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WO2023005953A1 - 载药单分子纳米聚合物、前药、胶束、药物递送系统及制备方法和用途 - Google Patents

载药单分子纳米聚合物、前药、胶束、药物递送系统及制备方法和用途 Download PDF

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WO2023005953A1
WO2023005953A1 PCT/CN2022/108093 CN2022108093W WO2023005953A1 WO 2023005953 A1 WO2023005953 A1 WO 2023005953A1 CN 2022108093 W CN2022108093 W CN 2022108093W WO 2023005953 A1 WO2023005953 A1 WO 2023005953A1
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drug
independently
molecule
nanopolymer
another
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French (fr)
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刘俊
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Jiaxing Qingzhun Medi Tech Co Ltd
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Jiaxing Qingzhun Medi Tech Co Ltd
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Priority to US18/292,836 priority Critical patent/US20250009895A1/en
Priority to CN202280038670.5A priority patent/CN118043077A/zh
Publication of WO2023005953A1 publication Critical patent/WO2023005953A1/zh
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K33/00—Medicinal preparations containing inorganic active ingredients
    • A61K33/24—Heavy metals; Compounds thereof
    • A61K33/243—Platinum; Compounds thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
    • A61K47/34—Macromolecular compounds obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyesters, polyamino acids, polysiloxanes, polyphosphazines, copolymers of polyalkylene glycol or poloxamers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
    • A61K47/59—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes
    • A61K47/60—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
    • A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/69—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
    • A61K47/6905—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a colloid or an emulsion
    • A61K47/6907—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a colloid or an emulsion the form being a microemulsion, nanoemulsion or micelle
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
    • A61K9/1075—Microemulsions or submicron emulsions; Preconcentrates or solids thereof; Micelles, e.g. made of phospholipids or block copolymers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A61K9/5107—Excipients; Inactive ingredients
    • A61K9/513—Organic macromolecular compounds; Dendrimers
    • A61K9/5146—Organic macromolecular compounds; Dendrimers obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyethylene glycol, polyamines, polyanhydrides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
    • C08G69/00—Macromolecular compounds obtained by reactions forming a carboxylic amide link in the main chain of the macromolecule
    • C08G69/48—Polymers modified by chemical after-treatment

Definitions

  • This application relates to the field of pharmaceutical technology and drug delivery systems, and relates to a drug-loaded single-molecule nanopolymer, prodrug, micelle, drug delivery system, preparation method and application, and in particular to an intracellular reducing microenvironment responsive activation type Dual-drug single-molecule nanopolymer prodrugs.
  • the present application also relates to the preparation method and application of the intracellular reducing microenvironment-responsive activated double-drug single-molecule nanopolymer prodrug.
  • Nano drug preparations have many advantages such as slow and controlled release and targeting. At present, nano drug preparation is a cutting-edge preparation technology with the core goal of precise cancer treatment. With the rapid development of nanotechnology, various biologically active molecules (chemical drugs, polypeptides, nucleic acids, etc.) can be stored in nanomaterials with various properties in various ways (such as molecular self-assembly). In particular, these nanomaterials can rely on the functional design of member molecules and the tiny regulation of assembly structures to become "smart transporters" in living organisms, which have the potential to overcome biological barriers at all levels and deliver bioactive molecules to target sites in a directional manner.
  • various biologically active molecules chemical drugs, polypeptides, nucleic acids, etc.
  • these nanomaterials can rely on the functional design of member molecules and the tiny regulation of assembly structures to become "smart transporters" in living organisms, which have the potential to overcome biological barriers at all levels and deliver bioactive molecules to target sites in a directional manner.
  • the drug release behavior of traditional nano-drug formulations has the problem that the local instantaneous drug concentration is difficult to reach an effective level.
  • traditional nano-preparations have high enrichment potential for tumor lesions, their slow drug release rate makes the killing effect of traditional nano-preparations on tumor cells even lower than that of free small molecule drugs.
  • the drug release behavior of traditional nano-preparations also occurs in the blood circulation, and the early leakage of the drug will reduce the bioavailability of the drug to the target lesion, resulting in the toxicity of non-target lesions.
  • nano-preparations or self-assembled nano-preparations also have the following obvious disadvantages: 1) poor colloidal stability, prone to structural dissociation under complex physiological conditions; 2) early leakage of drugs; and 3) complex preparation process, such as: Thin-film hydration, nano-precipitation, etc., need to remove uncoated drug molecules and auxiliary molecules (such as: organic solvents, etc.), which is difficult for large-scale mass production.
  • prodrug-based design strategies can not only solve the problem of premature drug leakage, but also rapidly release drugs in response to the tumor microenvironment or intracellular microenvironment.
  • nanopolymer prodrugs with higher in vivo and in vitro stability, improved drug loading and drug release properties, simple and efficient production process, and tolerance to ultrasound, lyophilization and reconstitution.
  • One object of the present application is to provide a drug-loaded single-molecule nanopolymer, which includes multiple polyamino acid chains, and the chains of the multiple polyamino acid chains are covalently linked by multiple divalent linkers L Pt to make the A plurality of polyamino acid chains constitute a non-linear skeleton, at least one end of the polyamino acid chain is connected with a hydrophilic polymer chain; wherein, the linear skeleton of the divalent linking group L Pt contains platinum atoms, and the platinum Atoms participate in the formation of platinum-based drug units, and the platinum-based drug units can be residues of active ingredients of platinum-based drugs or their prodrugs;
  • the side group of the polyamino acid chain is grafted with a second drug unit; wherein, the second drug unit may be a residue of an active ingredient of an antitumor drug or a prodrug thereof.
  • the drug-loaded single-molecule nanopolymer constructs multiple polyamino acid chains into a nonlinear skeleton through a divalent platinum-containing linker L Pt , at least one polyamino acid chain is connected to a hydrophilic polymer chain at the end, and the end of the L Pt
  • the platinum atom participates in the formation of the platinum-based drug unit (it may be the residue of the active ingredient of the platinum-based drug or its prodrug).
  • the drug-loaded single-molecule nanopolymer can be controlled to have a branched or moderately cross-linked three-dimensional structure, and further combined with the design of the position of the hydrophilic polymer chain at the end of the polyamino acid chain, the drug-loaded single molecule Molecular nanopolymers can form single-molecule nanopolymer micelles with a core-shell structure without self-assembly in aqueous media.
  • the hydrophilic polymer chains are distributed in the outer shell, and the drug ingredients are entrapped in the inner core.
  • the drug-loaded single-molecule nanopolymer can only be loaded with platinum drug units to form a platinum single-drug single-molecule nanopolymer; or the residue of its prodrug), the second drug unit can be grafted on the side group of the polyamino acid chain, and at this time, a double-drug single-molecule nanopolymer can be formed.
  • the relative content of the platinum drug unit and the second drug unit can be flexibly adjusted by controlling the feeding amount of the corresponding monomer.
  • the distribution density of L Pt can be adjusted by adjusting the feeding ratio of unbranched amino acid monomers and L Pt branched amino acid monomers. In the unbranched amino acid monomers, the amount of amino acid monomers containing the second drug unit can also be flexibly adjusted. Proportion.
  • the drug-loaded single-molecule nanopolymer has good stability in vivo and in vitro, good dispersibility, uniform particle size, no toxic and side effects, and does not release active pharmaceutical ingredients outside the cell but exhibits triggered release of active pharmaceutical ingredients inside the cell , in addition, it can be obtained by a preparation method with simple operation, mild reaction, low cost and environmental friendliness.
  • Another object of the present application is to provide a method for preparing a drug-loaded single-molecule nanopolymer, which includes the following steps: a platinum-containing compound having a structure such as formula (I-3), a structure such as formula (III-3) The monofunctional hydrophilic polymer shown, the optional structure of the drug compound shown in formula (II-3) and the optional compound shown in formula (IV-3) are mixed in an organic solvent to carry out ring opening Polymerization;
  • U 1 and U 2 are each independently a carbon-centered trivalent group
  • D Pt is a platinum-based drug unit (which may be a residue of a platinum-based drug active ingredient or its prodrug);
  • F 5 is -NH 2 , -COOH, Preferably -NH 2 ;
  • U 3 is independently a carbon center trivalent group
  • LR is independently a responsive linker
  • L 4 is independently a divalent linker or none
  • DT is a second drug unit (which can be an active ingredient of an antineoplastic drug or Residues of its prodrug); Among them, LR can undergo bond breaking under external stimuli;
  • PE is RE or protected RE, which does not have reactivity in the ring-opening polymerization reaction;
  • RE is independently H or R 0 ; wherein, R 0 is an end group not containing a drug unit;
  • the ring-opening polymerization reaction is carried out under anhydrous conditions
  • the reaction temperature of the ring-opening polymerization is 15-40° C., and more preferably, the reaction time of the ring-opening polymerization is 24-96 hours.
  • the polymerization utilizes ring-opening polymerization involving bis-N-carboxylic acid anhydride (NCA) to obtain single-molecule nanopolymers through a "one-pot method", which can form cores without self-assembly in aqueous media.
  • NCA bis-N-carboxylic acid anhydride
  • the micelles with a shell structure provide a drug delivery system that can release active pharmaceutical ingredients in response to the treatment of tumor diseases.
  • Another purpose of the present application is to provide an intracellular reducing microenvironment-responsive activated dual-drug single-molecule nanopolymer prodrug, which can be used as a platform technology for simultaneous delivery of two drug active ingredients.
  • Another object of the present application is to provide a method for preparing intracellular reducing microenvironment-responsive activated double-drug single-molecule nanopolymer prodrugs.
  • Another object of the present application is to provide a drug-loaded single-molecule nanopolymer micelle, whose composition is selected from any of the following: the aforementioned drug-loaded single-molecule nanopolymer, the drug-loaded single-molecule prepared by the aforementioned preparation method Nanopolymer, the aforementioned double-drug single-molecule nanopolymer prodrug, and the double-drug single-molecule nanopolymer prodrug prepared by the aforementioned preparation method; the drug-loaded single-molecule nanopolymer micelle has a core-shell structure, The outer shell structure is a hydrophilic layer formed by hydrophilic polymer chains, and the contained drug units are located in the inner core.
  • the drug-loaded single-molecule nanopolymer provided in this application can form nanopolymer micelles with a core-shell structure in situ during the polymerization reaction, including hydrophilic polymer chains located in the shell and drug units located in the core.
  • the platinum single-drug single-molecule nanopolymer provided by the present application can form nanopolymer micelles with a core-shell structure in situ during the polymerization reaction, including hydrophilic polymer chains located in the shell and platinum drug units located in the core.
  • the double-drug single-molecule nanopolymer provided by this application can form nanopolymer micelles with a core-shell structure in situ during the polymerization reaction, including a hydrophilic polymer chain located in the outer shell, a platinum drug unit located in the inner core, and a second polymer micelle. Two drug units.
  • Another object of the present application is to provide the use of the aforementioned drug-loaded single-molecule nanopolymer as a prodrug.
  • the drug-loaded single-molecule nanopolymer can enter the interior of cells, sense the intracellular microenvironment, release drug active ingredients in response, generate cytotoxicity, and inhibit the growth of tumor cells.
  • Another object of the present application is to provide a use of a double-drug single-molecule nanopolymer for delivery of active pharmaceutical ingredients or in the preparation of a drug delivery system.
  • the active pharmaceutical ingredient can be released from the aforementioned platinum drug unit and the aforementioned optional second drug unit.
  • the active ingredient of the drug can be an active ingredient of a platinum-based drug and an optional active ingredient of an antitumor drug.
  • Another object of the present application is to provide the aforementioned drug-loaded single-molecule nanopolymer, the drug-loaded single-molecule nanopolymer prepared by the aforementioned preparation method, the aforementioned double-drug single-molecule nanopolymer prodrug, or the aforementioned preparation method. Use of the obtained double-drug single-molecule nanometer polymer prodrug in the preparation of drugs for treating tumor diseases.
  • Another object of the present application is to provide a drug delivery system, which comprises a drug-loaded single-molecule nanopolymer micelle, the drug-loaded single-molecule nanopolymer micelle comprises the aforementioned drug-loaded single-molecule nanopolymer or the aforementioned preparation method The prepared drug-loaded single-molecule nanopolymer;
  • the hydrophilic polymer chain is located in the outer shell of the drug-loaded single-molecule nanopolymer micelle
  • Both the platinum drug unit and the second drug unit are located in the inner core of the drug-loaded single-molecule nanopolymer micelles.
  • Another object of the present application is to provide a drug delivery system, which comprises a double-drug single-molecule nanopolymer micelle, the double-drug single-molecule nanopolymer micelle comprises a polyamino acid linked to a hydrophilic polymer, wherein The ⁇ -carbon of the repeating unit of the polyamino acid is bonded to the prodrug part of the active ingredient of the platinum drug and the prodrug part of the active ingredient of the antineoplastic drug; preferably, the molecule of the active ingredient of the antineoplastic drug contains free hydroxyl, Free amino groups or a combination of both.
  • Another object of the present application is to provide a use of an intracellular reducing microenvironment-responsive activated double-drug single-molecule nanopolymer prodrug for the delivery of active pharmaceutical ingredients or in the preparation of a drug delivery system.
  • Another object of the present application is to provide the double NCA monomer of the active ingredient of platinum-based drugs and the single NCA monomer of the active ingredient of anti-tumor drugs in the preparation of single-molecule nanopolymer prodrugs or drug delivery systems; preferably, the The molecular structure of the active ingredients of the above-mentioned antineoplastic drugs contains free hydroxyl groups or free amino groups.
  • a biocompatible hydrophilic polymer with terminal amino or carboxyl groups and platinum-containing bis-N-carboxylic acid anhydride (NCA) monomers and active ingredients of antitumor drugs The single NCA monomer (preferably containing free hydroxyl group, free amino group or combination thereof in the molecule of the active ingredient of antineoplastic drugs, so as to be able to couple to the NCA terminal group) can form a single-molecule nanopolymer of core-shell structure in situ (can be used as Prodrugs, therefore, can also be recorded as unimolecular nanopolymer prodrugs), the polymer prodrugs can couple two kinds of antitumor drugs, good stability in vivo and in vitro, good dispersibility, uniform particle size, nontoxic Side effects, and no release of pharmaceutical active ingredients outside the cell, but trigger release of pharmaceutical active ingredients inside the cell, and its preparation method is simple and easy to operate, mild in reaction, low in cost and environmentally friendly.
  • NCA platinum-containing bis-N-carboxylic acid anhydride
  • the present application also provides the aforementioned drug-loaded single-molecule nanopolymer, the drug-loaded single-molecule nanopolymer prepared by the aforementioned preparation method, the aforementioned double-drug single-molecule nanopolymer prodrug, and the aforementioned double-drug single-molecule nanopolymer prepared by the aforementioned preparation method.
  • the present application also provides an intracellular reducing microenvironment-responsive activated double-drug single-molecule nanopolymer prodrug with a core-shell structure, a drug delivery system, and a preparation method and use thereof.
  • the present application overcomes said disadvantages of polymeric prodrugs in the conventional art.
  • the application provides a double-drug single-molecule nanopolymer prodrug with a core-shell structure, wherein the inner core contains a platinum-based drug molecular structural unit, a pharmaceutically active molecular structural unit, and a polyamino acid structural unit, and the structural units are shared by Valence bond connection; the shell is a biocompatible hydrophilic polymer (such as polyethylene glycol, etc.); preferably, the molecule of the pharmaceutically active molecule contains free hydroxyl groups, free amino groups or a combination of both.
  • the inner core contains a platinum-based drug molecular structural unit, a pharmaceutically active molecular structural unit, and a polyamino acid structural unit, and the structural units are shared by Valence bond connection
  • the shell is a biocompatible hydrophilic polymer (such as polyethylene glycol, etc.); preferably, the molecule of the pharmaceutically active molecule contains free hydroxyl groups, free amino groups or a combination of both.
  • the double-drug single-molecule nanopolymer prodrug of the present application can inhibit the non-specific reaction of the active ingredient of the drug in the blood circulation, and has the function of triggering the release of the active ingredient of the drug in response to the intracellular reducing microenvironment after entering the cell.
  • the application provides a double-drug single-molecule nanopolymer prodrug, which is composed of a hydrophilic polymer with a terminal amino group, a double NCA monomer of a platinum-based drug active ingredient, and an anti-tumor drug active ingredient.
  • a single NCA monomer is formed; preferably, the molecule of the active ingredient of the antineoplastic drug contains free hydroxyl group, free amino group or a combination of both.
  • the present application provides a dual-drug single-molecule nanopolymer prodrug comprising a polyamino acid linked to a hydrophilic polymer, wherein platinum is bonded to the alpha carbon of the repeating unit of the polyamino acid
  • the prodrug part of the drug-like active ingredient and the prodrug part of the antitumor drug active ingredient preferably, the molecule of the antitumor drug active ingredient contains free hydroxyl group, free amino group or a combination of both.
  • a drug delivery system which comprises a double-drug single-molecule nanopolymer micelle, and the double-drug single-molecule nanopolymer micelle comprises the aforementioned double-drug single-molecule nanopolymer or the aforementioned preparation method The prepared double-drug single-molecule nanopolymer;
  • the hydrophilic polymer chain is located in the outer shell of the double-drug single-molecule nanopolymer micelle;
  • Both the platinum-based drug unit and the second drug unit are located in the inner core of the double-drug single-molecule nanopolymer micelle.
  • the present application provides a drug delivery system, which comprises a double-drug single-molecule nanopolymer micelle, the polymer micelle has a core-shell structure, wherein the inner core contains a platinum-based drug molecular structural unit and a drug active molecular structural unit As well as polyamino acid structural units, the structural units are connected by covalent bonds; and the shell is a biocompatible hydrophilic polymer (such as polyethylene glycol, etc.); preferably, the molecule of the pharmaceutically active molecule Contains free hydroxyl groups, free amino groups or a combination of free hydroxyl groups and free amino groups.
  • the present application provides a drug delivery system, which comprises a double-drug single-molecule nanopolymer micelle, the polymer micelle is composed of a hydrophilic polymer with a terminal amino group, a platinum-based drug active ingredient double
  • the NCA monomer and the single NCA monomer of the active ingredient of the anti-tumor drug are formed; preferably, the molecule of the active ingredient of the anti-tumor drug contains a free hydroxyl group, a free amino group or a combination of the two.
  • the present application provides a drug delivery system comprising a double-drug single-molecule nanopolymer micelle, the double-drug single-molecule nanopolymer micelle comprising a polyamino acid linked to a hydrophilic polymer, wherein On the alpha carbon of the repeating unit of the polyamino acid, the prodrug part of the active ingredient of the platinum drug and the prodrug part of the active ingredient of the antineoplastic drug are bonded; preferably, the molecule of the active ingredient of the antineoplastic drug contains a free hydroxyl group , free amino groups or a combination of both.
  • the application provides a method for preparing the double-drug single-molecule nanopolymer prodrug of the application, the method comprising the steps of:
  • step (3) Under suitable reaction conditions, the monomer obtained in step (1) and step (2) is reacted with a hydrophilic polymer having terminal amino groups to obtain the double-drug single-molecule nanopolymer prodrug of the present application, and
  • the double-drug single-molecule nanopolymer prodrug of the present application is prepared by a one-step one-pot ring-opening polymerization method, that is, the present application provides a method for preparing a double-drug single-molecule nanopolymer prodrug, the method comprising follows the steps below:
  • the single NCA monomer of the active ingredient of antineoplastic drugs preferably, the molecular structure of the active ingredient of antineoplastic drugs contains free hydroxyl or free amino groups
  • the double NCA monomer of the active ingredient of platinum drugs The NCA monomer reacts with a hydrophilic polymer with terminal amino groups to obtain the double-drug single-molecule nanopolymer prodrug, and
  • This one-step, one-pot ring-opening polymerization method of the present application avoids many disadvantages of traditional self-assembled nano preparations.
  • the present application provides a bis-NCA monomer suitable for the preparation of platinum-based active ingredients of single-molecule nanopolymer prodrugs.
  • the present application provides a single NCA monomer suitable for preparing antitumor drug active ingredients containing free hydroxyl groups or free amino groups in the molecular structure of single-molecule nanopolymer prodrugs.
  • the present application provides a method for simultaneously delivering a pharmaceutical active ingredient to a target site, the method comprising preparing the target pharmaceutical active ingredient into a single-molecule nanopolymer prodrug and injecting an effective amount of the single-molecule nanopolymer prodrug Medicines are given to patients in need.
  • the pharmaceutical active of interest comprises a platinum-based pharmaceutical active or a prodrug thereof.
  • the present application provides a method for simultaneously delivering two active pharmaceutical ingredients to a target site, the method comprising preparing the two active active ingredients into a single-molecule nanopolymer prodrug and injecting an effective amount of the single-molecule Nanopolymer prodrugs are administered to patients in need.
  • the present application provides a method for simultaneously delivering two pharmaceutically active ingredients to a target site, the method comprising preparing the two pharmaceutically active ingredients into single-molecule nanopolymer prodrugs and preparing the prepared The single-molecule nanopolymer prodrug is administered to patients in need, wherein one of the two drug active components is a platinum drug, and the other is an anti-tumor drug active component containing free hydroxyl or free amino in the molecular structure.
  • the present application provides a polymer prodrug delivery system for delivering the active ingredient of a drug contained in a single NCA monomer of an active ingredient of an antineoplastic drug and a double NCA monomer of an active ingredient of a platinum drug to a target site
  • a polymer prodrug delivery system for delivering the active ingredient of a drug contained in a single NCA monomer of an active ingredient of an antineoplastic drug and a double NCA monomer of an active ingredient of a platinum drug to a target site
  • the molecular structure of the active ingredient of the antineoplastic drug contains free hydroxyl or free amino.
  • the application provides the single NCA monomer of the active ingredient of an antineoplastic drug and the use of the double NCA monomer of an active ingredient of a platinum-based drug in the preparation of a single-molecule nanopolymer prodrug micelle; preferably, the The molecular structure of the active ingredients of antineoplastic drugs contains free hydroxyl groups or free amino groups.
  • the application provides the use of the double-drug single-molecule nanopolymer prodrug in the preparation of a drug for the treatment of corresponding diseases, wherein the double-drug single-molecule nanopolymer prodrug comprises a hydrophilic polymer linking A polyamino acid, wherein the prodrug part of the active ingredient of the platinum drug and the prodrug part of the active ingredient of the antitumor drug are bonded to the ⁇ carbon of the repeating unit of the polyamino acid; preferably, the active ingredient of the antitumor drug
  • the molecule contains free hydroxyl groups, free amino groups or a combination of both.
  • the present application provides a method for treating tumors in a patient in need with combination therapy, the method administers a therapeutically effective amount of a single-molecule nanopolymer drug to the patient, wherein the single-molecule nanopolymer drug comprises A polyamino acid linked to a hydrophilic polymer, wherein the prodrug part of the active ingredient of a platinum-based drug and the prodrug part of an active ingredient of an antitumor drug are bonded to the ⁇ -carbon of the repeating unit of the polyamino acid; preferably, the The molecule of the active ingredient of the above-mentioned antineoplastic drug contains free hydroxyl group, free amino group or a combination of both.
  • the nanopolymer prodrug or polymer prodrug nanomicelle of the present application integrates the advantages of nano preparations (comprising: long blood circulation time, low liver organ uptake, lesion site target Potential for enrichment) and the advantages of prodrugs (reduce the early inactivation of active drugs, precise drug activation), and ultimately help to increase the spatiotemporal concentration of active drugs at the target site, thereby enhancing drug efficacy, while reducing the effect of drugs on non-active drugs. Potential toxic side effects at the targeted site.
  • the polymer prodrug nanomicelle of the present application has better stability than the polymer prodrug nanomicelle of the traditional technology.
  • the stability advantages of the polymer prodrug nanomicelles of the present application may be due to the following factors: full chemical bonding of the drug, resistance to physical treatments such as centrifugation, ultrafiltration, hydrothermal, ultrasonic, etc., to ensure that the nanomicelles
  • the production process of the polymer prodrug nanomicelle of the present application also has technical advantages. Without being limited by a specific theory, the technical advantages of the production process of the polymer prodrug nanomicelles of the present application may be due to the following factors: full chemical bonding of the drug, fine and controllable drug loading, breakthrough in the batch stability of the self-loading system Difficulties, and no free drug, can be stored in solution; a single nanoparticle is a single molecule, the freeze-drying and reconstitution process is simple, and the technical requirements are low; and the "one-pot method" synthesis of nanomicelles does not require complicated preparations such as film hydration and nanoprecipitation and purification process.
  • the polymer prodrug nanomicelles of the present application also have advantages in terms of drug loading and drug release.
  • the advantages of the polymer prodrug nanomicelles of the present application in terms of drug loading and drug release may be due to the following factors: full chemical bonding of the drug, no leakage of the drug outside (blood circulation, extracellular matrix); And intracellular triggered release, on the one hand, the release increases the concentration of drugs in time and space, strengthens the efficacy of drugs, and solves the disadvantages of passive and slow drug release of self-packaged nano-preparations; medicinal effect.
  • the polymer prodrug nanomicelle of the present application can contain double drugs, which has the following advantages: the targets of the double drugs are different, which overcomes drug resistance; consumes intracellular drug-resistant glutathione, which overcomes drug resistance; And platinum-based drugs and another anti-tumor drug such as camptothecin produce anti-tumor activities through different mechanisms of action, and have excellent synergistic effects.
  • Fig. 1 is a schematic diagram showing the composition of the double-drug (platinum drug unit + second drug unit) unimolecular nanopolymer prodrug micelle of the present application.
  • Figure 2 is a schematic diagram showing the composition of the double-drug (cisplatin+camptothecin) monomolecular nanopolymer prodrug micelles of the present application.
  • III-A, III-B, III-C and III-D in Fig. 3 are the mass spectrograms of the intermediate products used to synthesize the product of the present application respectively.
  • IV-A, IV-B, IV-C, IV-D and IV-E in Fig. 4 are 1 H NMR spectra of intermediate products used to synthesize the product of the present application respectively.
  • V-A and V-B in FIG. 5 are molecular exclusion chromatograms showing the molecular weights of the double-drug (cisplatin+camptothecin) single-molecule nanopolymer prodrug and its intermediate product of the present application, respectively.
  • Figure 6 is a dynamic light scattering (DLS) diagram characterizing the particle size and polydispersity index of the double-drug (cisplatin+camptothecin) unimolecular nanopolymer prodrug micelles and control micelles of the present application.
  • DLS dynamic light scattering
  • Fig. 7 is a transmission electron microscope image of the double-drug (cisplatin+camptothecin) unimolecular nanopolymer prodrug micelle (VII-A) and control micelle (VII-B) of the present application.
  • Fig. 8 is a transmission electron microscope image of the double-drug (cisplatin+camptothecin) unimolecular nanopolymer prodrug micelle (VIII-A) and control micelle (VIII-B) of the present application before and after freeze-drying.
  • Figure 9 shows the colloid dynamics characteristics of the double-drug (cisplatin+camptothecin) monomolecular nanopolymer prodrug micelles (A) and control micelles (B) of the present application tested by fluorescence correlation spectroscopy.
  • Fig. 10 is a small-angle X-ray scattering diagram characterizing the double-drug (cisplatin+camptothecin) monomolecular nanopolymer prodrug micelles of the present application.
  • Fig. 11 is a dynamic light scattering diagram characterizing the particle size distribution of the double-drug (cisplatin+camptothecin) single-molecule nanopolymer prodrug micelles and control micelles of the present application.
  • Fig. 12 is a high performance liquid chromatogram of the double-drug (cisplatin+camptothecin) single-molecule nanopolymer prodrug micelles and control micelles of the present application.
  • Figure 13 shows the drug release behavior of the double-drug (cisplatin+camptothecin) unimolecular nanopolymer prodrug micelles of the present application.
  • Figure 14 shows the simulated drug release behavior of control micelles in vitro.
  • Figure 15 shows the cytotoxicity of the double-drug (cisplatin+camptothecin) single-molecule nanopolymer prodrug micelles and control micelles of the present application.
  • Figure 16 shows the pharmacokinetic study results of the double-drug (cisplatin+camptothecin) single-molecule nanopolymer prodrug micelle (A) and control micelle (B) of the present application.
  • Figure 17 shows the parent drug accumulation test results of the double-drug (cisplatin+camptothecin) unimolecular nanopolymer prodrug micelles and control micelles of the present application.
  • Fig. 18 is the results of the tumor suppression test of the double-drug (cisplatin+camptothecin) single-molecule nanopolymer prodrug micelles and control micelles of the present application.
  • Fig. 19 is the 1 H NMR spectrum of the double-drug (cisplatin+camptothecin) monomolecular nanopolymer prodrug of the present application.
  • Figure 20 is a dynamic light scattering diagram characterizing the particle size and polydispersity index of the double-drug (cisplatin+paclitaxel) unimolecular nanopolymer prodrug micelles of the present application.
  • Figure 21 shows the drug release behavior of the double-drug (cisplatin+paclitaxel) unimolecular nanopolymer prodrug micelles of the present application.
  • Figure 22 shows the transmission electron microscope image of the double-drug (cisplatin+paclitaxel) unimolecular nanopolymer prodrug micelles of the present application.
  • Fig. 23 is a dynamic light scattering diagram of the particle size and polydispersity index of the double-drug (cisplatin + resiquimod) unimolecular nanopolymer prodrug micelles of the present application.
  • Figure 24 shows the drug release behavior of the double-drug (cisplatin+resiquimod) single-molecule nanopolymer prodrug micelles of the present application.
  • Figure 25 shows the transmission electron microscope image of the double-drug (cisplatin+resiquimod) single-molecule nanopolymer prodrug micelles of the present application.
  • Fig. 26 shows the results of gel permeation chromatography (SEC) test (A) and dynamic light scattering (DLS) test (B) of platinum single-drug single-molecule nanopolymer in Example 10 of the present application, using cisplatin.
  • SEC gel permeation chromatography
  • DLS dynamic light scattering
  • Figure 27 shows the transmission electron microscope (TEM) test images of platinum single-drug single-molecule nanopolymer micelles in Example 10 of the present application before (A) and after freeze-drying and reconstitution (B).
  • TEM transmission electron microscope
  • Fig. 28 shows the mass spectrum of NCA-DACHPt-NCA prepared in Example 11 of the present application.
  • Fig. 29 shows the results of gel permeation chromatography (SEC) test (A) and dynamic light scattering (DLS) test (B) of platinum single-drug single-molecule nanopolymer in Example 11 of the present application, using DACHPt.
  • SEC gel permeation chromatography
  • DLS dynamic light scattering
  • FIG. 30 shows the transmission electron microscope (TEM) test images of platinum single-drug single-molecule nanopolymer micelles in Example 11 of the present application before (A) and after freeze-drying and reconstitution (B).
  • TEM transmission electron microscope
  • Fig. 31 shows the 1 H NMR spectrum of PTX-ss-NCA prepared in Example 12 of the present application.
  • Figure 32 shows the DLS test results of the double-drug (cisplatin+paclitaxel) single-molecule nanopolymer prepared in Example 12 of the present application, (A) without ultrasonic treatment, (B) ultrasonic treatment.
  • Figure 33 shows the transmission electron microscope (TEM) of the double-drug (cisplatin+paclitaxel) unimolecular nanopolymer micelles prepared in Example 12 of the present application before (A) and after freeze-drying and reconstitution (B) test chart.
  • TEM transmission electron microscope
  • Figure 34 shows the results of drug release behavior of the double-drug (cisplatin+paclitaxel) single-molecule nanopolymer micelles prepared in Example 12 of the present application under different conditions.
  • Fig. 35 shows the 1 H NMR spectrum of R848-ss-NCA prepared in Example 13 of the present application.
  • Figure 36 shows the DLS test results of the double-drug (cisplatin+R848) single-molecule nanopolymer prepared in Example 13 of the present application, (A) without ultrasonic treatment, (B) ultrasonic treatment.
  • Figure 37 shows the transmission electron microscope (TEM) of the double-drug (cisplatin+R848) unimolecular nanopolymer micelles prepared in Example 13 of the present application before (A) and after freeze-drying and reconstitution (B) test chart.
  • TEM transmission electron microscope
  • Figure 38 shows the results of the drug release behavior of the double-drug (cisplatin+R848) single-molecule nanopolymer micelles prepared in Example 13 of the present application under different conditions.
  • Fig. 39 shows the 1 H NMR spectrum of MMAE-ss-NCA prepared in Example 14 of the present application.
  • FIG. 40 shows the DLS test results of the double-drug (cisplatin+MMAE) single-molecule nanopolymer prepared in Example 14 of the present application.
  • Figure 41 shows the transmission electron microscope (TEM) of the double-drug (cisplatin+MMAE) unimolecular nanopolymer micelles prepared in Example 14 of the present application before (A) and after freeze-drying and reconstitution (B) test chart.
  • TEM transmission electron microscope
  • Fig. 42 shows the drug release behavior results of the double-drug (cisplatin+MMAE) unimolecular nanopolymer micelles prepared in Example 14 of the present application under different conditions.
  • the terms “and/or”, “or/and”, “and/or” include any one of two or more of the associated listed items, and any of the associated listed items. and all combinations including any combination of any two of the relevant listed items, any more of the relevant listed items, or all of the relevant listed items. It should be noted that when at least three items are connected with at least two conjunctions selected from “and/or”, “or/and”, “and/or”, it should be understood that in this application, the technical solution Undoubtedly include the technical solutions that are all connected by "logic and”, and also undoubtedly include the technical solutions that are all connected by "logic or”. For example, "A and/or B” includes three parallel schemes of A, B and A+B.
  • the technical solution of "A, and/or, B, and/or, C, and/or, D” includes any one of A, B, C, and D (that is, all are connected by "logic or") technical solution), also includes any and all combinations of A, B, C, and D, that is, includes any combination of any two or any three of A, B, C, and D, and also includes A, B, and C , four combinations of D (that is, all use the technical scheme of "logic and" connection).
  • the units related to the data range are only followed by the right endpoint, it means that the units of the left endpoint and the right endpoint are the same.
  • 50 ⁇ 1000Da means that the units of the left endpoint 50 and the right endpoint 1000 are both Da.
  • water soluble polymer refers to any pharmaceutically acceptable biocompatible polymer that is soluble in water at room temperature.
  • water-soluble polymer having a terminal amino group refers to a water-soluble polymer as defined above in which one terminal in the molecular structure is an amino group (-NH 2 ).
  • auxiliary material used in the application refers to an auxiliary substance that can be included in the nanopolymer prodrug micelle composition of the application and does not cause obvious harmful pharmacological effects to patients, and it can be combined with " “Pharmaceutically acceptable excipient” or “pharmaceutically acceptable carrier” are used interchangeably.
  • therapeutically effective amount used in this application means the dosage of the Chinese medicine preparation of the application, which is sufficient to achieve the desired effect on the disease when the Chinese medicine preparation of the application is given to the subject to treat the novel coronavirus infectious disease. treatment effect.
  • the “therapeutically effective dose” can be adjusted according to the actual preparation form used, the symptoms and severity of the disease, and the age and body weight of the subject to be treated.
  • patient refers to a living organism suffering from or susceptible to a disease that can be prevented or treated by administering the double-drug single-molecule nanopolymer prodrug of the application, including humans and mammals, preferably humans .
  • tumor is understood in its broadest sense to mean an abnormal overgrowth of tissue.
  • Carcinoma or “cancer” refers to a malignant tumor.
  • Nano-medicine preparations such as Bind-14, NC-6300 are delivered through physical interactions (such as hydrophobic interactions, electrostatic interactions, etc.) between drug molecules and delivery molecules (amphiphilic molecules: lipid molecules, block copolymers, etc. ) is formed in a self-assembled manner. After the nanomedicine reaches the tumor lesion, the drug release is often achieved by passive diffusion. This slow drug release behavior will make it difficult for the local instantaneous drug concentration to reach an effective level. Therefore, although traditional nano-preparations have a high enrichment potential for tumor lesions, their slow drug release rate makes traditional nano-preparations even lower in killing effect on tumor cells than free small molecule drugs.
  • the platinum (Pt) in the drug-loaded single-molecule nanopolymer or in the double-drug single-molecule nanopolymer prodrug is tetravalent platinum, has an octahedral space structure, and has high chemical reaction inertia,
  • the chemical structure is stable in plasma and normal tissue, therefore, in the process of in vivo delivery, the systemic toxicity is small, and at the same time, there is no cross-resistance between tetravalent platinum and divalent platinum, and it enters into tumor cells, and the highly reducing environment can make Reduction of tetravalent platinum releases active divalent platinum species, which in turn produces cytotoxicity.
  • divalent platinum Compared with divalent platinum, divalent platinum has high chemical reactivity and can bind to proteins in plasma. Therefore, the bioavailability of divalent platinum is low; at the same time, divalent platinum can detoxify with thiol-containing biomolecules In addition, the cross-resistance of divalent platinum seriously restricts its clinical curative effect and long-term practicality.
  • a drug-loaded single-molecule nanopolymer which includes multiple polyamino acid chains, and the chains of the multiple polyamino acid chains are covalently linked by multiple divalent linkers L Pt to make all polyamino acid chains
  • the plurality of polyamino acid chains constitute a non-linear skeleton, at least one end of the polyamino acid chain is connected with a hydrophilic polymer chain; wherein, the linear skeleton of the divalent linking group L Pt contains platinum atoms, and the Platinum atoms participate in the formation of platinum-based drug units, and the platinum-based drug units can be residues of active ingredients of platinum-based drugs or their prodrugs;
  • the side group of the polyamino acid chain is grafted with a second drug unit; wherein, the second drug unit may be a residue of an active ingredient of an antitumor drug or a prodrug thereof.
