,500, 5,000, 6,000, 7,000, 8,000, 9,000, 10,000, 12,000, 15,000, 18,000, 20,000, 25,000, 30,000, 40,000, 50,000, 60,000, 70,000, 80,000, 90,000, 100,000 nt or bp, or a range formed by any two of these values as endpoints, can be selected as the cargo.

[0221] In some embodiments, the nucleic acid can be a nucleic acid of natural or synthetic origin. In some embodiments, the nucleic acid can include single-stranded nucleic acids and / or double-stranded nucleic acids. It should be understood that the nucleotide sequence of certain single-stranded nucleic acids (e.g., mRNA) can include self-complementary nucleotide segments that can fold back to form local double-stranded structures through base pairing. Such nucleic acids having local double-stranded segments are also within the scope of the “single-stranded nucleic acids” referred to in the present application.

[0222] In some embodiments, the nucleic acid can be an endogenous nucleic acid or an exogenous nucleic acid.

[0223] The complex can have a complex mass ratio of polycarbonate to nucleic acid ranging from 0.1 : 1 to 500: 1, for example, the complex mass ratio can be 0.1 : 1, 0.5: 1, 1: 1, 1.5: 1, 2: 1, 2.5: 1, 3: 1, 3.5: 1, 4: 1, 4.5: 1, 5: 1, 8: 1, 10: 1, 15: 1, 20: 1, 25: 1, 30: 1, 35: 1, 40: 1, 45: 1, 50: 1, 60: 1, 70: 1, 80: 1, 90: 1, 110: 1, 120: 1, 150: 1, 180: 1, 200: 1, 220: 1, 250: 1, 280: 1, 300: 1, 320: 1, 350: 1, 380: 1, 400: 1, 420: 1, 450: 1, 480: 1, 500: 1, or in the range formed by any two of the above-mentioned values as end values.

[0224] The complex can have an average particle size of between 10 and 1000 nm, for example, 10, 15, 20, 25, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 400, 410, 420, 430, 440, 450, 460, 470, 480, 490, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950, 1000 nm, or in the range formed by any two of the above-mentioned values as end values.

[0225] Preparation of polycarbonate nucleic acid complex

[0226] In another aspect, provided herein is a method of preparing a polycarbonate nucleic acid complex, comprising:

[0227] - providing a polycarbonate or a salt thereof, and at least one nucleic acid; and

[0228] - mixing the polycarbonate or the salt thereof with the at least one nucleic acid.

[0229] For brevity, see the above section "Polycarbonate nucleic acid complex" for relevant description of the polycarbonate or the salt thereof and the at least one nucleic acid.

[0230] Generally, the mixing is performed in an aqueous medium, preferably an acidic aqueous medium. In some embodiments, the aqueous medium can have a pH no higher than 7.4, for example, no higher than 6.5, no higher than 6, no higher than 5.5, or no higher than 5. In some embodiments, the aqueous medium can have a pH of 1 to 7.4, for example, 2 to 7, 2 to 6.5, 2 to 6, 2 to 5.5, or 2 to 5. In other embodiments, the mixing can be performed at a pH condition lower than the pKa of the polycarbonate or salt thereof. For example, the complex can be prepared by mixing the above aqueous medium containing the polycarbonate or salt thereof with the nucleic acid.

[0231] In some embodiments, the complex is in an aqueous medium having a pH no higher than 7.4, for example, no higher than 6.5, no higher than 6, no higher than 5.5, or no higher than 5. In some embodiments, the complex is in an aqueous medium having a pH of 2 to 7, 2 to 6, 2 to 5.5, or 2 to 5.

[0232] In some exemplary embodiments, the aqueous medium can be an acidic aqueous buffer solution, for example, a phosphate buffer, an acetic acid-sodium acetate buffer, a citric acid buffer, an acetic acid-ammonium acetate buffer, an oxalic acid buffer, and the like. In some embodiments, the acidic aqueous medium can contain an organic solvent, for example, DMSO, as needed, for example, no more than 10% w / w.

[0233] The mixing can be performed by adding the aqueous solution of the nucleic acid to the aqueous medium containing the polycarbonate dropwise, or vice versa. The mixing can also be facilitated by means of shaking, vortexing, stirring, inverting, microfluidics, and the like.

[0234] The mixing can be performed at a temperature that does not significantly affect the structure and properties of both the polycarbonate and the nucleic acid, for example, to a degree of destruction that does not reach 10% of the original level. In some embodiments, the mixing can be performed at room temperature.

[0235] After the mixing, the resulting mixture can be incubated for a period of time to allow better formation of the polycarbonate-nucleic acid complex.

[0236] The incubation can be performed at a temperature that does not significantly affect the structure and properties of both the polycarbonate and the nucleic acid, for example, to a degree of destruction that does not reach 10% of the original level. In some embodiments, the incubation can be performed at room temperature. In some embodiments, the resulting mixture is incubated at room temperature for 5 to 60 minutes, for example, 10 to 40 minutes, for example, 10 to 30 minutes, to produce the polycarbonate complex.

[0237] In another aspect, provided herein is a polycarbonate complex prepared by the above preparation method.

[0238] Compositions and products

[0239] In another aspect, provided herein is a composition comprising a polycarbonate nucleic acid complex described herein.

[0240] In some embodiments, the composition can be a pharmaceutical composition.

[0241] In other embodiments, the composition comprises a pharmaceutically acceptable carrier, and at least one polycarbonate nucleic acid complex described herein.

[0242] Carriers include liquid or solid filler substances, diluents, excipients, solvents or encapsulating materials, which are involved in carrying or transporting the polycarbonate or polycarbonate complex into or through the body of the subject. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient.

[0243] Some examples of substances that can be used as pharmaceutically acceptable carriers include polysaccharides and derivatives thereof, proteinaceous substances, oil and fat substances, inorganic fillers, solvents, adjuvants (such as injection diluents, buffers or carrier solutions), excipients, esters, high molecular materials, and the like. Specifically, these can include cellulose and derivatives thereof, such as sodium carboxymethylcellulose, ethyl cellulose, and cellulose acetate; polysaccharides, such as powdered tragacanth, alginic acid, agar, malt; proteinaceous substances, such as gelatin, collagen; inorganic fillers / buffers, such as talc, calcium carbonate, calcium phosphate, silicon dioxide, magnesium hydroxide, and aluminum hydroxide; aqueous solvents / diluents, such as water (e.g., pyrogen-free water), isotonic saline, Ringer's solution, pH buffered solutions, ethanol; hydrophilic polyols, such as glycols (e.g., propylene glycol), glycerol, sorbitol, mannitol; oil phase carriers / hydrophobic solvents, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; ester solvents / oily adjuvants, such as ethyl oleate and ethyl laurate; excipients, such as cocoa butter and suppository waxes; other high molecular materials, such as polyesters, polyamides, polycarbonates, and / or polyanhydrides; and other nontoxic compatible substances employed in pharmaceutical formulations. Such pharmaceutically acceptable carriers used in conjunction with the pharmaceutical active ingredients are well known in the art. As long as the conventionally used pharmaceutically acceptable carriers do not adversely react with the active ingredients (such as the polycarbonate complex described herein), they can be used in the compositions described herein. The compositions described herein can also include other active substances as desired.

[0244] It is understood that the relative amounts of the active ingredient, pharmaceutically acceptable carrier, and / or other ingredients in the compositions described above will vary, depending on the different applications for which the active ingredient (e.g., a polycarbonate nucleic acid complex as described herein) is suitable, and will depend on a number of factors, such as the age, sex, weight, and medical condition of the subject to whom the composition is to be administered, the route of administration of the composition, etc. By way of example, the compositions can comprise from 0.1% to 100% (w / w) of the active ingredient (e.g., a polycarbonate nucleic acid complex as described herein), such as 0.1%, 0.5%, 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100% (w / w), or a range formed by any two of these values.

[0245] In some embodiments, the composition can have a pH of no more than 7.4, such as no more than 6.5, no more than 6, no more than 5.5, or no more than 5. In some embodiments, the composition can have a pH of from 1 to 7.4, such as from 2 to 7, from 2 to 6.5, from 2 to 6, from 2 to 5.5, or from 2 to 5.

[0246] In some embodiments, the composition can further comprise other active substances.

[0247] In some embodiments, the composition is formulated in the form of an injection, an oral preparation, a topical preparation, an inhalant, or an implant, such as a solution, an emulsion, a suspension, a gel, an ointment, a patch, a capsule, a tablet, a powder, an aerosol, etc.

[0248] In another aspect, provided herein is an article of manufacture comprising:

[0249] - a polycarbonate nucleic acid complex as described herein;

[0250] - a composition as described herein; or

[0251] - a polycarbonate or salt thereof as described herein and at least one nucleic acid.

[0252] In some embodiments, the article of manufacture is for use in delivering at least one nucleic acid to a cell or subject. In some embodiments, the article of manufacture is for use in expressing at least one nucleic acid in a cell or subject.

[0253] In some embodiments, the article of manufacture comprises a polycarbonate or salt thereof and at least one nucleic acid. In such embodiments, the polycarbonate or salt thereof and the nucleic acid are physically separated from each other in the article of manufacture (e.g., packaged separately) for mixing and forming a complex prior to use.

[0254] In some embodiments, the product comprises the polycarbonate or salt thereof, at least one nucleic acid, polycarbonate nucleic acid complex, or composition comprising one or more of the same, packaged in a single unit dose or multiple unit dose units.

[0255] In some embodiments, the product can comprise the polycarbonate or salt thereof, at least one nucleic acid, polycarbonate nucleic acid complex, or composition comprising one or more of the same, packaged in a convenient fraction of a single unit dose (e.g., one-half or one-third of the dose).

[0256] In some embodiments, the product can further comprise one or more other components or ingredients, e.g., instructions for use; ancillary agents, e.g., markers, other therapeutic agents, etc.; apparatuses for use, e.g., apparatuses for withdrawing, mixing, metering, sealing, etc.

[0257] Methods of use and uses

[0258] In another aspect, there is provided a method of delivering at least one nucleic acid to a cell or subject, the method comprising:

[0259] - providing a complex or composition comprising the complex described herein, the complex being formed from the at least one nucleic acid and a polycarbonate or salt thereof described herein; and

[0260] - contacting the cell or the subject with the complex or composition.

[0261] In another aspect, there is provided a method of expressing at least one nucleic acid in a cell or subject, the method comprising:

[0262] - providing a complex or composition comprising the complex described herein, the complex being formed from the at least one nucleic acid and a polycarbonate or salt thereof described herein; and

[0263] - contacting the cell or the subject with the complex or composition.

[0264] - contacting the cell or the subject with the complex or composition.

[0265] In another aspect, there is provided the use of the polycarbonate complex or composition comprising the same for delivering at least one nucleic acid to a cell or subject.

[0266] In another aspect, there is provided the use of the polycarbonate complex or composition comprising the same for causing a cell or subject to express at least one nucleic acid.

[0267] In another aspect, there is provided the use of the polycarbonate nucleic acid complex or composition comprising the same in the manufacture of a product for delivering at least one nucleic acid to a cell or subject.

[0268] In another aspect, provided herein is use of a polycarbonate nucleic acid complex or a composition comprising the same in the manufacture of a product for expressing at least one nucleic acid in a cell or subject.

[0269] Generally, delivering or expressing at least one nucleic acid to a subject can be delivering or expressing the at least one nucleic acid to one or more body parts of the subject. The one or more body parts can include, for example, organs, muscles, subcutaneous tissue, joint cavities, bone and its microenvironment, blood, lymph, joint cavities, and other parts in which the at least one nucleic acid needs to be delivered.

[0270] In some embodiments, the cell can be an ex vivo cell. In some embodiments, the cell can be an adherent or suspension culture cell. In some embodiments, the cell can be a cell from or derived from an organ or tissue. In some embodiments, the cell can be a primary cell or an immortalized cell line.

[0271] In some embodiments, the subject can be an invertebrate or a vertebrate. In some embodiments, the subject can include a mammal, for example, a non-human mammal and a human.

[0272] In some embodiments, the contacting can be for a time period sufficient for the complex or the at least one nucleic acid contained therein to enter one or more body parts of the cell or subject, for example, 10 minutes or more.

[0273] In some embodiments, the at least one nucleic acid can be an endogenous or exogenous nucleic acid, for example, an endogenous or exogenous nucleic acid desired by or beneficial to the health of the subject.

[0274] In some embodiments, the delivering or expressing is in vitro delivering or expressing, or in vivo delivering or expressing.

[0275] In some embodiments, the delivering can be through an intra-arterial, intravenous, intraperitoneal, parenteral, intramuscular, subcutaneous, oral, topical, or inhalation route.

[0276] The polycarbonates described herein open up new avenues for the biological delivery of nucleic acids. The efficient delivery of nucleic acids to biological target sites (e.g., within cells) has been a significant challenge because nucleic acids are susceptible to inactivation or degradation by cationic interfering factors during delivery and need to overcome the barrier of cell membranes to enter the nucleus. The polycarbonate nanocarriers described herein are able to carry a wide range of molecular weights and lengths of different types of nucleic acids, with extremely high biological delivery adaptability. As carriers, the polycarbonates are able to provide effective protection, facilitate transmembrane transport, and timely degradation upon reaching a specific environment (e.g., a biological target site), thereby enabling the rapid release of nucleic acids to maximize their use. The degradation products of the polycarbonates (e.g., CO2, thiols, etc.) are biologically safe, greatly eliminating the problem of toxic side effects of existing carriers. At the same time, the preparation process of the polycarbonates is simple and friendly, the quality of the products is controllable, and is suitable for large-scale production and subsequent biological tests and industrial applications.

[0277] The unique properties of the polycarbonate nanomaterials make them suitable as a new generation of nucleic acid delivery carriers to achieve safe and effective nucleic acid delivery.

[0278] Embodiment / Combination

[0279] Embodiment A1: A polycarbonate nucleic acid complex comprising:

[0280] (a) a polycarbonate having a repeating unit of Formula I or a salt thereof:

[0281] wherein,

[0282] A comprises at least one nitrogen-containing group;

[0283] R 1 each independently selected from the group consisting of hydrogen, deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, substituted or unsubstituted ester, substituted or unsubstituted C 1-12 alkyl, substituted or unsubstituted C 1-12 alkoxy, substituted or unsubstituted C 1-12 alkylamino, substituted or unsubstituted C 1-12 alkyl ester, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 aryl, substituted or unsubstituted 3- to 10-membered heterocycloalkyl, and substituted or unsubstituted 5- to 14-membered heteroaryl;

[0284] a1 and a2 are each independently an integer from 1 to 16;

[0285] b1 and b2 are each independently an integer from 0 to 2; and

[0286] (b) at least one nucleic acid.

[0287] Embodiment A2: The polycarbonate nucleic acid complex of Embodiment A1, wherein the at least one nitrogen-containing group comprises at least one protonatable nitrogen-containing group.

[0288] Embodiment A3: The polycarbonate nucleic acid complex of Embodiment A1, wherein the at least one nitrogen-containing group is located on the backbone of the polycarbonate.

[0289] Embodiment A4: The polycarbonate nucleic acid complex of Embodiment A1, wherein at least one nitrogen atom of the at least one nitrogen-containing group is located on the backbone of the polycarbonate.

[0290] Embodiment A5: The polycarbonate nucleic acid complex of Embodiment A2, wherein the at least one protonatable nitrogen-containing group is located on the backbone of the polycarbonate.

[0291] Embodiment A6: The polycarbonate nucleic acid complex of Embodiment A2, wherein at least one nitrogen atom of the at least one protonatable nitrogen-containing group is located on the backbone of the polycarbonate.

[0292] Embodiment A7: The polycarbonate nucleic acid complex of Embodiment A2, wherein the at least one protonatable nitrogen-containing group has a group selected from the group consisting of an amino group, a nitrogen-containing cyclic group, and combinations thereof.

[0293] Embodiment A8: The polycarbonate nucleic acid complex of Embodiment A7, wherein the amino group is selected from the group consisting of a secondary amino group, a tertiary amino group, a quaternary amino group, and combinations thereof.