  • each occurrence of the active ingredient of the anti-tumor drug or its prodrug is independently connected to the amino acid repeating unit through a responsive linker LR , and the response Sexual linker LR can break bonds under external stimuli.
  • polyamino acid chain used in the present application means a polymer chain formed by sequentially linking -NH 2 ends and -COOH ends of a plurality of aminocarboxylic acid molecules through -CO-NH-bonds.
  • the " ⁇ amino acid” used in this application means NH 2 -CR C RE -COOH, where R C can be H or a non-hydrogen atom or group that does not affect the NCA ring-opened polymer, RE can be hydrogen or R 0 , wherein, R 0 is an end group not containing a drug unit. R 0 can also optionally be defined below.
  • amino acid as used in the present application means a compound containing at least one -NH and at least one -COOH, which can be a natural amino acid (such as lysine) or an unnatural amino acid (such as ornithine).
  • the amino acid unit constituting the structural unit of the polyamino acid chain of the present application may be an ⁇ amino acid unit.
  • Non-linear as used herein means a branched or cross-linked topology.
  • a divalent linker L Pt can be covalently connected with two polyamino acid chains to form two branch points, and by adjusting the relative ratio of L Pt and polyamino acid chains, the appropriate degree of branching can be controlled, specifically , can be regulated by adjusting the ratio of the number of L Pt to the total number of amino acid units in the drug-loaded single-molecule nanopolymer. The greater the average number of L Pts linked by a polyamino acid chain, the more branch points.
  • Too low a degree of branching leads to too high flexibility, a high degree of branching forms a cross-linked three-dimensional network, and too high a degree of cross-linking leads to high rigidity. Therefore, too few or too many branch points will lead to drug loading Unimolecular nanopolymers will affect the formation of nanomicelles and drug release properties.
  • Hydrophilic polymer chain or “hydrophilic polymer” as used herein means capable of swelling or dissolving in water.
  • the "polymer” used in this application has at least two structural units, and its molecular weight is not particularly limited, and may be greater than or equal to 1000 Da or less than or equal to 1000 Da.
  • hydrophilic polymer and hydrophilic polymer have the same meaning and can be used interchangeably.
  • hydrophilic polymer chain and hydrophilic polymer chain have the same meaning and can be used interchangeably.
  • the hydrophilic polymer chain can be connected to the N-terminal or C-terminal of the polyamino acid chain.
  • the hydrophilic polymer chain is connected to the C-terminus of the polyamino acid chain, and may be connected through an amide bond (-CONH-).
  • the hydrophilic polymer chain is connected to the N-terminus of the polyamino acid chain, and may be connected through an amide bond (-NH-CO-) or a carbamate group (-NH-COO-).
  • valence state is referred to as “valence” for short, and refers to the number of mutual combination of an atom or atomic group, group (root) of various elements with other atoms.
  • valence refers to the number of mutual combination of an atom or atomic group, group (root) of various elements with other atoms.
  • a “residue” of a substance generally refers to the remaining structure of the substance without at least one atom.
  • the state in which the platinum-containing substance is covalently linked to two adjacent atoms is also recorded as the residue of the platinum-containing substance. for example, Corresponding respectively residues.
  • the side group of the amino acid unit constituting the polyamino acid chain may be connected to L Pt to form a branch point, may be connected to a second drug unit, or may be a hydrogen atom or a free end group R 0 not connected to a drug unit.
  • a hydrophilic polymer chain may be linked to the end of the polyamino acid chain.
  • the chain length of the polyamino acid chain can be regulated through the introduction of terminal hydrophilic polymer chains, thereby adjusting the size of the drug-loaded single-molecule nanopolymer.
  • the drug-loaded single-molecule nanopolymer has suitable branching density, suitable drug loading, suitable ratio of different drugs and suitable polyamino acid chain length, so that the drug-loaded single-molecule nanopolymer has a suitable size and has a suitable
  • the size of nanomicelles including core size, shell thickness, particle size, average diameter, etc.
  • aqueous medium refers to an aqueous system, which may be water or an aqueous solution. It can be an in vitro system such as a buffer solution, an in vitro simulated solution, a cell culture medium, a tissue culture medium, or an in vivo system such as blood or tissue fluid.
  • the drug units in the drug-loaded single-molecule nanopolymer may only be platinum drug units, and in this case, it can be recorded as platinum single-drug single-molecule nanopolymer.
  • the second drug unit is optional and may or may not be contained.
  • the hydrophilicity and hydrophobicity of the second drug unit are not particularly limited, and may be a hydrophilic drug unit or a hydrophobic drug unit.
  • the second drug unit used in this application is different from the platinum-based drug unit, so that it can act on different targets.
  • the drug unit in the drug-loaded single-molecule nanopolymer includes a platinum drug unit and a second drug unit. At this time, it can be recorded as a double-drug single-molecule nanopolymer .
  • Figure 1 which also shows the drug release process in response to external stimulus conditions.
  • the platinum-based drug unit is the residue of cisplatin and the second drug unit is the residue of the active ingredient of camptothecin, and the schematic diagram of drug release in response to the intracellular reducing microenvironment is shown in FIG. 2 .
  • the drug-loaded single-molecule nanopolymer constructs multiple polyamino acid chains into a nonlinear skeleton through a divalent platinum-containing linker L Pt , at least one polyamino acid chain is connected to a hydrophilic polymer chain at the end, and the end of the L Pt
  • the platinum atom participates in the formation of the platinum-based drug unit (it may be the residue of the active ingredient of the platinum-based drug or its prodrug).
  • the drug-loaded single-molecule nanopolymer can be controlled to have a branched or moderately cross-linked three-dimensional structure, and further combined with the design of the position of the hydrophilic polymer chain at the end of the polyamino acid chain, the drug-loaded single molecule Molecular nanopolymers can form single-molecule nanopolymer micelles with a core-shell structure without self-assembly in aqueous media.
  • the hydrophilic polymer chains are distributed in the outer shell, and the drug ingredients are entrapped in the inner core.
  • the drug-loaded single-molecule nanopolymer can only be loaded with platinum drug units to form a platinum single-drug single-molecule nanopolymer; or the residue of its prodrug), the second drug unit can be grafted on the side group of the polyamino acid chain, and at this time, a double-drug single-molecule nanopolymer can be formed.
  • the relative content of the platinum drug unit and the second drug unit can be flexibly adjusted by controlling the feeding amount of the corresponding monomer.
  • the distribution density of L Pt can be adjusted by adjusting the feeding ratio of unbranched amino acid monomers and L Pt branched amino acid monomers. In the unbranched amino acid monomers, the amount of amino acid monomers containing the second drug unit can also be flexibly adjusted. Proportion.
  • the drug-loaded single-molecule nanopolymer has good stability in vivo and in vitro, good dispersibility, uniform particle size, no toxic and side effects, and does not release active pharmaceutical ingredients outside the cell but exhibits triggered release of active pharmaceutical ingredients inside the cell , in addition, it can be obtained by a preparation method with simple operation, mild reaction, low cost and environmental friendliness.
  • L Pt branched amino acid monomer refers to an amino acid monomer that participates in the branch point of the aforementioned nonlinear skeleton, such as the structural compound (carrying a platinum drug unit) shown in formula (I-3) herein.
  • non-branched amino acid monomer refers to an amino acid monomer that does not participate in the branch point that constitutes the aforementioned nonlinear skeleton, for example, the structural compound shown in formula (II-3) herein (carrying the second drug unit) , the compound of the structure shown in (IV-3).
  • a carbon-centered trivalent group means a trivalent group whose branching point is provided by a carbon atom.
  • each occurrence of U may be CRC, wherein R C may be H or a non-hydrogen atom or group that does not affect the NCA ring-opened polymer.
  • each U in all are CH.
  • any one of the indicated "*" ends is independently attached to the divalent linker L Pt , or to the monovalent side group R A .
  • any one of the indicated "*" ends is independently connected to said divalent linker L Pt , or to a drug-containing side chain containing said second drug unit.
  • alkyl refers to a monovalent alkyl group
  • alkylene refers to a divalent alkyl group
  • linking group refers to an atom or group with a valence state ⁇ 2
  • divalent linking group refers to the linking group whose valence is 2
  • end group refers to the atom or group whose valence is 1.
  • monovalent alkyl refers to the residue formed by the loss of any one hydrogen atom of the alkane compound
  • alkylene refers to the residue formed by the loss of any two hydrogen atoms of the alkane compound.
  • alkane compound here refers to It is a saturated hydrocarbon composed of carbon atoms and hydrogen atoms, which can be a chain (that is, without a ring) or a saturated ring (such as hexane). If there is no special description, it can preferably be a chain.
  • each occurrence of R is independently selected from side groups of the 19 natural amino acids (except proline), ionic forms of suitable side groups of the 19 natural amino acids (except proline ) , and ornithine Any of the side groups.
  • each occurrence of R is independently selected from any of the 19 natural amino acids (except proline ) and side groups of ornithine.
  • each occurrence of C 1-6 alkyl is independently C 1 alkyl, C 2 alkyl, C 3 alkyl, C 4 alkyl, C 5 alkyl or C 6 alkyl.
  • Suitable examples include, but are not limited to: methyl (Me, -CH 3 ), ethyl (Et, -CH 2 CH 3 ), 1-propyl (n-Pr, n-propyl, -CH 2 CH 2 CH 3 ), 2-propyl (i-Pr, i-propyl, -CH(CH 3 ) 2 ), 1-butyl (n-Bu, n-butyl, -CH 2 CH 2 CH 2 CH 3 ) , 2-methyl-1-propyl (i-Bu, i-butyl, -CH 2 CH(CH 3 ) 2 ), 2-butyl (s-Bu, s-butyl, -CH(CH 3 )CH 2 CH 3 ), 2-methyl-2-propyl (t-Bu, t-Bu,
  • each occurrence of C 1-6 alkylene is independently C 1 alkylene, C 2 alkylene, C 3 alkylene, C 4 alkylene, C 5 alkylene or C 6 alkylene.
  • Each occurrence of C 1-4 alkylene is independently C 1 alkylene, C 2 alkylene, C 3 alkylene or C 4 alkylene.
  • Suitable examples include, but are not limited to: methylene (-CH 2 -), 1,1-ethyl (-CH(CH 3 )-), 1,2-ethyl (-CH 2 CH 2 -), 1 ,1-Propyl (-CH(CH 2 CH 3 )-), 1,2-Propyl (-CH 2 CH(CH 3 )-), 1,3-Propyl (-CH 2 CH 2 CH 2 - ) and 1,4-butyl (-CH 2 CH 2 CH 2 CH 2 -).
  • each occurrence of R is independently a non-polar terminal group, such as C 1-6 alkyl, -LA - phenyl, -LA - SC 1-3 alkyl, further such as - CH 3 , -CH(CH 3 ) 2 , -CH 2 CH(CH 3 ) 2 , -CH(CH 3 )CH 2 CH 3 , -CH 2 CH 2 SCH 3 ,
  • each occurrence of R 0 is independently a polar uncharged end group such as -CH 2 -OH, -CH(OH)CH 3 , -CH 2 SH, -CH 2 CONH 2 or -CH2CH2CONH2 . _
  • each occurrence of R 0 is independently a non-polar end group or a polar uncharged end group.
  • each occurrence of R 0 is independently a hydrophilic end group (eg, a polar end group) or a hydrophobic end group (eg, a non-polar end group).
  • a hydrophilic end group eg, a polar end group
  • a hydrophobic end group eg, a non-polar end group.
  • the percentage of the number of platinum atoms in the divalent linker L Pt relative to the total number of amino acid units is 10% to 100%, preferably 10% % to 90%, another preferably 10% to 80%, another preferably 10% to 60%, another preferably 10% to 50%, another preferably 10% to 40%, another preferably 10% to 30%, Another preferred 15% to 25%, another preferred 18% to 22%, another preferred 15% to 80%, another preferred 15% to 60%, another preferred 15% to 50%, another preferred 15% ⁇ 40%, and preferably 15% ⁇ 30%.
  • the branch point density of the drug-loaded single-molecule nanopolymers can be tuned.
  • the percentage of the number of platinum atoms in the divalent linker L Pt relative to the total number of amino acid units can also be selected from any of the following percentages or the interval formed by any two percentages: 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96% , 97%, 98%, 99%, 100%, etc.
  • the ratio of the number of the second drug unit to the number of the platinum-based drug unit is (0-10):1, preferably (0 ⁇ 5):1, another preferably (0 ⁇ 3):1, another preferably (0 ⁇ 1):1, another preferably (0.5 ⁇ 10):1, another preferably (0.5 ⁇ 5):1, Another preferred ratio is (0.5-3):1, another preferred ratio is (1-5):1, another preferred ratio is (1-3):1, and further preferred configuration is (2-3):1.
  • the ratio of the number of the second drug unit to the number of the platinum-based drug unit can also be selected from any of the following ratios or intervals formed by any two: (0.1:1), 0.2:1) , (0.3:1), (0.4:1), (0.5:1), (0.6:1), (0.7:1), (0.8:1), (.9:1), (1:1), (1.1:1), (1.2:1), (1.3:1), (1.4:1), (1.5:1), (1.6:1), (1.8:1), (2:1), (2.5 :1), (2.6:1), (2.8:1), (3:1), (3.5:1), (4:1), (4.5:1), (5:1), (5.5:1 ), (6:1), (6.5:1), (7:1), (7.5:1), (8:1), (8.5:1), (9:1), (9.5:1), (10:1) etc.
  • the drug-loaded monomolecular nanopolymer does not contain a second drug unit.
  • the ratio of the number of the hydrophilic polymer chains to the number of platinum-based drug units is 1:(2-100), preferably 1:(10-60), preferably 1:(15-45), and preferably 1:(15-25).
  • the ratio of the number of the hydrophilic polymer chains to the number of platinum drug units can also be selected from any of the following ratios or intervals formed by any two: (1:2), (1 :3), (1:4), (1:5), (1:6), (1:7), (1:8), (1:9), (1:10), (1:11 ), (1:12), (1:13), (1:14), (1:15), (1:16), (1:18), (1:20), (1:22 ), (1:24), (1:25), (1:26), (1:28), (1:30), (1:35), (1:40), (1:45), (1 :55), (1:60), (1:65), (1:70), (1:75), (1:80), (1:85), (1:90), (1:95 ), (1:100), etc.
  • the drug-loaded monomolecular nanopolymer includes a tetravalent structural unit shown in formula (I), a monovalent structural unit shown in formula (III), and an optional divalent structural unit shown in formula (II).
  • Formula (I) appears each time, wherein, U 1 and U 2 are each independently a carbon-centered trivalent group, and D Pt is a platinum drug unit;
  • POL i is a hydrophilic polymer chain
  • L 5 is independently a divalent linking group or nothing
  • the drug-loaded monomolecular nanopolymer includes at least one of the divalent structural unit represented by formula (II) and the divalent structural unit represented by formula (IV).
  • the branch point density of the nonlinear structure can be appropriately reduced, and the inner core of the formed micelle is relatively loose, which can appropriately accelerate the release rate of the drug.
  • the drug-loaded monomolecular nanopolymer does not include the divalent structural unit represented by formula (II). At this time, a platinum single-drug single-molecule nanopolymer is formed.
  • the drug-loaded monomolecular nanopolymer does not include the divalent structural unit represented by formula (IV). In this case, all amino acid units are linked with drug units, at least platinum drug units.
  • the drug-loaded monomolecular nanopolymer includes a divalent structural unit represented by formula (II) and a divalent structural unit represented by formula (IV).
  • the polyamino acid chain consists of a tetravalent structural unit represented by formula (I) and a divalent structural unit represented by formula (II). At this time, all amino acid units are linked with drug units, or linked with platinum drug units (forming branch points), or linked with second drug units (without forming branch points, providing free drug-containing side chains). At this time, there is no need to add the monomer represented by formula (IV-3) to prepare the raw materials.
  • the polyamino acid chain consists of a tetravalent structural unit represented by formula (I) and a divalent structural unit represented by formula (IV).
  • the polyamino acid chain consists of a tetravalent structural unit represented by formula (I), a divalent structural unit represented by formula (II), and a divalent structural unit represented by formula (IV).
  • the wavy line Indicates the point of attachment of an atom or group.
  • Each occurrence independently contains the following structures:
  • U 10 is independently a trivalent hydrocarbon group, independently preferably a trivalent alkyl group; more preferably, It is independently a lysine or ornithine unit, when it is an ornithine unit, U 10 is >CH-CH 2 CH 2 CH 2 -*, when it is a lysine unit, U 10 is >CH-CH 2 CH 2 CH 2 CH 2 -*, where "*" points to D Pt .
  • independently is a lysine unit.
  • Each occurrence independently contains the following structures:
  • U 20 is independently a trivalent hydrocarbon group, independently preferably a trivalent alkyl group; more preferably, It is independently a lysine or ornithine unit, when it is an ornithine unit, U 10 is >CH-CH 2 CH 2 CH 2 -*, when it is a lysine unit, U 10 is >CH-CH 2 CH 2 CH 2 CH 2 -*, where "*" points to D Pt .
  • independently is a lysine unit.
  • a molecule of The structures are all the same, and U1 and U2 have the same structure at this time.
  • both U1 and U2 in one molecule are the same.
  • a molecule in The structures are all the same, at this time, the structures of U 10 and U 20 are the same.
  • both U 10 and U 20 in one molecule are the same.
  • each occurrence of formula (I) independently has the structure shown in formula (I-1):
  • U 10 and U 20 are independently as defined above;
  • R 11 and R 21 are each independently a divalent linking group, may be an alkylene group, may be independently preferably an alkylene group, may also be independently preferably a C 1-6 alkylene group, and may also independently be preferably a C 1-6 alkylene can also be independently more preferably methylene, 1,2-ethylene, 1,3-propylene, 1,4-butylene, 1,5-pentylene Or 1,6-hexylene, each independently more preferably methylene, 1,2-ethylene, 1,3-propylene or 1,4-butylene, each independently preferably 1,2-ethylene, 1,3-propylene or 1,4-butylene can also be independently preferably 1,2-ethylene or 1,3-propylene, and can also be independently Preferably it is 1,2-ethylene;
  • R 01 may be -(CH 2 ) q -, wherein q may be an integer selected from 1 to 6, further may be 1, 2, 3, 4, 5 or 6, may be preferably 1, 2, 3 or 4, Further can be 2.
  • each occurrence of D Pt is independently selected from residues of any one of cisplatin, carboplatin, nedaplatin, oxaliplatin, and lobaplatin.
  • each occurrence of formula (I) has the same structure.
  • Each occurrence independently contains the following structures:
  • U 30 is independently a trivalent hydrocarbon group, independently preferably a trivalent alkyl group; more preferably, It is independently a lysine or ornithine unit, when it is an ornithine unit, U 30 is >CH-CH 2 CH 2 CH 2 -*, when it is a lysine unit, U 30 is >CH-CH 2 CH 2 CH 2 CH 2 -*, where "*" points to D T .
  • independently is a lysine unit.
  • U3 in a molecule are all the same.
  • all U30s in a molecule are the same.
  • each occurrence of LR independently comprises a linker capable of cleavage under at least one of the following conditions: intracellular reducing conditions, reactive oxygen conditions, pH conditions, enzymatic solution and hydrolysis conditions.
  • the pH condition satisfies that the pH value is less than 6.8, more preferably the pH is 4.0-6.8.
  • the enzymatic hydrolysis conditions are selected from one or more of the following enzymes: MMP-2 enzyme and azoreductase.
  • the hydrolysis conditions are acidic hydrolysis conditions or basic hydrolysis conditions.
  • each occurrence of LR independently comprises one or more linkers in the following (a) group, (b) group, (c) group, (d) group and (e) group;
  • the groups in group (a) can respond to intracellular reducing conditions (such as glutathione environment), active oxygen conditions and other conditions.
  • Groups of group (b) may respond to reactive oxygen species (ROS) conditions and may belong to ROS responsive groups.
  • ROS reactive oxygen species
  • Group (c) groups may correspond to specific acidic pH conditions.
  • the groups in group (d) can be cleaved under the action of enzymes.
  • GPLGVRG peptide can be enzymatically hydrolyzed by MMP-2 enzyme.
  • Azo groups can be enzymatically cleaved under azoreductase conditions.
  • Group (e) can undergo hydrolysis.
  • groups in groups (a), (b), (c), (d) and (e) may be responsive to one or more stimulus conditions.
  • R 32 it is independently a divalent linking group, which may be an alkylene group , can be independently preferably an alkylene group, can also be independently preferably a C 1-6 alkylene group, can also independently be preferably a C 1-6 alkylene group, and can also independently be more preferably a methylene group, 1,2-ethylene, 1,3-propylene, 1,4-butylene, 1,5-pentylene or 1,6-hexylene can also be independently more preferably methylene, 1 , 2-ethylene, 1,3-propylene or 1,4-butylene, each independently preferably 1,2-ethylene, 1,3-propylene or 1,4-butylene group, each independently may be preferably 1,2-ethylene or 1,3-propylene, and each independently may be preferably 1,2-ethylene.
  • R 32 it is independently a divalent linking group, which may be an alkylene group , can be independently preferably an alkylene group, can also be independently preferably a C 1-6 alkylene group, can also independently be preferably a C
  • the link between LR and DT can be cleavable, so that the active pharmaceutical ingredient corresponding to DT or its prodrug can be released.
  • each occurrence of formula (II) has a structure shown in formula (II-1):
  • each occurrence of U 30 is independently defined as before;
  • R 32 and Z 4 are independently as defined above;
  • R 31 is independently a divalent linking group, may be an alkylene group, may be independently preferably an alkylene group, may also independently be preferably a C 1-6 alkylene group, and may also independently be preferably a C 1-6 alkylene groups, which can also be independently more preferably methylene, 1,2-ethylene, 1,3-propylene, 1,4-butylene, 1,5-pentylene or 1 , 6-hexylene, can also be independently more preferably methylene, 1,2-ethylene, 1,3-propylene or 1,4-butylene, can also be independently preferably 1,2 -Ethylene, 1,3-propylene or 1,4-butylene, each independently preferably 1,2-ethylene or 1,3-propylene, each independently preferably 1,2-Ethylene.
  • each occurrence of R 31 -L R -R 32 is independently -(CH 2 ) p1 -SS-(CH 2 ) p2 -, wherein p1 and p2 are each independently selected from 1 to
  • the integer of 6 may be 1, 2, 3, 4, 5 or 6 each independently, preferably 1, 2, 3 or 4 each independently, and may be 2 or 3 further independently.
  • each occurrence of R 31 -LR -R 32 is -(CH 2 ) 2 -SS-(CH 2 ) 2 -.
  • each occurrence of DT is independently selected from residues of any one of camptothecins, resiquimod, and paclitaxel.
  • the active pharmaceutical ingredient corresponding to DT or its prodrug should have a reactive group FT , or can be activated into a reactive group FT so that it can be modified with LR .
  • the reactive group FT may be one or more of functional groups such as hydroxyl, carboxyl, amino, and mercapto. In some embodiments, the reactive group FT is a free hydroxyl group, a free amino group, or a combination thereof.
  • a suitable Z4 linker can be selected according to the structural characteristics of the active pharmaceutical ingredient corresponding to DT or its prodrug.
  • the camptothecin-like compound includes camptothecin and derivatives or analogs thereof.
  • the camptothecins include irinotecan, topotecan, rubitecan, gemitecan, 9-aminocamptothecin, 9-nitrocamptothecin, and 7-ethyl -10-Hydroxycamptothecin.
  • each occurrence of formula (II) has the same structure.
  • each occurrence of L 5 is independently a divalent linking group, may be an alkylene group, may be independently preferably an alkylene group, and may also independently be preferably a C 1
  • the -6 alkylene group can also be independently preferably a C 1-6 alkylene group, and can also independently be more preferably a methylene group, 1,2-ethylene group, 1,3-propylene group, 1,4- Butylene, 1,5-pentylene or 1,6-hexylene can also be independently more preferably methylene, 1,2-ethylene, 1,3-propylene or 1,4-butylene
  • each occurrence of formula (III) has the same L5 and Z5 .
  • each occurrence of POL i independently comprises a hydrophilic polymer chain of any of the following: polyethylene glycol, poly(propylene glycol), copolymers of ethylene glycol and propylene glycol, poly (ethoxylated polyol), poly(enol), poly(vinylpyrrolidone), poly(hydroxyalkylmethacrylamide), poly(hydroxyalkylmethacrylate), poly(sugar), poly (alpha-hydroxy acid), poly(vinyl alcohol), polyphosphazene, polyoxazoline, poly(N-acryloylmorpholine), and any combination of the foregoing polymer chains.
  • the molecular weight of the hydrophilic polymer chain is selected from 50Da to 100kDa, preferably 100Da to 80kDa, preferably 500Da to 50kDa, preferably 500Da to 10kDa, and preferably 500Da-8000Da, preferably 500Da-6000Da, preferably 500Da-5000Da, preferably 1000Da-50kDa, preferably 1000Da-10kDa, preferably 1000Da-8000Da, preferably 1000Da-6000Da, and preferably 1000Da ⁇ 5000Da.
  • the molecular weight of the hydrophilic polymer chain can also be selected from any one or two of the following intervals: about 500Da, about 600Da, about 700Da, about 750Da, about 800Da, about 850Da, about 900Da, about 950Da, about 1000 Da, about 1100 Da, about 1200 Da, about 1300 Da, about 1400 Da, about 1500 Da, about 1600 Da, about 1800 Da, about 2000 Da, about 2200 Da, about 2400 Da, about 2500 Da, about 3000 Da, about 3500 Da, about 4000 Da, about 4500 Da, about 5000 Da, About 5500Da, about 6000Da, about 6500Da, about 7000Da, about 7500Da, about 8000Da, about 8500Da, about 9000Da, about 10000Da, etc., where "about” can mean ⁇ 10%, ⁇ 5%, ⁇ 2% or 0.
  • each occurrence of POL i independently comprises a polyethylene glycol segment; another preferably, the polyethylene glycol segment is mPEG, and another preferably, the poly The molecular weight of the ethylene glycol segment is selected from 50Da-100kDa, preferably 100Da-80kDa, preferably 500Da-50kDa, preferably 500Da-10kDa, preferably 500Da-8000Da, preferably 500Da-6000Da, and preferably 500Da-5000Da, preferably 1000Da-50kDa, preferably 1000Da-10kDa, preferably 1000Da-8000Da, preferably 1000Da-6000Da, preferably 2000Da-6000Da, preferably 4000Da-6000Da, and preferably 1000Da to 5000Da, preferably about 500Da, about 600Da, about 800Da, about 1000Da, about 1100Da, about 1200Da, about 1500Da, about 1600Da, about
  • the "molecular weight" of any POL i can independently represent a weight average molecular weight or a number average molecular weight.
  • the "molecular weight" of any one of POL i can independently represent a weight average molecular weight.
  • the "molecular weight" of any one of POL i can independently represent a number average molecular weight.
  • each occurrence of formula (III) has the same structure.
  • each occurrence of formula (IV) has a structure shown in formula (IV-1):
  • each occurrence of R E is independently a hydrogen atom or R 0 , wherein R 0 is an end group not containing a drug unit.
  • each occurrence of RE is independently R 0 .
  • R is as defined in any of the preceding embodiments.
  • the polyamino acid chain comprises amino acid units of the structure represented by formula (IV-1).
  • each occurrence of formula (IV) has the same structure.
  • each occurrence of formula (I) has the same structure; each occurrence of formula (III) has the same L 5 and Z 5 ; if any, formula (II) Each occurrence has the same structure; if any, the formula (IV) has the same structure each occurrence.
  • the drug-loaded single-molecule nanopolymer includes a tetravalent structural unit shown in formula (I-2), and a monovalent structure unit shown in formula (III-2) unit, an optional divalent structural unit shown in formula (II-2) and an optional divalent structural unit shown in formula (IV-1);
  • n11 and n21 are each independently 3 or 4, and n12 and n22 are each independently 1, 2, 3, 4 or 5;
  • n31 is independently 3 or 4
  • n32 is independently 2, 3 or 4
  • n33 is independently 2, 3 or 4;
  • n51 is independently 1, 2, 3 or 4;
  • p is independently a positive integer, preferably a positive integer less than or equal to 2500, another preferably a positive integer less than or equal to 2000, another preferably a positive integer less than or equal to 1500, another preferably a positive integer less than or equal to 1000, another preferably less than or equal to A positive integer equal to 800, another preferably a positive integer less than or equal to 600, another preferably a positive integer less than or equal to 500, another preferably a positive integer less than or equal to 400, another preferably a positive integer less than or equal to 300, another preferably a positive integer less than or equal to A positive integer equal to 250, preferably a positive integer less than or equal to 200, another preferably an integer selected from 2 to 2500, another preferably an integer selected from 3 to 2000, another preferably an integer selected from 5 to 1500, and another Preferably an integer selected from 5 to 1000, another preferably an integer selected from 5 to 800, another preferably an integer selected from 5 to 600, another preferably an integer selected from 5 to 500, and another preferably an integer
  • p is preferably a positive integer less than or equal to 500, another preferably a positive integer less than or equal to 400, another preferably a positive integer less than or equal to 300, another preferably a positive integer less than or equal to 250, another preferably less than or equal to A positive integer equal to 200, preferably an integer selected from 5 to 500, another preferably an integer selected from 5 to 400, another preferably an integer selected from 5 to 300, another preferably an integer selected from 5 to 250, and another It is preferably an integer selected from 5 to 200, another preferably an integer selected from 10 to 500, another preferably an integer selected from 10 to 400, another preferably an integer selected from 10 to 300, and another preferably selected from 10 to 500.
  • p can also be selected from any one of the following integers or the interval formed by any two integers: 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 45, 50, 55, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 215, 220, 225, 227, 240, 250, 260, 280, 300, 350, 400, 450, 500, etc.
  • p can also be an integer selected from any of the following ranges: 110-120, 100-120, 100-130, 100-140, 100-150, 90-120, 90-130, 90-140 , 90 ⁇ 150, 80 ⁇ 120, 80 ⁇ 130, 80 ⁇ 140, 80 ⁇ 150, etc.
  • p can also be an integer selected from any of the following ranges: 5-115, 5-114, 5-110, 5-100, 5-90, 5-88, 5-78, 5-78 , 5 ⁇ 777, 5 ⁇ 66, 5 ⁇ 65, 5 ⁇ 60, 5 ⁇ 55, 5 ⁇ 50, 5 ⁇ 45, 5 ⁇ 44, 5 ⁇ 40, 5 ⁇ 35, 5 ⁇ 34, 5 ⁇ 33, 5 ⁇ 30, 5 ⁇ 25, 5 ⁇ 20, 6 ⁇ 115, 6 ⁇ 114, 6 ⁇ 110, 6 ⁇ 100, 6 ⁇ 90, 6 ⁇ 88, 6 ⁇ 78, 6 ⁇ 78, 6 ⁇ 777, 6 ⁇ 66 , 6 ⁇ 65, 6 ⁇ 60, 6 ⁇ 55, 6 ⁇ 50, 6 ⁇ 45, 6 ⁇ 44, 6 ⁇ 40, 6 ⁇ 35, 6 ⁇ 34, 6 ⁇ 33, 6 ⁇ 30, 6 ⁇ 25, 6 ⁇ 20, 8 ⁇ 115, 8 ⁇ 114, 8 ⁇ 110, 8 ⁇ 100, 8 ⁇ 90, 8 ⁇ 88, 8 ⁇ 78, 8 ⁇ 78, 8 ⁇ 777, 8 ⁇ 66, 8 ⁇ 65, 8 ⁇ 60 , 8 ⁇ 55, 8
  • the divalent structural unit represented by formula (IV-1) is as defined above.
  • n11 and n21 are each independently 3 or 4, further, n11 and n21 are each independently 4.
  • n12 and n22 are each independently 2, 3, 4 or 5, may also be independently 2 or 3, may also be independently 2, and may also be independently 3.
  • n31 is independently 3 or 4, further independently may be 4.
  • n32 is independently 2, 3 or 4, further independently may be 2.
  • n33 is independently 2, 3 or 4, further independently may be 2.
  • n51 is independently 1, 2, 3 or 4, further independently 2, 3 or 4, further independently 2 or 3, also independently 2, and independently Land is 3.
  • n11 and n21 are each independently 4, and n12 and n22 are each independently 4.
  • n31 is independently 4, n32 is independently 2, and n33 is independently 2.
  • n51 is 2, 3 or 4 independently, and further can be 3 independently.
  • LR is -SS-.
  • Z 5 is -NH-.
  • LR is -SS- and Z is -NH-.
  • D Pt is cisplatin, oxaliplatin, or , DT is the residue of camptothecin, paclitaxel or resiquimod.
  • the molecular weight of drug-loaded single-molecule nanopolymers may be greater than 50kDa, further can be greater than 100kDa, further can be selected from 100kDa to 5000kDa, further can be selected from 150kDa to 5000kDa, further can be selected from 200kDa to 5000kDa, further can be selected from 250kDa to 5000kDa, and further can be selected from 300kDa to 5000kDa 5000kDa, further can be selected from 400kDa ⁇ 5000kDa, preferably 500kDa ⁇ 5000kDa, another preferably 500kDa ⁇ 4000kDa, another preferably 500kDa ⁇ 3000kDa, another preferably 500kDa ⁇ 2500kD
  • the molecular weight of the drug-loaded single-molecule nanopolymer can also be selected from any of the following molecular weights or the interval formed by any two molecular weights: 100kDa, 150kDa, 200kDa, 250kDa, 300kDa, 400kDa, 500kDa, 550kDa, 600kDa, 650kDa, 700kDa, 750kDa ⁇ 800kDa ⁇ 850kDa ⁇ 900kDa ⁇ 950kDa ⁇ 1000kDa ⁇ 1100kDa ⁇ 1200kDa ⁇ 1300kDa ⁇ 1400kDa ⁇ 1500kDa ⁇ 1600kDa ⁇ 1700kDa ⁇ 1800kDa ⁇ 1900kDa ⁇ 2000kDa ⁇ 2100kDa ⁇ 2200kDa ⁇ 2300kDa ⁇ 2400kDa ⁇ 2500kDa ⁇ 3000kDa ⁇ 3500 kDa ⁇ 4000kDa ⁇ 4500kDa ⁇ 5000kDa etc.
  • the number of platinum atoms in one molecule is greater than 40, further greater than 50, and may also be selected from 50 to 5000, further may be selected from 50 to 4000, and may be further selected from 50 to 5000.
  • 2000 further can be selected from 50-1500, further can be selected from 50-1000, further can be selected from 50-500, can also be selected from 60-2000, can also be selected from 60-1500, can also be selected from 60-1000, Can also be selected from 60-500, can also be selected from 80-2000, can also be selected from 80-1500, can also be selected from 80-1000, can also be selected from 80-500, can also be selected from 100-2000, can also be selected from Selected from 100-1500, can also be selected from 100-1000, can also be selected from 100-500, can also be selected from 150-2000, can also be selected from 150-1500, can also be selected from 150-1000, can also be selected from 150-500, can also be selected from 200-2000, can also be selected from 200-1500, can also be selected from 200-1000, can also be selected from 200-500, can also be selected from 250-2000, can also be selected from 250- 1500, can also be selected from 250-1000, can also be selected from 250-500, can also be selected from 300-
  • the number of platinum atoms in a molecule can also be selected from any one of the following values or the interval formed by any two values: 50, 60, 80, 100, 120, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, 800, 900, 1000, 1100, 1200, 1300, 1400, 1500, 1600, 1800, 2000, 2500, 3000, etc.
  • Controlling the molecular weight of the drug-loaded single-molecule nanopolymer in an appropriate range can control the size of the micelles formed by it to be suitable for pharmaceutical preparations.
  • the size of micelles in water, aqueous solutions or in vivo environments can be influenced.
  • the average diameter is about 25-45nm in 25°C water, and in some embodiments, the average diameter is about 30nm, 32nm, 34nm, 44nm, etc.
  • the "particle size of drug-loaded single-molecule nanopolymer micelles” refers to the average diameter or average particle size only when specified. Unless otherwise specified, the test temperature is 20-30°C, further 25°C.
  • the present application also provides a preparation method of a drug-loaded monomolecular nanopolymer, which includes the following steps: a platinum-containing compound having a structure shown in formula (I-3), a monomolecular compound having a structure shown in formula (III-3), The functionalized hydrophilic polymer, the optional drug compound with the structure shown in formula (II-3) and the optional compound shown in formula (IV-3) are mixed in an organic solvent to perform ring-opening polymerization;
  • PE is RE or protected RE , which is inert in the ring-opening polymerization reaction, that is, has no reactivity in the ring-opening polymerization reaction;
  • F 5 is -NH 2 , -COOH, Preferred is -NH 2 .
  • mPEG corresponds to CH 3 (OCH 2 CH 2 ) p -O-, and the definition of p is consistent with the above.
  • the ring-opening polymerization is performed under anhydrous conditions.
  • the reaction temperature of the ring-opening polymerization is 15-40° C., more preferably, the reaction time of the ring-opening polymerization is 24-96 hours.
  • the platinum-containing compound shown in formula (I-3) (can be recorded as NCA-Pt-NCA, a double NCA monomer), the drug compound (NCA-L R -D T , a single NCA monomer) and the compound represented by formula (IV-3) (which can be recorded as NCA-AA, a single NCA monomer) are both NCA functionalized amino acid monomers.