[0294] Embodiment A9: The polycarbonate nucleic acid complex of Embodiment A7, wherein the nitrogen-containing cyclic group is selected from the group consisting of a 3- to 10-membered saturated or unsaturated aliphatic or aromatic heterocyclic group having at least one ring nitrogen atom, a 5- to 20-membered heterocyclic group fused together from 3- to 10-membered saturated or unsaturated aliphatic or aromatic heterocyclic groups having at least one ring nitrogen atom, each other, or with an aliphatic or aromatic carbocyclic ring, and combinations thereof.

[0295] Embodiment A10: The polycarbonate nucleic acid complex of Embodiment A1, wherein the polycarbonate further comprises one or more additional groups selected from the group consisting of a redox-sensitive group, a hydrocarbon group, an oxygen-containing group, and combinations thereof.

[0296] Embodiment A11: The polycarbonate nucleic acid complex of Embodiment A10, wherein the one or more additional groups are located on the backbone and / or pendant groups of the polycarbonate.

[0297] Embodiment A12: The polycarbonate nucleic acid complex of Embodiment A10, wherein the one or more additional groups are located in the backbone of the polycarbonate.

[0298] Embodiment A13: The polycarbonate nucleic acid complex of Embodiment A10, wherein the one or more additional groups are located in the repeating unit of Formula I or other repeating unit.

[0299] Embodiment A14: The polycarbonate nucleic acid complex of Embodiment A10, wherein the polycarbonate comprises at least one redox-sensitive group.

[0300] Embodiment A15: The polycarbonate nucleic acid complex of Embodiment A14, wherein the at least one redox-sensitive group comprises one or more of a single sulfur bond, a single selenium bond, a disulfide bond, a diselenide bond, a trisulfide bond, a triselenide bond, a tetrasulfide bond, a kethathiol bond.

[0301] Embodiment A16: The polycarbonate nucleic acid complex of Embodiment A10, wherein the polycarbonate comprises at least one hydrocarbyl group.

[0302] Embodiment A17: The polycarbonate nucleic acid complex of Embodiment A16, wherein the at least one hydrocarbyl group comprises one or more of a saturated or unsaturated, substituted or unsubstituted, chain hydrocarbyl group and a cyclic hydrocarbyl group.

[0303] Embodiment A18: The polycarbonate nucleic acid complex of Embodiment A10, wherein the polycarbonate comprises at least one oxygen-containing group.

[0304] Embodiment A19: The polycarbonate nucleic acid complex of Embodiment A18, wherein the at least one oxygen-containing group comprises one or more of an ether group, an epoxy group, a ketone group.

[0305] Embodiment A20: The polycarbonate nucleic acid complex of Embodiment A1, wherein the polycarbonate or salt thereof has a repeating unit of Formula VI:

[0306] wherein,

[0307] X and Y each independently comprise at least one nitrogen-containing group and / or at least one redox-sensitive group, with the proviso that at least one of X and Y comprises at least one protonatable nitrogen-containing group, and X is different from Y;

[0308] L is a bond, C 1-16 hydrocarbylene, or carbonate bond;

[0309] R 1Each group is independently selected from the following groups when it appears: hydrogen, deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, substituted or unsubstituted ester group, substituted or unsubstituted C group. 1-12 Alkyl, substituted or unsubstituted C 1-12 Alkoxy, substituted or unsubstituted C 1-12 Alkylamino, substituted or unsubstituted C 1-12 Alkyl ester group, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 Aryl, substituted or unsubstituted 3- to 10-membered heterocyclic alkyl and substituted or unsubstituted 5- to 14-membered heteroaryl;

[0310] p1 to p4 are each an independent integer from 0 to 2;

[0311] q1 and q4 are each independent integers from 1 to 16; and

[0312] q2 and q3 are each independent integers from 0 to 16.

[0313] Embodiment A21: The polycarbonate nucleic acid complex according to Embodiment A1, wherein the polycarbonate has a weight-average molecular weight (M) of 0.5 to 100 kDa. w ).

[0314] Implementation Method A22: The polycarbonate nucleic acid complex according to Implementation Method A1, wherein the at least one nucleic acid is selected from the group consisting of deoxyribonucleic acid (DNA), ribonucleic acid (RNA), peptide nucleic acid (PNA), locked nucleic acid (LNA), and any combination thereof.

[0315] Implementation A23: The polycarbonate nucleic acid complex according to Implementation A1, wherein the at least one nucleic acid comprises a single-stranded nucleic acid and / or a double-stranded nucleic acid.

[0316] Implementation A24: The polycarbonate nucleic acid complex according to Implementation A1, wherein the composite mass ratio of the polycarbonate or its salt to the at least one nucleic acid ranges from 0.1:1 to 500:1.

[0317] Embodiment A24: The polycarbonate nucleic acid complex according to Embodiment A1, wherein the complex has an average particle size between 10 and 1000 nm.

[0318] Implementation Method B1: A method for preparing polycarbonate, the method comprising:

[0319] Polymerization reaction is carried out using at least one monomer having Formula II and at least one monomer having Formula III:

[0320] In Formula II and Formula III,

[0321] A 1 and A 2 each independently comprises one or more groups independently having Formula IV:

[0322] wherein,

[0323] M is selected from the group consisting of a nitrogen-containing group, a redox-sensitive group, an alkylene group, an oxygen-containing group, and combinations thereof;

[0324] x1and x2are each independently an integer from 0 to 2;

[0325] y1and y2are each independently an integer from 0 to 16;

[0326] R 1 each occurrence is independently selected from the group consisting of hydrogen, deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, substituted or unsubstituted ester, substituted or unsubstituted C 1-12 alkyl, substituted or unsubstituted C 1-12 alkoxy, substituted or unsubstituted C 1-12 alkylamino, substituted or unsubstituted C 1-12 alkyl ester, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 aryl, substituted or unsubstituted 3- to 10-membered heterocycloalkyl, and substituted or unsubstituted 5- to 14-membered heteroaryl;

[0327] wherein, at least one of A 1 and A 2 has a nitrogen-containing group;

[0328] wherein, for A 1 and A 2 when comprising two or more groups independently having Formula IV, the two or more groups independently having Formula IV are connected to each other via L M , wherein L M is a bond, a carbonate bond, or a C 1-16 alkylene group; and

[0329] wherein, the wavy line represents a bonding position to an adjacent atom.

[0330] Embodiment B2: The method of Embodiment B1, wherein A 1 comprises one group having Formula IV.

[0331] Embodiment B3: The method according to Embodiment B1, wherein A 2 comprises one group of Formula IV.

[0332] Embodiment C1: A polycarbonate obtained by the method according to Embodiment B1.

[0333] Embodiment D1: A method for preparing a polycarbonate nucleic acid complex, comprising:

[0334] - providing a polycarbonate or a salt thereof, and at least one nucleic acid; and

[0335] - mixing the polycarbonate or the salt thereof with the at least one nucleic acid.

[0336] Embodiment D2: The method according to Embodiment D1, wherein the mixing is carried out in an aqueous medium, preferably an acidic aqueous medium.

[0337] Embodiment D3: The method according to Embodiment D1, wherein the at least one nucleic acid and the polycarbonate nucleic acid complex are defined according to any one of Embodiments A1 to A24.

[0338] Embodiment E1: A polycarbonate complex obtained by the method according to Embodiment D1.

[0339] Embodiment F1: A composition comprising a polycarbonate nucleic acid complex according to Embodiment A1.

[0340] Embodiment F2: The composition according to Embodiment F1, wherein the composition is a pharmaceutical composition.

[0341] Embodiment F3: The composition according to Embodiment F2, wherein the composition further comprises a pharmaceutically acceptable carrier.

[0342] Embodiment F4: The composition according to Embodiment F3, wherein the pharmaceutically acceptable carrier comprises one or more of polysaccharides and derivatives thereof, proteinaceous substances, oil and fat substances, inorganic fillers, solvents, adjuvants, excipients, esters, high molecular materials.

[0343] Embodiment F5: The composition according to Embodiment F1, wherein the composition comprises 0.1% to 100% (w / w) of the polycarbonate or the salt thereof, or the complex, by weight thereof.

[0344] Embodiment F6: The composition according to Embodiment F1, wherein the composition has a pH of not higher than 7.4, not higher than 6.5, not higher than 6, not higher than 5.5, or not higher than 5.

[0345] Embodiment F7: The composition according to Embodiment F1, wherein the composition further comprises other active substances.

[0346] Embodiment F8: The composition according to Embodiment D1, wherein the composition is formulated in the form of an injection, an oral preparation, a topical preparation, an inhalant or an implant, such as a solution, an emulsion, a suspension, a gel, an ointment, a patch, a capsule, a tablet, a powder, an aerosol, and the like dosage forms.

[0347] Embodiment G1: A product comprising:

[0348] a polycarbonate nucleic acid complex according to Embodiment A1;

[0349] a composition according to Embodiment F1; or

[0350] the polycarbonate or salt thereof and the at least one nucleic acid.

[0351] Embodiment G2: The product according to Embodiment G1, for use in delivering the at least one nucleic acid to a cell or subject, or for use in expressing the at least one nucleic acid in a cell or subject.

[0352] Embodiment G3: The product according to Embodiment G1, comprising the polycarbonate or salt thereof and the at least one nucleic acid physically separated from each other.

[0353] Embodiment G4: The product according to Embodiment G1, further comprising other components or constituents selected from the group consisting of instructions for use, auxiliary agents, handling implements, and any combination thereof.

[0354] Embodiment H1: A method of delivering at least one nucleic acid to a cell or subject, the method comprising:

[0355] - providing a polycarbonate nucleic acid complex according to Embodiment A1 or a composition according to Embodiment F1, the complex being formed from the at least one nucleic acid complexed with the polycarbonate or salt thereof; and

[0356] - contacting the cell or the subject with the complex or composition.

[0357]

[0358] Embodiment I1, a method of expressing at least one nucleic acid in a cell or subject, the method comprising:

[0359] - providing a polycarbonate nucleic acid complex according to Embodiment A1 or a composition according to Embodiment F1, the complex being formed from the at least one nucleic acid complexed with the polycarbonate or salt thereof; and

[0360] ​a complex formed from the at least one nucleic acid and the polycarbonate or salt thereof;

[0361] contacting the cell or the subject with the complex or composition.

[0362] Embodiment J1, use of a polycarbonate nucleic acid complex according to Embodiment A1 or a composition according to Embodiment F1 for delivering at least one nucleic acid to a cell or a subject.

[0363] Embodiment K1, use of a polycarbonate nucleic acid complex according to Embodiment A1 or a composition according to Embodiment F1 for causing a cell or a subject to express at least one nucleic acid.

[0364] Embodiment L1: use of a polycarbonate nucleic acid complex according to Embodiment A1 or a composition according to Embodiment F1 in the manufacture of a product for delivering at least one nucleic acid to a cell or a subject.

[0365] Embodiment M1: use of a polycarbonate nucleic acid complex according to Embodiment A1 or a composition according to Embodiment F1 in the manufacture of a product for causing a cell or a subject to express at least one nucleic acid.

[0366] Embodiment N1: the method according to Embodiment H1 or I1 or the use according to any one of Embodiments J1 to M1, wherein the cell is an ex vivo cell.

[0367] Embodiment N2: the method according to Embodiment H1 or I1 or the use according to any one of Embodiments J1 to M1, wherein the subject is an invertebrate or a vertebrate.

[0368] Embodiment N3: the method according to Embodiment H1 or I1 or the use according to any one of Embodiments J1 to M1, wherein the subject is a mammal.

[0369] Embodiment N4: the method according to Embodiment H1 or I1, wherein the contacting is for a duration sufficient for the complex or at least one nucleic acid contained therein to enter one or more body parts of the cell or subject.

[0370] Embodiment N5: the method according to Embodiment H1 or the use according to Embodiment J1 or L1, wherein the delivery is in vivo delivery or in vitro delivery.

[0371] Embodiment N6: the method according to Embodiment H1 or the use according to Embodiment J1 or L1, wherein the delivery is effected by intra-arterial, intravenous, intraperitoneal, parenteral, intramuscular, subcutaneous, oral, inhalation, topical routes.

[0372] Examples

[0373] The following examples further describe and demonstrate embodiments in accordance with the present application. These examples are given solely for the purpose of illustration and are not to be construed as the only ways in which the application can be practiced, as the application can be practiced in many ways.

[0374] Materials and Instruments

[0375] The instruments used in the examples are summarized in Table 1 below:

[0376] Table 1. Instruments used in the examples

[0377] The materials used in the examples are summarized in Table 2 below:

[0378] Table 2. Materials used in the examples

[0379] Example 1: Monomer preparation and characterization

[0380] Preparation Example A1: 2,2'-((3-methoxypropyl)azanediyl)bis(1-ethanol) (A013)

[0381] Diethanolamine (1.46 g, 9.54 mmol), 1-bromo-3-methoxypropane (1.56 g, 14.84 mmol) and anhydrous potassium carbonate (1.58 g, 11.44 mmol) were weighed. Dry acetonitrile (30 mL) was then added and the reaction was carried out at 85 °C for 18 hours. After the reaction was cooled to room temperature, acetonitrile was removed by distillation under reduced pressure. Saturated brine (85 mL) was added to the residue after rotary evaporation and extracted with ethyl acetate (170 mL, 3X). The organic phase was collected and dried over anhydrous sodium sulfate, and the solvent was removed by distillation under reduced pressure and dried to obtain monomer A013 (1.20 g) with a yield of 71.1%.

[0382] 1 H NMR (700 MHz, CDC13) δ 3.62 (t, CH2N(CH2CH2OH)2), 3.50 (t, CH3OCH2CH2), 3.33 (s, CH3OCH2CH2), 2.68 (m, CH3OCH2CH2CH2N(CH2CH2OH)2), 1.78 (m, CH3OCH2CH2CH2N).

[0383] HRMS (ESI, m / z): C8H 19 NO3, [M+H] + Calculated: 178.1443; Found: 178.1428.

[0384] Preparation Example A2: 2,2'-((3-(1H-pyrrolo-1-yl)propyl)azadiyl)bis(1-ethanol) (A017)

[0385] The process was the same as described in Preparation Example A1, except that 1-bromo-3-methoxypropane was replaced with 1-(3-bromopropyl)pyrrole, and the extraction solvent was replaced with dichloromethane instead of ethyl acetate. The yield of monomer A017 after post-treatment was 76.0%.

[0386] 1 H NMR(700MHz,CDCl3)δ6.67(t,NCHCH),6.14(t,NCHCH),3.94(t,CH2NCH2CH2OH)2),3.60(t,CH 2NCH2CH2OH)2),2.65(t,CH2N(CH2CH2OH)2),2.54(t,NCH2CH2CH2N),1.99(m,NCH2CH2CH2N).

[0387] HRMS(ESI,m / z):C 11 H 20 N₂O₂, [M+H] + Calculated value: 213.1603; Measured value: 213.1612.

[0388] Preparation Example B1: (Methylazonyl)bis(ethane-2,1-diyl)bis(1H-imidazol-1-carboxylic acid ester) (B003)

[0389] Under ice bath conditions, A003 (N-methyldiethanolamine, 20.0 g, 1 eq) was dissolved in 200 mL of DCM solvent, and CDI (68.1 g, 2.5 eq) was added. The mixture was brought back to room temperature and stirred at room temperature for 1 hour. After the reaction was completed, water was added to quench the reaction. The mixture was extracted three times with 200 mL of DCM, and the organic phases were combined and washed three times with pure water (500 mL × 3). The organic phases were dried over anhydrous sodium sulfate, filtered, and evaporated to dryness to obtain B003 yellow solid (40 g, 77.6%).

[0390] 1 H NMR (700MHz, CDCl3) δ8.12(t,J=1.1Hz,2H),7.39(t,J=1.5Hz,2H),7.08–7.04(m,2H),4.49(t,J=5.7Hz,4H),2.88(t,J=5.7Hz,4H),2.43(s,3H).

[0391] Preparation Example B2: Piperazine-1,4-diylbis(ethane-2,1-diyl)bis(1H-imidazol-1-carboxylate) (B004)

[0392] Reference the procedure of Preparation Example B1, except replace A003 with A004 (1,4-bis(2-hydroxyethyl)piperazine) to give B004 as a yellow oil in 96.0% yield.