  • N-carboxylic acid anhydride functional group is denoted as NCA.
  • the polymerization utilizes ring-opening polymerization involving bis-N-carboxylic acid anhydride (NCA) to obtain single-molecule nanopolymers through a "one-pot method", which can form cores without self-assembly in aqueous media.
  • NCA bis-N-carboxylic acid anhydride
  • the micelles with a shell structure provide a drug delivery system that can release active pharmaceutical ingredients in response to the treatment of tumor diseases.
  • the monomer shown in formula (I-3) is a kind of branched amino acid monomer of the present application, and the platinum atom is used as a bridge to connect two NCA functional groups.
  • the monomer can form a nonlinear skeleton through ring-opening polymerization, providing non-linear Branching points in linear backbones.
  • Both the monomers represented by formula (II-3) and the monomers represented by formula (IV-3) are non-branched amino acid monomers in the present application.
  • One end of the monomer represented by formula (II-3) is an NCA functional group, and the other end carries the second drug unit DT .
  • This monomer can participate in the formation of polyamino acid chains through ring-opening polymerization, but does not provide branch points in the nonlinear backbone .
  • the monomer shown in formula (IV-3) is NCA functionalized amino acid, contains NCA functional group, and does not contain other reactive groups (referring to participate in the reactivity of ring-opening polymerization reaction), in ring-opening polymerization reaction, only NCA participates in the reaction, and this monomer participates in the formation of polyamino acid chains, but does not provide branching points in the nonlinear backbone.
  • the monomer shown in formula (III-3) can play the role of end-capping agent, and the more the amount is, the easier it is to obtain shorter polyamino acid chains and smaller drug-loaded single-molecule nanopolymers.
  • the amount of the indicated monomers can adjust the molecular size of the drug-loaded single-molecule nanopolymer, and at the same time control the drug-loading amount of each single-molecule nanopolymer.
  • the mole percentage of the monomer represented by formula (I-3) in all amino acid monomers can be 15% to 100%, preferably 15% to 90% %, another preferably 15% to 80%, another preferred 15% to 60%, another preferred 15% to 50%, another preferred 15% to 40%, another preferred 15% to 30%, another preferred 20% to 80%, preferably 20% to 60%, preferably 20% to 50%, preferably 20% to 40%, and preferably 20% to 30%.
  • the mole percentage of the monomer represented by formula (I-3) in all amino acid monomers can also be selected from any of the following percentages or the interval formed by any two percentages: 15%, 16%, 17%, 18%, 19% %, 20%, 22%, 24%, 25%, 26%, 28%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, 100%, etc.
  • the molar ratio of the monomer shown in formula (II-3) relative to the monomer shown in formula (I-3) can refer to the number of the second drug unit and the number of platinum drug units Quantity ratio.
  • the molar ratio of the monomer shown in formula (II-3) relative to the monomer shown in formula (I-3) can be (0 ⁇ 10): 1, preferably (0 ⁇ 5): 1, another preferably (0 ⁇ 3):1, another preferably (0 ⁇ 1):1, another preferably (0.5 ⁇ 10):1, another preferably (0.5 ⁇ 5):1, another preferably (0.5 ⁇ 3):1, Another preferred ratio is (1-5):1, another preferred ratio is (1-3):1, and further preferred ratio is (2-3):1.
  • the molar ratio of the monomer shown in formula (II-3) relative to the monomer shown in formula (I-3) can be selected from any of the following ratios or the interval formed by any two: (0.1:1), 0.2:1 ), (0.3:1), (0.4:1), (0.5:1), (0.6:1), (0.7:1), (0.8:1), (.9:1), (1:1) , (1.1:1), (1.2:1), (1.3:1), (1.4:1), (1.5:1), (1.6:1), (1.8:1), (2:1), ( 2.5:1), (2.6:1), (2.8:1), (3:1), (3.5:1), (4:1), (4.5:1), (5:1), (5.5: 1), (6:1), (6.5:1), (7:1), (7.5:1), (8:1), (8.5:1), (9:1), (9.5:1) , (10:1), etc.
  • the molar ratio of the monomer shown in formula (IV-3) relative to the monomer shown in formula (I-3) can be numerically compared with the amount of the hydrophilic polymer chain and the platinum drug The ratio of the number of units.
  • the molar ratio of the monomer shown in formula (IV-3) relative to the monomer shown in formula (I-3) can be 1:(2 ⁇ 100), preferably 1:(10 ⁇ 60), and another preferably 1: (15-45), and preferably 1:(15-25).
  • the molar ratio of the monomer shown in formula (IV-3) relative to the monomer shown in formula (I-3) can also be selected from any of the following ratios or the interval formed by any two: (1:2), (1 :3), (1:4), (1:5), (1:6), (1:7), (1:8), (1:9), (1:10), (1:11 ), (1:12), (1:13), (1:14), (1:15), (1:16), (1:18), (1:20), (1:22 ), (1:24), (1:25), (1:26), (1:28), (1:30), (1:35), (1:40), (1:45), (1 :55), (1:60), (1:65), (1:70), (1:75), (1:80), (1:85), (1:90), (1:95 ), (1:100), etc.
  • the double-drug single-molecule nanopolymer prodrug according to the present application which comprises a polyamino acid linked to a hydrophilic polymer, wherein the prodrug of a platinum-based drug active ingredient is bonded to the ⁇ -carbon of the repeating unit of the polyamino acid Moieties and prodrug moieties of the active ingredients of antineoplastic drugs; preferably, the molecules of the active ingredients of antineoplastic drugs contain free hydroxyl groups, free amino groups or a combination of both.
  • the double-drug single-molecule nanopolymer prodrug comprises a random copolyamino acid backbone linked to a hydrophilic polymer.
  • the hydrophilic polymer is selected from poly(alkylene glycol) (e.g., polyethylene glycol (“PEG”), poly(propylene glycol) ( "PPG"), copolymers of ethylene glycol and propylene glycol, etc.), poly(ethoxylated polyols), poly(enols), poly(vinylpyrrolidone), poly(hydroxyalkylmethacrylamides), poly (hydroxyalkyl methacrylate), poly(sugar), poly(alpha-hydroxy acid), poly(vinyl alcohol), polyphosphazene, polyoxazoline (“POZ”), poly(N-acryl phylloline) and any combination of these substances.
  • PEG polyethylene glycol
  • PPG poly(propylene glycol)
  • POZ polyoxazoline
  • the hydrophilic polymer is selected from polyethylene glycol ("PEG"), preferably from polyethylene glycol terminated with a methoxy group at one end.
  • PEG polyethylene glycol
  • the molecular weight of the hydrophilic polymer is not particularly limited, such as PEG50-5000, PEG50-6000, PEG50-7000, PEG50-8000, PEG500-2000, PEG500-4000, PEG500-6000, PEG500-8000, PEG500-10000, PEG500 -20000, PEG1000-20000, PEG1000-50000 or PEG1000-80000 can be used in this application, and the unit is Da.
  • PEG500-4000 it means that the molecular weight is 500-4000Da.
  • the platinum drugs include cisplatin, carboplatin, nedaplatin, oxaliplatin and lobaplatin.
  • Their chemical structures, preparation methods and pharmacological actions are all known in the art.
  • cisplatin is a cell cycle non-specific anticancer drug with the following structure:
  • the active ingredient of the platinum drug is cisplatin.
  • the active ingredients of antitumor drugs containing free hydroxyl groups, free amino groups or a combination of the two in the molecule are selected from: Camptophylla Bases, including camptothecin and its derivatives or analogs, such as irinotecan, topotecan, rubitecan, gemitecan, 9-aminocamptothecin and 9-nitrocamptothecin, 7 -Ethyl-10-hydroxycamptothecin (SN38), etc.; tumor immune activators resiquimod (Resiquimod, R-848), tiramod, etc.; paclitaxel (PTX), epirubicin, Cetaxel, docetaxel, pemetrexed, auristatin methyl E, gemcitabine, dexamethasone, etc.; and protein kinase inhibitors sorafenib, dasatini
  • camptothecin belongs to the class of DNA topoisomerase I inhibitors and has the following structure:
  • Resiquimod (R-848) is an immune response regulator with the activity of promoting tumor immunity. Its structural formula is as follows:
  • Paclitaxel is an alkaloid extracted from Taxus genus, which belongs to cell cycle specific antineoplastic drugs. It promotes tubulin polymerization, inhibits depolymerization, maintains tubulin stability, and inhibits cell mitosis.
  • the chemical name of paclitaxel is 5 ⁇ ,20-epoxy-1,2 ⁇ ,4,7 ⁇ ,10 ⁇ ,13 ⁇ -hexahydroxytaxane-11-en-9-one-4,10-diacetate-2-benzene Formate-13[(2'R,3'S)-N-benzoyl-3-phenylisoserine ester], the structural formula is as follows:
  • the active ingredient of the antitumor drug containing free hydroxyl group, free amino group or a combination of the two in the molecule is camptothecin or Camptothecin.
  • the double-drug single-molecule nanopolymer prodrug has the following structure:
  • p can be selected from 1-500, m can be selected from 0-100, n can be selected from 1-100, and k can be selected from 1-1000, but they are not limited to this range.
  • p corresponds to the degree of polymerization of polyethylene glycol units, and the definition of p can also refer to other parts of this paper.
  • the number m of the second drug unit in one molecule may be selected from 0-5000, further may be selected from 0-4000, further may be selected from 0-2000, further may be selected from 0-1500, and further may be selected from selected from 0 to 1000, further selected from 0 to 500, further selected from 10 to 2000, selected from 10 to 1500, selected from 10 to 1000, selected from 10 to 500, and selected from 20-2000, can also be selected from 20-1500, can also be selected from 20-1000, can also be selected from 20-500, can also be selected from 40-2000, can also be selected from 40-1500, can also be selected from 40- 1000, can also be selected from 40-500, can also be selected from 50-2000, can also be selected from 50-1500, can also be selected from 50-1000, can also be selected from 50-500, can also be selected from 80-2000, Can also be selected from 80-1500, can also be selected from 80-1000, can also be selected from 80-500, can also be selected from 100-2000, can also be
  • the number m of the second drug unit in one molecule can also be selected from any of the following values or the interval formed by any two: 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 18, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, 100, 120, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, 800, 900, 1000, 1100, 1200, 1300, 1400, 1500, 1600, 1800, 2000, 2500, 3000, etc.
  • the amino acid unit in the general formula is lysine, and the lysine unit in the general formula forms a nonlinear skeleton through a Pt-containing linker, which can contain multiple polylysine chains, corresponding to k
  • the amino acid unit and the amino acid unit corresponding to n can be located on different polylysine chains, and different polylysine chains can be connected to some lysine units corresponding to m, and the C of different polylysine chains
  • the ends can each be capped with a polyethylene glycol segment.
  • Random in the general formula means that the amino acid units are randomly aggregated.
  • the double-drug single-molecule nanopolymer prodrug has the following structure:
  • m can be selected from 0-100
  • n can be selected from 1-100
  • k can be selected from 1-1000, but they are not limited to this range.
  • the drug delivery system of the present application comprises a double-drug single-molecule nanopolymer micelle
  • the polymer micelle comprises a polyamino acid linked to a hydrophilic polymer, wherein the ⁇ -carbon of the repeating unit of the polyamino acid is bonded
  • the prodrug part of the active ingredient of the platinum drug and the prodrug part of the active ingredient of the antineoplastic drug are combined; preferably, the molecule of the active ingredient of the antineoplastic drug contains a free hydroxyl group, a free amino group or a combination of the two.
  • the dual-drug single-molecule nanopolymer prodrug comprises a random copolyamino acid backbone linked to a hydrophilic polymer.
  • the hydrophilic polymer is selected from poly(alkylene glycol) (e.g., polyethylene glycol (“PEG”), poly(propylene glycol) (“PPG”), ethylene glycol (“PPG”), Copolymers of diol and propylene glycol, etc.), poly(ethoxylated polyol), poly(enol), poly(vinylpyrrolidone), poly(hydroxyalkylmethacrylamide), poly(hydroxyalkylmethyl acrylate), poly(sugar), poly(alpha-hydroxy acid), poly(vinyl alcohol), polyphosphazene, polyoxazoline (“POZ”), poly(N-acryloylmorpholine), poly-2- Methacryloyloxyethyl phosphorylcholine (PMPC) and any combination of these substances.
  • PEG polyethylene glycol
  • PPG poly(propylene glycol)
  • PPG ethylene glycol
  • Copolymers of diol and propylene glycol, etc. poly(ethoxy
  • said hydrophilic polymer is selected from polyethylene glycol (“PEG”), preferably methoxy-terminated polyethylene glycol.
  • the platinum drug is selected from cisplatin, carboplatin, nedaplatin, oxaliplatin and lobaplatin.
  • the active ingredient of the platinum drug is cisplatin.
  • the active ingredient of the antitumor drug containing free hydroxyl group, free amino group or a combination of the two in the molecule is a camptothecin compound, including camptothecin and its derivatives or analogs, For example, irinotecan, topotecan, rubitecan, gemitecan, 9-aminocamptothecin, 9-nitrocamptothecin, 7-ethyl-10-hydroxycamptothecin (SN38), etc.; Quinimod (Resiquimod, R-848) and paclitaxel (Paclitaxel, PTX).
  • camptothecin compound including camptothecin and its derivatives or analogs,
  • camptothecin and its derivatives or analogs for example, irinotecan, topotecan, rubitecan, gemitecan, 9-aminocamptothecin, 9-nitrocamptothecin, 7-ethyl-10-hydroxycamptothecin (SN38), etc
  • the active ingredient of the antitumor drug containing free hydroxyl group, free amino group or a combination of both in the molecule is camptothecin.
  • the double-drug single-molecule nanopolymer prodrug has the structure shown above as P100 or P200.
  • the double-drug single-molecule nanopolymer prodrug has the following structure:
  • p is selected from 1-500
  • m can be selected from 0-100
  • n can be selected from 1-100
  • k can be selected from 1-1000, but they are not limited to this range.
  • the double-drug single-molecule nanopolymer prodrug has the structure shown in the aforementioned formula P101 or P201.
  • the double-drug single-molecule nanopolymer prodrug has the following structure:
  • m can be selected from 0-100
  • n can be selected from 1-100
  • k can be selected from 1-1000, but they are not limited to this range.
  • the double-drug single-molecule nanopolymer prodrug provided by the application can be prepared by a method comprising the following steps:
  • step (3) Under suitable reaction conditions, the monomer obtained in step (1) and step (2) is reacted with a hydrophilic polymer having terminal amino groups to obtain a double-drug single-molecule nanopolymer prodrug; and
  • the single NCA monomer of the antitumor drug active ingredient containing free hydroxyl or free amino in the molecular structure is synthesized by the method shown below:
  • Boc-Lyc-OtBu can be prepared by the method of following formula:
  • the single NCA monomer of the antitumor drug active ingredient containing free hydroxyl or free amino in the molecular structure is synthesized by the method shown below:
  • the double NCA monomer of the platinum-based drug active ingredient is synthesized by the method shown below:
  • the double-drug single-molecule nanopolymer prodrug of the present application is synthesized by the method (one-pot method) as follows:
  • p is selected from 1-500
  • m can be selected from 0-100
  • n can be selected from 1-100
  • k can be selected from 1-1000, but they are not limited to this range.
  • the double-drug single-molecule nanopolymer prodrug of the present application is synthesized by a method comprising the following steps:
  • the dried hydrophilic polymer is dissolved in an anhydrous organic solvent such as DMF, and the mono-NCA monomer and the platinum-based active ingredient of an antineoplastic drug containing free hydroxyl or free amino in the molecular structure are dissolved.
  • the bis-NCA monomer of the active ingredient of the drug is dissolved in the same organic solvent, and the resulting solution is slowly added to the reaction system drop by drop, the reaction tube is sealed, taken out from the glove box, and continuously stirred and reacted in the oil bath for a sufficient time;
  • reaction product was slowly dropped into glacial ether to obtain a white precipitate, and the supernatant was discarded to obtain a purified product;
  • the obtained product was vacuum-dried, and the dried solid was dissolved in a suitable solvent (for example, DMSO), placed in a dialysis bag (MWCO: 100kDa), dialyzed in ultrapure water for several days (changing the water several times during the period), and frozen After drying, the final product, the nanopolymer micelles, was collected.
  • a suitable solvent for example, DMSO
  • the double-drug single-molecule nanopolymer prodrug of the present application is synthesized by a method comprising the following steps:
  • the polyethylene glycol after drying is dissolved in DMF, and the mono-NCA monomer of the anti-tumor drug active ingredient containing free hydroxyl or free amino in the molecular structure and the double NCA single NCA monomer of the platinum-based drug active ingredient are mixed.
  • the solution was dissolved in the same organic solvent, and the resulting solution was slowly added to the reaction system drop by drop, the reaction tube was sealed, taken out from the glove box, and kept stirring in the oil bath for a sufficient time;
  • reaction product was slowly dropped into glacial ether to obtain a white precipitate, and the supernatant was discarded to obtain a purified product;
  • the obtained product was vacuum-dried, and the dried solid was dissolved in DMSO, placed in a dialysis bag (MWCO: 100kDa), dialyzed in ultrapure water for two days (water was changed 5 times), and after freeze-drying, the final product was collected. That is, nanopolymer micelles.
  • a drug-loaded single-molecule nanopolymer micelle the composition of which is selected from any one of the following: the aforementioned drug-loaded single-molecule nanopolymer, the drug-loaded single-molecule prepared by the aforementioned preparation method Nanopolymer, the aforementioned double-drug single-molecule nanopolymer prodrug, and the double-drug single-molecule nanopolymer prodrug prepared by the aforementioned preparation method; the drug-loaded single-molecule nanopolymer micelle has a core-shell structure, The outer shell structure is a hydrophilic layer formed by hydrophilic polymer chains, and the contained drug units are located in the inner core.
  • the drug-loaded single-molecule nanopolymer can be controlled to have a branched or moderately cross-linked three-dimensional structure, and further combined with the design of the position of the hydrophilic polymer chain at the end of the polyamino acid chain, the drug-loaded single molecule Molecular nanopolymers can form single-molecule nanopolymer micelles with a core-shell structure without self-assembly in aqueous media.
  • the hydrophilic polymer chains are distributed in the outer shell, and the drug ingredients are entrapped in the inner core.
  • a drug delivery system which comprises a drug-loaded single-molecule nanopolymer micelle, and the drug-loaded single-molecule nanopolymer micelle comprises the aforementioned drug-loaded single-molecule nanopolymer or the aforementioned preparation
  • the drug-loaded single-molecule nanopolymer prepared by the method preferably, the hydrophilic polymer chain is located in the shell of the drug-loaded single-molecule nanopolymer micelle; the platinum drug unit and the second drug The units are all located in the inner core of the drug-loaded single-molecule nanopolymer micelles.
  • the size of the drug-loaded single-molecule nanopolymer micelles is involved.
  • the test temperature is 20-30°C, further 25°C.
  • the size and morphology of the drug-loaded monomolecular nanopolymer micelles can be characterized by the test methods in the examples below.
  • the particle size of the drug-loaded single-molecule nanopolymer micelles refers to the average diameter or average particle size only when specified.
  • test condition "in water” can be pure water or aqueous solution.
  • aqueous solutions are buffer solutions (such as PBS solution), physiological simulated solutions, and the like.
  • the particle size or particle size range of the drug-loaded single-molecule nanopolymer micelles is selected from 10 to 120 nm, preferably 10 to 110 nm, another preferably 10 to 100 nm, another preferably 10 to 80 nm, and another preferably 10-50nm, another preferably 10-40nm, another preferably 10-30nm, another preferably 15-120nm, another preferably 15-110nm, another preferably 15-100nm, another preferably 15-80nm, another preferably 15 ⁇ 50nm, another preferably 15 ⁇ 40nm, another preferably 15 ⁇ 30nm, another preferably 20 ⁇ 120nm, preferably 20 ⁇ 110nm, another preferably 20 ⁇ 100nm, another preferably 20 ⁇ 80nm, another preferably 20 ⁇ 70nm , another preferably 20-50nm, another preferably 20-40nm, another preferably 25-120nm, another preferably 25-110nm, another preferably 25-100nm, another preferably 25-80nm, another preferably 25
  • the particle size of the drug-loaded single-molecule nanopolymer micelles can also be selected from any one or two of the following intervals: 15nm, 16nm, 18nm, 20nm, 25nm, 30nm, 35nm, 40nm, 45nm, 50nm, 55nm, 60nm , 65nm, 70nm, 80nm, 90nm, 100nm, 110nm, 120nm, etc.
  • the test temperature may be 20-30°C, further 25°C. It can be the test result of dynamic light scattering in water, or the test result of transmission electron microscope.
  • the particle size of the drug-loaded single-molecule nanopolymer micelles tested by transmission electron microscopy is ⁇ 120nm, further ⁇ 100nm, further ⁇ 90nm, further ⁇ 80nm, further ⁇ 70nm, further ⁇ 60nm, further ⁇ 50nm.
  • the average diameter of the drug-loaded single-molecule nanopolymer micelles is selected from 15-50 nm, may also be 15-40 nm, may also be 20-40 nm, and may also be 25-35 nm.
  • the test temperature may be 20-30°C, further 25°C. It can be the test result of dynamic light scattering in water, or the test result of transmission electron microscope.
  • the radius of the micelle inner core is 5-50 nm, may also be 5-45 nm, may also be 5-40 nm, may also be 5-35 nm, may also be 5-30 nm, may also be 5-25 nm, 5-20nm, 5-15nm, 5-10nm, 10-50nm, 10-45nm, 10-40nm, 10-35nm, or 5-10nm. It may be 10 to 30 nm, may be 10 to 25 nm, may be 10 to 20 nm, or may be 6 to 8 nm.
  • the radius of the micelle core can also be selected from any one of the following sizes or the interval formed by any two: 5nm, 6nm, 7nm, nm, 9nm, 10nm, 11nm, 12nm, 13nm, 14nm, 15nm, 16nm, 17nm, 18nm, 19nm , 20nm, 22nm, 24nm, 25nm, 30nm, 35nm, 40nm, 45nm, 50nm, etc.
  • the test temperature may be 20-30°C, further 25°C.
  • the results can be tested in water. Further can be small angle X-ray scattering (SAXS) test results.
  • SAXS small angle X-ray scattering
  • the thickness of the micelle shell in water at 25°C is 5-40nm, may also be 5-35nm, may also be 5-30nm, may also be 5-25nm, may also be 5-20nm, or may It may be 5 to 15 nm, may be 10 to 40 nm, may be 10 to 35 nm, may be 10 to 30 nm, may be 10 to 25 nm, may be 10 to 20 nm, or may be 8 to 12 nm.
  • the thickness of the micelle shell in water at 25°C can also be selected from any of the following sizes or intervals consisting of any two: 5nm, 6nm, 7nm, nm, 9nm, 10nm, 11nm, 12nm, 13nm, 14nm, 15nm, 16nm, 17nm, 18nm, 19nm, 20nm, 22nm, 24nm, 25nm, 30nm, 35nm, 40nm, etc.
  • SAXS small angle X-ray scattering
  • a drug delivery system which comprises a double-drug single-molecule nanopolymer micelle, and the double-drug single-molecule nanopolymer micelle comprises a polyamino acid linked to a hydrophilic polymer, wherein On the alpha carbon of the repeating unit of the polyamino acid, the prodrug part of the active ingredient of the platinum drug and the prodrug part of the active ingredient of the antineoplastic drug are bonded; preferably, the molecule of the active ingredient of the antineoplastic drug contains a free hydroxyl group , free amino groups or a combination of both.
  • hydrophilic polymer is selected from the group consisting of polyethylene glycol, poly(propylene glycol), copolymers of ethylene glycol and propylene glycol, poly(ethoxylated polyols), poly(enol) , poly(vinylpyrrolidone), poly(hydroxyalkylmethacrylamide), poly(hydroxyalkylmethacrylate), poly(sugar), poly(alpha-hydroxy acid), poly(vinyl alcohol), poly Phosphazene, polyoxazoline, poly(N-acryloylmorpholine), and any combination of these.
  • the platinum drug is selected from one or more of cisplatin, carboplatin, nedaplatin, oxaliplatin and lobaplatin.
  • the active ingredient of an antineoplastic drug containing a free hydroxyl group, a free amino group, or a combination of the two is selected from one or more of camptothecin compounds, resiquimod, and paclitaxel;
  • the camptothecin compound includes camptothecin and its derivatives or analogs, more preferably, the camptothecin compound includes irinotecan, topotecan, rubitecan, gemitecan , 9-aminocamptothecin, 9-nitrocamptothecin and 7-ethyl-10-hydroxycamptothecin.
  • Another aspect of the present application also provides the use of double NCA monomers of platinum-based drug active ingredients and single NCA monomers of anti-tumor drug active ingredients in the preparation of single-molecule nanopolymer prodrugs or drug delivery systems; preferably, The molecular structure of the active ingredient of the antitumor drug contains free hydroxyl group or free amino group.
  • Another object of the present application is to provide the use of the aforementioned drug-loaded single-molecule nanopolymer as a prodrug.
  • the drug-loaded single-molecule nanopolymer can enter the interior of cells, sense the intracellular microenvironment, release drug active ingredients in response, generate cytotoxicity, and inhibit the growth of tumor cells.
  • the drug release mechanism is as follows: (a) In the highly reducing microenvironment of the cell, tetravalent platinum is reduced to remove the ligand linked to the polyamino acid, thereby realizing the release of the active species of divalent platinum in the cell. Selective release. (b) In the highly reducing environment in the cell, especially the reduction reaction with the high concentration of glutathione in the cell, the disulfide bond is broken to generate a free sulfhydryl group, which then attacks the carbonate or urethane bond connected to the anti-tumor drug , so as to realize the selective release of anti-tumor active drugs in cells.
  • the molecular mechanism of releasing the active Pt(II) drug from tetravalent platinum can be as follows:
  • Another aspect of the present application also provides the aforementioned drug-loaded single-molecule nanopolymer, the drug-loaded single-molecule nanopolymer prepared by the aforementioned preparation method, the aforementioned double-drug single-molecule nanopolymer prodrug, and the aforementioned preparation method.
  • Tumor diseases may include but not limited to lung cancer, gastric cancer, bladder cancer, ovarian cancer, testicular cancer, endometrial cancer, bone cancer, sarcoma, cervical cancer, esophageal cancer, liver cancer, colorectal cancer, head and neck cancer, chorionic epithelial cancer Carcinoma, malignant mole, non-Hodgkin's lymphoma and acute and chronic myelogenous leukemia. It can also include but not limited to lung cancer, esophageal cancer, head and neck tumors,
  • the drug-loaded single-molecule nanopolymer or the nanopolymer prodrug of dual drug active ingredients can be used for the treatment of lung cancer, gastric cancer, bladder cancer, ovarian cancer, testicular cancer, endometrial cancer, bone cancer, sarcoma, cervical cancer, Esophageal cancer, liver cancer, colorectal cancer, head and neck cancer, choriocarcinoma, malignant mole, non-Hodgkin's lymphoma, acute and chronic myelogenous leukemia, etc.
  • the nano-preparation of the present application can also be used to increase the sensitivity of tumor cells to radiotherapy, and the simultaneous administration of radiotherapy can strengthen the control of local progression of lung cancer, esophageal cancer, and head and neck tumors.
  • the present application provides a use of the nanopolymer of dual drug active ingredients according to the present application in the preparation of a drug for treating the tumor.
  • the present application provides a method of combined drug treatment of said tumor in a patient in need thereof, the method administers a therapeutically effective amount of the single-molecule nanopolymer prodrug of the present application or its formulation to the patient .
  • the dosage form of the pharmaceutical preparation according to the present application can be any dosage form clinically applicable to the treatment of the disease, including solutions, suspensions, gels, lyophilized powders, capsules or tablets, etc.
  • the dosage form of the pharmaceutical preparation of the present application is a dosage form suitable for injection (such as intravenous infusion).
  • the formulation for injection may be presented in unit dosage form, e.g. in ampoules, vials, prefilled syringe or multi-dose container.
  • the pharmaceutical formulation according to the present application may also contain at least one pharmaceutically acceptable excipient, such as isotonic agent, wetting agent, One or more of solvents, emulsifiers, preservatives, buffers, acidifying groups, alkalizing agents, antioxidants, chelating agents, coloring agents, complexing agents, flavoring agents, suspending agents and lubricants.
  • excipients are known in the art, and those skilled in the art can select suitable one or more excipients to add to the pharmaceutical preparation of the present application according to the content of the present application.
  • the pharmaceutical formulation according to the present application can be administered to a patient in need by oral, intramuscular injection, intraperitoneal injection, intravenous injection and subcutaneous injection to treat the patient's disease, such as the above-mentioned tumors.
  • Clinicians in the relevant field can select and determine the dosage regimen of the nanopolymer micelles of the present application or its preparations to provide the desired therapeutic effect according to the nature of the disease to be treated, the time of treatment, and the age and physical condition of the patient .
  • the desired dose may conveniently be administered in a single dose or in multiple doses administered at appropriate intervals, eg once, two or more appropriate doses per day.
  • the nanopolymer micelles or preparations thereof of the present application can be administered in combination with other chemotherapeutic agents and/or radiation.
  • the measurement parameters related to raw material components may have slight deviations within the weighing accuracy range unless otherwise specified. Involves temperature and time parameters, allowing for acceptable deviations due to instrumental test accuracy or operational accuracy.
  • the raw materials, experimental reagents and experimental instruments used in the following examples can be purchased from the market, the reaction conditions used are known in the art, and the identification or assay methods used are commonly used in the art Methods.
  • Diboc is di-tert-butoxycarbonyl dicarbonate
  • DMAP is 4-dimethylaminopyridine
  • THF is tetrahydrofuran
  • DMF is N,N-dimethylformamide
  • EDC is 1- (3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride
  • NHS means N-hydroxysuccinimide
  • BTC means bis(trichloromethyl)carbonate, Bis(trichloromethyl)carbonate
  • Tween 80 means Tween 80
  • DACHPt means (1,2-diaminocyclohexane) platinum dichloride
  • CPT means camptothecin
  • GSH means glutathione.
  • the "molecular weight" of polyethylene glycol refers to the number average molecular weight unless otherwise specified.
  • the molecular weight of the single-molecule nanopolymer is related to the number-average molecular weight unless otherwise specified.
  • the mass spectrometry test conditions are: the substance to be detected is prepared as a 1 mg/mL dichloromethane or methanol solution, 0.5 ⁇ L of the solution is added dropwise to the sample stage, and after drying at room temperature, the sample stage is sent to the ion source Tests were performed (Bruker REFLEX Model III MALDI-TOF-MS).
  • the 1 H NMR test conditions are as follows: the substance to be detected is configured as a 10 mg/mL CDCl3_ solution, and the 1 H NMR spectrum is established using a Bruker AVANCE 500 III superconducting pulse Fourier transform nuclear magnetic resonance spectrometer , the test temperature is 25°C, the number of scans is 64, and the internal standard is tetramethylsilane (TMS).
  • TMS tetramethylsilane
  • % (w/v) means mass volume percentage
  • % (v/v) means volume ratio
  • Embodiment 1 The preparation of the drug active molecule prodrug containing free hydroxyl group, free amino group or the combination of both in the molecule.
  • N-Boc-N'-Cbz-L-Lys (2g, 5.26mmol) was dissolved in chloroform (15mL) and mixed with sodium bicarbonate solution (12mL 0.45mmol/L). Stir for 5 minutes under nitrogen protection, then add Diboc (di-tert-butoxycarbonyl dicarbonate) in chloroform (1.22 g, 5.5 mmol, 9 mL) dropwise, reflux for 90 minutes, and cool to room temperature. The organic phase was separated, and the aqueous phase was extracted with chloroform. The combined organic phases were evaporated to dryness under reduced pressure, and N-Boc-N'-Cbz-Lys-OtBu was obtained by column chromatography.
  • Camptothecin 500mg, 1.43mmol was dispersed in anhydrous dichloromethane and dissolved (80mL), in an ice bath, under nitrogen protection, anhydrous dichloromethane (3mL) containing triphosgene (157mg, 0.53mmol) was added, Continue to stir under ice bath for 30 minutes, add anhydrous dichloromethane (10 mL) dissolved with DMAP (4-dimethylaminopyridine, 560 mg, 4.6 mmol) until camptothecin is completely dissolved, and continue stirring reaction 1 under ice bath Hours, transferred to room temperature and continued stirring reaction in the dark for 1 hour.
  • DMAP 4-dimethylaminopyridine
  • CPT-ss-OH (52.8mg, 0.1mmol) was dispersed into anhydrous dichloromethane (15mL), and under N2 environment, 1mL of anhydrous dichloromethane dissolved with triphosgene (13.2mg, 0.045mmol) was added, in Stir in an ice bath for 30 minutes, then add anhydrous dichloromethane (2 mL) dissolved with DMAP (39 mg, 0.32 mmol) until completely dissolved, continue to stir and react in an ice bath for 1 hour, then turn to room temperature and continue to avoid light and stir for reaction 1 Hour.
  • Boc-Lys-OtBu (45.3mg, 0.15mmol) was added in anhydrous dichloromethane (1mL) under the protection of N 2 , mixed evenly, and stirred at room temperature for 24 hours in the dark. After the reaction was completed, 50 mL of dichloromethane was added, and washed three times with 0.1M HCl aqueous solution, saturated NaCl and water successively, the organic phase was collected and dried with anhydrous Na 2 SO 4 , separated by column chromatography to obtain light yellow crystals Boc-Lys-OtBu-ss-CPT.
  • Boc-Lys-OtBu-ss-CPT powder (85.7mg, 0.1mmol) was dissolved in dichloromethane (2mL), mixed with trifluoroacetic acid (2mL), reacted at room temperature for 2 hours, evaporated under reduced pressure to remove the solvent, and added di Methyl chloride was dissolved and washed with saturated sodium bicarbonate, and the organic phase was collected and dried to obtain Lys-ss-CPT.
  • Lys-ss-CPT (0.1mmol) was dissolved in dichloromethane (2mL), mixed with dichloromethane (2mL) containing triphosgene (0.2mmol), reacted at room temperature for 2 hours, distilled off the solvent under reduced pressure, added Dissolve tetrahydrofuran (60°C), add an appropriate amount of n-hexane, crystallize in a refrigerator at 4°C, and collect white needle-like crystal NCA-Lys-ss-CPT.
  • Cisplatin (1.0g, 3.33mmol) was dispersed in distilled water, added with 30% H 2 O 2 (20ml), stirred at 70°C in the dark for 5h until clear, cooled to room temperature and placed in a 4°C refrigerator for recrystallization. After crystallization and filtration, the filter cake was washed with ice water, ethanol and diethyl ether in sequence, and dried to obtain compound (1) as yellow crystals.
  • N-Boc-N'-Cbz-L-Lys (2g, 5.26mmol) was dissolved in chloroform (15mL) and mixed with sodium bicarbonate solution (12mL 0.45mmol/L). Stir for 5 minutes under nitrogen protection, then add Diboc in chloroform solution (1.22 g, 5.5 mmol, 9 mL) dropwise, reflux for 90 minutes, and cool to room temperature. The organic phase was separated, and the aqueous phase was extracted with chloroform. The combined organic phases were evaporated to dryness under reduced pressure, and N-Boc-N ⁇ -Cbz-Lys-OtBu was obtained by column chromatography.
  • Example 3 The application of the drug-loaded single-molecule nanopolymer (a double-drug single-molecule nanopolymer, which can be used as a nanopolymer prodrug) and the preparation of nanomicelles thereof
  • the drug-loaded single-molecule nanopolymer has the tetravalent structural unit shown in the aforementioned formula (I-2), the monovalent structural unit shown in the formula (III-2), and the formula (II-2) The divalent structural unit shown;
  • each occurrence of n11 and n21 is 4, and each occurrence of n12 and n22 is 2;
  • each occurrence of n31 is 4, each occurrence of n32 is 2, each occurrence of n33 is 2; each occurrence of L R is -SS-;
  • Every occurrence of n51 is 3; every occurrence of Z 5 is NH;
  • D Pt is the residue of cisplatin
  • D T is the residue of camptothecin
  • p is approximately equal to 113, and the corresponding mPEG molecular weight is approximately 5000 Da (number average molecular weight).
  • the monomers represented by the formula (I-3), the monomers represented by the formula (III-3) and the monomers represented by the formula (II-3) are used to prepare the drug-loaded single-molecule nanopolymer.
  • the structure of the monomer shown in formula (I-3) is the NCA-Pt-NCA prepared in Example 2, and the structure of the monomer shown in Formula (II-3) is the NCA-Lys-ss prepared in Example 1 -CPT, the structure of the monomer shown in formula (III-3) is
  • the reaction equation for preparing the drug-loaded single-molecule nanopolymer is as follows (the prepared drug-loaded single-molecule nanopolymer is denoted as P101):
  • m can be selected from 0-100
  • n can be selected from 1-100
  • k can be selected from 1-1000, but they are not limited to this range.
  • Methoxy-polyethylene glycol-amino (MeO-PEG-NH 2 , 0.1g, 0.01mmol) was dissolved in benzene (3mL), stirred until PEG was completely dissolved, frozen in liquid nitrogen, and dried in vacuum with cold hydrazine for 6 hours.
  • the dried polyethylene glycol was dissolved in anhydrous DMF (2mL), stirred evenly, and NCA-Lys-ss-CPT (0.50mmol) and NCA-Pt-NCA (0.18mmol) were dissolved in anhydrous DMF (2 mL) was slowly added dropwise to the reaction system, the reaction tube was sealed, taken out from the glove box, and placed in an oil bath at 35°C for 72 hours with continuous stirring.