[0393] 1 H NMR (700 MHz, CDC13) δ 8.14 (d, J = 1.3 Hz, 2H), 7.43 (t, J = 1.5 Hz, 2H), 7.14 - 7.05 (m, 2H), 4.52 (t, J = 5.8 Hz, 4H), 2.77 (t, J = 5.8 Hz, 4H), 2.56 (s, 8H).

[0394] Preparation Example B3: (Ethylazanediyl)bis(ethane-2, 1-diyl) bis(1H-imidazole-1- carboxylate) (B005)

[0395] Reference the procedure of Preparation Example B1, except replace A003 with A005 (N- ethyldiethanolamine) to give B005 as a yellow oil in 55.0% yield.

[0396] 1 H NMR (700 MHz, CDC13) δ 8.14 (d, J = 1.3 Hz, 2H), 7.43 (t, J = 1.5 Hz, 2H), 7.14 - 7.05 (m, 2H), 4.52 (t, J = 5.8 Hz, 4H), 2.77 (t, J = 5.8 Hz, 4H), 2.56 (s, 8H).

[0397] Preparation Example B4: (Butylazanediyl)bis(ethane-2, 1-diyl) bis(1H-imidazole-1- carboxylate) (B007)

[0398] Reference the procedure of Preparation Example B1, except replace A003 with A007 (N- butyldiethanolamine) to give B007 as a yellow oil in 73.7% yield.

[0399] 1 H NMR (700 MHz, CDC13) δ 8.14 (d, J = 1.3 Hz, 2H), 7.43 (t, J = 1.5 Hz, 2H), 7.14 - 7.05 (m, 2H), 4.52 (t, J = 5.8 Hz, 4H), 2.77 (t, J = 5.8 Hz, 4H), 2.56 (s, 8H).

[0400] Preparation B5: 3-(1-(2-((1H-imidazole-1-carbonyl)oxy)ethyl)piperidin-4-yl)propyl 1H- imidazole-1-carboxylate (B021)

[0401] Reference to the procedure of Preparation B1, except that A003 was replaced by A021 (3-(1-(2-hydroxyethyl)piperidin-4-yl)propan-1-ol) to give B021 as a yellow solid in 66.8% yield.

[0402] 1 H NMR (700 MHz, CDC13) δ 8.12 (dt, J = 4.0, 1.1 Hz, 2H), 7.41 (dt, J = 5.1, 1.5 Hz, 2H), 7.08 - 7.05 (m, 2H), 4.50 (t, J = 5.8 Hz, 2H), 4.39 (t, J = 6.7 Hz, 2H), 2.74 (t, J = 5.8 Hz, 2H), 2.07 (td, J = 11.4, 2.4 Hz, 2H), 1.82 - 1.77 (m, 2H), 1.69 (dt, J = 12.8, 2.2 Hz, 2H), 1.37 - 1.18 (m, 7H).

[0403] Preparation B6: M008

[0404] 2-Hydroxyethyl disulfide (1 g, 6.48 mmol) and diphenyl carbonate (2.08 g, 9.72 mmol) were dissolved in anhydrous toluene (400 mL), then Novozyme 435 (N-435) equivalent to 2.08 g of diphenyl carbonate was added. After the reaction solution was reacted at 70°C under nitrogen for 12 hours, the lipase was removed by filtration. The collected filtrate was concentrated under reduced pressure to obtain a solid, and the solid was washed with methanol to obtain a crude product. The crude product was recrystallized with ethyl acetate to obtain white crystals of M008 in a yield of about 60%.

[0405] 1 H-NMR (400 MHz, CDC13) δ 4.37 (m, OCOCH2CH2SS), 3.05 (m, OCOCH2CH2SS).

[0406] Preparation B7: M009

[0407] A003 (0.77 g, 6.48 mmol) and diphenyl carbonate (2.08 g, 9.72 mmol) were dissolved in anhydrous toluene (400 mL), and then 2.08 g of Novozyme 435 (N-435) was added. After the reaction solution was reacted at 70°C under nitrogen for 12 hours, the lipase was removed by filtration. The collected filtrate was concentrated under reduced pressure to obtain a solid, which was separated by a silica gel column (the eluent of the column chromatography was methanol and ethyl acetate at a volume ratio of 1 / 1). The crude product was recrystallized with ethyl acetate to obtain white crystals of M009 at a yield of 43%.

[0408] 1 H-NMR (700 MHz, CDC13): δ 4.2 (t, OCOCH2CH2NCH3), 2.69 (t, OCOCH2CH2NCH3), 2.34 (s, OCOCH2CH2NCH3).

[0409] Example 2: Polymer preparation

[0410] The monomers used in the polymer preparation examples are summarized in Table 3 below.

[0411] Table 3. Monomers used in the polymer preparation examples

[0412] Preparation example P1: Polymer 250

[0413] Polymer 250 contains the following repeating unit:

[0414] The starting material A003 (48 mg, 1 eq), B003 (148 mg, 1.2 eq), and cesium fluoride (6.0 mg, 0.1 eq) were dissolved in 2 mL of ethyl acetate solution and placed in a 60°C reaction for 15 hours. After the reaction was completed, it was returned to room temperature, 2 mL of the reaction solution was precipitated with 50 mL of ether, centrifuged at 4000 x g for 5 minutes, the supernatant was removed, the organic phase was evaporated to obtain a light yellow solid product. 20 mL of acidic aqueous solution (pH = 5) was added for dissolution and ultrafiltration (MWCO: 1000), after 10 volumes of ultrafiltration, it was concentrated to 10 mL, filtered with a 0.2 μm filter and freeze-dried to obtain white solid 250 (105 mg, yield 80.6%). Mw: 9.0 KDa.

[0415] Preparation example P2: Polymer 251

[0416] Polymer 251 contains the following repeating unit:

[0417] Referring to the procedure of Preparation P1, starting material A004 (57 mg, 1 eq), B004 (142 mg, 1.2 eq), and cesium fluoride (5.0 mg, 0.1 eq) gave 251 (155 mg, yield 82.4.0%) as white crystalline granules. Mw: 10.5 KDa.

[0418] Preparation P3: Polymer 269

[0419] Polymer 269 contains the following repeating unit:

[0420] Referring to the procedure of Preparation P1, starting material A006 (102.1 mg, 1 eq), B003 (260 mg, 1.2 eq), and cesium fluoride (10.3 mg, 0.1 eq) gave 269 (100 mg, yield 40.5%) as white oil. Mw: 4.4 KDa.

[0421] Preparation P4: Polymer 273

[0422] Polymer 273 contains the following repeating unit:

[0423] Starting material A010 (100 mg, 1 eq), B003 (134 mg, 1.2 eq), and cesium fluoride (5.2 mg, 0.1 eq) were dissolved in 1 mL of ethyl acetate solution and left to react at 60 °C for 15 hours. After the reaction was completed, it was brought to room temperature, 2 mL of ethyl acetate was added, washed with pure water three times (3 X 3 mL), the organic phase was dried with anhydrous sodium sulfate, filtered, and the ethyl acetate was evaporated to obtain 273 (63 mg, yield 26.9%) as a light yellow oil. Mw: 3.5 KDa.

[0424] Preparation P5: Polymer 276

[0425] Polymer 276 contains the following repeating unit:

[0426] Referring to the procedure of Preparation P1, starting material A004 (106 mg, 1 eq), B003 (222 mg, 1.2 eq), and cesium fluoride (9.4 mg, 0.1 eq) gave 276 (35 mg, yield 15.2%) as white oil. Mw: 6.3 KDa.