  • the reaction product was slowly dropped into glacial ether to obtain a white precipitate, the supernatant was discarded and the above operation was repeated three times to obtain a purified product.
  • the product was vacuum dried in a vacuum pan for 6 hours.
  • the dried solid was dissolved in DMSO (dimethyl sulfoxide, 2 mL), placed in a dialysis bag (MWCO: 100 kDa), dialyzed in ultrapure water for two days (change water 5 times), and after freeze-drying, the final product was collected (P101).
  • DMSO dimethyl sulfoxide
  • m can be selected from 0-100
  • n can be selected from 1-100
  • k can be selected from 1-1000, but they are not limited to this range.
  • the paclitaxel and cisplatin double-drug polymer prodrug nanomicelles were prepared by substituting paclitaxel and resiquimod respectively bundles and resiquimod and cisplatin double-drug polymer prodrug nanomicelles.
  • Embodiment 4 Characterization of the double-drug single-molecule polymer prodrug nanomicelle of the present application
  • the lyophilized product was dissolved in water (1 mg/mL), and the molecular weight distribution of the product was characterized by GPC (superdex200). As shown in V-A in Figure 5 and V-B in Figure 5, a large molecular weight product was successfully synthesized. The molecular weight after polymerization was quantified by molecular exclusion chromatography, and the results are shown in V-A and V-B in FIG. 5 .
  • the lyophilized product was dissolved in water (1 mg/mL), and the molecular weight of the quantitative product was about 1030 kDa by using analytical ultra-high speed centrifugation technology. The molecular weight after polymerization was quantified by analytical ultracentrifugation.
  • the molecular weight of PEG is 5kDa (p is about 113 in the formula (III-2)), and the molecular weight of the nanoparticles after the reaction is 1030kDa.
  • the lyophilized product was dissolved in water (1 mg/mL), and the diffusion time of the product quantified by fluorescence correlation spectroscopy (FCS) was about 7600 ⁇ s.
  • FCS fluorescence correlation spectroscopy
  • the molecular weight after polymerization was quantified by fluorescence correlation spectroscopy.
  • the nano-preparation (0.01mg/mL) PBS buffer solution (10mM, pH 7.4) was configured, and the particle size and polydispersity index (PDI) of the nano-preparation were characterized by a dynamic light scattering instrument.
  • the results are shown in FIG. 6 . According to Fig.
  • the average particle diameter of the double-drug single-molecule nanopolymer micelles of the present application is 33.6 nanometers, and the particle diameter scope is 21.5 ⁇ 52.7 nanometers, and the polydispersity index PDI of particle size distribution is about 0.05;
  • the particle diameter ranges from 32.3 to 264.1 nanometers, the average particle diameter is about 96.7 nanometers, and the polydispersity index PDI of the particle diameter is greater than >0.1, specifically in the range of 0.18 to 0.30.
  • Nanoparticles obtained by chemical polymerization have a high degree of uniformity in size, while those obtained by self-assembly are not uniform in size.
  • the double-drug single-molecule polymer prodrug nanomicelle obtained by chemical polymerization has excellent colloidal solution stability, is resistant to freeze-drying and reconstitution, and has a stable structure.
  • the nanoparticles obtained by self-assembly are not resistant to freeze-drying and reconstitution, and the original structure cannot be obtained after reconstitution.
  • FCS Fluorescence correlation spectroscopy
  • the double-drug monomolecular polymer prodrug nanomicelle solution (0.1mg/mL) and the self-assembled nanomicelle solution (0.1mg/mL) were mixed and diluted with water, and the colloidal kinetics was tested by fluorescence correlation spectroscopy (FCS) academic features. The results are shown in FIG. 9 .
  • the double-drug monomolecular polymer prodrug nanomicelle solution (0.1 mg/mL) was sent to small-angle X-ray scattering (SAXS) test. The results are shown in Figure 10.
  • the synthesized product is a nano-sized micellar structure
  • the radius of the inner core is about 6.4 nanometers
  • the thickness of the outer shell PEG layer is about 9.8 nanometers.
  • the diameter of the micelles is about 32.4 nm.
  • the average particle size is 33.6 nm, the particle size range is 21.5-52.7 nm, and the polydispersity index PDI of the particle size distribution is about 0.05;
  • the average particle size is 33.9 nanometers, the particle size range is 22.1-53.4 nanometers, and the polydispersity index PDI of particle size distribution is about 0.05; Excellent resistance to ultrasonic treatment, stable particle size.
  • the average particle size is about 96.7 nm, the particle size range is 32.3-264.1 nm, and the polydispersity index PDI of the particle size is about 0.18; after 60min ultrasonic treatment, the average particle size is about 342 nanometers, the particle size range is 7.6-464.1 nanometers, and the polydispersity index PDI of the particle size is about 0.64; therefore, the self-assembled nano-preparation cisplatin@PEG-PGlu( ss-CPT) cannot withstand ultrasonic treatment, and the structure is obviously deformed.
  • Example 5 The application of double-drug single-molecule polymer prodrug nanomicelles responds to the intracellular reducing microenvironment (research on the release of active original drugs)
  • the tetravalent platinum in the unimolecular nanopolymer of the present application undergoes a reduction reaction in the cell, and the molecular mechanism of releasing the active Pt (II) drug can be as follows:
  • the concentration of CPT in the dialyzate was determined according to HPLC ( Figure 12), the mobile phase was methanol and deionized water (20–100%, v/v), the flow rate was 1.0 mL/min, 25°C, and the absorption wavelength was 370 nm.
  • the concentration of Pt in the dialyzate was determined by ICP-MS. The results are shown in FIG. 13 .
  • the double-drug single-molecule polymer prodrug nanomicelles of the present application formed by chemical polymerization will not release the active drug in advance outside the cell, and exhibit the function of triggering the release of the active drug inside the cell.
  • micellar cisplatin@PEG-PGlu(ss-CPT), CPT@PEG-PLA in the extracellular microenvironment pH 7.4, GSH (0mM).
  • the PBS solution (10mM) containing the above nano-preparation was injected into the dialysis bag (MWCO : 10kDa) and immersed in the above two PBS solutions (28mL, 10mM, containing 0.5% (w/v) Tween 80), incubated at 37°C for 48 hours under gentle shaking (100RPM). Extract 1mL at predetermined interval time points Release medium, and supplement 1mL fresh blank medium.
  • Example 6 The cytotoxicity of the double-drug single-molecule polymer prodrug nanomicelle of the present application
  • cytotoxicity of cisplatin, cisplatin@PEG-PGlu, cisplatin@PEG-PGlu(ss-CPT), CPT@PEG-PLA, free CPT&cisplatin double drug and the single molecular nano-prodrug of the application was tested by MTT method analyze.
  • A549 cells were seeded in 96-well plates (100 ⁇ L) at a density of 10 5 cells/mL. Incubate for 24 hours in a cell culture incubator at 37°C and 5% CO2 , discard the old medium, and then add 100 ⁇ L of drug preparations containing different concentrations to each well. 0% blank control, after cultured for 48h, the viability of the cells was detected.
  • CPT camptothecin
  • Pt platinum
  • a large amount of double-drug self-assembled nano-preparation cisplatin@PEG-PGlu(ss-CPT) and free cisplatin were used as the control group.
  • blood was collected at different time points, and the Pt content was determined by ICP-MS, in which ultracentrifugation (10000G) of the plasma was used to separate free small molecule cisplatin and nano-preparations containing Pt.
  • the blood circulation time of the single-molecule nano-prodrug is significantly prolonged, and the single-molecule nano-prodrug is in the blood circulation.
  • the free cisplatin drug will not leak in advance; and for the self-assembled nano-preparation as a control, compared with the free cisplatin solution control group, the blood circulation time of the self-assembled nano-preparation is significantly prolonged, and the self-assembled nano-preparation in the blood circulation There will be premature leakage of free cisplatin drug.
  • Example 8 The tumor drug accumulation test of the double-drug single-molecule polymer prodrug nanomicelle of the present application
  • the tumor model used in this experiment was subcutaneous transplantation of A549 lung cancer.
  • the drug preparation was injected into female Bal b/c nude mice (8 weeks) by tail vein injection.
  • the injection dose was 200 microliters, CPT 5mg/kg, Pt 1.8mg/kg, in which cisplatin was used as the reference test group, and the same amount of double-drug self-assembled nano-preparation cisplatin@PEG-PGlu(ss-CPT) was used as the control group.
  • the tumor mass was dissected, concentrated nitric acid was used to fully dissolve the tumor, and the Pt content in the tumor was determined by ICP-MS. The results are shown in FIG. 17 .
  • the drug concentration in the tumor site was lower in the cisplatin control group, and the nano-preparation could be significantly enriched in the tumor site by relying on the EPR effect of the nano-preparation on the tumor.
  • the enrichment amount of the tumor drug gradually increased.
  • the enrichment amount of the single-molecule nano-prodrug prepared by this application is significantly better than that of the double-drug self-assembled nano-preparation cisplatin@PEG-PGlu(ss-CPT).
  • the possible reasons include that the single-molecule nano-prodrug is better in blood circulation.
  • the stability of colloidal structure prevents the early leakage of drugs.
  • Example 9 In vivo tumor suppressor efficacy evaluation of double-drug single-molecule polymer prodrug nanomicelles of the application
  • the tumor model used in this experiment was subcutaneous transplantation of A549 lung cancer.
  • the drug preparation was injected into female Bal b/c nude mice (8 weeks) by tail vein injection.
  • the injection dose was 200 microliters, CPT 5mg/kg, Pt 1.8mg/kg, in which PBS was used as the reference test group, the same amount of single-drug self-assembled nanomicelle preparations CPT@PEG-PLA and cisplatin@PEG-PGlu were used as the control group, and the same amount of CPT&cisplatin
  • the molecular mixed solution was used as the control group, and the same amount of double-drug self-assembled nano-preparation cisplatin@PEG-PGlu(ss-CPT) was used as the control group.
  • the results are shown in FIG. 18 .
  • the single-molecule nano-prodrug mice grew in good condition, and the tumor growth was significantly inhibited. It not only effectively reduces the toxicity of chemotherapy drugs, but also significantly improves the antitumor efficacy of traditional self-assembled nano-agents. Therefore, the single-molecule nano-prodrug of the present application has excellent clinical application prospects.
  • Embodiment 10 Platinum monodrug (cisplatin) unimolecular nanopolymer and preparation of micelles thereof
  • Methoxypolyethylene glycol propylamine (MeO-PEG-NH 2 , molecular weight 5kDa, 0.1g, 0.01mmol, monofunctional hydrophilic polymer) was dissolved in benzene (3mL), stirred until PEG was completely dissolved, and frozen in liquid nitrogen , then dried in vacuum with cold hydrazine for 6 hours.
  • the dried polyethylene glycol was dissolved in anhydrous DMF (2mL), stirred evenly, and NCA-Pt-NCA (prepared by the method of Example 2, 0.18mmol) was dissolved in anhydrous DMF (2mL) , slowly added dropwise to the reaction system, sealed the reaction tube, took it out from the glove box, placed it in an oil bath at 35°C and continued stirring for 72 hours.
  • the reaction product was slowly dropped into glacial ether to obtain a white precipitate, the supernatant was discarded and the above operation was repeated three times to obtain a purified product.
  • the product was vacuum dried in a vacuum pan for 6 hours.
  • the dried solid was dissolved in DMSO (2 mL), placed in a dialysis bag (MWCO: 100 kDa), dialyzed in ultrapure water for two days (water was changed 5 times), and the final product was collected after freeze-drying.
  • the dynamic light scattering (DLS) test results are shown in Figure 26(B).
  • the average particle size of the polymerized product is about 30.4 nm, the particle size range is 23.8-41.1 nm, and the polydispersity index PDI of the particle size distribution is about 0.05.
  • the test result of transmission electron microscope (TEM) is shown in Fig. 27.
  • the polymerized product is uniform spherical with a diameter of less than 50 nm and an average diameter of about 30 nm, which is basically consistent with the DLS test result. After freeze-drying and reconstitution treatment, the size and shape are stable.
  • cisplatin single-molecule nanopolymer nanoparticles with uniform size can be obtained by chemical polymerization, which can be used as a prodrug.
  • Embodiment 11 Platinum single drug (DACHPt) unimolecular nanopolymer and preparation of micelles thereof
  • NCA-Pt-NCA NCA-DACHPt-NCA
  • DACHPt ((1,2-diaminocyclohexane) platinum dichloride, 3.8 g, 10 mmol) was dispersed in distilled water, 30% H 2 O 2 (60 mL) was added, and stirred at 70° C. in the dark for 5 h until clear, After cooling down to room temperature, place in a refrigerator at 4°C for recrystallization. After crystallization and filtration, the filter cake was washed with ice water, ethanol and diethyl ether in sequence, and dried to obtain compound crystals (1).
  • NCA-DACHPt-NCA The mass spectrum of NCA-DACHPt-NCA is shown in FIG. 28 .
  • the 1 H NMR test can be performed with reference to the method in this example.
  • mPEG-CH 2 CH 2 CH 2 NH 2 which can be abbreviated as mPEG-NH 2 , 5kDa, 0.05g, 0.01mmol
  • benzene 3mL
  • the reactant was transferred to an anhydrous and oxygen-free glove box, and the dried polyethylene glycol was dissolved in anhydrous DMF (2mL), magnetically stirred evenly, and NCA-DACHPt-NCA (0.4mmol) was completely dissolved in anhydrous DMF (20 mL), then slowly added dropwise to the reaction system containing mPEG-NH 2 , connected to a balloon to collect the carbon dioxide product, closed the reaction system, took it out from the glove box, and placed it in an oil bath at 35°C for 72 hours with continuous stirring. The reaction product was slowly dropped into glacial ether to obtain a white precipitate, the supernatant was discarded and the above operation was repeated three times to obtain a purified product.
  • the product was vacuum dried in a vacuum pan for 6 hours.
  • the dried solid was dissolved in DMSO (10 mL), placed in a dialysis bag (MWCO: 100 kDa), dialyzed in ultrapure water for two days (water was changed 5 times), and after freeze-drying, the final polymer product (P102) was collected.
  • the dynamic light scattering (DLS) test results are shown in Figure 29(B).
  • the average particle size of the polymerized product is about 34 nm, the particle size range is 24.2-43.5 nm, and the polydispersity index of the particle size distribution is about 0.05.
  • the test results of transmission electron microscopy (TEM) are shown in Figure 30.
  • the polymerized product is uniform spherical, with a diameter of less than 50 nanometers and an average diameter of about 30 nanometers, which is consistent with the DLS results, and compared before and after freeze-drying and reconstitution treatment, the size The shape is stable.
  • nanoparticle of DACHPt single-molecule nanopolymer with uniform size can be obtained by chemical polymerization, which can be used as a prodrug.
  • Paclitaxel (PTX, 854mg, 1.0mmol) was dispersed in anhydrous dichloromethane and dissolved (200mL), in an ice bath, under nitrogen protection, anhydrous dichloromethane (5mL) containing triphosgene (297mg, 1.0mmol) was added, Continue to stir in the ice bath for 30 minutes, add DMAP (488mg, 4.0mmol) in anhydrous dichloromethane (10mL) until the paclitaxel is completely dissolved, continue to stir and react in the ice bath for 1 hour, turn to room temperature and continue to stir in the dark React for 1 hour.
  • DMAP 488mg, 4.0mmol
  • PTX-ss-OH as raw material instead of CPT-ss-OH, refer to the method of 1.4 to 1.6 in Example 1 to prepare PTX-ss-NCA.
  • the 1 H NMR spectrum of PTX-ss-NCA is shown in FIG. 31 .
  • 7.2-8.1ppm represents the three benzene ring peaks of paclitaxel; the peak at 3.4ppm proves the successful bonding of paclitaxel derivatives and NCA-Lysine; 1.2-1.9ppm represents the side chain n-butyl peak of lysine.
  • Methoxypolyethylene glycol propylamine (MeO-PEG-NH 2 , 5kDa, 0.2g, 0.01mmol) was dissolved in benzene (3mL), magnetically stirred until the PEG was completely dissolved, frozen in liquid nitrogen, and vacuum-dried with cold hydrazine for 6 hours .
  • the reaction product was slowly dropped into glacial ether to obtain a white precipitate, the supernatant was discarded and the above operation was repeated three times to obtain a purified product.
  • the product was vacuum dried in a vacuum pan for 6 hours.
  • the dried solid was dissolved in DMSO (12 mL), placed in a dialysis bag (MWCO: 100 kDa), dialyzed in ultrapure water for two days (water was changed 5 times), and after freeze-drying, the final polymer product (P103) was collected.
  • test results without ultrasonic treatment and with ultrasonic treatment are shown in (A) and (B) in Figure 32, respectively, where the ultrasonic parameters (1.0MHz, 9.9W) and ultrasonic time (60min).
  • the average particle diameter of the polymerized product is about 46.4 nanometers, and the particle diameter ranges from 34.6 to 61.8 nanometers, which proves the success of the polymerization reaction, and can withstand ultrasonic and other treatments, and the particle diameter remains unchanged.
  • the transmission electron microscope (TEM) test was carried out. After dialysis, the solution was freeze-dried three times and reconstituted (1 mg/mL). The test results before and after reconstitution are shown in (A) and (B) in Figure 33, respectively. After polymerization, the product is uniform spherical, with an average diameter of about 40 nanometers, which is basically the same size as DLS, and after freeze-drying and reconstitution treatment, the size and shape are stable.
  • the concentration of PTX in the dialyzate was determined by HPLC, the mobile phase was methanol and deionized water (20%-100% (v/v), volume ratio), the flow rate was 1.0mL/min, 25°C, and the absorption wavelength was 270nm.
  • R848-ss-OH was prepared by referring to the method of 1.3. in Example 1.
  • R848-ss-NCA was prepared by using R848-ss-OH as the raw material instead of CPT-ss-OH, referring to the method from 1.4 to 1.6 in Example 1.
  • the 1 H NMR spectrum of R848-ss-NCA is shown in FIG. 35 .
  • above 7.2ppm represents the biphenyl peak of R848; the peak at 3.4ppm proves the successful bonding of R848 derivatives and NCA-Lysine; 1.2-1.9ppm (except the single peak at 1.45ppm) represents the side chain n-butyl peak of Lysine.
  • Methoxypolyethylene glycol propylamine (MeO-PEG-NH 2 , 5kDa, 0.1g, 0.01mmol) was dissolved in benzene (3mL), magnetically stirred until the PEG was completely dissolved, frozen in liquid nitrogen, and vacuum-dried with cold hydrazine for 6 hours .
  • the product was vacuum dried in a vacuum pan for 6 hours.
  • the dried solid was dissolved in DMSO (12 mL), placed in a dialysis bag (MWCO: 100 kDa), dialyzed in ultrapure water for two days (water changes 5 times), and after freeze-drying, the final product was collected (P104).
  • test results without ultrasonic treatment and with ultrasonic treatment are shown in (A) and (B) in Figure 36, respectively, where the ultrasonic parameters (1.0MHz, 9.9W) and ultrasonic time (60min).
  • the average particle diameter of the polymerized product is about 44.9 nanometers, and the particle diameter ranges from 32.8 to 66.2 nanometers, which proves the success of the polymerization reaction, and can withstand ultrasonic and other treatments, and the particle diameter remains unchanged.
  • the transmission electron microscope (TEM) test was carried out. After dialysis, the solution was freeze-dried three times and reconstituted (1 mg/mL). The test results before and after reconstitution are shown in (A) and (B) in Figure 37, respectively. After polymerization, the product is uniform spherical, with an average diameter of about 40 nanometers, which is basically the same size as DLS, and after freeze-drying and reconstitution treatment, the size and shape are stable.
  • MMAE-ss-OH was prepared by referring to the method of 1.3. in Example 1.
  • MMAE (718mg, 1.0mmol) was dissolved in anhydrous dichloromethane (20mL), in an ice bath, anhydrous dichloromethane (5mL) containing triphosgene (297mg, 1.0mmol) was added under nitrogen protection, and the Stir in the bath for 30 minutes, add DMAP (488mg, 4.0mmol) in anhydrous dichloromethane (10mL) until MMAE is completely dissolved, continue to stir and react in an ice bath for 1 hour, turn to room temperature and continue to avoid light and stir for 1 hour .
  • MMAE-ss-NCA was prepared by using MMAE-ss-OH as the raw material instead of CPT-ss-OH, referring to the method of 1.4 to 1.6 in Example 1.
  • the 1 H NMR spectrum of MMAE-ss-NCA is shown in FIG. 39 .
  • above 7.6ppm represents the benzene ring peak of MMAE; the peak at 3.4ppm proves the successful bonding of MMAE derivatives and NCA-Lysine; 1.2-1.9ppm represents the side chain n-butyl peak of Lysine.
  • Methoxy-polyethylene glycol propylamine (MeO-PEG-NH 2 , 5kDa, 0.1g, 0.01mmol) was dissolved in benzene (3mL), magnetically stirred until the PEG was completely dissolved, frozen in liquid nitrogen, then vacuum-dried with cold hydrazine6 Hour.
  • the reactant is transferred into an anhydrous and oxygen-free glove box, and the polyethylene glycol after freeze-drying is dissolved in anhydrous DMF (2mL), stirred evenly by magnetic force, NCA-Pt-NCA (0.2mmol, the method of embodiment 2 Preparation) and MMAE-ss-NCA (0.2mmol) were dissolved in anhydrous DMF (30mL), and then slowly added dropwise to the reaction system containing MeO-PEG-NH 2 , the carbon dioxide product was collected by connecting a balloon, and the reaction system was closed. Take it out from the glove box, place it in an oil bath at 35°C and keep stirring for 72h.
  • the reaction product was slowly dropped into glacial ether to obtain a white precipitate, the supernatant was discarded and the above operation was repeated three times to obtain a purified product.
  • the product was vacuum dried in a vacuum pan for 6 hours.
  • the dried solid was dissolved in DMSO (12 mL), placed in a dialysis bag (MWCO: 100 kDa), dialyzed in ultrapure water for two days (water was changed 5 times), and after lyophilization, the final polymer product (P105) was collected.
  • the results of the DLS test (without sonication) are shown in Figure 40.
  • the average particle diameter of the polymerized product is about 43.9 nanometers, and the particle diameter ranges from 30.5 to 58.4 nanometers, which proves the success of the polymerization reaction.
  • the transmission electron microscope (TEM) test was carried out. After dialysis, the solution was freeze-dried three times and reconstituted (1 mg/mL). The test results before and after reconstitution are shown in (A) and (B) in Figure 41, respectively. After polymerization, the product is uniform spherical, with an average diameter of about 40 nanometers, which is basically the same size as DLS, and after freeze-drying and reconstitution treatment, the size and shape are stable.
  • MMAE concentration of MMAE in the dialyzate was determined by HPLC, the mobile phase was methanol and deionized water (20–100%, (v/v)), the flow rate was 1.0 mL/min, 25°C, and the absorption wavelength was 280 nm.

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Abstract

一种载药单分子纳米聚合物，其通过二价含铂连接基LPt将多条聚氨基酸链构建成非线性骨架，至少一条聚氨基酸链的端部连接有亲水性聚合物链，LPt中的铂原子参与构成铂类药物单元，聚氨基酸链的侧基还可选地接枝有第二药物单元。该载药单分子纳米聚合物可作为前药，可构成胶束或药物递送系统，还涉及制备方法和用途。

Description

载药单分子纳米聚合物、前药、胶束、药物递送系统及制备方法和用途
本申请要求于2021年7月27日提交于中国专利局、申请号为CN2021108603562发明名称为“双药单分子纳米聚合物前药及其制备方法和用途”的中国专利申请的优先权，其全部内容通过引用结合在本申请中。
技术领域
本申请涉及药物技术领域及药物递送系统，涉及一种载药单分子纳米聚合物、前药、胶束、药物递送系统及制备方法和用途，还特别涉及一种胞内还原微环境响应活化型双药单分子纳米聚合物前药。本申请还涉及所述胞内还原微环境响应活化型双药单分子纳米聚合物前药的制备方法及用途。
背景技术
纳米药物制剂具有缓控释、靶向等诸多优点。目前，纳米药物制剂是以癌症精准治疗为核心目标的尖端制剂技术。随着纳米技术的高速发展，各种生物活性分子(化学药物、多肽、核酸等)得以通过多种方式(如：分子自组装)存储于性状多样的纳米材料中。尤其是，这些纳米材料可以依靠组员分子的功能化设计以及组装结构的精妙调控，成为生物体内的“智能运输机”，具有克服各级生物屏障、定向输送生物活性分子至靶部位的潜力。目前，已解决了传统药物制剂的诸多弊端：降低了药物毒副作用，提高了药物对病灶部位的生物利用度。过去数十年中，纳米药物递送系统在肿瘤靶向治疗中的研究取得了十分显著的进展，目前已有多种制剂(脂质体Doxil TM、白蛋白Abraxane TM等)在全球上市。抗癌纳米粒的直径在10～100纳米范围内，肿瘤血管可以渗漏大分子，被称为“EPR效应”(增强渗透和驻留效应)。纳米粒能够从血管里渗漏出来并在肿瘤内蓄积，还可在细胞外间隙进行扩散。
然而，传统纳米药物制剂(如Bind-14，NC-6300)的药释行为存在局部瞬时药物浓度难以达到有效水平的问题。传统纳米制剂虽然对于肿瘤病灶具有较高富集潜力，然而其缓慢的药释速率导致传统纳米制剂对肿瘤细胞杀伤效果甚至低于游离小分子药物。同时，传统纳米制剂的药释行为也发生在血液循环中，这种药物早期泄漏会降低药物对靶病灶的生物利用度，造成非靶病灶的毒性。另外，传统纳米制剂或自组装纳米制剂还具有以下明显缺点：1)胶体稳定性差，在复杂的生理条件下容易发生结构解离；2)药物容易早期泄漏；以及3)制备工艺复杂，如：薄膜水化、纳米沉淀法等，需要去除未包裹药物分子及辅助分子(如：有机溶剂等)，大规模量产难度大。
针对肿瘤微环境选择性控释以及药物过早泄漏的问题，将药物分子通过化学键合的方式构建前药体系是一条可行的方式，前药是指药物经过化学结构修饰后得到的在体外无活性或活性较小的一类药物,在体内能够经物理、化学或酶激活后释放出活性药物而 发挥药效。基于前药的设计策略除了可以解决药物提前泄漏问题，同时可以响应肿瘤微环境或胞内微环境迅速释药。
已有研究把化疗药物键合到聚合物高分子上，形成聚合物前药分子，然后通过聚合物前药分子的自组装形成纳米粒子。例如中国专利文献CN109908084A，CN101203549B，CN100457185C，CN100344293C和“金滔，羟喜树碱MePEG-PLA纳米粒的制备及其体外抗肿瘤研究，浙江中医药大学，硕士论文，2013-05-01”公开了聚合物前药纳米粒子。但是，这些纳米粒子在血液循环中同样会结构解离，在体内不稳定，同样面临工艺复杂，不耐受超声、冻干复溶等问题。
因此，对体内体外稳定性更高、具有改进的载药和释药性能、生产工艺简单高效且耐受超声、冻干复溶等处理的纳米聚合物前药存在未满足的需求。
发明内容
本申请的一个目的在于提供一种载药单分子纳米聚合物，其包含多条聚氨基酸链，所述多条聚氨基酸链的链间通过多个二价连接基L Pt共价相连使所述多条聚氨基酸链构成非线性骨架，至少一条所述聚氨基酸链的端部连接有亲水性聚合物链；其中，所述二价连接基L Pt的线性骨架中包含铂原子，所述铂原子参与构成铂类药物单元，所述铂类药物单元可以为铂类药物活性成分或其前药的残基；
可选地，所述聚氨基酸链的侧基接枝有第二药物单元；其中，所述第二药物单元可以为抗肿瘤药物活性成分或其前药的残基。
该载药单分子纳米聚合物通过二价含铂连接基L Pt将多条聚氨基酸链构建成非线性骨架，至少一条聚氨基酸链的端部连接有亲水性聚合物链，L Pt中的铂原子参与构成铂类药物单元(可以为铂类药物活性成分或其前药的残基)。通过调节L Pt的分布密度可控制该载药单分子纳米聚合物具有支化或适度交联的三维结构，进一步结合亲水性聚合物链在聚氨基酸链端部的位置设计，使载药单分子纳米聚合物能够在水性介质中无需自组装就可形成具有核壳结构的单分子纳米聚合物胶束，亲水性聚合物链分布于外壳，药物成分被包载于内核。该载药单分子纳米聚合物可以仅装载铂类药物单元，构成铂单药单分子纳米聚合物；该载药单分子纳米聚合物可以还接枝第二药物单元(可以为抗肿瘤药物活性成分或其前药的残基)，该第二药物单元可以接枝在聚氨基酸链的侧基，此时可构成双药单分子纳米聚合物。铂类药物单元及第二药物单元的相对含量可以通过控制相应单体的投料量灵活调节。L Pt的分布密度可通过调节非支化氨基酸单体与L Pt支化氨基酸单体的投料比例进行调节，在非支化氨基酸单体中还可以灵活调节包含第二药物单元的氨基酸单体所占比例。该载药单分子纳米聚合物在体内和体外稳定性好，分散性好，粒径均一，无毒副作用，而且在胞外不会释放药物活性成分而在胞内呈现出触发性释放药物活性成分，此外，可采用操作简便、反应温和、成本低廉和环境友好的制备方法获得。
本申请的另一个目的在于提供一种载药单分子纳米聚合物的制备方法，其包括如下步骤：将结构如式(I-3)所示的含铂化合物、结构如式(III-3)所示的单官能化亲水聚合物、可选的结构如式(II-3)所示的药物化合物以及可选的如式(IV-3)所示化合物在有机溶剂中混合，进行开环聚合反应；
Figure PCTCN2022108093-appb-000001
其中，U 1和U 2各自独立地为碳中心三价基团，D Pt为铂类药物单元(可以为铂类药物活性成分或其前药的残基)；
mPEG为甲氧基聚乙二醇链段(通过O与L 5连接)；L 5独立地为二价连接基或无；Z 5独立地为-NH-或-C(＝O)-；
F 5为-NH 2、-COOH、
Figure PCTCN2022108093-appb-000002
优选为-NH 2；
U 3独立地为碳中心三价基团，L R独立地为响应性连接基，L 4独立地为二价连接基或无，D T为第二药物单元(可以为抗肿瘤药物活性成分或其前药的残基)；其中，L R能够在外界刺激下发生键断裂；
P E为R E或被保护的R E，在所述开环聚合反应中不具备反应性；R E独立地为H或R 0；其中，R 0为不含药物单元的端基；
进一步优选地，所述开环聚合反应于无水条件下进行；
更进一步优选地，开环聚合反应温度为15～40℃，更优选地，开环聚合反应时间为24～96h。该聚合反应利用双N-羧酸酐(NCA)参与的开环聚合反应，通过“一锅法”得到单分子纳米聚合物，该单分子纳米聚合物可以在水性介质中无需自组装即可形成核壳结构的胶束，提供可响应性释放药物活性成分的药物递送系统，用于肿瘤疾病的治疗。
本申请的另一个目的在于提供一种胞内还原微环境响应活化型双药单分子纳米聚合物前药，该纳米聚合物前药可以用作同时递送两种药物活性成分的平台技术。
本申请的另一个目的在于提供一种用于制备胞内还原微环境响应活化型双药单分子纳米聚合物前药的方法。
本申请的另一目的在于提供一种载药单分子纳米聚合物胶束，其组成选自如下任一种：前述的载药单分子纳米聚合物，前述的制备方法制备得到的载药单分子纳米聚合物，前述的双药单分子纳米聚合物前药，和前述的制备方法制得的双药单分子纳米聚合物前药；所述载药单分子纳米聚合物胶束具有核壳结构，外壳结构为亲水性聚合物链形成的亲水层，所包载的药物单元位于内核中。
本申请提供的载药单分子纳米聚合物可以在聚合反应的同时原位形成具有核壳结构的纳米聚合物胶束，包括位于外壳的亲水聚合物链以及位于内核的药物单元。
本申请提供的铂单药单分子纳米聚合物可以在聚合反应的同时原位形成具有核壳结构的纳米聚合物胶束，包括位于外壳的亲水聚合物链以及位于内核的铂类药物单元。
本申请提供的双药单分子纳米聚合物可以在聚合反应的同时原位形成具有核壳结构的纳米聚合物胶束，包括位于外壳的亲水聚合物链以及位于内核的铂类药物单元及第二药物单元。
本申请的再一个目的在于提供前述载药单分子纳米聚合物作为前药的用途。该载药单分子纳米聚合物能够进入到细胞内部，感知细胞内微环境，响应性地释放药物活性成分，产生细胞毒性，抑制肿瘤细胞的生长。
本申请的再一个目的在于提供一种双药单分子纳米聚合物用于递送药物活性成分的用途或在制备药物递送系统中的用途。药物成活性分可以释放自前述铂类药物单元以及前述可选的第二药物单元。药物活性成分可以为铂类药物活性成分及可选的抗肿瘤药物活性成分。
本申请的再一个目的在于提供前述载药单分子纳米聚合物，前述的制备方法制备得到的载药单分子纳米聚合物，前述的双药单分子纳米聚合物前药，或前述的制备方法制备得到的双药单分子纳米聚合物前药在制备用于治疗肿瘤疾病的药物中的用途。
本申请的再一目的在于提供一种药物递送系统，它包含载药单分子纳米聚合物胶束，该载药单分子纳米聚合物胶束包含前述载药单分子纳米聚合物或前述的制备方法制备得到的载药单分子纳米聚合物；
优选地，
所述亲水性聚合物链位于所述载药单分子纳米聚合物胶束的外壳；
所述铂类药物单元以及所述第二药物单元均位于所述载药单分子纳米聚合物胶束的内核。
本申请的再一目的在于提供一种药物递送系统，它包含双药单分子纳米聚合物胶束，该双药单分子纳米聚合物胶束包含与亲水性聚合物连接的聚氨基酸，其中在该聚氨基酸的重复单元的α碳上键合了铂类药物活性成分的前药部分和抗肿瘤药物活性成分的前药部分；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
本申请的再一个目的在于提供一种胞内还原微环境响应活化型双药单分子纳米聚合物前药用于递送药物活性成分的用途或在制备药物递送系统中的用途。
本申请的另一个目的在于提供铂类药物活性成分的双NCA单体和抗肿瘤药物活性成分的单NCA单体在制备单分子纳米聚合物前药或药物递送系统中的用途；优选地，所述抗肿瘤药物活性成分的分子结构中含有游离羟基或游离氨基。
本申请人经过广泛而深入的研究，出乎意料地发现生物相容性具有末端氨基或羧基的亲水性聚合物与含铂的双N-羧酸酐(NCA)单体和抗肿瘤药物活性成分的单NCA单体(抗肿瘤药物活性成分的分子中优选含有游离羟基、游离氨基或其组合，从而能够偶联上NCA端基)可以原位形成核壳结构的单分子纳米聚合物(可作为前药，因此，也可记为单分子纳米聚合物前药)，该聚合物前药能够偶联两种抗肿瘤药物，在体内和体外稳定性好，分散性好，粒径均一，无毒副作用，以及在胞外不会释放药物活性成分而 在胞内呈现出触发性释放药物活性成分，且其制备方法操作简便、反应温和、成本低廉和环境友好。
本申请还提供前述载药单分子纳米聚合物，前述的制备方法制备得到的载药单分子纳米聚合物，前述的双药单分子纳米聚合物前药，前述的制备方法制得的双药单分子纳米聚合物前药，前述载药单分子纳米聚合物胶束，或前述的药物递送系统在制备用于治疗肿瘤疾病的药物中的用途。
本申请还提供一种具有核壳结构的胞内还原微环境响应活化型双药单分子纳米聚合物前药、药物递送系统及其制备方法和用途。
本申请克服了传统技术中聚合物前药的所述缺点。
下面从不同方面概述本申请，这些方面及其任何变化形式中所描述的发明相互独立又彼此关联，一起构成本申请的内容。
一方面，本申请提供具有核壳结构的双药单分子纳米聚合物前药，其中内核含有铂类药物分子结构单元和药物活性分子结构单元以及聚氨基酸结构单元，所述结构单元之间通过共价键连接；外壳为生物相容性亲水性聚合物(如，聚乙二醇等)；优选地，所述药物活性分子的分子中含有游离羟基、游离氨基或两者的组合。本申请的双药单分子纳米聚合物前药可以抑制药物活性成分在血液循环中的非特异性反应，进入细胞内部后具有响应胞内还原微环境而触发释放所述药物活性成分的功能。
在一些实施方案中，本申请提供一种双药单分子纳米聚合物前药，其由具有末端氨基的亲水性聚合物、铂类药物活性成分的双NCA单体和抗肿瘤药物活性成分的单NCA单体形成；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
在一些实施方案中，本申请提供一种双药单分子纳米聚合物前药，其包含与亲水性聚合物连接的聚氨基酸，其中在该聚氨基酸的重复单元的α碳上键合了铂类药物活性成分的前药部分和抗肿瘤药物活性成分的前药部分；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
本申请的另一方面，提供一种药物递送系统，它包含双药单分子纳米聚合物胶束，该双药单分子纳米聚合物胶束包含前述双药单分子纳米聚合物或前述的制备方法制备得到的双药单分子纳米聚合物；
优选地，所述亲水性聚合物链位于所述双药单分子纳米聚合物胶束的外壳；
所述铂类药物单元以及所述第二药物单元均位于所述双药单分子纳米聚合物胶束的内核。
另一方面，本申请提供一种药物递送系统，它包含双药单分子纳米聚合物胶束，该聚合物胶束具有核壳结构，其中内核含有铂类药物分子结构单元和药物活性分子结构单元以及聚氨基酸结构单元，所述结构单元之间通过共价键连接；且外壳为生物相容性亲水性聚合物(如，聚乙二醇等)；优选地，所述药物活性分子的分子中含有游离羟基、游离氨基或游离羟基与游离氨基的组合。
在一些实施方案中，本申请提供一种药物递送系统，它包含双药单分子纳米聚合物胶束，该聚合物胶束由具有末端氨基的亲水性聚合物、铂类药物活性成分的双NCA单 体和抗肿瘤药物活性成分的单NCA单体形成；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
在一些实施方案中，本申请提供一种药物递送系统，它包含双药单分子纳米聚合物胶束，该双药单分子纳米聚合物胶束包含与亲水性聚合物连接的聚氨基酸，其中在该聚氨基酸的重复单元的α碳上键合了铂类药物活性成分的前药部分和抗肿瘤药物活性成分的前药部分；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
再一方面，本申请提供一种制备本申请双药单分子纳米聚合物前药的方法，该方法包括如下步骤：
(1)在合适反应条件下，合成抗肿瘤药物活性成分的单NCA单体，优选地，所述抗肿瘤药物活性成分的分子结构中含有游离羟基或游离氨基，
(2)在合适反应条件下，合成铂类药物活性成分的双NCA单体，
(3)在合适反应条件下，使步骤(1)和步骤(2)得到的单体与具有末端氨基的亲水性聚合物反应，得到本申请的双药单分子纳米聚合物前药，以及
(4)分离和纯化等处理所得到的双药单分子纳米聚合物前药。
在一些实施方案中，通过一步一锅开环聚合法制备本申请的双药单分子纳米聚合物前药，即本申请提供一种制备双药单分子纳米聚合物前药的方法，该方法包括如下步骤：
(1)在合适反应条件下，使抗肿瘤药物活性成分的单NCA单体(优选地，所述抗肿瘤药物活性成分的分子结构中含有游离羟基或游离氨基)、铂类药物活性成分的双NCA单体与具有末端氨基的亲水性聚合物一起反应，得到所述的双药单分子纳米聚合物前药，以及
(2)分离和纯化等处理所得到的双药单分子纳米聚合物前药。
本申请的这种一步一锅开环聚合法规避了传统自组装纳米制剂的诸多弊端。
再一方面，本申请提供一种适用于制备单分子纳米聚合物前药的铂类药物活性成分的双NCA单体。
再一方面，本申请提供一种适用于制备单分子纳米聚合物前药的分子结构中含有游离羟基或游离氨基的抗肿瘤药物活性成分的单NCA单体。
再一方面，本申请提供一种将药物活性成分同时递送至目标部位的方法，该方法包括将目标药物活性成分制备成单分子纳米聚合物前药以及将有效量的该单分子纳米聚合物前药给予有需要的患者。
在一些实施方案中，目标药物活性成分包括铂类药物活性成分或其前药。
再一方面，本申请提供一种将两种药物活性成分同时递送至目标部位的方法，该方法包括将这两种药物活性成分制备成单分子纳米聚合物前药以及将有效量的该单分子纳米聚合物前药给予有需要的患者。
在一些实施方案中，本申请提供一种将两种药物活性成分同时递送至目标部位的方法，该方法包括将这两种药物活性成分制备成单分子纳米聚合物前药以及将所制得的单分子纳米聚合物前药给予有需要的患者，其中所述两种药物活性成分中的一种是铂类药物，另一种为分子结构中含有游离羟基或游离氨基的抗肿瘤药物活性成分。
再一方面，本申请提供抗肿瘤药物活性成分的单NCA单体和铂类药物活性成分的双NCA单体在制备用于将所含药物活性成分递送至目标部位的聚合物前药给药系统中的用途；优选地，所述抗肿瘤药物活性成分的分子结构中含有游离羟基或游离氨基。
在一些实施方案中，本申请提供抗肿瘤药物活性成分的单NCA单体和铂类药物活性成分的双NCA单体在制备单分子纳米聚合物前药胶束中的用途；优选地，所述抗肿瘤药物活性成分的分子结构中含有游离羟基或游离氨基。
再一方面，本申请提供双药单分子纳米聚合物前药在制备用于治疗相应疾病的药物中的用途，其中所述的双药单分子纳米聚合物前药包含与亲水性聚合物连接的聚氨基酸，其中在该聚氨基酸的重复单元的α碳上键合了铂类药物活性成分的前药部分和抗肿瘤药物活性成分的前药部分；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
再一方面，本申请提供一种联合用药治疗有需要的患者中肿瘤的方法，该方法将治疗有效量的单分子纳米聚合物药物给予所述患者，其中所述的单分子纳米聚合物药物包含与亲水性聚合物连接的聚氨基酸，其中在该聚氨基酸的重复单元的α碳上键合了铂类药物活性成分的前药部分和抗肿瘤药物活性成分的前药部分；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
本申请人经实验研究出乎意料地发现，本申请的纳米聚合物前药或聚合物前药纳米胶束整合了纳米制剂的优点(包括：长血液循环时间，低肝脏器官摄取，病灶部位靶向富集的潜力)以及前药的优点(降低活性药物的早期失活，精准的药物活化)，最终有利于提高活性药物在靶向部位的时空浓度，从而增强药效，同时降低药物对于非靶向部位的潜在毒副作用。
本申请的聚合物前药纳米胶束相对于传统技术的聚合物前药纳米胶束具有更好的稳定性。不受特定理论的限制，导致本申请聚合物前药纳米胶束的稳定性优势可能是如下因素：药物全化学键合，耐受离心、超滤、水热、超声等物理处理，保证纳米胶束的化学组成稳定性；喜树碱等药物间疏水作用形成内核，确保核壳的胶束结构；以及以铂为桥交联聚氨基酸实现内核的交联，实现了胶束结构的稳定性。
另外，本申请聚合物前药纳米胶束的生产工艺也具有技术优势。不受特定理论的限制，导致本申请聚合物前药纳米胶束的生产工艺具有所述技术优势可能是如下因素：药物全化学键合，载药量精细可控，突破自主装体系批次稳定性难题，且无游离药物，可溶液储存；单个纳米粒子即单个分子，冻干复溶工艺简单，技术要求低；以及“一锅法”合成纳米胶束，无需薄膜水化，纳米沉淀等复杂制备和纯化工艺。
再有，本申请聚合物前药纳米胶束的在载药和释药方面同样具有优势。不受特定理论的限制，导致本申请聚合物前药纳米胶束在载药和释药方面具有优势可能是如下因素：药物全化学键合，保外(血液循环、细胞外基质)无药物泄漏；以及胞内触发释放，一方面释放提高时空药物浓度，强化药物功效，解决自助装纳米制剂被动缓慢释药的弊端，另一方面，双药均响应统一胞内微环境，协同释放，最大化双药功效。
最后，本申请体的聚合物前药纳米胶束可以包含双药，具有以下优势：双药靶点不同，克服了抗药性；消耗胞内耐药性谷胱甘肽，克服了耐药性；以及铂类药物和另一种 抗肿瘤药物如喜树碱分别以不同的作用机理产生抗肿瘤活性，具有优异的协同作用。
本申请的一个或多个实施例的细节在下面的附图和描述中提出。本申请的其他特征、目的和优点将从说明书、附图以及权利要求书变得明显。
附图说明
为了更清楚地说明本申请实施例中的技术方案、更完整地理解本申请及其有益效果，下面将对实施例描述中所需要使用的附图作简单的介绍。显而易见地，下面描述中的附图仅仅是本申请的一些实施例，对本领域技术人员来说，在不付出创造性劳动的前提下，还可以根据这些附图获得其它的附图。
图1是显示本申请双药(铂类药物单元+第二药物单元)单分子纳米聚合物前药胶束组成的示意图。
图2是显示本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束组成的示意图。
图3中的III-A、III-B、III-C和III-D分别为用于合成本申请产物的中间产物的质谱图。
图4中的IV-A、IV-B、IV-C、IV-D和IV-E分别为用于合成本申请产物的中间产物的 1H NMR谱图。
图5中的V-A和V-B分别为显示本申请双药(顺铂+喜树碱)单分子纳米聚合物前药及其中间产物的分子量的分子排阻色谱图。
图6为表征本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束和对照胶束的粒径和多分散指数的动态光散射(DLS)图。
图7为本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束(VII-A)和对照胶束(VII-B)的透射电子显微镜图像。
图8为本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束(VIII-A)和对照胶束(VIII-B)的冻干前后的透射电子显微镜图像。
图9为使用荧光关联光谱法测试的本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束(A)和对照胶束(B)的胶体动力学特征。
图10为表征本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束的小角X射线散射图。
图11为表征本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束与对照胶束的粒径分布的动态光散射图。
图12为本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束与对照胶束的高效液相色谱图。
图13显示了本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束的释药行为。
图14显示了对照胶束的体外模拟释药行为。
图15显示了本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束和对照胶束的细胞毒性。
图16显示了本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束(A)和对照胶束(B)的药物动力学研究结果。
图17显示了本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束和对照胶束的母体药物累积量试验结果。
图18是本申请双药(顺铂+喜树碱)单分子纳米聚合物前药胶束和对照胶束的抑癌试验结果。
图19是本申请双药(顺铂+喜树碱)单分子纳米聚合物前药的 1H NMR谱图。
图20是表征本申请双药(顺铂+紫杉醇)单分子纳米聚合物前药胶束的粒径和多分散指数的动态光散射图。
图21显示了本申请双药(顺铂+紫杉醇)单分子纳米聚合物前药胶束的释药行为。
图22显示了本申请双药(顺铂+紫杉醇)单分子纳米聚合物前药胶束的透射电子显微镜图像。
图23是本申请双药(顺铂+瑞喹莫德)单分子纳米聚合物前药胶束的粒径和多分散指数的动态光散射图。
图24显示了本申请双药(顺铂+瑞喹莫德)单分子纳米聚合物前药胶束的释药行为。
图25显示了本申请双药(顺铂+瑞喹莫德)单分子纳米聚合物前药胶束的透射电子显微镜图像。
图26表示本申请实施例10中铂单药单分子纳米聚合物的凝胶渗透色谱(SEC)测试(A)及动态光散射(DLS)测试(B)的结果图，采用顺铂。
图27表示本申请实施例10中铂单药单分子纳米聚合物胶束在冻干复溶前(A)及冻干复溶后(B)的透射电镜(TEM)测试图。
图28表示本申请实施例11中制备的NCA-DACHPt-NCA的质谱图。
图29表示本申请实施例11中铂单药单分子纳米聚合物的凝胶渗透色谱(SEC)测试(A)及动态光散射(DLS)测试(B)的结果图，采用DACHPt。
图30表示本申请实施例11中铂单药单分子纳米聚合物胶束在冻干复溶前(A)及冻干复溶后(B)的透射电镜(TEM)测试图。
图31表示本申请实施例12中制备的PTX-ss-NCA的 1H NMR谱图。
图32表示本申请实施例12中制备的双药(顺铂+紫杉醇)单分子纳米聚合物的DLS测试结果，(A)不超声处理，(B)超声处理。
图33表示本申请实施例12中制备的双药(顺铂+紫杉醇)单分子纳米聚合物胶束在冻干复溶前(A)及冻干复溶后(B)的透射电镜(TEM)测试图。
图34表示本申请实施例12中制备的双药(顺铂+紫杉醇)单分子纳米聚合物胶束在不同条件下的释药行为结果图。
图35表示本申请实施例13中制备的R848-ss-NCA的 1H NMR谱图。
图36表示本申请实施例13中制备的双药(顺铂+R848)单分子纳米聚合物的DLS测试结果，(A)不超声处理，(B)超声处理。
图37表示本申请实施例13中制备的双药(顺铂+R848)单分子纳米聚合物胶束在冻干复溶前(A)及冻干复溶后(B)的透射电镜(TEM)测试图。
图38表示本申请实施例13中制备的双药(顺铂+R848)单分子纳米聚合物胶束在不同条件下的释药行为结果图。
图39表示本申请实施例14中制备的MMAE-ss-NCA的 1H NMR谱图。
图40表示本申请实施例14中制备的双药(顺铂+MMAE)单分子纳米聚合物的DLS测试结果。
图41表示本申请实施例14中制备的双药(顺铂+MMAE)单分子纳米聚合物胶束在冻干复溶前(A)及冻干复溶后(B)的透射电镜(TEM)测试图。
图42表示本申请实施例14中制备的双药(顺铂+MMAE)单分子纳米聚合物胶束在不同条件下的释药行为结果图。
具体实施方式
上文已从一般方面概述了本申请，下面将结合实施例进一步详细描述本申请。
下面结合附图、实施方式和实施例，对本申请作进一步详细的说明。应理解，这些实施方式和实施例仅用于说明本申请而不用于限制本申请的范围，提供这些实施方式和实施例的目的是使对本申请公开内容理解更加透彻全面。还应理解，本申请可以以许多不同的形式来实现，并不限于本文所描述的实施方式和实施例，本领域技术人员可以在不违背本申请内涵的情况下作各种改动或修改，得到的等价形式同样落于本申请的保护范围。例如，作为一个实施方式的部分而说明或描述的特征可以以合适的方式组合于另一实施方式中，以产生新的实施方式。此外，在下文的描述中，给出了大量具体的细节以便提供对本申请更为充分地理解，应理解，本申请可以无需一个或多个这些细节而得以实施。
术语
本文所使用的术语“和/或”、“或/和”、“及/或”的选择范围包括两个或两个以上相关所列项目中任一个项目，也包括相关所列项目的任意的和所有的组合，所述任意的和所有的组合包括任意的两个相关所列项目、任意的更多个相关所列项目、或者全部相关所列项目的组合。需要说明的是，当用至少两个选自“和/或”、“或/和”、“及/或”的连词组合连接至少三个项目时，应当理解，在本申请中，该技术方案毫无疑问地包括均用“逻辑与”连接的技术方案，还毫无疑问地包括均用“逻辑或”连接的技术方案。比如，“A及/或B”包括A、B和A+B三种并列方案。又比如，“A，及/或，B，及/或，C，及/或，D”的技术方案，包括A、B、C、D中任一项(也即均用“逻辑或”连接的技术方案)，也包括A、B、C、D的任意的和所有的组合，也即包括A、B、C、D中任两项或任三项的组合，还包括A、B、C、D的四项组合(也即均用“逻辑与”连接的技术方案)。
本申请中，涉及“多个”、“多种”等描述，如无特别限定，指在数量上大于等于2。比如，“一种或多种”从数量上等于1或≥2，可以为一种、两种或更多种。
本文中所使用的“其组合”、“其任意组合”、“其任意组合方式”等中包括所列项目中任两个或任两个以上项目的所有合适的组合方式。
本文中，“合适的组合方式”、“合适的方式”、“任意合适的方式”等中所述“合适”，以能够实施本申请的技术方案、解决本申请的技术问题、实现本申请预期的技术效果为准。
本文中，“优选”、“更好”、“更佳”、“为宜”、“另优选地”等仅为描述效果更好的实 施方式或实施例，应当理解，并不构成对本申请保护范围的限制。如果一个技术方案中出现多处“优选”，如无特别说明，且无矛盾之处或相互制约关系，则每项“优选”各自独立。在一个技术方案中，当“优选”与一个或多个“另优选”同时出现时，其中的任两个或任意更多个“优选”之间可以不结合，也可以结合形成不同的特征。
本申请中，“进一步”、“更进一步”、“特别”等用于描述目的，表示内容上的差异，但并不应理解为对本申请保护范围的限制。
本申请中所使用的术语“含有”、“包含”和“包括”是同义词，其是包容性或开放式的，不排除额外的、未被引述的成员、元素或方法步骤。
本申请中，以开放式描述的技术特征中，包括所列举特征组成的封闭式技术方案，也包括包含所列举特征的开放式技术方案。
本申请中用端点表示的数值范围包括该范围内所包含的所有数值及分数，以及所引述的端点。
本申请中，涉及到数值区间(也即数值范围)，如无特别说明，可选的数值分布在上述数值区间内视为连续，且包括该数值范围的两个数值端点(即最小值及最大值)，以及这两个数值端点之间的每一个数值。该数值区间中的“数值”可以为任意的定量值，比如数字、百分比、比例等。“数值区间”允许广义地包括百分比区间，比例区间，比值区间等定量区间。如无特别说明，当数值区间仅仅指向该数值区间内的整数时，包括该数值范围的两个端点整数，以及两个端点之间的每一个整数。此外，当提供多个范围描述特征或特性时，可以合并这些范围。换言之，除非另有指明，否则本文中所公开之范围应理解为包括其中所归入的任何及所有的子范围。
本申请中，涉及数据范围的单位，如果仅在右端点后带有单位，则表示左端点和右端点的单位是相同的。比如，50～1000Da表示左端点50和右端点1000的单位都是Da。
在本申请提及的所有文献都在本申请中引用作为参考，就如同每一篇文献被单独引用作为参考那样。除非和本申请的发明目的、技术方案或二者相冲突，否则，本申请涉及的引用文献以全部内容、全部目的被引用。本申请中涉及引用文献时，相关技术特征、术语、名词、短语等在引用文献中的定义也一并被引用。本申请中涉及引用文献时，被引用的相关技术特征的举例、优选方式也可作为参考纳入本申请中，但以能够实施本申请为限。应当理解，当引用内容与本申请中的描述相冲突时，以本申请为准或者适应性地根据本申请的描述进行修正。
为准确理解本申请中所使用的术语，下面特别定义部分术语的含义。对于在此没有特别定义的术语，它们具有本领域技术人员普遍理解和接受的含义。如果在此所定义的某个术语的含义与本领域技术人员普遍理解和接受的含义不一致，则该术语的含义以在此所定义的含义为准。
本申请中使用的术语“水溶性聚合物”是指在室温可溶于水的药学上可接受的任何生物相容性聚合物。
本申请中使用的术语“具有末端氨基的水溶性聚合物”是指分子结构中的一个末端是氨基(-NH 2)的如上所定义的水溶性聚合物。
本申请中使用的术语“药学上可接受的辅料”是指可以被包含在本申请的纳米聚合 物前药胶束组合物中并且不对患者造成明显有害药理学作用的辅助物质，它可与“药学上可接受的赋形剂”或“药学上可接受的载体”互换使用。
本申请中所用的术语“治疗有效量”意指本申请的中药制剂的用量，在对受试者给予本申请的中药制剂治疗新型冠状病毒感染性疾病时该用量足以实现所期望的对疾病的治疗效果。“治疗有效量”可以根据实际所用的制剂形式、疾病的症状及其严重性和所治疗受试者的年龄、体重等进行调整。
本申请中所用的术语“患者”是指患有或易患有可以通过施用本申请的双药单分子纳米聚合物前药来预防或治疗的疾病的活生物，包括人类和哺乳动物，优选人类。
在本申请中，“肿瘤”以其最宽的含义理解，指非正常过度生长的组织。“癌”或“癌症”指恶性肿瘤。
本说明书中提及水溶性聚合物(如PEG)的分子量时，除非另外规定，否则所有对分子量的提及均指重均分子量。
本申请中公开的所有数值范围包括其端值在内，且包括没有明确列出的该范围内的任何小范围。
传统纳米药物制剂(如Bind-14，NC-6300)通过药物分子与递送分子(两亲性分子：脂质分子，嵌段共聚物等)之间的物理作用(如：疏水作用、静电作用等)以自组装的方式形成，纳米药剂在到达肿瘤病灶后，药物释放往往依靠被动扩散的方式实现，这种缓慢药释行为会造成局部瞬时药物浓度难以达到有效水平。因此，传统纳米制剂虽然对于肿瘤病灶具有较高富集潜力，然而其缓慢的药释速率导致传统纳米制剂对肿瘤细胞杀伤效果甚至低于游离小分子药物。同时，传统纳米制剂的药释行为也发生在血液循环中，这种药物早期泄漏会降低药物对靶病灶的生物利用度，造成非靶病灶的毒性。因此，如何控制药物在病灶部位选择性地控释和快释是提升纳米药物制剂高效治疗肿瘤的关键。
在本申请的一些实施方案中，在载药单分子纳米聚合物或在双药单分子纳米聚合物前药中的铂(Pt)为四价铂，具有八面体空间结构，化学反应惰性高，在血浆及正常组织化学结构稳定，因此，在体内输送过程中，系统性毒副反应小，同时，四价铂与二价铂无交叉耐药性，进入肿瘤细胞内，高还原性环境能使四价铂还原释放出活性二价铂物种，进而产生细胞毒性。相比于二价铂，二价铂的化学反应学活性高，能与血浆中的蛋白等结合，因此，二价铂的生物利用度低；同时，二价铂能与含巯基生物分子发生解毒反应，造成铂药的体内失活；另外，二价铂交叉耐药性严重制约了其临床的疗效和长期实用。
在本申请的一个方面，提供一种载药单分子纳米聚合物，其包含多条聚氨基酸链，所述多条聚氨基酸链的链间通过多个二价连接基L Pt共价相连使所述多条聚氨基酸链构成非线性骨架，至少一条所述聚氨基酸链的端部连接有亲水性聚合物链；其中，所述二价连接基L Pt的线性骨架中包含铂原子，所述铂原子参与构成铂类药物单元，所述铂类药物单元可以为铂类药物活性成分或其前药的残基；
可选地，所述聚氨基酸链的侧基接枝有第二药物单元；其中，所述第二药物单元可以为抗肿瘤药物活性成分或其前药的残基。
在载药单分子纳米聚合物的一些实施方案中，所述抗肿瘤药物活性成分或其前药的 残基每次出现，独立地通过响应性连接基L R连接至氨基酸重复单元，所述响应性连接基L R能够在外界刺激下发生键断裂。
本申请所用的“聚氨基酸链”表示由多个氨基羧酸分子的-NH 2端和-COOH端通过-CO-NH-键依次连接而成的聚合物链。在一些较佳的实施例中，所述聚氨基酸链由α氨基酸单元组成，进一步地，所述聚氨基酸链的主链由-NH-C-C(＝O)-构成。更进一步地，所述第二药物单元每次出现，独立地连接到相应的α氨基酸单元的α碳。本申请所用的“α氨基酸”表示具有NH 2-CR CR E-COOH，其中R C可以为H或者为不影响NCA开环聚合物的非氢原子或基团，R E可以为氢或R 0，其中，R 0为不含药物单元的端基。R 0还可以选择下文中的定义。
本申请所用的“氨基酸”表示含有至少一个-NH 2和至少一个-COOH的化合物，可以为天然氨基酸(如赖氨酸)，也可以为非天然氨基酸(如鸟氨酸)。构成本申请聚氨基酸链的结构单元的氨基酸单元可以为α氨基酸单元。
本申请所用“非线性”表示支化的或者交联的拓扑结构。本申请中，一个二价连接基L Pt可以与两条聚氨基酸链共价连接，形成两个支化点，通过调节L Pt与聚氨基酸链的相对比例，可以控制合适支化程度，具体地，可以通过调节载药单分子纳米聚合物中L Pt数量相对于氨基酸单元总数量的比例来调控。一条聚氨基酸链连接的L Pt的平均数量越多，支化点越多。支化程度太低导致柔性太高，支化程度较高时形成交联的三维网络，交联度太高会导致刚性较大，因此，支化点的数量过少或过多都会导致载药单分子纳米聚合物都会影响纳米胶束的形成及药物释放性质。
本申请所用的“亲水性聚合物链”或“亲水性聚合物”指能够在水中溶胀或溶解。本申请所用“聚合物”具有至少两个结构单元，其分子量没有特别限定，可以大于等于1000Da，也可以小于等于1000Da。本申请中，“亲水性聚合物”与“亲水聚合物”具有相同含义，可以互换使用。本申请中，“亲水性聚合物链”与“亲水聚合物链”具有相同含义，可以互换使用。
在本申请中，亲水性聚合物链可以连接在聚氨基酸链的N端或C端。
在一些实施方式中，亲水性聚合物链连接在聚氨基酸链的C端，可通过酰胺键(-CONH-)连接。
在一些实施方式中，亲水性聚合物链连接在聚氨基酸链的N端，可通过酰胺键(-NH-CO-)或氨基甲酸酯基(-NH-COO-)连接。
在本申请中，“价态”简称“价”，是各种元素的一个原子或原子团、基(根)与其他原子相互化合的数目。本领域技术人员可以理解“一价基团”、“二价基团”、“三价基团”、……等价态所定义基团的含义。
在本申请中，某一物质的“残基”通常指该物质失去至少一个原子所剩余的结构。比如，-NH-C(CH 2CH 2CH 2CH2NH-)-C(＝O)-为赖氨酸的三价残基。此外，含铂物质与两个相邻原子(如O等)共价连接后的状态也记为该含铂物质的残基。比如，
Figure PCTCN2022108093-appb-000003
分别对应
Figure PCTCN2022108093-appb-000004
的残基。
构成聚氨基酸链的氨基酸单元的侧基可以连接L Pt而形成支化点，可以连接第二药物单元，还可以为不连接药物单元的氢原子或游离端基R 0。
聚氨基酸链的端部可以连接有亲水性聚合物链。制备载药单分子纳米聚合物时，通过端基亲水性聚合物链的引入，可以调控聚氨基酸链的链长，从而调节载药单分子纳米聚合物的尺寸。