[0427] Preparation P6: Polymer 281

[0428] Polymer 281 contains the following repeating unit:

[0429] Preparation P7: Polymer 282

[0430] Preparation P7: Polymer 282

[0431] Polymer 282 contains the following repeating unit:

[0432] Preparation P7: Polymer 282

[0433] Preparation P8: Polymer 283

[0434] Polymer 283 contains the following repeating unit:

[0435] Preparation P7: Polymer 282

[0436] Preparation P9: Polymer 288

[0437] Polymer 288 contains the following repeating unit:

[0438] Preparation P7: Polymer 282

[0439] Preparation P10: Polymer 293

[0440] Polymer 293 contains the following repeating unit:

[0441] Preparation P7: Polymer 282

[0442] Preparation Pll: Polymer 294

[0443] Polymer 294 contains the following repeating unit:

[0444] A similar procedure as described in Preparation P4, starting from A007 (100 mg, 1 eq), B007 (260 mg, 1.2 eq), and cesium fluoride (9.4 mg, 0.1 eq) afforded 294 (80 mg, 22.2% yield) as a light yellow oil. Mw: 6.0 KDa.

[0445] Preparation P12: Polymer 298

[0446] Polymer 298 contains the following repeating unit:

[0447] A similar procedure as described in Preparation P1, starting from A032 (100.4 mg, 1 eq), B007 (207 mg, 1.2 eq), and cesium fluoride (7.4 mg, 0.1 eq) afforded 298 (160 mg, 64.8% yield) as a white crystalline solid. Mw: 5.0 KDa.

[0448] Preparation P13: Polymer 299

[0449] Polymer 299 contains the following repeating unit:

[0450] A similar procedure as described in Preparation P4, starting from A021 (101 mg, 1 eq), B007 (230 mg, 1.2 eq), and cesium fluoride (8.2 mg, 0.1 eq) afforded 299 (100 mg, 41.4% yield) as a light yellow oil. Mw: 1.2 KDa.

[0451] Preparation P14: Polymer 300

[0452] Polymer 300 contains the following repeating unit:

[0453] A similar procedure as described in Preparation P4, starting from A022 (253 mg, 1 eq), B007 (510 mg, 1.2 eq), and cesium fluoride (18.3 mg, 0.1 eq) afforded 300 (210 mg, 37.2% yield) as a light yellow oil. Mw: 23.6 KDa.

[0454] Preparation P15: Polymer 301

[0455] Polymer 301 contains the following repeating unit:

[0456] Following the procedure of Preparation Example P1, the raw materials were A023 (98.5 mg, 1 eq), B007 (260 mg, 1.2 eq), and cesium fluoride (9.3 mg, 0.1 eq), yielding a white crystalline solid 301 (120 mg, yield 46.7%). Mw: 8.5 kDa.

[0457] Preparation Example P16: Polymer 303

[0458] Polymer 303 contains the following repeating units:

[0459] Following the procedure of Preparation Example P1, the raw materials were A032 (100 mg, 1 eq), B005 (192 mg, 1.2 eq), and cesium fluoride (7.4 mg, 0.1 eq), yielding a white, fluffy solid 303 (150 mg, yield 69.5%). Mw: 5.7 kDa.

[0460] Preparation Example P17: Polymer 307

[0461] Polymer 307 contains the following repeating units:

[0462] Following the procedure of Preparation Example P1, the raw materials were A013 (99 mg, 1 eq), B004 (150 mg, 1.2 eq), and cesium fluoride (8.6 mg, 0.1 eq), yielding a white crystalline solid 307 (120 mg, yield 47.0%). Mw: 3.2 kDa.

[0463] Preparation Example P18: Polymer 328

[0464] Polymer 328 contains the following repeating units:

[0465] Following the procedure of Preparation Example P1, the raw materials were A031 (100 mg, 1 eq), B004 (326 mg, 1.2 eq), and cesium fluoride (11.4 mg, 0.1 eq), yielding a white crystalline solid 328 (36 mg, yield 11.8%). Mw: 3.3 kDa.

[0466] Preparation Example P19: Polymer 329

[0467] Polymer 329 contains the following repeating units:

[0468] Following the procedure of Preparation Example P1, the raw materials were A032 (102 mg, 1 eq), B004 (255 mg, 1.2 eq), and cesium fluoride (8.6 mg, 0.1 eq), yielding a white crystalline solid 329 (120 mg, yield 45.9%). Mw: 3.5 kDa.

[0469] Preparation P20: Polymer 435

[0470] Polymer 435 contains the following repeating unit:

[0471] Following the procedure described in Preparation P1, starting from A017 (101 mg, 1 eq), B004 (210 mg, 1.2 eq), and cesium fluoride (7.2 mg, 0.1 eq), afforded 435 (130 mg, 55.5% yield) as a white crystalline solid. Mw: 9.3 KDa.

[0472] Preparation P21: Polymer 474

[0473] Polymer 474 contains the following repeating unit:

[0474] Following the procedure described in Preparation P1, starting from A006 (100 mg, 1 eq), B021 (305 mg, 1.2 eq), and cesium fluoride (10.0 mg, 0.1 eq), lyophilized to afford 474 (40 mg, 13.6% yield) as a white crystalline solid. Mw: 1.9 KDa.

[0475] Preparation P22: Polymer 475

[0476] Polymer 475 contains the following repeating unit:

[0477] Following the procedure described in Preparation P4, starting from A007 (102 mg, 1 eq), B021 (280 mg, 1.1 eq), and cesium fluoride (9.4 mg, 0.1 eq), afforded 475 (90 mg, 32.1% yield) as a light yellow oil. Mw: 2.1 KDa.

[0478] Preparation P23: Polymer 545

[0479] Polymer 545 contains the following repeating unit:

[0480] Following the procedure described in Preparation P1, starting from A021 (100 mg, 1 eq), B021 (210 mg, 1.05 eq), and cesium fluoride (10.0 mg, 0.1 eq), afforded 545 (128 mg, 54.7% yield) as a white solid. Mw: 7.1 KDa.

[0481] Preparation P24: Polymer 547

[0482] Polymer 547 contains the following repeating unit:

[0483] The starting material A023 (103 mg, 1 eq), B021 (248 mg, 1.05 eq), and cesium fluoride (9.3 mg, 0.1 eq) were dissolved in 1 mL of dichloromethane solution and placed in a 60 °C reaction for 15 hours. After the reaction was completed, it was restored to room temperature, 1 mL of the reaction was precipitated with 25 mL of ether, centrifuged at 4000xg for 5 minutes, the supernatant was removed, and redissolved in 1 mL of dichloromethane, precipitated with 25 mL of ether, centrifuged at 4000xg for 5 minutes, the supernatant was removed, and the organic phase was evaporated to obtain a light yellow solid 547 (150 mg, yield 57.5%). Mw: 17.3 KDa.

[0484] Preparation Example P25: Polymer 550

[0485] Polymer 550 contains the following repeating units:

[0486] Referring to the process of Preparation Example P24, the starting material is A003 (99 mg, 1 eq), B005 (287 mg, 1.07 eq), and cesium fluoride (12.8 mg, 0.1 eq) to obtain a light yellow solid 550 (110 mg, yield 41.6%). Mw: 15.2 KDa.

[0487] Preparation Example P26: Polymer E004

[0488] Polymer E004 contains the following repeating units:

[0489] The starting material M004 (172 mg, 1 eq), M005 (525 mg, 1 eq) was dissolved in 2 mL of tetrahydrofuran solution and placed in a 50 °C reaction for 48 hours. After the reaction was completed, it was restored to room temperature, 2 mL of the reaction was precipitated with 50 mL of ether, centrifuged at 4000xg for 5 minutes, the supernatant was removed, and the organic phase was evaporated to obtain a light yellow solid product. Redissolved in 2 mL of dichloromethane, precipitated with 50 mL of ether, centrifuged at 4000xg for 5 minutes, the supernatant was removed, and the organic phase was evaporated to obtain a light yellow solid E004 (383 mg, yield 55%). Mw: 6.2 KDa.

[0490] Preparation Example P27: Polymer C004

[0491] Polymer C004 contains the following repeating units:

[0492] The starting material M006 (525 mg, 1 eq) and M007 (152 mg, 1 eq) were dissolved in 1 mL of a mixture solution of methanol and water (methanol / water = 9 / 1) and placed in a 60 °C reaction under light protection and nitrogen protection for 5 days. After 5 days, 16 mg of M007 was added to continue stirring the reaction for 2 days. After the reaction was completed, it was returned to room temperature, 20 mL of 0.1 M aqueous hydrochloric acid (HC1) was added for dissolution, and ultrafiltration (MWCO: 1000) was performed overnight with a pH = 4 HC1 aqueous solution. The volume of ultrafiltration was 200 mL, and after concentration to 10 mL, filtration was performed with a 0.2 μιη filter, and lyophilization was performed to obtain fluffy white solid C004 (323 mg, yield 47.7%). Mw: 5.5 KDa.