通过综合控制连接L Pt的氨基酸单元的数量、连接第二药物单元的氨基酸单元的数量，带游离端基(不连接药物单元)的氨基酸单元的数量以及亲水性聚合物链的数量，可以控制载药单分子纳米聚合物具有合适的支化密度、合适的载药量、合适的不同药物比例以及合适的聚氨基酸链长，从而使载药单分子纳米聚合物具有合适的尺寸，并具有合适的柔性和亲疏水单元比例，进一步结合亲水性聚合物链在聚氨基酸链端部的位置设计，使载药单分子纳米聚合物能够在水性介质中无需自组装就可形成纳米尺度(如直径20～120nm，进一步如30～120nm)的单分子聚合物胶束，亲水性聚合物链形成外壳，药物成分被包裹于内核。纳米胶束的尺寸(包括内核尺寸、外壳厚度、粒径、平均直径等)可以通过包括但不限于动态光散射(DLS)、小角X射线散射(SAXS)、透射电镜(TEM)法等方法直接测得或根据测试结果换算得到。
本申请所用的“水性介质”指含水体系，可以为水或水溶液。可以为缓冲溶液、体外模拟液、细胞培养液、组织培养液等体外体系，还可以为血液、组织液等体内体系。
该载药单分子纳米聚合物中的药物单元可以仅为铂类药物单元，此时，可记为铂单药单分子纳米聚合物。
该载药单分子纳米聚合物中，第二药物单元是可选的，可以含有，也可以不含有。在本申请中，如无特别限定，第二药物单元的亲疏水性没有特别限定，可以为亲水性药物单元，也可以为疏水性药物单元。
在载药单分子纳米聚合物的一些实施方案中，本申请所用的第二药物单元不同于铂类药物单元，从而可以作用于不同的靶点。
在载药单分子纳米聚合物的一些实施方案中，该载药单分子纳米聚合物中的药物单元包括铂类药物单元和第二药物单元，此时，可记为双药单分子纳米聚合物。可参考图1，图中还显示了响应于外界刺激条件的药物释放过程。铂类药物单元为顺铂的残基及第二药物单元为喜树碱活性成分的残基，并响应于胞内还原微环境的药物释放示意图如图2所示。
该载药单分子纳米聚合物通过二价含铂连接基L Pt将多条聚氨基酸链构建成非线性骨架，至少一条聚氨基酸链的端部连接有亲水性聚合物链，L Pt中的铂原子参与构成铂类药物单元(可以为铂类药物活性成分或其前药的残基)。通过调节L Pt的分布密度可控制该载药单分子纳米聚合物具有支化或适度交联的三维结构，进一步结合亲水性聚合物链 在聚氨基酸链端部的位置设计，使载药单分子纳米聚合物能够在水性介质中无需自组装就可形成具有核壳结构的单分子纳米聚合物胶束，亲水性聚合物链分布于外壳，药物成分被包载于内核。该载药单分子纳米聚合物可以仅装载铂类药物单元，构成铂单药单分子纳米聚合物；该载药单分子纳米聚合物可以还接枝第二药物单元(可以为抗肿瘤药物活性成分或其前药的残基)，该第二药物单元可以接枝在聚氨基酸链的侧基，此时可构成双药单分子纳米聚合物。铂类药物单元及第二药物单元的相对含量可以通过控制相应单体的投料量灵活调节。L Pt的分布密度可通过调节非支化氨基酸单体与L Pt支化氨基酸单体的投料比例进行调节，在非支化氨基酸单体中还可以灵活调节包含第二药物单元的氨基酸单体所占比例。该载药单分子纳米聚合物在体内和体外稳定性好，分散性好，粒径均一，无毒副作用，而且在胞外不会释放药物活性成分而在胞内呈现出触发性释放药物活性成分，此外，可采用操作简便、反应温和、成本低廉和环境友好的制备方法获得。
在本申请中，“L Pt支化氨基酸单体”指参与构成前述非线性骨架的支化点的氨基酸单体，如本文中式(I-3)所示结构化合物(携带铂类药物单元)。
在本申请中，“非支化氨基酸单体”指不参与构成前述非线性骨架的支化点的氨基酸单体，比如，本文中式(II-3)所示结构化合物(携带第二药物单元)、(IV-3)所示结构的化合物。
在双药单分子纳米聚合物的一些实施方案中，任一条所述聚氨基酸链的主链由多个
Figure PCTCN2022108093-appb-000005
所示结构通过-C(＝O)-NH-键依次键连而成，任一个所述
Figure PCTCN2022108093-appb-000006
中的U独立地为碳中心三价基团，任一个所示“*”端独立地连接到所述二价连接基L Pt，或者连接到氢原子或一价侧基R A；所述一价侧基R A中为含有所述第二药物单元的含药侧链，或者为不含药物单元的端基R 0。
本申请中，“碳中心三价基团”表示由碳原子提供支化点的三价基团。
在一些实施方案中，U每次出现，可以为CR C，其中，R C可以为H或者为不影响NCA开环聚合物的非氢原子或基团。
在一些实施方案中，每一个
Figure PCTCN2022108093-appb-000007
中的U均为CH。
在一些实施方案中，任一个所示“*”端独立地连接到所述二价连接基L Pt，或者连接到所述一价侧基R A。
在另一些实施方案中，任一个所示“*”端独立地连接到所述二价连接基L Pt，或者连接到含有所述第二药物单元的含药侧链。
本申请中，如无其他限定，“烷基”指一价烷基，“亚烷基”指二价烷基，“连接基”指价态≥2的原子或基团，“二价连接基”指价态为2的连接基，“端基”指价态为1的 原子或基团。在本申请中，“一价烷基”指烷烃化合物失去任意一个氢原子形成的残基，“亚烷基”指烷烃化合物失去任意两个氢原子形成的残基，这里的“烷烃化合物”指的是由碳原子和氢原子构成的饱和烃，可以为链式的(即不含环)，也可以含饱和环(如己烷)，如果没有特别说明，可以优选为链式的。
在一些实施方案中，R 0每次出现，独立地选自19种天然氨基酸(脯氨酸除外)的侧基、19种天然氨基酸(脯氨酸除外)合适侧基的离子形式以及鸟氨酸的侧基中任一种。
在一些实施方案中，R 0每次出现，独立地选自19种天然氨基酸(脯氨酸除外)以及鸟氨酸的侧基中任一种。
在一些实施方案中，R 0每次出现，独立地选自如下任一种基团：C 1-6烷基、-L A-COOH、-L A-NH 2、-L A-OH、-L A-SH、-L A-CONH 2、-L A-咪唑基、-L A-NHC(＝NH)NH 2、-L A-苯基、-L A-吲哚基和-L A-S-C 1-3烷基；其中，任一个L A独立地选自C 1-6亚烷基，独立地优选为C 1-4亚烷基，进一步独立地优选为亚甲基、1,2-亚乙基、1,3-亚丙基或1,4-亚丁基。
本申请中，C 1-6烷基每次出现，独立地为C 1烷基、C 2烷基、C 3烷基、C 4烷基、C 5烷基或C 6烷基。合适的实例包括但不限于：甲基(Me、-CH 3)、乙基(Et、-CH 2CH 3)、1-丙基(n-Pr、n-丙基、-CH 2CH 2CH 3)、2-丙基(i-Pr、i-丙基、-CH(CH 3) 2)、1-丁基(n-Bu、n-丁基、-CH 2CH 2CH 2CH 3)、2-甲基-1-丙基(i-Bu、i-丁基、-CH 2CH(CH 3) 2)、2-丁基(s-Bu、s-丁基、-CH(CH 3)CH 2CH 3)、2-甲基-2-丙基(t-Bu、t-丁基、-C(CH 3) 3)、1-戊基(n-戊基、-CH 2CH 2CH 2CH 2CH 3)、2-戊基(-CH(CH3)CH2CH2CH3)、3-戊基(-CH(CH 2CH 3) 2)、2-甲基-2-丁基(-C(CH 3) 2CH 2CH 3)、3-甲基-2-丁基(-CH(CH 3)CH(CH 3) 2)、3-甲基-1-丁基(-CH 2CH 2CH(CH 3) 2)、2-甲基-1-丁基(-CH 2CH(CH 3)CH 2CH 3)、1-己基(-CH 2CH 2CH 2CH 2CH 2CH 3)、2-己基(-CH(CH 3)CH 2CH 2CH 2CH 3)、3-己基(-CH(CH 2CH 3)(CH 2CH 2CH 3))、2-甲基-2-戊基(-C(CH 3) 2CH 2CH 2CH 3)、3-甲基-2-戊基(-CH(CH 3)CH(CH 3)CH 2CH 3)、4-甲基-2-戊基(-CH(CH 3)CH 2CH(CH 3) 2)、3-甲基-3-戊基(-C(CH 3)(CH 2CH 3) 2)、2-甲基-3-戊基(-CH(CH 2CH 3)CH(CH 3) 2)、2,3-二甲基-2-丁基(-C(CH 3) 2CH(CH 3) 2)和3,3-二甲基-2-丁基(-CH(CH 3)C(CH 3) 3。又如，C 1-3烷基每次出现，独立地为C 1烷基、C 2烷基或C 3烷基。
本申请中，C 1-6亚烷基每次出现，独立地为C 1亚烷基、C 2亚烷基、C 3亚烷基、C 4亚烷基、C 5亚烷基或C 6亚烷基。C 1-4亚烷基每次出现，独立地为C 1亚烷基、C 2亚烷基、C 3亚烷基或C 4亚烷基。合适的实例包括但不限于：亚甲基(-CH 2-)、1,1-乙基(-CH(CH 3)-)、1,2-乙基(-CH 2CH 2-)、1,1-丙基(-CH(CH 2CH 3)-)、1,2-丙基(-CH 2CH(CH 3)-)、1,3-丙基(-CH 2CH 2CH 2-)和1,4-丁基(-CH 2CH 2CH 2CH 2-)。
在一些实施方案中，R 0每次出现，独立地选自如下任一种基团：-CH 3、-CH(CH 3) 2、-CH 2CH(CH 3) 2、-CH(CH 3)CH 2CH 3、-CH 2CH 2SCH 3、
Figure PCTCN2022108093-appb-000008
Figure PCTCN2022108093-appb-000009
-CH 2-OH、-CH(OH)CH 3、-CH 2SH、-CH 2CONH 2、-CH 2CH 2CONH 2、-CH 2CH 2CH 2NH 2及其离子形式、-CH 2CH 2CH 2CH 2NH 2及其离子形式、- CH 2CH 2CH 2NHC(＝NH)NH 2及其离子形式、
Figure PCTCN2022108093-appb-000010
及其离子形式、-CH 2COOH及其离子形式和-CH 2CH 2COOH或及离子形式。
在一些实施方案中，R 0每次出现，独立地为非极性端基，如C 1-6烷基、-L A-苯基、-L A-S-C 1-3烷基，进一步如-CH 3、-CH(CH 3) 2、-CH 2CH(CH 3) 2、-CH(CH 3)CH 2CH 3、-CH 2CH 2SCH 3、
Figure PCTCN2022108093-appb-000011
在一些实施方案中，R 0每次出现，独立地为极性端基，如-L A-COOH、-L A-NH 2、-L A-OH、-L A-SH、-L A-CONH 2、-L A-咪唑基、-L A-NHC(＝NH)NH 2或-L A-吲哚基，进一步如
Figure PCTCN2022108093-appb-000012
-CH 2-OH、-CH(OH)CH 3、-CH 2SH、-CH 2CONH 2、-CH 2CH 2CONH 2、-CH 2CH 2CH 2NH 2及其离子形式、-CH 2CH 2CH 2CH 2NH 2及其离子形式、-CH 2CH 2CH 2NHC(＝NH)NH 2及其离子形式、
Figure PCTCN2022108093-appb-000013
及其离子形式、-CH 2COOH及其离子形式和-CH 2CH 2COOH或及离子形式。
在一些实施方案中，R 0每次出现，独立地为极性不带电荷的端基，如-CH 2-OH、-CH(OH)CH 3、-CH 2SH、-CH 2CONH 2或-CH 2CH 2CONH 2。
在一些实施方案中，R 0每次出现，独立地为非极性端基或极性不带电荷的端基。
在一些实施方案中，R 0每次出现，独立地为亲水性端基(如极性端基)或疏水性端基(如非极性端基)。R 0越疏水，形成的内核越紧密，在水性介质中的纳米聚合物会比较小，药物释放速率会慢一些。反之，R 0越亲水，置于水性介质中时，内核会比较松散，药物释放速率会快一些。
在载药单分子纳米聚合物的一些实施方案中，一个分子中，所述二价连接基L Pt中的铂原子的数量相对于氨基酸单元总数量的百分比为10％～100％，优选为10％～90％，另优选为10％～80％，另优选为10％～60％，另优选为10％～50％，另优选为10％～40％，另优选为10％～30％，另优选为15％～25％，另优选为18％～22％，另优选为15％～80％，另优选为15％～60％，另优选为15％～50％，另优选为15％～40％，另优选为15％～30％。通过控制二价连接基L Pt中的铂原子的数量相对于氨基酸单元总数量的百分比，可以调节载药单分子纳米聚合物的支化点密度。一个分子中，所述二价连接基L Pt中的铂原子的数量相对于氨基酸单元总数量的百分比还可以选自如下任一种百分数或者任两种百分数构成的区间：10％、15％、20％、25％、30％、35％、40％、45％、50％、55％、60％、65％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％、100％等。
在载药单分子纳米聚合物的一些实施方案中，一个分子中，所述第二药物单元的数量与所述铂类药物单元的数量之比为(0～10):1，优选为(0～5):1，另优选为(0～3):1，另优选为(0～1):1，另优选为(0.5～10):1，另优选为(0.5～5):1，另优选为(0.5～3):1，另优选为(1～5):1，另优选为(1～3):1，另优选为(2～3):1。通过控制第二药物单元的数量与铂类药物单元的数 量之比，可以调节两种不同药物的释放量。一个分子中，所述第二药物单元的数量与所述铂类药物单元的数量之比还可以选自如下任一种比例或任两种构成的区间：(0.1:1)、0.2:1)、(0.3:1)、(0.4:1)、(0.5:1)、(0.6:1)、(0.7:1)、(0.8:1)、(.9:1)、(1:1)、(1.1:1)、(1.2:1)、(1.3:1)、(1.4:1)、(1.5:1)、(1.6:1)、(1.8:1)、(2:1)、(2.5:1)、(2.6:1)、(2.8:1)、(3:1)、(3.5:1)、(4:1)、(4.5:1)、(5:1)、(5.5:1)、(6:1)、(6.5:1)、(7:1)、(7.5:1)、(8:1)、(8.5:1)、(9:1)、(9.5:1)、(10:1)等。
一些实施方案中，载药单分子纳米聚合物不含有第二药物单元。
在载药单分子纳米聚合物的一些实施方案中，一个分子中，所述亲水性聚合物链的数量与所述铂类药物单元的数量之比为1:(2～100)，优选为1:(10～60)，另优选为1:(15～45)，另优选为1:(15～25)。通过控制亲水性聚合物链的数量与铂类药物单元的数量，可以调节载药单分子纳米聚合物的聚氨基酸链长，从而控制单分子聚合物的尺寸，进而控制水性介质中纳米聚合物胶束的尺寸。一个分子中，所述亲水性聚合物链的数量与所述铂类药物单元的数量之比还可以选自如下任一种比例或任两种构成的区间：(1:2)、(1:3)、(1:4)、(1:5)、(1:6)、(1:7)、(1:8)、(1:9)、(1:10)、(1:11)、(1:12)、(1:13)、(1:14)、(1:15)、(1:16)、(1:18)、(1:20)、(1:22)、(1:24)、(1:25)、(1:26)、(1:28)、(1:30)、(1:35)、(1:40)、(1:45)、(1:55)、(1:60)、(1:65)、(1:70)、(1:75)、(1:80)、(1:85)、(1:90)、(1:95)、(1:100)等。
在载药单分子纳米聚合物的一些实施方案中，包括式(I)所示的四价结构单元，式(III)所示的一价结构单元、可选的式(II)所示的二价结构单元以及可选的式(IV)所示的二价结构单元；
Figure PCTCN2022108093-appb-000014
式(I)每次出现，其中，U 1和U 2各自独立地为碳中心三价基团，D Pt为铂类药物单元；
式(III)每次出现，其中，POL i为亲水性聚合物链；L 5独立地为二价连接基或无；Z 5独立地为-NH-或-C(＝O)-；
式(II)每次出现，其中，U 3独立地为碳中心三价基团，L R独立地为响应性连接基，L 4独立地为二价连接基，D T为第二药物单元；其中，L R能够在外界刺激下发生键断裂；
式(IV)每次出现，其中，U 6独立地为碳中心三价基团，R E独立地为H或R 0；其中，R 0为不含药物单元的端基。
在一些实施方式中，所述载药单分子纳米聚合物包括式(II)所示的二价结构单元以 及式(IV)所示的二价结构单元中的至少一种。通过引入这两种结构单元中的至少一种，可以适当降低非线性结构的支化点密度，所形成的胶束的内核相对松散，可适当加快释放药物的速率。
在一些实施方式中，式(II)所示的二价结构单元以及式(IV)所示的二价结构单元中仅存在其中的一种。
在一些实施方式中，所述载药单分子纳米聚合物不包括式(II)所示的二价结构单元。此时构成铂单药单分子纳米聚合物。
在一些实施方式中，所述载药单分子纳米聚合物不包括式(IV)所示的二价结构单元。此时，所有的氨基酸单元均连接有药物单元，至少连接有铂类药物单元。
在一些实施方式中，所述载药单分子纳米聚合物包括式(II)所示的二价结构单元以及式(IV)所示的二价结构单元。
在一些实施方式中，聚氨基酸链由式(I)所示的四价结构单元和式(II)所示的二价结构单元组成。此时，所有的氨基酸单元均连接有药物单元，或连接铂类药物单元(形成支化点)，或连接第二药物单元(不形成支化点，提供游离的含药侧链)。此时，制备原料无需加入式(IV-3)所示单体。
在一些实施方式中，聚氨基酸链由式(I)所示的四价结构单元和式(IV)所示的二价结构单元组成。
在一些实施方式中，聚氨基酸链由式(I)所示的四价结构单元、式(II)所示的二价结构单元和式(IV)所示的二价结构单元组成。
在本申请中，以波浪线
Figure PCTCN2022108093-appb-000015
表示原子或基团的连接位点。
在载药单分子纳米聚合物的一些实施方案中，
Figure PCTCN2022108093-appb-000016
每次出现，独立地包含如下结构：
Figure PCTCN2022108093-appb-000017
其中，U 10独立地为三价烃基，独立地优选为三价烷基；更优选地，
Figure PCTCN2022108093-appb-000018
独立地为赖氨酸或鸟氨酸单元，其为鸟氨酸单元时，U 10为>CH-CH 2CH 2CH 2-*，其为赖氨酸单元时，U 10为>CH-CH 2CH 2CH 2CH 2-*，其中的“*”指向D Pt。
一些实施方案中，
Figure PCTCN2022108093-appb-000019
独立地为赖氨酸单元。
在载药单分子纳米聚合物的一些实施方案中，
Figure PCTCN2022108093-appb-000020
每次出现，独立地包含如下结构：
Figure PCTCN2022108093-appb-000021
其中，U 20独立地为三价烃基，独立地优选为三价烷基；更优选地，
Figure PCTCN2022108093-appb-000022
独立地为赖氨酸或鸟氨酸单元，其为鸟氨酸单元时，U 10为>CH-CH 2CH 2CH 2-*，其为赖氨酸单元时，U 10为>CH-CH 2CH 2CH 2CH 2-*，其中的“*”指向D Pt。
一些实施方案中，
Figure PCTCN2022108093-appb-000023
独立地为赖氨酸单元。
在一些实施方案中，一个分子中的
Figure PCTCN2022108093-appb-000024
结构均相同，此时U 1和U 2结构均相同。
在一些实施方案中，一个分子中的U 1和U 2均相同。
在一些实施方案中，一个分子中
Figure PCTCN2022108093-appb-000025
结构均相同，此时U 10和U 20结构均相同。
在一些实施方案中，一个分子中的U 10和U 20均相同。
在载药单分子纳米聚合物的一些实施方案中，D Pt与相邻基团形成如下结构的主链：-C(＝O)-O-Pt-O-C(＝O)-或-C(＝O)-NH-O-Pt-O-NH-C(＝O)-。
在载药单分子纳米聚合物的一些实施方案中，D Pt与相邻基团形成如下结构的主链：-C(＝O)-O-Pt-O-C(＝O)-。
在载药单分子纳米聚合物的一些实施方案中，式(I)每次出现，独立地具有式(I-1)所示结构：
Figure PCTCN2022108093-appb-000026
其中，U 10和U 20分别独立地如前所定义；
Z 11和Z 21各自独立地为无、-C(＝O)-或-C(＝O)-O-*，还可以各自独立地为-C(＝O)-或-C(＝O)-O-*，其中的“*”指向D Pt；
R 11和R 21各自独立地为二价连接基，可以为亚烃基，可以各自独立地优选为亚烷基，还可以各自独立地优选为C 1-6亚烃基，还可以各自独立地优选为C 1-6亚烷基，还可以各自独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基、1,4-亚丁基、1,5-亚戊基或1,6-亚己基，还可以各自独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，还可以各自独立地优选为1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，还可以各自独立地优选为1,2-亚乙基或1,3-亚丙基，还可以各自独立地优选为1,2-亚乙基；
X 11和X 21各自独立地为-C(＝O)-O-*或-C(＝O)-NH-O-*，还可以各自独立地为-C(＝O)-O-*，其中的“*”指向D Pt。
在一些实施方案中，Z 11-R 11-X 11和Z 21-R 21-X 21可以各自独立地优选为-C(＝O)-R 01-C(＝O)-O-*、-C(＝O)-NH-R 01-C(＝O)-O-*、-C(＝O)-R 01-C(＝O)-NH-O-*或-C(＝O)-NH-R 01-C(＝O)-NH-O-*，进一步可以为-C(＝O)-R 01-C(＝O)-O-*，其中的“*”指向D Pt；其中，R 01的定义与R 11或R 21一致。R 01可以为-(CH 2) q-，其中q可以为选自1～6的整数，进一步可以为1、2、3、4、5或6，可以优选为1、2、3或4，进一步可以为2。
在载药单分子纳米聚合物的一些实施方案中，D Pt每次出现，独立地选自顺铂、卡铂、奈达铂、奥沙利铂和洛铂中任一种的残基。
在载药单分子纳米聚合物的一些实施方案中，式(I)每次出现，具有相同结构。
在载药单分子纳米聚合物的一些实施方案中，
Figure PCTCN2022108093-appb-000027
每次出现，独立地包含如下结构：
Figure PCTCN2022108093-appb-000028
其中，U 30独立地为三价烃基，独立地优选为三价烷基；更优选地，
Figure PCTCN2022108093-appb-000029
独立地为赖氨酸或鸟氨酸单元，其为鸟氨酸单元时，U 30为>CH-CH 2CH 2CH 2-*，其为赖氨酸单元时，U 30为>CH-CH 2CH 2CH 2CH 2-*，其中的“*” 指向D T。一些实施方案中，
Figure PCTCN2022108093-appb-000030
独立地为赖氨酸单元。
在一些实施方案中，一个分子中的U 3均相同。
在一些实施方案中，一个分子中的U 30均相同。
在载药单分子纳米聚合物的一些实施方案中，L R每次出现，独立地包含能够在如下至少一种条件下发生断裂的连接基：胞内还原条件、活性氧条件、pH条件、酶解条件和水解条件。
在一些实施方案中，所述pH条件满足pH值小于6.8，进一步优选pH为4.0～6.8。
在一些实施方案中，所述酶解条件选自如下的一种或多种酶：MMP-2酶和偶氮还原酶。
在一些实施方案中，所述水解条件为酸性水解条件或碱性水解条件。
在一些实施方案中，L R每次出现，独立地包含如下(a)组、(b)组、(c)组、(d)组和(e)组中的一种或多种连接基；
(a)组：-S-S-；
(b)组：草酸酯基、硼酸酯基、酮缩硫醇基、硫醚基、单硒基、二硒基、二价碲基、噻唑啉酮基、硼酸基和3～7元脯氨酸低聚链；
(c)组：缩醛基和腙键；
(d)组：GPLGVRG肽段和偶氮基；
(e)组：-C(＝O)-O-和-O-C(＝O)-。
其中，(a)组的基团可以响应于胞内还原条件(如谷胱甘肽环境)、活性氧条件等条件。
(b)组的基团可以响应于活性氧(ROS)条件，可属于ROS响应基团。
(c)组的基团可相应于特定的酸性pH条件。
(d)组的基团可在酶作用下断裂。GPLGVRG肽段可在MMP-2酶作用下酶解。偶氮基可在偶氮还原酶条件下酶解。
(e)组的基团可发生水解。
需要理解的时，(a)组、(b)组、(c)组、(d)组和(e)组中的基团可以响应于一种或多种刺激条件。
在一些实施方案中，L R每次出现，独立地包含如下的一种或多种连接基：-S-S-、草酸酯基、芳基硼酸酯基、缩醛基、腙键、GPLGVRG肽段、偶氮基、-C(＝O)-O-和-O-C(＝O)-；另优选地，所述芳基硼酸酯基为苯基硼酸酯基。
在一些实施方案中，(b)组的ROS响应基团可以包含如下基团中的一种或多种：酮缩硫醇基(-S-C(CH 3) 2-S-)、硫醚键(-S-)、单硒键(-Se-)、二硒键(-Se-Se-)、二价碲(-Te-)、草酸酯基(-OC(＝O)-C(＝O)-O-)、噻唑啉酮基
Figure PCTCN2022108093-appb-000031
硼酸酯基(如
Figure PCTCN2022108093-appb-000032
)、硼酸基(如-B(OH) 2)和脯氨酸低聚链。
在一些实施方案中，脯氨酸低聚链的结构如
Figure PCTCN2022108093-appb-000033
所示，其中np为选自3～8的整数。在其中的一些实施例中，脯氨酸低聚链中的脯氨酸单元数选自3～7(如3、4、5、5或7)。在其中的一些实施例中，n＝7。
ROS响应基团的举例如-Ar-OC(＝O)-C(＝O)-O-，其中，Ar为亚芳基，举例如亚苯基，进一步举例如1,4-亚苯基。
在一些实施方案中，-L 4-D T每次出现，独立地包括Z 4-D T，其中，Z 4每次出现，独立地为化学键或者选自如下任一种基团：-C(＝O)-、-O-、-S-、-O-C(＝O)-*、-NH-C(＝O)-*、和-NH-，其中，“*”所示端指向D T。
在一些实施方案中，-L 4-D T每次出现，其结构独立为-R 32-Z 4-D T，其中，R 32每次出现，独立地为二价连接基，可以为亚烃基，可以各自独立地优选为亚烷基，还可以各自独立地优选为C 1-6亚烃基，还可以独立地优选为C 1-6亚烷基，还可以独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基、1,4-亚丁基、1,5-亚戊基或1,6-亚己基，还可以独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，还可以各自独立地优选为1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，还可以各自独立地优选为1,2-亚乙基或1,3-亚丙基，还可以各自独立地优选为1,2-亚乙基。
在一些实施方案中，D T与Z 4独立地形成如下任一种连接基：-C(＝O)-O-、-O-C(＝O)-、-O-C(＝O)-O-、-O-C(＝O)-NH-、-NH-C(＝O)-O-、-C(＝O)-NH-和-NH-C(＝O)-。
在一些实施方案中，D T与Z 4独立地形成如下任一种连接基：-C(＝O)-O-、-O-C(＝O)-、-O-C(＝O)-O-、-O-C(＝O)-NH-和-NH-C(＝O)-O-。
在一些实施方案中，D T与Z 4独立地形成-O-C(＝O)-O-连接方式。
在一些实施方案中，L R和D T之间的连接方式可以是可断裂的，从而可以释放出D T对应的药物活性成分或其前药。可断裂的连接方式的举例如：-C(＝O)-O-、-O-C(＝O)-、-O-C(＝O)-O-、-O-C(＝O)-NH-、-NH-C(＝O)-O-。
在一些实施方案中，式(II)每次出现，具有式(II-1)所示结构：
Figure PCTCN2022108093-appb-000034
其中，U 30每次出现，独立地如前所定义；
R 32和Z 4每次出现，分别独立地如前所定义；
Z 3每次出现，独立地为无、-C(＝O)-或-C(＝O)-O-*，还可以独立地为-C(＝O)-或-C(＝O)-O-*，还可以独立地优选为-C(＝O)-O-*，还可以独立地优选为-C(＝O)-，其中的“*”指向 R 31；
R 31每次出现，独立地为二价连接基，可以为亚烃基，可以各自独立地优选为亚烷基，还可以各自独立地优选为C 1-6亚烃基，还可以独立地优选为C 1-6亚烷基，还可以独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基、1,4-亚丁基、1,5-亚戊基或1,6-亚己基，还可以独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，还可以各自独立地优选为1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，还可以各自独立地优选为1,2-亚乙基或1,3-亚丙基，还可以各自独立地优选为1,2-亚乙基。
在一些实施方案中，R 31-L R-R 32每次出现，独立地为-(CH 2) p1-S-S-(CH 2) p2-，其中，p1和p2各自独立地为选自1～6的整数，进一步可以各自独立地为1、2、3、4、5或6，可以各自独立地优选为1、2、3或4，进一步可以各自独立地为2或3。
在一些实施方案中，R 31-L R-R 32每次出现，均为-(CH 2) 2-S-S-(CH 2) 2-。
在一些实施方案中，Z 3-R 31-L R-R 32每次出现，均为-C(＝O)-(CH 2) 2-S-S-(CH 2) 2-*或-C(＝O)-O-(CH 2) 2-S-S-(CH 2) 2-*，其中的“*”指向D T。
在一些实施方案中，D T每次出现，独立地选自喜树碱类化合物、瑞喹莫德和紫杉醇中任一种的残基。
应当理解，D T对应的药物活性成分或其前药应当具有反应性基团F T，或者可以被活化为反应性基团F T，从而能够被修饰上L R。反应性基团F T可以为羟基、羧基、氨基、巯基等官能团中的一种或多种。在一些实施方案中，反应性基团F T为游离羟基、游离氨基或者其组合。当D T对应的药物活性成分或其前药中具有≥2个的反应性基团时，可以保护其中的部分反应基团使得一个D T与一个L R相连接。反应性基团F T可以利用常规的偶合反应形成共价连接基，比如，羟基可以反应形成醚键(-O-)、酯基(-O-C(＝O)-)、碳酸酯基(-O-C(＝O)-O-)等，羧基可以反应形成酰胺键(-C(＝O)-NH-)等，氨基可以反应形成二价氨基(-NH-)、酰胺键(-C(＝O)-NH-)、氨基甲酸酯基(-NH-C(＝O)-O-)等。本领域可以根据D T对应的药物活性成分或其前药的结构特点选择合适Z 4连接基。
在一些实施方案中，所述喜树碱类化合物包括喜树碱及其衍生物或类似物。
在一些实施方案中，所述喜树碱类化合物包括依立替康、拓扑替康、卢比替康、吉咪替康、9-氨基喜树碱、9-硝基喜树碱和7-乙基-10-羟基喜树碱。
在一些实施方案中，式(II)每次出现，具有相同结构。
在一些实施方案中，式(III)中，Z 5每次出现，独立地为-NH-、-C(＝O)-或*-O-C(＝O)-，其中的“*”指向L 5。
在一些实施方案中，式(III)中，L 5每次出现，独立地为二价连接基，可以为亚烃基，可以各自独立地优选为亚烷基，还可以各自独立地优选为C 1-6亚烃基，还可以独立地优选为C 1-6亚烷基，还可以独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基、1,4-亚丁基、1,5-亚戊基或1,6-亚己基，还可以独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，还可以各自独立地优选为1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，还可以各自独立地优选为1,2-亚乙基或1,3-亚丙基，还可以各自独立地优选为1,2-亚乙基。
在一些实施方案中，式(III)每次出现，具有相同的L 5和Z 5。
在一些实施方案中，式(III)中，POL i每次出现，独立地包含如下任一种亲水聚合物 链：聚乙二醇、聚(丙二醇)、乙二醇和丙二醇的共聚物、聚(乙氧基化多元醇)、聚(烯醇)、聚(乙烯基吡咯烷酮)、聚(羟烷基甲基丙烯酰胺)、聚(羟烷基甲基丙烯酸酯)、聚(糖)、聚(α-羟基酸)、聚(乙烯醇)、聚膦腈、聚噁唑啉、聚(N-丙烯酰基吗啉)以及前述聚合物链的任意组合。
在一些实施方案中，式(III)中，所述亲水聚合物链的分子量选自50Da～100kDa，另优选为100Da～80kDa，另优选为500Da～50kDa，另优选为500Da～10kDa，另优选为500Da～8000Da，另优选为500Da～6000Da，另优选为500Da～5000Da，另优选为1000Da～50kDa，另优选为1000Da～10kDa，另优选为1000Da～8000Da，另优选为1000Da～6000Da，另优选为1000Da～5000Da。所述亲水聚合物链的分子量还可以选自如下任一种或任两种构成的区间：约500Da、约600Da、约700Da、约750Da、约800Da、约850Da、约900Da、约950Da、约1000Da、约1100Da、约1200Da、约1300Da、约1400Da、约1500Da、约1600Da、约1800Da、约2000Da、约2200Da、约2400Da、约2500Da、约3000Da、约3500Da、约4000Da、约4500Da、约5000Da、约5500Da、约6000Da、约6500Da、约7000Da、约7500Da、约8000Da、约8500Da、约9000Da、约10000Da等，其中的“约”可表示±10％、±5％、±2％或0。
在一些实施方案中，式(III)中，POL i每次出现，独立地包含聚乙二醇链段；另优选地，所述聚乙二醇链段为mPEG，另优选地，所述聚乙二醇链段的分子量选自50Da～100kDa，另优选为100Da～80kDa，另优选为500Da～50kDa，另优选为500Da～10kDa，另优选为500Da～8000Da，另优选为500Da～6000Da，另优选为500Da～5000Da，另优选为1000Da～50kDa，另优选为1000Da～10kDa，另优选为1000Da～8000Da，另优选为1000Da～6000Da，另优选为2000Da～6000Da，另优选为4000Da～6000Da，另优选为1000Da～5000Da，另优选为约500Da、约600Da、约800Da、约1000Da、约1100Da、约1200Da、约1500Da、约1600Da、约2000Da、约2200Da、约2500Da、约3000Da、约3500Da、约4000Da、约4400Da、约4500Da、约5000Da、约5500Da、约6000Da、约6500Da、约7000Da、约8000Da、约9000Da、约10kDa、约12kDa、约15kDa、约20kDa、约25kDa、约30kDa、约35kDa、约40kDa或约40kDa，其中的“约”可表示±10％、±5％、±2％或0。
式(III)中，任一种POL i的“分子量”可以独立地表示重均分子量或数均分子量。
式(III)中，任一种POL i的“分子量”可以独立地表示重均分子量。
式(III)中，任一种POL i的“分子量”可以独立地表示数均分子量。
在一些实施方案中，式(III)每次出现，具有相同结构。
在载药单分子纳米聚合物的一些实施方案中，式(IV)每次出现，具有式(IV-1)所示结构：
Figure PCTCN2022108093-appb-000035
其中，R E每次出现，独立地为氢原子或R 0，其中R 0为不含药物单元的端基。
在一些实施方案中，R E每次出现，独立地为R 0。
在一些实施方案中，R 0如前文任一实施方案中所定义。
在一些实施方案中，聚氨基酸链中包含式(IV-1)所示结构的氨基酸单元。
在一些实施方案中，式(IV)每次出现，具有相同结构。
在载药单分子纳米聚合物的一些实施方案中，式(I)每次出现，具有相同结构；式(III)每次出现，具有相同的L 5和Z 5；如有，式(II)每次出现，具有相同结构；如有，式(IV)每次出现，具有相同结构。此时，制备载药单分子纳米聚合物时仅需针对相应结构单元提供单一种类的原料即可。
在载药单分子纳米聚合物的一些实施方案中，所述载药单分子纳米聚合物包括式(I-2)所示的四价结构单元，式(III-2)所示的一价结构单元、可选的式(II-2)所示的二价结构单元以及可选的式(IV-1)所示的二价结构单元；
Figure PCTCN2022108093-appb-000036
优选地，
n11和n21各自独立地为3或4，n12和n22各自独立地为1、2、3、4或5；
n31独立地为3或4，n32独立地为2、3或4，n33独立地为2、3或4；
n51独立地为1、2、3或4；
p独立地为正整数，优选为小于等于2500的正整数，另优选为小于等于2000的正整数，另优选为小于等于1500的正整数，另优选为小于等于1000的正整数，另优选为小于等于800的正整数，另优选为小于等于600的正整数，另优选为小于等于500的正整数，另优选为小于等于400的正整数，另优选为小于等于300的正整数，另优选为小于等于250的正整数，另优选为小于等于200的正整数，另优选为选自2～2500的整数，另优选为选自3～2000的整数，另优选为选自5～1500的整数，另优选为选自5～1000的整数，另优选为选自5～800的整数，另优选为选自5～600的整数，另优选为选自5～500 的整数，另优选为选自5～400的整数，另优选为选自5～300的整数，另优选为选自5～250的整数，另优选为选自5～200的整数，另优选为选自5～1500的整数，另优选为选自5～1000的整数，另优选为选自5～800的整数，另优选为选自10～600的整数，另优选为选自10～500的整数，另优选为选自10～400的整数，另优选为选自10～300的整数，另优选为选自10～250的整数，另优选为选自10～200的整数，另优选为选自20～600的整数，另优选为选自20～500的整数，另优选为选自20～400的整数，另优选为选自20～300的整数，另优选为选自20～250的整数，另优选为选自20～200的整数；另优选为选自50～500的整数，另优选为选自50～400的整数，另优选为选自50～300的整数，另优选为选自50～250的整数，另优选为选自50～200的整数；另优选为选自100～500的整数，另优选为选自100～400的整数，另优选为选自100～300的整数，另优选为选自100～250的整数，另优选为选自100～200的整数，另优选为选自100～150的整数。
在一些实施方案中，p优选为小于等于500的正整数，另优选为小于等于400的正整数，另优选为小于等于300的正整数，另优选为小于等于250的正整数，另优选为小于等于200的正整数，优选为选自5～500的整数，另优选为选自5～400的整数，另优选为选自5～300的整数，另优选为选自5～250的整数，另优选为选自5～200的整数，另优选为选自10～500的整数，另优选为选自10～400的整数，另优选为选自10～300的整数，另优选为选自10～250的整数，另优选为选自10～200的整数，另优选为选自20～500的整数，另优选为选自20～400的整数，另优选为选自20～300的整数，另优选为选自20～250的整数，另优选为选自20～200的整数；另优选为选自50～500的整数，另优选为选自50～400的整数，另优选为选自50～300的整数，另优选为选自50～250的整数，另优选为选自50～200的整数；另优选为选自100～500的整数，另优选为选自100～400的整数，另优选为选自100～300的整数，另优选为选自100～250的整数，另优选为选自100～200的整数，另优选为选自100～150的整数。
在一些实施方案中，p还可以选自如下任一个整数或任两个整数构成的区间：3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、45、50、55、60、70、80、90、100、110、120、130、140、150、160、170、180、190、200、210、215、220、225、227、240、250、260、280、300、350、400、450、500等。
在一些实施方案中，p还可以选自如下任一范围内的整数：110～120，100～120，100～130，100～140，100～150，90～120，90～130，90～140，90～150，80～120，80～130，80～140，80～150等。
在一些实施方案中，p还可以选自如下任一范围内的整数：5～115，5～114，5～110，5～100，5～90，5～88，5～78，5～78，5～777，5～66，5～65，5～60，5～55，5～50，5～45，5～44，5～40，5～35，5～34，5～33，5～30，5～25，5～20，6～115，6～114，6～110，6～100，6～90，6～88，6～78，6～78，6～777，6～66，6～65，6～60，6～55，6～50，6～45，6～44，6～40，6～35，6～34，6～33，6～30，6～25，6～20，8～115，8～114，8～110，8～100，8～90，8～88，8～78，8～78，8～777，8～66，8～65，8～60，8～55，8～50，8～45，8～44，8～40，8～35，8～34，8～33，8～30，8～25，8～20，10～115，10～114，10～110，10～100，10～90，10～88，10～78，10～78， 10～777，10～66，10～65，10～60，10～55，10～50，10～45，10～44，10～40，10～35，10～34，10～33，10～30，10～25，10～20，15～115，15～114，15～110，15～100，15～90，15～88，15～78，15～78，15～777，15～66，15～65，15～60，15～55，15～50，15～45，15～44，15～40，15～35，15～34，20～115，20～114，20～110，20～100，20～90，20～88，20～78，20～78，20～777，20～66，20～65，20～60，20～55，20～50，20～45，20～44，20～40，20～35，20～34等。
式(IV-1)所示的二价结构单元如前文所定义。
在一些实施方式中，n11和n21各自独立地为3或4，进一步地，n11和n21各自独立地为4。
在一些实施方案中，n12和n22各自独立地为2、3、4或5，还可以为各自独立地为2或3，还可以为各自独立地为2，还可以为各自独立地为3。
在一些实施方案中，n31独立地为3或4，进一步地可以独立地为4。
在一些实施方案中，n32独立地为2、3或4，进一步地可以独立地为2。
在一些实施方案中，n33独立地为2、3或4，进一步地可以独立地为2。
在一些实施方案中，n51独立地为1、2、3或4，进一步地可以独立地为2、3或4，更进一步可以独立地为2或3，还可以独立地为2，还可以独立地为3。
在一些实施方式中，n11和n21各自独立地为4，n12和n22各自独立地为4。
在一些实施方式中，n31独立地为4，n32独立地为2，n33独立地为2。
在一些实施方式中，n51独立地为2、3或4，进一步可以独立地为3。
在一些实施方式中，L R为-S-S-。
在一些实施方式中，Z 5为-NH-。
在一些实施方式中，L R为-S-S-，Z 5为-NH-。
在一些实施方式中，D Pt为顺铂、奥沙利铂或
Figure PCTCN2022108093-appb-000037
的残基，D T为喜树碱、紫杉醇或瑞喹莫德的残基。
在载药单分子纳米聚合物的一些实施方案中，载药单分子纳米聚合物的分子量(可以为重均分子量或数均分子量，可以优选为重均分子量，也可以优选为数均分子量)可大于50kDa，进一步可大于100kDa，更进一步可选自100kDa～5000kDa，更进一步可选自150kDa～5000kDa，更进一步可选自200kDa～5000kDa，更进一步可选自250kDa～5000kDa，更进一步可选自300kDa～5000kDa，更进一步可选自400kDa～5000kDa，优选为500kDa～5000kDa，另优选为500kDa～4000kDa，另优选为500kDa～3000kDa，另优选为500kDa～2500kDa，另优选为500kDa～2000kDa，另优选为500kDa～1500kDa，另优选为600kDa～1500kDa，另优选为800kDa～1200kDa。载药单分子纳米聚合物的分子量还可以选自如下任一种分子量或任两种分子量构成的区间：100kDa、150kDa、200kDa、250kDa、300kDa、400kDa、500kDa、550kDa、600kDa、650kDa、700kDa、750kDa、800kDa、850kDa、900kDa、950kDa、1000kDa、1100kDa、1200kDa、1300kDa、1400kDa、1500kDa、1600kDa、1700kDa、1800kDa、1900kDa、2000kDa、2100kDa、2200kDa、2300kDa、2400kDa、2500kDa、3000kDa、3500 kDa、4000kDa、4500kDa、5000kDa等。
在载药单分子纳米聚合物的一些实施方案中，一个分子中的铂原子数量大于40，进一步大于50，还可以选自50～5000，进一步可选自50～4000，进一步可选自50～2000，进一步可选自50～1500，进一步可选自50～1000，进一步可选自50～500，还可以选自60～2000，还可以选自60～1500，还可以选自60～1000，还可以选自60～500，还可以选自80～2000，还可以选自80～1500，还可以选自80～1000，还可以选自80～500，还可以选自100～2000，还可以选自100～1500，还可以选自100～1000，还可以选自100～500，还可以选自150～2000，还可以选自150～1500，还可以选自150～1000，还可以选自150～500，还可以选自200～2000，还可以选自200～1500，还可以选自200～1000，还可以选自200～500，还可以选自250～2000，还可以选自250～1500，还可以选自250～1000，还可以选自250～500，还可以选自300～400。一个分子中的铂原子数量还可以选自如下任一个数值或任两个数值构成的区间：50、60、80、100、120、150、200、250、300、350、400、450、500、550、600、650、700、750、800、900、1000、1100、1200、1300、1400、1500、1600、1800、2000、2500、3000等。
控制载药单分子纳米聚合物的分子量在合适的范围，可以将其形成的胶束控制在适制为药物制剂的尺寸。比如，可以影响到在水中、水溶液中或体内环境中的胶束尺寸。以数均分子量约为1000kDa为例，一些实施例中，25℃水中，平均直径约为25～45nm，其中一些实施例中，平均直径约为30nm、32nm、34nm、44nm等。
本申请中，关于“载药单分子纳米聚合物胶束的粒径”，仅在特别说明时指平均直径或平均粒径。如无特别说明，测试温度在20～30℃，进一步为25℃。
本申请还提供一种载药单分子纳米聚合物的制备方法，其包括如下步骤：将结构如式(I-3)所示的含铂化合物、结构如式(III-3)所示的单官能化亲水聚合物、可选的结构如式(II-3)所示的药物化合物以及可选的如式(IV-3)所示化合物在有机溶剂中混合，进行开环聚合反应；
Figure PCTCN2022108093-appb-000038
其中，
P E为R E或被保护的R E，在所述开环聚合反应中表现为惰性，也即在所述开环聚合反应中不具备反应性；
U 1、U 2、D Pt、U 3、L R、L 4、D T、mPEG、L 5和R E的定义分别与前述一致；
F 5为-NH 2、-COOH、
Figure PCTCN2022108093-appb-000039
优选为-NH 2。
本申请中，如无其他说明，mPEG对应CH 3(OCH 2CH 2) p-O-，p的定义与前述一致。
在一些实施方案中，所述开环聚合反应于无水条件下进行。
在一些实施方案中，开环聚合反应温度为15～40℃，更优选地，开环聚合反应时间为24～96h。
式(I-3)所示的含铂化合物(可记为NCA-Pt-NCA，一种双NCA单体)、结构如式(II-3)所示的药物化合物(NCA-L R-D T，一种单NCA单体)以及如式(IV-3)所示化合物(可记为NCA-AA，一种单NCA单体)均为NCA官能化的氨基酸单体。
本申请中，“N-羧酸酐”官能团记为NCA。“N-羧酸酐”官能团指的是含有-NH-C(＝O)-O-C(＝O)-结构的官能团。如无其他说明，优选为环状结构，进一步可以优选为五元环(如
Figure PCTCN2022108093-appb-000040
)。
该聚合反应利用双N-羧酸酐(NCA)参与的开环聚合反应，通过“一锅法”得到单分子纳米聚合物，该单分子纳米聚合物可以在水性介质中无需自组装即可形成核壳结构的胶束，提供可响应性释放药物活性成分的药物递送系统，用于肿瘤疾病的治疗。
式(I-3)所示单体为本申请的一种支化氨基酸单体，以铂原子为桥连接两个NCA官能团，该单体经开环聚合反应，可形成非线性骨架，提供非线性骨架中的支化点。
式(II-3)所示单体和式(IV-3)所示单体均为本申请中的非支化氨基酸单体。
式(II-3)所示单体一端为NCA官能团，另一端携带第二药物单元D T，该单体经开环聚合可参与形成聚氨基酸链，但不提供非线性骨架中的支化点。
式(IV-3)所示单体为NCA官能化的氨基酸，包含NCA官能团，且不包含其他的反应性基团(指参与开环聚合反应的反应性)，在开环聚合反应中，仅NCA参与反应，该单体参与形成聚氨基酸链，但不提供非线性骨架中的支化点。
可以理解，在式(I-3)所示单体的基础上，引入式(II-3)所示单体和式(IV-3)所示单体中的至少一种，可以在聚氨基酸链中起到间隔式(I-3)所示单体中的支化点的作用，从而调节聚氨基酸链中的支化点分布密度，调节载药单分子纳米聚合物的整体的支化或交联情况。
式(III-3)所示单体可起到封端剂的作用，用量越多，越容易获得较短的聚氨基酸链以及尺寸较小载药单分子纳米聚合物，通过调节式(III-3)所示单体的用量多少，可以调节载药单分子纳米聚合物的分子大小，同时还可控制每个单分子纳米聚合物的载药量。
在一些实施方案中，式(I-3)所示单体在所有氨基酸单体(具体为NCA官能化的氨基酸单体)中的摩尔百分比可以为15％～100％，优选为15％～90％，另优选为15％～80％，另优选为15％～60％，另优选为15％～50％，另优选为15％～40％，另优选为15％～30％，另另优选为20％～80％，另优选为20％～60％，另优选为20％～50％，另优选为20％～40％， 另优选为20％～30％。式(I-3)所示单体在所有氨基酸单体中的摩尔百分比还可以选自如下任一种百分数或者任两种百分数构成的区间：15％、16％、17％、18％、19％、20％、22％、24％、25％、26％、28％、30％、35％、40％、45％、50％、55％、60％、65％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％、100％等。
在一些实施方案中，式(II-3)所示单体相对于式(I-3)所示单体的摩尔比，在数值上，可以参考第二药物单元的数量与铂类药物单元的数量之比。式(II-3)所示单体相对于式(I-3)所示单体的摩尔比可以为(0～10):1，优选为(0～5):1，另优选为(0～3):1，另优选为(0～1):1，另优选为(0.5～10):1，另优选为(0.5～5):1，另优选为(0.5～3):1，另优选为(1～5):1，另优选为(1～3):1，另优选为(2～3):1。式(II-3)所示单体相对于式(I-3)所示单体的摩尔比可以选自如下任一种比例或任两种构成的区间：(0.1:1)、0.2:1)、(0.3:1)、(0.4:1)、(0.5:1)、(0.6:1)、(0.7:1)、(0.8:1)、(.9:1)、(1:1)、(1.1:1)、(1.2:1)、(1.3:1)、(1.4:1)、(1.5:1)、(1.6:1)、(1.8:1)、(2:1)、(2.5:1)、(2.6:1)、(2.8:1)、(3:1)、(3.5:1)、(4:1)、(4.5:1)、(5:1)、(5.5:1)、(6:1)、(6.5:1)、(7:1)、(7.5:1)、(8:1)、(8.5:1)、(9:1)、(9.5:1)、(10:1)等。
在一些实施方案中，式(IV-3)所示单体相对于式(I-3)所示单体的摩尔比，在数值上，可以参考亲水性聚合物链的数量与铂类药物单元的数量之比。式(IV-3)所示单体相对于式(I-3)所示单体的摩尔比可以为1:(2～100)，优选为1:(10～60)，另优选为1:(15～45)，另优选为1:(15～25)。式(IV-3)所示单体相对于式(I-3)所示单体的摩尔比还可以选自如下任一种比例或任两种构成的区间：(1:2)、(1:3)、(1:4)、(1:5)、(1:6)、(1:7)、(1:8)、(1:9)、(1:10)、(1:11)、(1:12)、(1:13)、(1:14)、(1:15)、(1:16)、(1:18)、(1:20)、(1:22)、(1:24)、(1:25)、(1:26)、(1:28)、(1:30)、(1:35)、(1:40)、(1:45)、(1:55)、(1:60)、(1:65)、(1:70)、(1:75)、(1:80)、(1:85)、(1:90)、(1:95)、(1:100)等。
根据本申请的双药单分子纳米聚合物前药，其包含与亲水性聚合物连接的聚氨基酸，其中在该聚氨基酸的重复单元的α碳上键合了铂类药物活性成分的前药部分和抗肿瘤药物活性成分的前药部分；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
在双药单分子纳米聚合物前药的一些实施方案中，所述的双药单分子纳米聚合物前药包含与亲水性聚合物连接的无规共聚氨基酸主链。
在双药单分子纳米聚合物前药的一些实施方案中，所述的亲水性聚合物选自聚(烷二醇)(如，聚乙二醇(“PEG”)、聚(丙二醇)(“PPG”)、乙二醇和丙二醇的共聚物等)、聚(乙氧基化多元醇)、聚(烯醇)、聚(乙烯基吡咯烷酮)、聚(羟烷基甲基丙烯酰胺)、聚(羟烷基甲基丙烯酸酯)、聚(糖)、聚(α-羟基酸)、聚(乙烯醇)、聚膦腈、聚噁唑啉(“POZ”)、聚(N-丙烯酰基吗啉)以及这些物质的任意组合。
在双药单分子纳米聚合物前药的优选实施方案中，所述的亲水性聚合物选自聚乙二醇(“PEG”)，优选选自一端为甲氧基封端的聚乙二醇。该亲水性聚合物的分子量没有特别限制，例如PEG50-5000、PEG50-6000、PEG50-7000、PEG50-8000、PEG500-2000、PEG500-4000、PEG500-6000、PEG500-8000、PEG500-10000、PEG500-20000、PEG1000-20000、PEG1000-50000或PEG1000-80000等均可以用于本申请，单位为Da。其中，以PEG500-4000为例，表示分子量位500～4000Da。
在载药单分子纳米聚合物及双药单分子纳米聚合物前药的一些实施方案中，所述的铂类药物包括顺铂、卡铂、奈达铂、奥沙利铂和洛铂。它们的化学结构、制备方法和药理学作用本领域中都是已知的。例如，顺铂属于细胞周期非特异性抗癌药物，具有下式结构：
Figure PCTCN2022108093-appb-000041
在载药单分子纳米聚合物及双药单分子纳米聚合物前药的优选实施方案中，所述的铂类药物活性成分是顺铂。
在载药单分子纳米聚合物及双药单分子纳米聚合物前药的一些实施方案中，所述分子中含有游离羟基、游离氨基或两者的组合的抗肿瘤药物活性成分选自：喜树碱类化合物，包括喜树碱及其衍生物或类似物，例如依立替康、拓扑替康、卢比替康、吉咪替康、9-氨基喜树碱和9-硝基喜树碱、7-乙基-10-羟基喜树碱(SN38)等；肿瘤免疫激活剂瑞喹莫德(Resiquimod，R-848)、替拉莫德等；紫杉醇(Paclitaxel，PTX)、表柔比星、多西紫杉醇、多西他赛、培美曲塞、甲基澳瑞他汀E、吉西他滨、低塞米松等；以及蛋白激酶抑制剂索拉非尼、达沙替尼等。
这些抗肿瘤药物活性成分的化学结构、制备方法和药理学作用都是本领域中已知的。例如，喜树碱属于DNA拓扑异构酶I抑制剂，具有下式结构：
Figure PCTCN2022108093-appb-000042
瑞喹莫德(Resiquimod，R-848)是免疫应答调节剂，具有促进肿瘤免疫的活性，其结构式如下：
Figure PCTCN2022108093-appb-000043
紫杉醇(Paclitaxel，PTX)是从红豆杉属植物中提取的生物碱，属于细胞周期特异性抗肿瘤药物，通过促进微管蛋白聚合抑制解聚，保持微管蛋白稳定，抑制细胞有丝分裂。紫杉醇的化学名称为5β,20-环氧-1,2α,4,7β,10β,13α-六羟基紫杉烷-11-烯-9-酮-4,10-二乙酸酯-2-苯甲酸酯-13[(2'R,3'S)-N-苯甲酰-3-苯基异丝氨酸酯]，结构式如下：
Figure PCTCN2022108093-appb-000044
在载药单分子纳米聚合物及双药单分子纳米聚合物前药的优选实施方案中，所述分子中含有游离羟基、游离氨基或两者的组合的抗肿瘤药物活性成分是喜树碱是喜树碱。
在载药单分子纳米聚合物及双药单分子纳米聚合物前药的一些实施方案中，所述的双药单分子纳米聚合物前药具有如下结构：
Figure PCTCN2022108093-appb-000045
其中，p可以选自1-500，m可以选自0-100，n可以选自1-100，k可以选自1-1000，但均不限于此范围。其中，p对应聚乙二醇单元的聚合度，p的定义还可参考本文的其他部分。
该通式中，m可对应于一个分子中的第二药物单元数量；n可对应一个分子中的铂 原子(L Pt中的铂)数量；k可对应一个分子中的mPEG链段数量。
在一些实施方案中，一个分子中的第二药物单元的数量m可选自0～5000，进一步可选自0～4000，进一步可选自0～2000，进一步可选自0～1500，进一步可选自0～1000，进一步可选自0～500，还可以选自10～2000，还可以选自10～1500，还可以选自10～1000，还可以选自10～500，还可以选自20～2000，还可以选自20～1500，还可以选自20～1000，还可以选自20～500，还可以选自40～2000，还可以选自40～1500，还可以选自40～1000，还可以选自40～500，还可以选自50～2000，还可以选自50～1500，还可以选自50～1000，还可以选自50～500，还可以选自80～2000，还可以选自80～1500，还可以选自80～1000，还可以选自80～500，还可以选自100～2000，还可以选自100～1500，还可以选自100～1000，还可以选自100～500，还可以选自500～1500，还可以选自600～1500，还可以选自800～1500，还可以选自800～1200。一个分子中的第二药物单元的数量m还可以选自如下任一种数值或任两个构成的区间：0、1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、18、20、25、30、35、40、45、50、60、70、80、90、100、120、150、200、250、300、350、400、450、500、550、600、650、700、750、800、900、1000、1100、1200、1300、1400、1500、1600、1800、2000、2500、3000等。
需要说明的是，该通式中的氨基酸单元为赖氨酸，通式中的赖氨酸单元通过含Pt的连接基形成非线性骨架，其中可以包含多条聚赖氨酸链，与k对应的氨基酸单元以及与n对应的氨基酸单元可以位于不同的聚赖氨酸链上，不同的聚赖氨酸链可以各自连接一些与m对应的赖氨酸单元，不同的聚赖氨酸链的C端可以各自被聚乙二醇链段所封端。通式中的“random”表示各氨基酸单元之间是无规聚合的。
在载药单分子纳米聚合物及双药单分子纳米聚合物前药的具体实施方案中，所述的双药单分子纳米聚合物前药具有如下结构：
Figure PCTCN2022108093-appb-000046
Figure PCTCN2022108093-appb-000047
其中，m可以选自0-100，n可以选自1-100，k可以选自1-1000，但均不限于此范围。
根据本申请的药物递送系统，它包含双药单分子纳米聚合物胶束，该聚合物胶束包含与亲水性聚合物连接的聚氨基酸，其中在该聚氨基酸的重复单元的α碳上键合了铂类药物活性成分的前药部分和抗肿瘤药物活性成分的前药部分；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
在药物递送系统的一些实施方案中，所述的双药单分子纳米聚合物前药包含与亲水性聚合物连接的无规共聚氨基酸主链。
在药物递送系统的一些实施方案中，所述的亲水性聚合物选自聚(烷二醇)(如，聚乙二醇(“PEG”)、聚(丙二醇)(“PPG”)、乙二醇和丙二醇的共聚物等)、聚(乙氧基化多元醇)、聚(烯醇)、聚(乙烯基吡咯烷酮)、聚(羟烷基甲基丙烯酰胺)、聚(羟烷基甲基丙烯酸酯)、聚(糖)、聚(α-羟基酸)、聚(乙烯醇)、聚膦腈、聚噁唑啉(“POZ”)、聚(N-丙烯酰基吗啉)、聚2-甲基丙烯酰氧基乙基磷酰胆碱(PMPC)以及这些物质的任意组合。
在药物递送系统的优选实施方案中，所述的亲水性聚合物选自聚乙二醇(“PEG”)，优选为甲氧基封端的聚乙二醇。
在药物递送系统的一些实施方案中，所述的铂类药物选自顺铂、卡铂、奈达铂、奥沙利铂和洛铂。
在药物递送系统的优选实施方案中，所述的铂类药物活性成分是顺铂。
在药物递送系统的一些实施方案中，所述分子中含有游离羟基、游离氨基或两者的组合的抗肿瘤药物活性成分是喜树碱类化合物，包括喜树碱及其衍生物或类似物，例如依立替康、拓扑替康、卢比替康、吉咪替康、9-氨基喜树碱和9-硝基喜树碱、7-乙基-10-羟基喜树碱(SN38)等；瑞喹莫德(Resiquimod，R-848)和紫杉醇(Paclitaxel，PTX)。
在药物递送系统的优选实施方案中，所述分子中含有游离羟基、游离氨基或两者的组合的抗肿瘤药物活性成分是喜树碱。
在药物递送系统的一些实施方案中，所述的双药单分子纳米聚合物前药具有前述P100或P200所示结构。
在药物递送系统的一些实施方案中，所述的双药单分子纳米聚合物前药具有如下结构：
Figure PCTCN2022108093-appb-000048
其中，p选自1-500，m可以选自0-100，n可以选自1-100，k可以选自1-1000，但均不限于此范围。
在药物递送系统的一些具体实施方案中，所述的双药单分子纳米聚合物前药具有前述式P101或P201所示结构。
在药物递送系统的一些具体实施方案中，所述的双药单分子纳米聚合物前药具有如下结构：
Figure PCTCN2022108093-appb-000049
其中，m可以选自0-100，n可以选自1-100，k可以选自1-1000，但均不限于此范围。
本申请提供的双药单分子纳米聚合物前药可以通过包括如下步骤的方法制得：
(1)在合适反应条件下，合成抗肿瘤药物活性成分的单NCA单体；优选地，所述抗肿瘤药物活性成分的分子结构中含有游离羟基或游离氨基；
(2)在合适反应条件下，合成铂类药物活性成分的双NCA单体；
(3)在合适反应条件下，使步骤(1)和步骤(2)得到的单体与具有末端氨基的亲水性聚合物反应，得到双药单分子纳米聚合物前药；以及
(4)分离和纯化等处理所得到的双药单分子纳米聚合物前药。
在一些优选实施方案中，所述分子结构中含有游离羟基或游离氨基的抗肿瘤药物活性成分的单NCA单体通过如下所示的方法合成：
Figure PCTCN2022108093-appb-000050
其中的Boc-Lyc-OtBu可通过下式的方法制备得到：
Figure PCTCN2022108093-appb-000051
在一个优选实施方案中，所述分子结构中含有游离羟基或游离氨基的抗肿瘤药物活性成分的单NCA单体通过如下所示的方法合成：
Figure PCTCN2022108093-appb-000052
在一些优选实施方案中，所述铂类药物活性成分的双NCA单体通过如下所示的方法合成：
Figure PCTCN2022108093-appb-000053
在一些优选实施方案中，本申请的双药单分子纳米聚合物前药通过如下所示的方法(一锅法)合成：
Figure PCTCN2022108093-appb-000054
其中，p选自1-500，m可以选自0-100，n可以选自1-100，k可以选自1-1000，但均不限于此范围。
在一些实施方案中，本申请的双药单分子纳米聚合物前药通过包括如下步骤的方法合成：
将具有末端氨基的亲水性聚合物(例如聚乙二醇)溶解于有机溶剂例如苯中，冷冻，然后冷肼真空干燥；
在手套箱中，将干燥后的所述亲水性聚合物溶解于无水有机溶剂例如DMF中，将分子结构中含有游离羟基或游离氨基的抗肿瘤药物活性成分的单NCA单体和铂类药物活性成分的双NCA单体溶于相同有机溶剂中，将所得溶液逐滴缓慢加到反应体系中，密封反应管，从手套箱中取出，在油浴中持续搅拌反应足够时间；
将反应产物缓慢滴入冰乙醚中，得到白色沉淀，弃去上清液，获得纯化的产物；
将获得的产物真空干燥，干燥后的固体溶解于适合的溶剂(例如，DMSO)中，置入透析袋(MWCO：100kDa)，在超纯水中透析数天(期间换水多次)，冷冻干燥后，收集最终产物，即纳米聚合物胶束。
在具体实施方案中，本申请的双药单分子纳米聚合物前药通过包括如下步骤的方法合成：
将具有游离氨基的甲氧基封端的聚乙二醇溶解于苯中，冷冻，然后冷肼真空干燥；
在手套箱中，干燥后的所述聚乙二醇溶解于DMF中，将分子结构中含有游离羟基或游离氨基的抗肿瘤药物活性成分的单NCA单体和铂类药物活性成分的双NCA单体溶于相同有机溶剂中，将所得溶液逐滴缓慢加入到反应体系中，密封反应管，从手套箱中取出，在油浴中持续搅拌反应足够时间；
将反应产物缓慢滴入冰乙醚中，得到白色沉淀，弃去上清液，获得纯化的产物；
将获得的产物真空干燥，干燥后的固体溶解于DMSO中，置入透析袋(MWCO：100kDa)中，在超纯水中透析两天(换水5次)，冷冻干燥后，收集最终产物，即纳米聚合物胶束。
本申请的另一个方面，还提供一种载药单分子纳米聚合物胶束，其组成选自如下任一种：前述载药单分子纳米聚合物，前述的制备方法制备得到的载药单分子纳米聚合物，前述的双药单分子纳米聚合物前药，和前述的制备方法制得的双药单分子纳米聚合物前药；所述载药单分子纳米聚合物胶束具有核壳结构，外壳结构为亲水性聚合物链形成的亲水层，所包载的药物单元位于内核中。
通过调节L Pt的分布密度可控制该载药单分子纳米聚合物具有支化或适度交联的三维结构，进一步结合亲水性聚合物链在聚氨基酸链端部的位置设计，使载药单分子纳米聚合物能够在水性介质中无需自组装就可形成具有核壳结构的单分子纳米聚合物胶束，亲水性聚合物链分布于外壳，药物成分被包载于内核。
本申请的另一个方面，还提供一种药物递送系统，它包含载药单分子纳米聚合物胶束，该载药单分子纳米聚合物胶束包含前述载药单分子纳米聚合物或前述的制备方法制备得到的载药单分子纳米聚合物；优选地，所述亲水性聚合物链位于所述载药单分子纳米聚合物胶束的外壳；所述铂类药物单元以及所述第二药物单元均位于所述载药单分子 纳米聚合物胶束的内核。
本申请中，涉及载药单分子纳米聚合物胶束的尺寸，如无特别说明，测试温度在20～30℃，进一步为25℃。
载药单分子纳米聚合物胶束的尺寸和形貌表征，可以采用下文实施例中的测试方法。
本申请中，涉及载药单分子纳米聚合物胶束的粒径，仅在特别说明时指平均直径或平均粒径。
本申请中，涉及胶束尺寸测定时，测试条件“水中”可以为纯水，也可以为水溶液。水溶液的举例如缓冲液(如PBS溶液)、生理模拟液等。
在一些实施方案中，载药单分子纳米聚合物胶束粒径或粒径范围选自10～120nm，优选为10～110nm，另优选为10～100nm，另优选为10～80nm，另优选为10～50nm，另优选为10～40nm，另优选为10～30nm，另优选为15～120nm，另优选为15～110nm，另优选为15～100nm，另优选为15～80nm，另优选为15～50nm，另优选为15～40nm，另优选为15～30nm，另优选为20～120nm，优选为20～110nm，另优选为20～100nm，另优选为20～80nm，另优选为20～70nm，另优选为20～50nm，另优选为20～40nm，另优选为25～120nm，优选为25～110nm，另优选为25～100nm，另优选为25～80nm，另优选为25～50nm，另优选为25～40nm，另优选为25～35nm，另优选为30～35nm。载药单分子纳米聚合物胶束粒径还可以选自如下任一种或任两种构成的区间：15nm、16nm、18nm、20nm、25nm、30nm、35nm、40nm、45nm、50nm、55nm、60nm、65nm、70nm、80nm、90nm、100nm、110nm、120nm等。测试温度可以在20～30℃，进一步为25℃。可以为水中的动态光散射测试结果，也可以为透射电镜测试结果。
在一些实施方案中，透射电镜法(可参考实施例4)测试的载药单分子纳米聚合物胶束的粒径≤120nm，进一步≤100nm，更进一步≤90nm，更进一步≤80nm，更进一步≤70nm，更进一步≤60nm，更进一步≤50nm。
在一些实施方案中，载药单分子纳米聚合物胶束的平均直径选自15～50nm，还可以为15～40nm，还可以为20～40nm，还可以为25～35nm。测试温度可以在20～30℃，进一步为25℃。可以为水中的动态光散射测试结果，也可以为透射电镜测试结果。
在一些实施方案中，胶束内核半径为5～50nm，还可以为5～45nm，还可以为5～40nm，还可以为5～35nm，还可以为5～30nm，还可以为5～25nm，还可以为5～20nm，还可以为5～15nm，还可以为5～10nm，还可以为10～50nm，还可以为10～45nm，还可以为10～40nm，还可以为10～35nm，还可以为10～30nm，还可以为10～25nm，还可以为10～20nm，还可以为6～8nm。胶束内核半径还可以选自如下任一种尺寸或任两种构成的区间：5nm、6nm、7nm、nm、9nm、10nm、11nm、12nm、13nm、14nm、15nm、16nm、17nm、18nm、19nm、20nm、22nm、24nm、25nm、30nm、35nm、40nm、45nm、50nm等。测试温度可以在20～30℃，进一步为25℃。可以为水中测试结果。进一步可以为小角X射线散射(SAXS)测试结果。
在一些实施方案中，25℃时水中的胶束外壳厚度为5～40nm，还可以为5～35nm，还可以为5～30nm，还可以为5～25nm，还可以为5～20nm，还可以为5～15nm，还可以为10～40nm，还可以为10～35nm，还可以为10～30nm，还可以为10～25nm，还可以为 10～20nm，还可以为8～12nm。25℃时水中的胶束外壳厚度还可以选自如下任一种尺寸或任两种构成的区间：5nm、6nm、7nm、nm、9nm、10nm、11nm、12nm、13nm、14nm、15nm、16nm、17nm、18nm、19nm、20nm、22nm、24nm、25nm、30nm、35nm、40nm等。可以为水中测试结果。进一步可以为小角X射线散射(SAXS)测试结果。
本申请的另一个方面，还提供一种药物递送系统，它包含双药单分子纳米聚合物胶束，该双药单分子纳米聚合物胶束包含与亲水性聚合物连接的聚氨基酸，其中在该聚氨基酸的重复单元的α碳上键合了铂类药物活性成分的前药部分和抗肿瘤药物活性成分的前药部分；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
在一些实施方案中，其中所述的亲水性聚合物选自聚乙二醇、聚(丙二醇)、乙二醇和丙二醇的共聚物、聚(乙氧基化多元醇)、聚(烯醇)、聚(乙烯基吡咯烷酮)、聚(羟烷基甲基丙烯酰胺)、聚(羟烷基甲基丙烯酸酯)、聚(糖)、聚(α-羟基酸)、聚(乙烯醇)、聚膦腈、聚噁唑啉、聚(N-丙烯酰基吗啉)以及这些物质的任意组合。
在一些实施方案中，其中所述的铂类药物选自顺铂、卡铂、奈达铂、奥沙利铂和洛铂中的一种或多种。
在一些实施方案中，其中所述分子中含有游离羟基、游离氨基或两者的组合的抗肿瘤药物活性成分选自喜树碱类化合物、瑞喹莫德和紫杉醇中的一种或多种；优选地，所述喜树碱类化合物包括喜树碱及其衍生物或类似物，更优选地，所述喜树碱类化合物包括依立替康、拓扑替康、卢比替康、吉咪替康、9-氨基喜树碱、9-硝基喜树碱和7-乙基-10-羟基喜树碱。
因此，根据上述方法以及在以下实施例中举例说明的方法，本领域技术人员能够轻易地制备得到本申请的载药单分子纳米聚合物胶束或双药物活性成分的纳米聚合物前药胶束。
本申请的另一个方面，还提供铂类药物活性成分的双NCA单体和抗肿瘤药物活性成分的单NCA单体在制备单分子纳米聚合物前药或药物递送系统中的用途；优选地，所述抗肿瘤药物活性成分的分子结构中含有游离羟基或游离氨基。
本申请的再一个目的在于提供前述载药单分子纳米聚合物作为前药的用途。该载药单分子纳米聚合物能够进入到细胞内部，感知细胞内微环境，响应性地释放药物活性成分，产生细胞毒性，抑制肿瘤细胞的生长。
一些实施方案中，药物释放机理为：(a)在细胞内高还原性微环境，四价铂经还原，能够脱掉与聚氨基酸连接的配体，从而实现二价铂活性物种在细胞内的选择性释放。(b)在细胞内高还原环境下，尤其与细胞内高浓度的谷胱甘肽发生还原反应，二硫键断裂，产生游离巯基，巯基进而攻击与抗肿瘤药物连接的碳酸酯或氨酯键，从而实现抗肿瘤活性药物在细胞内的选择性释放。
以第二药物单元为喜树碱(CPT)的残基、响应性连接基L R为二硫键，且式(II)所示结构单元包括如下结构为例，释放活性CPT药物的分子机理可以如下：
Figure PCTCN2022108093-appb-000055
以铂类药物单元为顺铂的残基，且式(I)所示结构单元包括如下结构为例，由四价铂释放活性Pt(II)药物的分子机理可以如下：
Figure PCTCN2022108093-appb-000056
具体可以如下：
Figure PCTCN2022108093-appb-000057
本申请的另一个方面，还提供前述载药单分子纳米聚合物，前述的制备方法制备得到的载药单分子纳米聚合物，前述的双药单分子纳米聚合物前药，前述的制备方法制得的双药单分子纳米聚合物前药，前述载药单分子纳米聚合物胶束，或前述的药物递送系统在制备用于治疗肿瘤疾病的药物中的用途。
肿瘤疾病可以包括但不限于肺癌、胃癌、膀胱癌、卵巢癌、睾丸癌、子宫内膜癌、骨癌、肉瘤、宫颈癌、食管癌、肝癌、结直肠癌、头颈部癌、绒毛膜上皮癌、恶性葡萄胎、非霍奇金淋巴瘤和急性及慢性粒细胞性白血病等。还可包括但不限于肺癌、食管癌、头颈部肿瘤，
在此基础上，本领域技术人员根据本说明书中所述的方法和本领域中已知的技术可以容易地进一步制备得到本申请的载药单分子纳米聚合物胶束或纳米聚合物前药胶束的所期望剂型的制剂。
本申请的载药单分子纳米聚合物或双药物活性成分的纳米聚合物前药可用于治疗肺癌、胃癌、膀胱癌、卵巢癌、睾丸癌、子宫内膜癌、骨癌、肉瘤、宫颈癌、食管癌、肝癌、结直肠癌、头颈部癌、绒毛膜上皮癌、恶性葡萄胎、非霍奇金淋巴瘤和急性及慢性粒细胞性白血病等。本申请的纳米制剂也可用于增加肿瘤细胞对放疗的敏感性，放疗同时给药可加强对肺癌、食管癌、头颈部肿瘤的局部进展的控制。
在一些实施方案中，本申请提供一种根据本申请的双药物活性成分的纳米聚合物在制备用于治疗所述肿瘤的药物中的用途。
在另一些实施方案中，本申请提供一种联合用药治疗有需要的患者中所述肿瘤的方法，该方法将治疗有效量的本申请的单分子纳米聚合物前药或其制剂给予所述患者。
本领域技术人员使用本领域中已知的方法可以将本申请的双药物活性成分的纳米聚合物前药配制成任何所需的药物制剂。根据本申请的药物制剂的剂型可以是临床上适 用于治疗所述疾病的任何剂型，包括溶液、混悬液、凝胶、冻干粉、胶囊或片剂等。优选地，本申请药物制剂的剂型是适用于注射(例如静脉输注)的剂型。
如果将根据本申请的纳米聚合物胶束配制用于通过注射给药，例如通过静脉内输注，则该用于注射的制剂可以以单位剂型存在，存在于例如安瓿、西林瓶、预灌封注射器或多剂量容器中。
本领域技术人员理解，根据本申请的药物制剂除包含根据本申请的纳米聚合物胶束以外还可以根据需要包含至少一种药学上可接受的赋形剂，例如等渗剂、湿润剂、增溶剂、乳化剂、防腐剂、缓冲剂、酸化基、碱化剂、抗氧化剂、螯合剂、着色剂、络合剂、矫味剂、助悬剂以及润滑剂中的一种或多种。这些赋形剂是本领域中已知的，本领域技术人员根据本申请的内容可以选择合适的一种或多种赋形剂添加到本申请的药物制剂中。
可以通过口服、肌内注射、腹膜内注射、静脉内注射和皮下注射等途径将根据本申请的药物制剂施用于有需要的患者以治疗该患者的疾病，例如上述的肿瘤。
有关领域的临床医生可以根据所治疗的疾病的性质、治疗的时间以及患者的年龄和身体状况，选择和确定本申请的纳米聚合物胶束或其制剂的给药方案以提供所需的治疗效果。可以使用单一剂量或以合适间隔给予多次剂量方便地给予所需剂量，例如每天给药1次、2次或更多次合适的剂量。
本领域技术人员可以根据所含药物活性成分的种类和量轻易地确定本申请的纳米聚合物胶束或其制剂的适用剂量。
在所述肿瘤的治疗中，可以与其它化学治疗剂和/或辐射联合给予本申请的纳米聚合物胶束或其制剂。
实施例
为进一步说明本申请，提供下面的实施例。这些实施例仅用于举例说明本申请，本申请的范围不限于所提供的实施例。
下面将结合实施例对本申请的实施方案进行详细描述。应理解，这些实施例仅用于说明本申请而不用于限制本申请的范围。下列实施例中未注明具体条件的实验方法，优先参考本申请中给出的指引，还可以按照本领域的实验手册或常规条件，还可以按照制造厂商所建议的条件，或者参考本领域已知的实验方法。
下述的具体实施例中，涉及原料组分的量度参数，如无特别说明，可能存在称量精度范围内的细微偏差。涉及温度和时间参数，允许仪器测试精度或操作精度导致的可接受的偏差。
除非特别说明，否则下面实施例中使用的原料、实验试剂和实验仪器均可从市场购得，所使用的反应条件为本领域中已知，并且所使用的鉴定或测定方法为本领域中常用的方法。
以下各例中，如无其他说明，Diboc为二叔丁氧羰基二碳酸酯；DMAP为4-二甲氨基吡啶；THF为四氢呋喃；DMF为N,N-二甲基甲酰胺；EDC为1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐；NHS表示N-羟基琥珀酰亚胺；BTC表示双(三氯甲基)碳酸酯， Bis(trichloromethyl)carbonate；Tween 80表示吐温80；DACHPt表示(1,2-二氨基环己烷)二氯化铂；CPT对应喜树碱；GSH表示谷胱甘肽。
以下各例中，涉及聚乙二醇的“分子量”，如无其他说明，指数均分子量。
以下各例中，涉及单分子纳米聚合物的分子量，如无其他说明，指数均分子量。
以下各例中，如无其他说明，指反应式中的“rt”表示室温。
以下各例中，如无特别说明，质谱测试条件为：待检测物配置成1mg/mL二氯甲烷或甲醇溶液，滴加0.5μL溶液至样品台，室温干燥后，将样品台送入离子源进行测试(Bruker REFLEX III型MALDI-TOF-MS)。
以下各例中，如无特别说明， 1H NMR测试条件为：待检测物配置成10mg/mL CDCl3_溶液，采用Bruker AVANCE 500Ⅲ型超导脉冲傅里叶变换核磁共振波谱仪建立 1H NMR图谱，测试温度25℃，扫描次数64次，内标为四甲基硅烷(TMS)。
以下各例中，％(w/v)表示质量体积百分比，％(v/v)表示体积比。
实施例1.分子中含有游离羟基、游离氨基或两者的组合的药物活性分子前药的制备。
Figure PCTCN2022108093-appb-000058
1.1.N-Boc-N`-Cbz-Lys-OtBu的合成
Figure PCTCN2022108093-appb-000059
N-Boc-N`-Cbz-L-Lys(2g，5.26mmol)溶解于氯仿(15mL)，与碳酸氢钠溶液(12mL 0.45mmol/L)混合。在氮气保护下搅拌5分钟，后逐滴加入内含Diboc(二叔丁氧羰 基二碳酸酯)的氯仿溶液(1.22g，5.5mmol，9mL)，回流反应90分钟，冷却至室温。分离有机相，并用氯仿萃取水相，合并有机相减压蒸干溶剂，柱层析得到N-Boc-N`-Cbz-Lys-OtBu。
该中间产物的 1H NMR谱图见图4中IV-A。
1.2.Boc-Lys-OtBu的合成
Figure PCTCN2022108093-appb-000060
N-Boc-N`-Cbz-Lys-OtBu(436.5mg，1.0mmol)溶解于甲醇(5mL)溶解，加入10％Pd/C(53.2mg，0.05mmol)，充分混匀后，氢气置换出反应瓶中空气，并在气球压力下进行氢化反应，室温搅拌反应2小时，通过硅藻土过滤分离不溶物，并用甲醇洗脱，减压蒸馏除去溶剂，得到黄色油状物。
该中间产物的质谱图参见图3中III-A。该中间产物的 1H NMR谱图参见图4中IV-B。
1.3.CPT-ss-OH的合成
Figure PCTCN2022108093-appb-000061
喜树碱(500mg，1.43mmol)分散于无水二氯甲烷溶解(80mL)，在冰浴中，在氮气保护下加入含有三光气(157mg，0.53mmol)的无水二氯甲烷(3mL)，继续在冰浴下搅拌30分钟，加入溶解有DMAP(4-二甲氨基吡啶，560mg，4.6mmol)的无水二氯甲烷(10mL)至喜树碱完全溶解，在冰浴下继续搅拌反应1小时，转入室温继续避光搅拌反应1小时。在N 2保护下加入溶有2-羟乙基二硫化物(1.75mL，14.3mmol)的无水THF(四氢呋喃，15mL)，混合均匀后避光室温搅拌反应24小时。待反应结束后，再加入50mL二氯甲烷，并依次用0.1M HCl水溶液、饱和的NaCl和水各洗3次，收集有机相并用无水Na 2SO 4干燥，通过旋转蒸发仪减压蒸馏除去有机溶剂。得到的黄色固体通过氯仿/甲醇(3/10,v/v)重结晶纯化，得到淡黄色晶体CPT-ss-OH。
该中间产物的质谱图见图3中III-B。该中间产物的 1H NMR谱图见图4中IV-C。
1.4.Boc-Lys-OtBu-ss-CPT的合成
Figure PCTCN2022108093-appb-000062
CPT-ss-OH(52.8mg，0.1mmol)分散至无水二氯甲烷(15mL)，在N 2环境下，加入1mL溶解有三光气(13.2mg，0.045mmol)的无水二氯甲烷，在冰浴下搅拌30分钟，随后加入溶解有DMAP(39mg，0.32mmol)的无水二氯甲烷(2mL)至完全溶解，在冰浴下继续搅拌反应1小时，转入室温继续避光搅拌反应1小时。在N 2保护下加入溶有Boc-Lys-OtBu(45.3mg，0.15mmol)的无水二氯甲烷(1mL)，混合均匀后避光室温搅拌反应24小时。待反应结束后，加入50mL二氯甲烷，并依次用0.1M HCl水溶液、饱和的NaCl和水各洗3次，收集有机相并用无水Na 2SO 4干燥，通过柱层析分离得到淡黄色晶体Boc-Lys-OtBu-ss-CPT。
该中间产物的 1H NMR谱图见图4中IV-D。
1.5.Lys-ss-CPT的合成
Figure PCTCN2022108093-appb-000063
Boc-Lys-OtBu-ss-CPT粉末(85.7mg，0.1mmol)溶解于用二氯甲烷(2mL)溶解，与三氟乙酸(2mL)混合，室温反应2小时，减压蒸馏去除溶剂，加入二氯甲烷溶解，并用饱和的碳酸氢钠洗涤，收集有机相，干燥得到Lys-ss-CPT。
1.6.NCA-Lys-ss-CPT的合成
Figure PCTCN2022108093-appb-000064
Lys-ss-CPT(0.1mmol)溶解于用二氯甲烷(2mL)溶解，与含有三光气(0.2mmol)的二氯甲烷(2mL)混合，室温反应2个小时，减压蒸馏去除溶剂，加入四氢呋喃溶解(60℃)，加入适量正己烷，在4℃冰箱结晶，收集白色针状晶体NCA-Lys-ss-CPT。
该喜树碱前药的质谱图见图3中III-C。
实施例2.铂类药物活性成分前药的制备。
Figure PCTCN2022108093-appb-000065
2.1.化合物(4)的合成
Figure PCTCN2022108093-appb-000066
顺铂(1.0g，3.33mmol)分散于蒸馏水中，加入30％H 2O 2(20ml)，70℃下避光搅拌5h至澄清，降至室温后置于4℃冰箱进行重结晶。析晶后过滤，依次使用冰水、乙醇、乙醚洗涤滤饼，干燥得到黄色晶体化合物(1)。
称取化合物(1)(0.5mg，1.5mmol)与琥珀酸酐(0.62g，6.15mmol)溶解于DMF(N,N-二甲基甲酰胺，20ml)中，70℃下避光搅拌过夜。减压蒸馏除去溶剂，依次使用甲醇、丙酮洗涤后干燥得到淡黄色化合物(4)。
化合物(4)的 1H NMR谱图见图4中IV-E。
2.2.N-Boc-N`-Cbz-Lys-OtBu和Boc-Lys-OtBu的合成
Figure PCTCN2022108093-appb-000067
2.3.N-Boc-N`-Cbz-Lys-OtBu的合成
Figure PCTCN2022108093-appb-000068
N-Boc-N`-Cbz-L-Lys(2g，5.26mmol)溶解于氯仿(15mL)，与碳酸氢钠溶液(12mL 0.45mmol/L)混合。在氮气保护下搅拌5分钟，后逐滴加入内含Diboc的氯仿溶液(1.22g，5.5mmol，9mL)，回流反应90分钟，冷却至室温。分离有机相，并用氯仿萃取水相，合并有机相减压蒸干溶剂，柱层析得到N-Boc-N`-Cbz-Lys-OtBu。
2.4.Boc-Lys-OtBu的合成
Figure PCTCN2022108093-appb-000069
N-Boc-N`-Cbz-Lys-OtBu(436.5mg，1.0mmol)溶解于甲醇(5mL)溶解，加入10％Pd/C(53.2mg，0.05mmol)，充分混匀后，氢气置换出反应瓶中空气，并在气球压力下进行氢化反应，室温搅拌反应2小时，通过硅藻土过滤分离不溶物，并用甲醇洗脱，减压蒸馏除去溶剂，得到黄色油状物。
2.5.化合物(5)的合成
Figure PCTCN2022108093-appb-000070
化合物(4)(0.12mmol)溶解于无水DMF(5mL)，与含有EDC(1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐，0.15mmol)，DMAP(0.05mmol)的无水DMF溶液(5mL)混合，反应30min，加入含有化合物(3)(0.1mmol)的无水DMF溶液(5mL)混合，在室温下搅拌反应6分小时。通过柱层析分离得到淡黄色固体(5)。
2.6.化合物(6)的合成
Figure PCTCN2022108093-appb-000071
化合物(5)(0.1mmol)溶解于DMF(2mL)溶解，与三氟乙酸(2mL)混合，室温反应2小时，减压蒸馏去除溶剂，减压蒸馏除去溶剂，依次使用甲醇、丙酮洗涤后干燥得到淡黄色化合物(6)。
2.7.化合物(7)的合成
Figure PCTCN2022108093-appb-000072
化合物(6)(0.1mmol)溶解于DMF(2mL)溶解，与含有三光气(0.2mmol)的DMF(2mL)混合，室温反应2个小时，减压蒸馏去除溶剂，加入四氢呋喃溶解(60℃)，加入适量正己烷，在4℃冰箱结晶，收集白色针状晶体(7)NCA-Pt-NCA。化合物(7)的质谱图见图3中III-D。
实施例3.本申请载药单分子纳米聚合物(一种双药单分子纳米聚合物，可作为纳米聚 合物前药)及其纳米胶束的制备
本例中，载药单分子纳米聚合物具有前文所述式(I-2)所示的四价结构单元，式(III-2)所示的一价结构单元、以及式(II-2)所示的二价结构单元；
Figure PCTCN2022108093-appb-000073
进一步地，在本例中，n11和n21每次出现均为4，n12和n22每次出现均为2；
n31每次出现均为4，n32每次出现均为2，n33每次出现均为2；L R每次出现均为-S-S-；
n51每次出现均为3；Z 5每次出现均为NH；
更进一步地，D Pt为顺铂的残基，D T为喜树碱的残基；
更进一步地，p约等于113，对应的mPEG的分子量约为5000Da(数均分子量)。
此时，采用式(I-3)所示单体、式(III-3)所示单体和式(II-3)所示单体制备载药单分子纳米聚合物。
进一步地，式(I-3)所示单体的结构为实施例2制备的NCA-Pt-NCA，式(II-3)所示单体的结构为实施例1制备的NCA-Lys-ss-CPT，式(III-3)所示单体的结构为
Figure PCTCN2022108093-appb-000074
制备载药单分子纳米聚合物的反应方程式如下(制备的载药单分子纳米聚合物记为P101)：
Figure PCTCN2022108093-appb-000075
其中，m可以选自0-100，n可以选自1-100，k可以选自1-1000，但均不限于此范围。
甲氧基-聚乙二醇-氨基(MeO-PEG-NH 2，0.1g，0.01mmol)溶解于苯(3mL)，搅拌至PEG完全溶解，液氮冷冻，接冷肼真空干燥处理6小时。然后手套箱中，干燥后的聚乙二醇溶解于无水DMF(2mL)，搅拌均匀，取NCA-Lys-ss-CPT(0.50mmol)和NCA-Pt-NCA(0.18mmol)溶于无水DMF(2mL)中，逐滴缓慢加入到反应体系中，密封反应管，从手套箱中取出，置于35℃油浴中持续搅拌反应72h。将反应产物缓慢滴入冰乙醚中，得到白色沉淀，弃去上清液并重复上述操作三次，获得纯化的产物。产物置于真空锅中真空干燥6小时。干燥后的固体溶解于DMSO(二甲基亚砜，2mL)，置入透析袋(MWCO：100kDa)中，在超纯水中透析两天(换水5次)，冷冻干燥后，收集最终产物(P101)。
Figure PCTCN2022108093-appb-000076
其中，m可以选自0-100，n可以选自1-100，k可以选自1-1000，但均不限于此范围。
使用上文关于制备喜树碱和顺铂双药聚合物前药纳米胶束的方法，用紫杉醇和瑞喹莫德分别代替喜树碱，制得了紫杉醇和顺铂双药聚合物前药纳米胶束以及瑞喹莫德和顺铂双药聚合物前药纳米胶束。
紫杉醇和顺铂双药聚合物前药纳米胶束以及瑞喹莫德和顺铂双药聚合物前药纳米胶束的动态光散射特性和释药特性见图20-25。
实施例4.本申请双药单分子聚合物前药纳米胶束的表征
4.1.本申请中使用的对照胶束
A：CPT@PEG-PLA
(参照以下论文：金滔，羟喜树碱MePEG-PLA纳米粒的制备及其体外抗肿瘤研究，浙江中医药大学，硕士论文，2013-05-01)
B：顺铂@PEG-Gluc(ss-CPT)
(参照以下专利文献：一种铂交联喜树碱前药胶束纳米药物及其制备方法和应用，CN109908084A)
C：顺铂@PEG-PGlu
(参照专利文献：CN101203549B，CN100457185C，CN100344293C)
D：顺铂(市购)
4.2.理化表征：分子量，GPC(SEC)等
将冻干的产物溶解于水中(1mg/mL)，利用GPC(superdex200)表征产物分子量分布，如图5中V-A和图5中V-B所示，大分子量产物成功合成。利用分子排阻色谱法定量聚合后分子量，结果如图5中V-A和V-B中所示。
将冻干的产物溶解于水中(1mg/mL)，利用分析型超高速离心技术，定量产物分子量约为1030kDa。利用分析型超速离心定量聚合后的分子量。
结果：PEG的分子量为5kDa(式(III-2)中p约为113)，反应后纳米粒子的分子量为1030kDa。
将冻干的产物溶解于水中(1mg/mL)，利用荧光相关光谱仪(FCS)定量产物的扩散时间约为7600μs。利用荧光相关谱定量聚合后的分子量。
结果：PEG的扩散时间为121μs，反应后纳米粒子的扩散时间为7600μs。
4.3.粒径表征：
配置纳米制剂(0.01mg/mL)PBS缓冲溶液(10mM，pH 7.4)，通过动态光散射仪对纳米制剂的粒径和多分散指数(PDI)进行表征。结果如图6中所示。根据图6，本申请的双药单分子纳米聚合物胶束的平均粒径为33.6纳米，粒径范围为21.5～52.7纳米，粒径分布的多分散指数PDI约为0.05；而对照胶束的粒径范围为32.3～264.1纳米，平均粒径约为96.7纳米，粒径的多分散指数PDI大于>0.1，具体在0.18～0.30范围内。
4.4.TEM形貌测试：
配置纳米制剂水溶液(0.01mg/mL)，将TEM铜网浸润在溶液中，使溶液完全浸没铜网，沉淀30min，镊子取出铜网，用滤纸吸去多余溶液，自然风干后，用质量分数2％的磷钨酸水溶液进行负染2min，再次自然风干，干燥后将铜网置于透射电子显微镜进行观察和拍照。结果如图7(VII-A，及VII-B)中所示。
化学聚合得到的纳米粒子具有高度尺寸均一性，而自组装得到的纳米粒子尺寸均一性不高。
透析后溶液经过3次冻干，复溶(1mg/mL)，比较透析后以及3次往返冻干复溶处理后的形貌测试，双药单分子聚合物前药纳米胶束保持原始形貌，而通过自组装形成的纳米制剂无法耐受冻干复溶工艺。结果如图8(VIII-A，VIII-B)中所示。
结论：化学聚合获得的双药单分子聚合物前药纳米胶束具有优异的胶体溶液稳定性，耐受冻干复溶，结构稳定。自组装得到的纳米粒子不耐受冻干复溶，复溶后无法获得原始结构。
4.5.荧光关联光谱法(FCS)测试胶体动力学特征.