[0493] Preparation Example P28: Polymer R003

[0494] Polymer R003 contains the following repeating unit:

[0495] The starting material benzyl alcohol (0.01 g, 0.088 mmol), M008 (0.35 g, 0.98 mmol) and M009 (1 g, 3.45 mmol) were dissolved in 6 mL of anhydrous toluene solution, vacuumed for 10 minutes to remove trace amounts of water in the system and replaced with nitrogen, and 100 mg of dried Novozym435 was placed in a 70 °C reaction for 24 hours. After the reaction was completed, it was returned to room temperature, diluted with 2 mL of dichloromethane, filtered to remove the enzyme (Novozym435), and dried by settling with anhydrous ether three times to obtain white solid R003 (0.87 g, yield 65%). Mw: 8.7 KDa.

[0496] Example 3: Polymer molecular weight characterization

[0497] Aqueous solutions of polymers 250, 251, 269, 276, 298, 301, 303, 307, 328, 329, 435, 474, 475, 545, 547, 550, C004, R003 were prepared in 0.3 M sodium acetate buffer (pH 5) (5 mg / mL). Aqueous solutions of polymers 273, 281, 282, 283, 288, 293, 294, 299, 300, E004 were prepared in 0.3 M sodium acetate buffer (pH 5) with 5% DMSO (5 mg / mL). The samples were filtered through 0.2 μΜ filters and the molecular weights were determined using aqueous size exclusion chromatography with a differential refractive index detector and a multi-angle static light scattering detector (SEC-RI-MALS) instrument; the mobile phase was 0.3 M sodium acetate buffer (pH 5); the gel permeation chromatography column was an Ultrahydrogel 1000 (Waters), the sample injector temperature was 4 °C, the column temperature was set to 30 °C, the flow rate was 0.6 mL / min, and the dn / dc was set to 0.1659.

[0498] Example 4: Preparation of polymer-nucleic acid complexes

[0499] The following exemplary methods for preparing complexes of the above-mentioned polymers with nucleic acids are provided in this example.

[0500] Method 1: An aqueous solution of a polymer was prepared in 20 mM sodium citrate buffer, pH 5 (0.8 mg / mL). An aqueous solution of Luc-mRNA (the CDS sequence is shown in SEQ ID No.: 1 (Figure 6) and the full-length sequence is shown in SEQ ID No.: 2 (Figure 7)) or pCMV-luc plasmid DNA (PF0461, PlasmidFactory) was prepared in 20 mM sodium citrate buffer, pH 5 (40 μg / mL). The above-prepared polymer solution and the Luc-mRNA solution or the pCMV-luc solution were mixed thoroughly in a 1:1 volume ratio and left to stand at room temperature for 10 minutes to obtain a polymer / Luc-mRNA complex or a polymer / pCMV-luc complex. Complexes of each of polymers 250, 251, 269, 276, 298, 301, 303, 307, 328, 329, 435, 474, 475, 545, 547, 550, linear polyethylenimine (L-PEI, 919012, Merck), C004, R003 with Luc-mRNA were prepared using Method 1. Complexes of each of linear polyethylenimine (L-PEI, 919012, Merck), polymers 547, 550 with pCMV-luc plasmid were prepared using Method 1.

[0501] Method 2: Prepare a DMSO-containing aqueous solution of the polymer (1% DMSO, 0.8 mg / mL) with 20 mM pH 5 sodium citrate buffer. Prepare an aqueous solution of Luc-mRNA (CDS sequence shown in SEQ ID No.: 1 (Figure 6), full-length sequence shown in SEQ ID No.: 2 (Figure 7)) or pCMV-luc plasmid DNA (PF0461, PlasmidFactory) (40 pg / mL) with 20 mM pH 5 sodium citrate buffer. Mix the polymer solution and the Luc-mRNA solution or the pCMV-luc solution uniformly at a volume ratio of 1:1, and stand at room temperature for 10 minutes to obtain the polymer-Luc-mRNA or polymer-pCMV-luc plasmid complex. The complex of polymer 273, 281, 282, 283, 288, 293, 294, 299, 300, E004, respectively, with Luc-mRNA was prepared by method 2. The complex of polymer 283 with pCMV-luc plasmid was prepared by method 2.

[0502] Figure 1 shows a schematic diagram of the formation of a polycarbonate nucleic acid complex from a polycarbonate according to the present disclosure and a nucleic acid.

[0503] Example 5: Performance characterization of polymer-nucleic acid complex

[0504] A. Measurement of nanoparticle average particle size, particle dispersion index (PDI), and Zeta surface potential of polymer-nucleic acid complex

[0505] Add 200 pL of the polymer / Luc-mRNA or polymer / pCMV-luc complex prepared in Example 4 to a micro-sample cell (ZEN0040). Use a nanoparticle size analyzer (Zetasizer Pro, Malvern Panalytical) to measure the nanoparticle size and particle dispersion index of the sample, with a temperature setting of 25°C and a 173° laser angle. Take 1 mL of the polymer / Luc-mRNA or polymer / pCMV-luc complex prepared in Example 4 with a 1 mL syringe to a disposable folded capillary sample cell (DTS1070), and use the Zetasizer Pro to measure the surface potential (Zeta) of the sample.

[0506] B. Measurement of encapsulation efficiency of polymer-nucleic acid complex

[0507] Take the polymer-nucleic acid complex prepared in Example 4, 400 ng of Luc-mRNA or pCMV-luc, and 10x DNA buffer (P022-02-AA, Vazyme) to pre-mix at a volume ratio of 9:1, and take 20 pL of the pre-mix to load onto a 1% pre-made agarose gel (G661012, Thermo ScientificTM ) and run in 1xTEA running buffer (abs9260, Axygen (Shanghai) Biotech Co., Ltd.) at 150V (1645050, 1704486, Bio-RAD) for 15 min. Then the gel was taken out and imaged by UV imager (4600SF, Tanon). The gel brightness of unencapsulated Luc-mRNA or pCMV-luc was used for normalization. After normalization, if the sample had no bright band, the encapsulation efficiency of the complex was recorded as 100%. The results are shown in Table 4 and Table 5.

[0508] Table 4. Characterization results of polymer / Luc-mRNA complex

[0509] Table 5. Characterization results of polymer / pCMV-luc complex

[0510] Example 6: Cell safety and transfection performance of the complex

[0511] The safety and transfection performance of the complex were verified by mRNA transfection of healthy human primary bronchial epithelial cells (HBE, ID: BML31M2, cell passages 5-11) and plasmid transfection of HEK-293T (CBP60439, ATCC, cell passages 4-7). The culture conditions and medium formula of the primary bronchial epithelial cells HBE were set according to the reference (Eur Respir J 2022; 59: 2100671), and the HEK-293T was cultured in DMEM medium (GIBCO Cat: 10566016) containing 10% fetal bovine serum (GIBCO Cat: A5256701).

[0512] 3x10 4 HBE cells or 2x10 4 HEK-293T cells were seeded in a 96-well cell culture plate (Cat: 167425, Thermo Scientific TM ) and incubated at 37°C, 5% CO2 incubator for 24 hours. 5 μL of PBS (PB180237, Procell, negative control) or 5 μL of the polymer / Luc-mRNA complex or polymer / pCMV-luc complex prepared in Example 4 (including polymer L-PEI, polymer E004, polymer C004, polymer R003, each complexed with Luc-mRNA or pCMV-luc as a comparative example) was directly added to the cell culture solution of each well of the 96-well plate, and the plate was placed in a 37°C, 5% CO2 incubator for further incubation for 15 hours (Luc-mRNA) or 48 hours (pCMV-luc).

[0513] Cell viability was tested to reflect the safety performance of the complexes. Cell culture medium (PneumaCult-ALI basal medium, Cat: 05002, Stemcell Technologies) and Prestoblue solution in Prestoblue HS Cell Viability Assay Kit (P50201, Invitrogen) were mixed at 9 / 1 volume ratio to make Prestoblue mixture. The medium in each well of the 96-well cell culture plate was removed by pipette, 100 μΐ^of Prestoblue mixture was added, and the plate was incubated in a 37°C, 5% CO2 incubator for 30 minutes. Then, 80 μΐ^from each well was removed to a 96-well black plate (Cat: 237107, Thermo Scientific TM ), and the fluorescence reading was tested by Varioskan LUX Multifunctional Microplate Reader (VLBL00GD1, Thermo Scientific TM ) with excitation set at 560 nm and emission set at 590 nm. The reading of the culture well with PBS addition was set as 100%. The reading of the complex transfection well was normalized to the PBS sample reading. The results were shown as relative cell viability (%), as shown in Figure 3 (after Luc-mRNA transfection) and Figure 5 (after pCMV-luc transfection). The results showed good biosafety of the polymer-nucleic acid complex on cells.

[0514] The residual Prestoblue solution in the 96-well cell culture plate was removed, and 100 μΐ^of IX cell lysis solution (E1531, Promega) was added to each well, which was incubated at 4°C for 10 minutes. 20 μΐ^of lysis solution was taken to a white 96-well plate (Cat: 236107, Thermo Scientific TM ), and 50 μΐ^of Luciferase 1000 Assay Reagent (E4550, Promega) was added, which was incubated at room temperature for 5 minutes. The bioluminescence signal (RLU) was read by Varioskan LUX Multifunctional Microplate Reader. The results are shown in Figure 2 (after Luc-mRNA transfection) and Figure 4 (after pCMV-luc transfection). The results showed that the polymer-nucleic acid complex could effectively transfect cells and successfully express the loaded nucleic acid, and the transfection efficiency was significantly higher than that of the negative control and all the comparative examples.

[0515] Unless expressly stated otherwise, all dimensions herein involved and their specific numerical values should not be understood as strictly limited to the recited exact values. Rather, each numerical value should be construed as including the value itself and a functionally equivalent range of variation around that value. For example, a dimension disclosed as "10 nm" should be understood to mean "about 10 nm."

[0516] All documents cited herein, including any cross-referenced or related patents or patent applications and any patent or patent applications to which this application claims priority or benefit thereof, are hereby incorporated by reference in their entirety. No admission is made that any of the references constitute prior art and none should be taken as an acknowledgement or any form of suggestion that this reference is many alone, or in any combination, teach, suggest, or disclose any aspect of the present application. In addition, the reference herein to any document or other source should not be taken as a suggestion that the present application is not entitled to antedate such document or source by virtue of prior application. Further, the references herein to any standards or conventions are not intended to limit the scope of the application to that which is presently considered proper according to those standards or conventions.

[0517] While the application has been described and illustrated with specific embodiments, it is understood that various modifications, substitutions, and changes can be made by those skilled in the art without departing from the spirit and scope of the application. Accordingly, the specification is intended to embrace all modifications and equivalent arrangements that fall within the scope of the appended claims.

Claims

1. A polycarbonate nucleic acid complex comprising: (a) a polycarbonate having repeat units of the formula I or salts thereof: wherein, A comprises at least one nitrogen-containing group; R 1 Each group is independently selected from the following groups when it appears: hydrogen, deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, substituted or unsubstituted ester group, substituted or unsubstituted C group. 1-12 Alkyl, substituted or unsubstituted C 1-12 Alkoxy, substituted or unsubstituted C 1-12 Alkylamino, substituted or unsubstituted C 1-12 Alkyl ester group, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 Aryl, substituted or unsubstituted 3- to 10-membered heterocyclic alkyl and substituted or unsubstituted 5- to 14-membered heteroaryl; a1 and a2 are each independently an integer from 1 to 16; b1 and b2 are each independently an integer from 0 to 2; and (b) at least one nucleic acid.

2. The polycarbonate nucleic acid complex of claim 1, wherein, The at least one nitrogen-containing group comprises at least one protonatable nitrogen-containing group.

3. The polycarbonate nucleic acid complex of claim 1 or 2, wherein, The at least one nitrogen-containing group is located in the main chain of the polycarbonate; in particular, at least one nitrogen atom of the at least one nitrogen-containing group is located in the main chain of the polycarbonate; more particularly, the at least one protonatable nitrogen-containing group is located in the main chain of the polycarbonate; more particularly, at least one nitrogen atom of the at least one protonatable nitrogen-containing group is located in the main chain of the polycarbonate.

4. The polycarbonate nucleic acid complex of claim 2, wherein, The at least one protonatable nitrogen-containing group has a group selected from the group consisting of an amino group, a nitrogen-containing cyclic group, and combinations thereof; in particular, the amino group is selected from the group consisting of a secondary amino group, a tertiary amino group, a quaternary amino group, and combinations thereof; in particular, the nitrogen-containing cyclic group is selected from the group consisting of a 3- to 10-membered saturated or unsaturated aliphatic or aromatic heterocyclic group having at least one ring nitrogen atom, a 5- to 20-membered heterocyclic group fused with each other or with an aliphatic or aromatic carbocyclic group of a 3- to 10-membered saturated or unsaturated aliphatic or aromatic heterocyclic group having at least one ring nitrogen atom, and combinations thereof.

5. The polycarbonate nucleic acid complex of claim 1, wherein, The nucleic acid comprises one or more selected from the group consisting of a deoxyribonucleic acid (DNA), a ribonucleic acid (RNA), a peptide nucleic acid (PNA), a locked nucleic acid (LNA), and combinations thereof.

6. The polycarbonate nucleic acid complex of claim 1, wherein, The complexing mass ratio of the polycarbonate or salt thereof to the at least one nucleic acid ranges from 0.1:1 to 500:

1.

7. A method of preparing a polycarbonate, the method comprising: polymerizing at least one monomer having formula II with at least one monomer having formula III: in formula II and formula III, A 1 and A 2 each independently contains one or more groups independently of one another having the formula IV: wherein, M is selected from the group consisting of a nitrogen-containing group, a redox- sensitive group, an alkylene group, an oxygen-containing group, and combinations thereof; x1 and x2 are each independently an integer from 0 to 2; y1 and y2 are each independently an integer from 0 to 16; R 1 Each group is independently selected from the following groups when it appears: hydrogen, deuterium, tritium, hydroxyl, halogen, carboxyl, nitro, amino, cyano, substituted or unsubstituted ester group, substituted or unsubstituted C group. 1-12 Alkyl, substituted or unsubstituted C 1-12 Alkoxy, substituted or unsubstituted C 1-12 Alkylamino, substituted or unsubstituted C 1-12 Alkyl ester group, substituted or unsubstituted C 3-14 cycloalkyl, substituted or unsubstituted C 6-14 Aryl, substituted or unsubstituted 3- to 10-membered heterocyclic alkyl and substituted or unsubstituted 5- to 14-membered heteroaryl; wherein A 1 and A 2 has at least one nitrogen-containing group; wherein, for A 1 and A 2 when comprising two or more groups independently of Formula IV, the two or more groups independently of Formula IV are connected to each other via L M wherein L M is a bond, a carbonate linkage, or a C 1-16 alkylene group; and wherein the wavy line represents a bonding position with an adjacent atom.

8. A method of preparing the polycarbonate nucleic acid complex according to claim 1, the method comprising: - providing a polycarbonate or salt thereof, and at least one nucleic acid; and - mixing the polycarbonate or salt thereof with the at least one nucleic acid.

9. The method of claim 8, wherein, The mixing is performed in an aqueous medium, preferably an acidic aqueous medium.

10. A composition comprising the polycarbonate nucleic acid complex according to claim 1; in particular, the composition is a pharmaceutical composition.

11. A method of delivering at least one nucleic acid to a cell or a subject or expressing at least one nucleic acid in a cell or a subject, the method comprising: - providing the polycarbonate nucleic acid complex according to claim 1 or the composition according to claim 10, the complex being formed by complexing the at least one nucleic acid with the polycarbonate or salt thereof; and - contacting the cell or the subject with the complex or composition.

12. Use of the polycarbonate nucleic acid complex according to claim 1 or the composition according to claim 10 for delivering at least one nucleic acid to a cell or a subject.

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