将双药单分子聚合物前药纳米胶束溶液(0.1mg/mL)与自组装形成的纳米胶束溶液(0.1mg/mL)与水混合稀释，利用荧光关联光谱法(FCS)测试胶体动力学特征。结果如图9中所示。
结果显示，化学聚合得到双药单分子聚合物前药纳米胶束耐受稀释，稀释10000倍，仍保持原始结构。自组装得到的纳米制剂不耐受稀释，稀释100后，自组装结构解离。
4.6.小角X射线散射(SAXS)测试
将双药单分子聚合物前药纳米胶束溶液(0.1mg/mL)送小角X射线散射(SAXS)测试。结果如图10所示。
结果现实显示合成的产物是具有纳米尺寸的胶束结构，内核的半径约为6.4纳米，外壳PEG层厚度约为9.8纳米。胶束的直径约为32.4纳米。
4.7.胶体稳定性
药物制剂在水溶液中的分散以及血液循环中会遭受机械剪切力的作用，因此，纳米制剂的胶体稳定性对于药物制剂的药代动力学及其产业化工艺意义重大。本试验考察本申请双药单分子聚合物前药纳米胶束以及自组装纳米制剂顺铂@PEG-PGlu(ss-CPT)在超声处理下，利用动态光散射测试其胶体结构的稳定。其中，超声参数(1.0MHz，9.9W)，超声时间(0min，5min，60min)，将超声后的溶液送动态光散射仪分析纳米制剂的粒径分布。结果如图11中所示。
结论：经化学聚合得到的单分子双药纳米胶束能够极好的耐受超声处理，粒径保持稳定，相反，自组装纳米制剂顺铂@PEG-PGlu(ss-CPT)无法耐受超声处理，结构明显变形。
根据图11，对于本申请的双药单分子纳米聚合物胶束，在超声前，平均粒径为33.6纳米，粒径范围为21.5-52.7纳米，粒径分布的多分散指数PDI约为0.05；在经过60min超声处理后，平均粒径为33.9纳米，粒径范围为22.1-53.4纳米，粒径分布的多分散指数PDI约为0.05；因此，本申请的双药单分子纳米聚合物胶束能够极好的耐受超声处理，粒径保持稳定。
对于自组装纳米制剂顺铂@PEG-PGlu(ss-CPT)对照胶束，在超声前，平均粒径约为96.7纳米，粒径范围为32.3～264.1纳米，粒径的多分散指数PDI约为0.18；在经过60min超声处理后，平均粒径约为342纳米，粒径范围为7.6～464.1纳米，粒径的多分散指数PDI约为0.64；因此，自组装纳米制剂顺铂@PEG-PGlu(ss-CPT)无法耐受超声处理， 结构明显变形。
实施例5.本申请双药单分子聚合物前药纳米胶束响应胞内还原微环境(释放活性原药的研究)
5.1.药物释放机理
释放活性CPT药物的分子机理
Figure PCTCN2022108093-appb-000077
本申请的单分子纳米聚合物中的四价铂在细胞内经过还原反应，释放活性Pt(II)药物的分子机理可如下所示：
Figure PCTCN2022108093-appb-000078
5.2.体外模拟释药
考察本申请双药单分子聚合物前药纳米胶束在不同条件下的释药行为：(1)胞外微环境：pH 7.4，GSH(0mM)、(2)胞内微环境：pH 7.4，GSH(10mM)。将含有上述纳米制剂的PBS溶液(10mM)注入透析袋(MWCO：10000)并浸入上述两种PBS溶液(10mM，28mL，含0.5％(w/v)Tween 80)中，然后在摇晃(100RPM)下，以37℃孵育48小时。在预定的间隔时间点提取1mL释放介质，并补充1mL新鲜的空白介质。根据HPLC(图12)测定透析物中CPT的浓度，流动相为甲醇和去离子水(20–100％，v/v)，流速为1.0mL/min，25℃，吸收波长为370nm。通过ICP-MS测定透析物中的Pt的浓度。结果如图13中所示。
结论：通过化学聚合形成的本申请双药单分子聚合物前药纳米胶束在胞外不会提前释放活性药物，胞内呈现出触发性释放活性药物的功能。
5.3.体外模拟释药
对照胶束顺铂@PEG-PGlu(ss-CPT)、CPT@PEG-PLA在胞外微环境(pH 7.4，GSH(0mM)。将含有上述纳米制剂的PBS溶液(10mM)注入透析袋(MWCO：10kDa)并浸入上述两种PBS溶液(28mL，10mM，含0.5％(w/v)Tween 80)中，在温和的摇晃(100RPM)下，37℃孵育48小时。在预定间隔时间点提取1mL释放介质，并补充1mL新鲜的空白介质。通过HPLC测定透析物中CPT的浓度，流动相为甲醇和去离子水(20％-100％，v/v)，流速为1.0mL/min，25℃，吸收波长为370nm。通过ICP-MS测定透析物中的Pt浓度。结果如图14中所示。
结论：自组装纳米制剂在细胞外同样会持续性的释放药物。
实施例6.本申请双药单分子聚合物前药纳米胶束的细胞毒性
使用MTT法对顺铂、顺铂@PEG-PGlu、顺铂@PEG-PGlu(ss-CPT)、CPT@PEG-PLA、游离CPT&顺铂双药以及本申请单分子纳米前药的细胞毒性进行分析。以10 5个/mL的密度将A549细胞接种于96孔板(100μL)。在37℃、5％CO 2的细胞培养箱中孵育24h，弃去旧培养基，然后每孔加入100μL含有不同浓度的药物制剂，不加药物组为100％对照，不含细胞的对照孔为0％空白对照,培养48h后，检测细胞的存活率。检测时弃掉细胞的培养液,每孔加入100μL MTT溶液(0.5mg/mL)继续培养4h，小心吸去培养液并加入DMSO(100μL/孔)溶解蓝紫色结晶物，稍微振荡，使用酶标仪测定在570nm处的吸光度值(参比波长630nm)。细胞存活率的计算公式：细胞存活率(％)＝[(OD样品孔-OD空白组)/(OD对照孔-OD空白组)]×100％。结果如图15中所示。
实施例7.本申请双药单分子聚合物前药纳米胶束的药物动力学研究
采用尾静脉注射放将药物制剂注入雌性Bal b/c小鼠(8周)，注射剂量；200微升，喜树碱(CPT)5mg/kg，铂(Pt)1.8mg/kg，其中以等量的双药自组装纳米制剂顺铂@PEG-PGlu(ss-CPT)以及游离顺铂为对照组。静脉注射后，不同时间点取血，利用ICP-MS测定Pt含量，其中对血浆超速离心处理(10000G)用于分取游离小分子顺铂以及内含Pt的纳米制剂。结果表明，相比于游离顺铂溶液对照组，纳米制剂的血液循环时间呈现出显著性的延长，其中本申请制备的单分子纳米前药在血液循环中不会出现药物早期泄漏(即：血液循环中Pt总量与血液循环中纳米制剂Pt量一致)，而通过自主装形成的纳米制剂顺铂@PEG-PGlu(ss-CPT)在血液循环中会出现药物的提前泄漏(即：即：血液循环中Pt总量高于血液循环中纳米制剂Pt量)。结果如图16(图16中A，图16中B)中所示。
由所示实验结果可以得出结论：对于本申请的单分子纳米前药，相比游离顺铂溶液对照组，单分子纳米前药血液循环时间显著性延长，且单分子纳米前药在血液循环中不会提前泄漏游离顺铂药物；而对于作为对照的自组装纳米制剂，相比游离顺铂溶液对照组，自组装纳米制剂的血液循环时间显著性延长，且自组装纳米制剂在血液循环中会提前泄漏游离顺铂药物。
实施例8.本申请双药单分子聚合物前药纳米胶束的肿瘤药物累积量试验
本试验中使用的肿瘤模型为A549肺癌皮下移植瘤，采用尾静脉注射给药方式将药 物制剂注入雌性Bal b/c nude小鼠(8周)，注射剂量；200微升，CPT 5mg/kg，Pt 1.8mg/kg，其中以顺铂为参照试验组，以等量的双药自组装纳米制剂顺铂@PEG-PGlu(ss-CPT)对照组。静脉注射后的不同时间点，解剖取得肿瘤块，浓硝酸充分溶解肿瘤，肿瘤内Pt含量通过ICP-MS测定。结果如图17中所示。
结果表明，顺铂对照组在肿瘤部位药物浓度较低，纳米制剂能够依靠纳米制剂对于肿瘤的EPR效应显著性地富集在肿瘤部位，随着时间延长，肿瘤药物富集量逐渐升高。其中本申请制备的单分子纳米前药富集量明显优于装双药自组装纳米制剂顺铂@PEG-PGlu(ss-CPT)，可能的原因包括单分子纳米前药在血液循环中更优异的胶体结构稳定性，防止药物早期泄漏性能。
实施例9.本申请双药单分子聚合物前药纳米胶束的活体抑癌功效评价
本试验中使用的肿瘤模型为A549肺癌皮下移植瘤，采用尾静脉注射给药方式将药物制剂注入雌性Bal b/c nude小鼠(8周)，注射剂量；200微升，CPT 5mg/kg，Pt 1.8mg/kg，其中以PBS为参照试验组，以等量的单药自组装纳米胶束制剂CPT@PEG-PLA、顺铂@PEG-PGlu为对照组，以等量的CPT&顺铂小分子混合溶液为对照组，以等量的双药自组装纳米制剂顺铂@PEG-PGlu(ss-CPT)对照组。结果如图18中所示。
结果表明，在抑制肿瘤试验过程中单分子纳米前药小鼠生长状态良好，肿瘤生长得到了明显的抑制，单分子纳米前药显著性优于其他对照试验组，说明本申请单分子纳米前药不仅有效地降低了化疗药物的毒性，还显著提升了传统自组装纳米制剂的抑瘤效能。因此，本申请单分子纳米前药具有优异的临床应用前景。
实施例10.铂单药(顺铂)单分子纳米聚合物及其胶束制备
10.1.铂单药单分子纳米聚合物的制备
甲氧基聚乙二醇丙胺(MeO-PEG-NH 2，分子量为5kDa，0.1g，0.01mmol，单官能化亲水聚合物)溶解于苯(3mL)，搅拌至PEG完全溶解，液氮冷冻，接冷肼真空干燥处理6小时。然后手套箱中，干燥后的聚乙二醇溶解于无水DMF(2mL)，搅拌均匀，取NCA-Pt-NCA(采用实施例2的方法制备，0.18mmol)溶于无水DMF(2mL)中，逐滴缓慢加入到反应体系中，密封反应管，从手套箱中取出，置于35℃油浴中持续搅拌反应72h。将反应产物缓慢滴入冰乙醚中，得到白色沉淀，弃去上清液并重复上述操作三次，获得纯化的产物。产物置于真空锅中真空干燥6小时。干燥后的固体溶解于DMSO(2mL)，置入透析袋(MWCO：100kDa)中，在超纯水中透析两天(换水5次)，冷冻干燥后，收集最终产物。
10.2.铂单药单分子纳米聚合物或其胶束的表征
参考实施例4的方法进行测试。
凝胶渗透色谱(SEC)结果如图26(A)图所示，与单官能化亲水聚合物mPEG-NH 2(5kDa)相比，聚合后的单分子纳米聚合物的分子量明显增大，证实了聚合反应的成功进行，制得的单分子纳米聚合物数均分子量超过2000kDa。
动态光散射(DLS)测试结果如图26(B)图所示，聚合后产物的平均粒径约30.4纳米，粒径范围为23.8～41.1纳米，粒径分布的多分散指数PDI约为0.05。
透射电镜(TEM)的测试结果如图27所示，聚合后产物呈均匀的球形，直径在50 纳米以下，平均直径约30纳米，与DLS测试结果基本一致。经过冻干复溶处理后，尺寸和形貌均稳定。
可见，通过化学聚合的方法能够得到尺寸均一的顺铂单分子纳米聚合物的纳米粒子，可作为前药。
实施例11.铂单药(DACHPt)单分子纳米聚合物及其胶束制备
11.1.NCA-Pt-NCA(NCA-DACHPt-NCA)的制备
Figure PCTCN2022108093-appb-000079
DACHPt((1,2-二氨基环己烷)二氯化铂，3.8g，10mmol)分散于蒸馏水中，加入30％的H 2O 2(60mL)，70℃下避光搅拌5h至澄清，降至室温后置于4℃冰箱进行重结晶。析晶后过滤，依次使用冰水、乙醇、乙醚洗涤滤饼，干燥得到化合物晶体(1)。称取化合物(1)(0.414mg，1.0mmol)与琥珀酸酐(0.62g，6.15mmol)溶解于DMF(20mL)中，70℃下避光搅拌过夜。减压蒸馏除去溶剂，依次使用甲醇、丙酮洗涤后干燥得到化合物(2)。
化合物(2)(0.1mmol)溶解于无水DMF(5.0mL)，与含有EDC(0.15mmol)，DMAP(0.05mmol)的无水DMF溶液(5.0mL)混合，反应30min，加入含有化合物(3)(0.1mmol)的无水DMF溶液(5.0mL)混合，在室温下搅拌反应6小时。通过柱层析分离得到淡黄色固体(4)。
化合物(4)(0.1mmol)溶解于DMF(2.0mL)溶解，与三氟乙酸(2.0mL)混合，室温反应2小时，减压蒸馏去除溶剂，减压蒸馏除去溶剂，依次使用甲醇、丙酮洗涤后干燥得到淡黄色化合物(5)。
化合物(5)(0.1mmol)溶解于DMF(2.0mL)溶解，与含有三光气(0.2mmol)的DMF(2.0mL)混合，室温反应2个小时，减压蒸馏去除溶剂，加入四氢呋喃溶解(60℃)，加入适量正己烷，在4℃冰箱结晶，收集白色针状晶体(6)NCA-Pt-NCA，具体记为NCA-DACHPt-NCA。
NCA-DACHPt-NCA的质谱图如图28所示。
其他实施例部分，如无特殊说明，可参考本例的方法进行 1H NMR测试。
11.2.铂单药单分子纳米聚合物(P102)的制备
甲氧基聚乙二醇丙胺(mPEG-CH 2CH 2CH 2NH 2，可简记为mPEG-NH 2，5kDa，0.05g，0.01mmol)溶解于苯(3mL)，磁力搅拌至PEG完全溶解，液氮冷冻，接冷肼真空干燥处理6小时。然后将反应物转移到无水无氧的手套箱内，干燥后的聚乙二醇溶解于无水DMF(2mL)，磁力搅拌均匀，NCA-DACHPt-NCA(0.4mmol)完全溶于无水DMF(20mL)中，继而逐滴缓慢加入到含mPEG-NH 2的反应体系中，连接气球收集二氧化碳产物，封闭反应体系，从手套箱中取出，置于35℃油浴中持续搅拌反应72h。将反应产物缓慢滴入冰乙醚中，得到白色沉淀，弃去上清液并重复上述操作三次，获得纯化的产物。产物置于真空锅中真空干燥6小时。干燥后的固体溶解于DMSO(10mL)，置入透析袋(MWCO：100kDa)中，在超纯水中透析两天(换水5次)，冷冻干燥后，收集最终聚合产物(P102)。
Figure PCTCN2022108093-appb-000080
11.3.铂单药单分子纳米聚合物及其胶束的表征
凝胶渗透色谱(SEC)结果如图29(A)所示，与单官能化亲水聚合物mPEG-NH 2(5kDa)相比，聚合后的单分子纳米聚合物产物的分子量变大，证实了聚合反应的成功，制得的单分子纳米聚合物数均分子量超过2000kDa。
动态光散射(DLS)测试结果如图29(B)所示，聚合后产物的平均粒径约34纳米，粒径范围为24.2～43.5纳米，粒径分布的多分散指数约为0.05。
透射电镜(TEM)的测试结果如图30所示，聚合后产物呈均匀的球形，直径50纳米以下，平均直径约30纳米，与DLS结果一致，并且经过冻干复溶处理前后进行比较，尺寸形貌稳定。
可见，通过化学聚合的方法能够得到尺寸均一的DACHPt单分子纳米聚合物的纳米粒子，可作为前药。
实施例12.双药单分子纳米聚合物及其胶束制备(顺铂+紫杉醇)
12.1.PTX-ss-OH的制备
紫杉醇(PTX，854mg，1.0mmol)分散于无水二氯甲烷溶解(200mL)，在冰浴中，在氮气保护下加入含有三光气(297mg，1.0mmol)的无水二氯甲烷(5mL)，继续在冰浴下搅拌30分钟，加入溶解有DMAP(488mg，4.0mmol)的无水二氯甲烷(10mL)至紫杉醇完全溶解，在冰浴下继续搅拌反应1小时，转入室温继续避光搅拌反应1小时。在N 2保护下加入溶有2-羟乙基二硫化物(17mL，15mmol)的无水THF(15mL)，混合均匀后避光室温搅拌反应24小时。待反应结束后，再加入50mL二氯甲烷，并依次用0.1M HCl水溶液、饱和的NaCl和水各洗3次，收集有机相并用无水Na 2SO 4干燥，通过旋转蒸发仪减压蒸馏除去有机溶剂。得到的黄色固体通过二氯甲烷/甲醇硅胶柱纯化，最后通过旋蒸淡黄色产物PTX-ss-OH。
Figure PCTCN2022108093-appb-000081
12.2.PTX-ss-OH的制备
以PTX-ss-OH为原料代替CPT-ss-OH，参考实施例1中1.4至1.6的方法，制备PTX-ss-NCA。
Figure PCTCN2022108093-appb-000082
PTX-ss-NCA的 1H NMR谱图如图31所示。其中，7.2-8.1ppm代表紫杉醇的3个苯环峰；3.4ppm处的峰证明紫杉醇衍生物与NCA-Lysine的成功键合；1.2-1.9ppm代表赖氨酸的侧链正丁基峰。
12.3.双药单分子纳米聚合物(P103)的制备
甲氧基聚乙二醇丙胺(MeO-PEG-NH 2，5kDa，0.2g，0.01mmol)溶解于苯(3mL)，磁力搅拌至PEG完全溶解，液氮冷冻，接冷肼真空干燥处理6小时。然后将反应物转移到无水无氧的手套箱内，冷冻干燥后的聚乙二醇溶解于无水DMF(2mL)，搅拌均匀，NCA-Pt-NCA(0.2mmol，采用实施例2的方法制备)和PTX-ss-NCA(0.2mmol)充分溶于无水DMF(30mL)中，继而逐滴缓慢加入到含MeO-PEG-NH 2的反应体系中，连 接气球收集二氧化碳产物，封闭反应体系，从手套箱中取出，置于35℃油浴中持续搅拌反应72h。将反应产物缓慢滴入冰乙醚中，得到白色沉淀，弃去上清液并重复上述操作三次，获得纯化的产物。产物置于真空锅中真空干燥6小时。干燥后的固体溶解于DMSO(12mL)，置入透析袋(MWCO：100kDa)中，在超纯水中透析两天(换水5次)，冷冻干燥后，收集最终聚合产物(P103)。
12.4.双药单分子纳米聚合物及其胶束的表征
参考实施例4的方法进行表征。
进行DLS测试时，不进行超声处理和进行超声处理的测试结果分别如图32中(A)、(B)所示，其中，超声参数(1.0MHz，9.9W)，超声时间(60min)。聚合后产物的平均粒径约46.4纳米，粒径范围为34.6～61.8纳米，证明了聚合反应的成功，并且能够耐受超声等处理，粒径保持不变。
进行透射电镜(TEM)测试，透析后溶液经过3次冻干，复溶(1mg/mL)。复溶前后的测试结果分别如图33中(A)、(B)所示。聚合后产物呈均匀的球形，平均直径约40纳米，与DLS尺寸基本一致，并且经过冻干复溶处理，尺寸形貌稳定。
可见，通过化学聚合的方法能够得到尺寸均一、结构稳定的单分子聚合物纳米粒子。
12.5.药物释放行为考察
考察本例制备的双药单分子纳米聚合物(顺铂+PTX)在不同条件下的释药行为：(1)胞外微环境：pH 7.4，GSH(0mM)、(2)胞内微环境：pH 7.4，GSH(10mM)。将含有上述纳米制剂的PBS溶液(10mM)注入透析袋(MWCO：10000)并浸入上述两种PBS溶液(10mM，28mL，含0.5％(w/v)Tween 80)中，然后在摇晃(100RPM)下，以37℃孵育48小时。在预定的间隔时间点提取1mL释放介质，并补充1mL新鲜的空白介质。通过HPLC测定透析物中PTX的浓度，流动相为甲醇和去离子水(20％～100％(v/v)，体积比)，流速为1.0mL/min，25℃，吸收波长为270nm。
测试结果可参阅图34。结果发现：通过化学聚合形成的双药单分子纳米聚合物(顺铂+PTX)作为前药，在胞外(0mM GSH)不会提前释放活性药物，胞内(10mM GSH)呈现出触发性释放活性药物的功能。
实施例13.双药单分子纳米聚合物及其胶束制备(顺铂+R848)
13.1.R848-ss-OH的制备
以R848为原料，参考实施例1中1.3.的方法制备R848-ss-OH。
Figure PCTCN2022108093-appb-000083
13.2.R848-ss-NCA的制备
以R848-ss-OH为原料代替CPT-ss-OH，参考实施例1中1.4至1.6的方法，制备R848-ss-NCA。
Figure PCTCN2022108093-appb-000084
R848-ss-NCA的 1H NMR谱图如图35所示。其中，7.2ppm以上代表R848的联苯峰；3.4ppm处的峰证明R848衍生物与NCA-Lysine的成功键合；1.2-1.9ppm(除1.45ppm处单个高峰)代表Lysine的侧链正丁基峰。
13.3.双药单分子纳米聚合物(P104)的制备
甲氧基聚乙二醇丙胺(MeO-PEG-NH 2，5kDa，0.1g，0.01mmol)溶解于苯(3mL)，磁力搅拌至PEG完全溶解，液氮冷冻，接冷肼真空干燥处理6小时。然后手套箱中，干燥后的聚乙二醇溶解于无水DMF(2mL)，搅拌均匀，NCA-Pt-NCA(0.2mmol，采用实施例2的方法制备)和R848-ss-NCA(0.2mmol)溶于无水DMF(30mL)中，继而逐滴缓慢加入到含MeO-PEG-NH 2的反应体系中，连接气球收集二氧化碳产物，封闭反应体系，从手套箱中取出，置于35℃油浴中持续搅拌反应72h。将反应产物缓慢滴入冰乙醚中，得到白色沉淀，弃去上清液并重复上述操作三次，获得纯化的产物。产物置于真空锅中真空干燥6小时。干燥后的固体溶解于DMSO(12mL)，置入透析袋(MWCO：100kDa)中，在超纯水中透析两天(换水5次)，冷冻干燥后，收集最终产物(P104)。
13.4.双药单分子纳米聚合物及其胶束的表征
参考实施例4的方法进行表征。
进行DLS测试时，不进行超声处理和进行超声处理的测试结果分别如图36中(A)、(B)所示，其中，超声参数(1.0MHz，9.9W)，超声时间(60min)。聚合后产物的平均粒径约44.9纳米，粒径范围为32.8～66.2纳米，证明了聚合反应的成功，并且能够耐受超声等处理，粒径保持不变。
进行透射电镜(TEM)测试，透析后溶液经过3次冻干，复溶(1mg/mL)。复溶前后的测试结果分别如图37中(A)、(B)所示。聚合后产物呈均匀的球形，平均直径约40纳米，与DLS尺寸基本一致，并且经过冻干复溶处理，尺寸形貌稳定。
可见，通过化学聚合的方法能够得到尺寸均一、结构稳定的单分子聚合物纳米粒子。
13.5.药物释放行为考察
考察本例制备的双药单分子纳米聚合物(顺铂+R848)在不同条件下的释药行为：(1)胞外微环境：pH 7.4，GSH(0mM)、(2)胞内微环境：pH 7.4，GSH(10mM)。将含有上述纳米制剂的PBS溶液(10mM)注入透析袋(MWCO：10000)并浸入上述两种PBS溶液(10mM，28mL，含0.5％(w/v)Tween 80)中，然后在摇晃(100RPM)下，以37℃孵育48小时。在预定的间隔时间点提取1mL释放介质，并补充1mL新鲜的空白介质。通过HPLC测定透析物中R848的浓度，流动相为甲醇和去离子水(20–100％(v/v)，体积比)，流速为1.0mL/min，25℃，吸收波长为320nm。
测试结果可参阅图38。结果发现，通过化学聚合形成的双药单分子纳米聚合物(顺 铂+R848)作为前药，在胞外(0mM GSH)不会提前释放活性药物，胞内(10mM GSH)呈现出触发性释放活性药物的功能。
实施例14.双药单分子纳米聚合物及其胶束制备(顺铂+MMAE)
14.1.MMAE-ss-OH的制备
以MMAE为原料，参考实施例1中1.3.的方法制备MMAE-ss-OH。
MMAE(718mg，1.0mmol)溶解于无水二氯甲烷(20mL)，在冰浴中，在氮气保护下加入含有三光气(297mg，1.0mmol)的无水二氯甲烷(5mL)，继续在冰浴下搅拌30分钟，加入溶解有DMAP(488mg，4.0mmol)的无水二氯甲烷(10mL)至MMAE完全溶解，在冰浴下继续搅拌反应1小时，转入室温继续避光搅拌反应1小时。在N 2保护下加入溶有2-羟乙基二硫化物(17mL，15mmol)的无水THF(15mL)，混合均匀后避光室温搅拌反应24小时。待反应结束后，再加入50mL二氯甲烷，并依次用0.1M HCl水溶液、饱和NaCl和水各洗3次，收集有机相并用无水Na 2SO 4干燥，通过旋转蒸发仪减压蒸馏除去有机溶剂。得到的黄色固体通过二氯甲烷/甲醇硅胶柱纯化，最后通过旋蒸淡黄色产物MMAE-ss-OH。
Figure PCTCN2022108093-appb-000085
14.2.MMAE-ss-NCA的制备
以MMAE-ss-OH为原料代替CPT-ss-OH，参考实施例1中1.4至1.6的方法，制备MMAE-ss-NCA。
Figure PCTCN2022108093-appb-000086
MMAE-ss-NCA的 1H NMR谱图如图39所示。其中，7.6ppm以上代表MMAE的苯环峰；3.4ppm处的峰证明MMAE衍生物与NCA-Lysine的成功键合；1.2-1.9ppm代表Lysine的侧链正丁基峰。
14.3.双药单分子纳米聚合物(P105)的制备(顺铂+MMAE)
甲氧基-聚乙二醇丙胺(MeO-PEG-NH 2，5kDa，0.1g，0.01mmol)溶解于苯(3mL)，磁力搅拌至PEG完全溶解，液氮冷冻，接冷肼真空干燥处理6小时。然后将反应物转移入无水无氧的手套箱中，冻干后的聚乙二醇溶解于无水DMF(2mL)，磁力搅拌均匀，NCA-Pt-NCA(0.2mmol，实施例2的方法制备)和MMAE-ss-NCA(0.2mmol)溶于无水DMF(30mL)中，继而逐滴缓慢加入到含MeO-PEG-NH 2的反应体系中，连接气 球收集二氧化碳产物，封闭反应体系，从手套箱中取出，置于35℃油浴中持续搅拌反应72h。将反应产物缓慢滴入冰乙醚中，得到白色沉淀，弃去上清液并重复上述操作三次，获得纯化的产物。产物置于真空锅中真空干燥6小时。干燥后的固体溶解于DMSO(12mL)，置入透析袋(MWCO：100kDa)中，在超纯水中透析两天(换水5次)，冷冻干燥后，收集最终聚合产物(P105)。
14.4.双药单分子纳米聚合物(顺铂+MMAE)及其胶束的表征
参考实施例4的方法进行表征。
进行DLS测试(未超声处理)结果如图40所示。聚合后产物的平均粒径约43.9纳米，粒径范围为30.5～58.4纳米，证明了聚合反应的成功。
进行透射电镜(TEM)测试，透析后溶液经过3次冻干，复溶(1mg/mL)。复溶前后的测试结果分别如图41中(A)、(B)所示。聚合后产物呈均匀的球形，平均直径约40纳米，与DLS尺寸基本一致，并且经过冻干复溶处理，尺寸形貌稳定。
可见，通过化学聚合的方法能够得到尺寸均一、结构稳定的单分子聚合物纳米粒子。
14.5.药物释放行为考察
考察本例制备的双药单分子纳米聚合物(顺铂+MMAE)在不同条件下的释药行为：(1)胞外微环境：pH 7.4，GSH(0mM)、(2)胞内微环境：pH 7.4，GSH(10mM)。将含有上述纳米制剂的PBS溶液(10mM)注入透析袋(MWCO：10000)并浸入上述两种PBS溶液(10mM，28mL，含0.5％(w/v)Tween 80)中，然后在摇晃(100RPM)下，以37℃孵育48小时。在预定的间隔时间点提取1mL释放介质，并补充1mL新鲜的空白介质。通过HPLC测定透析物中MMAE的浓度，流动相为甲醇和去离子水(20–100％，(v/v))，流速为1.0mL/min，25℃，吸收波长为280nm。
测试结果可参阅图42。结果发现，通过化学聚合形成的双药单分子纳米聚合物(顺铂+MMAE)作为前药，在胞外(0mM GSH)不会提前释放活性药物，胞内(10mM GSH)呈现出触发性释放活性药物的功能。
以上各实施例的各技术特征可以进行任意的组合，为使描述简洁，未对上述实施例中的各个技术特征所有可能的组合都进行描述，然而，只要这些技术特征的组合不存在矛盾，都应当认为是本说明书记载的范围。
以上各实施例仅表达了本申请的几种实施方式，其描述较为具体和详细，但并不能因此而理解为对本申请保护范围的限制。应当指出的是，对于本领域的普通技术人员来说，在不脱离本申请构思的前提下，还可以做出若干变形和改进，这些都属于本申请的保护范围。此外应理解，在阅读了本申请的上述讲授内容之后，本领域技术人员可以对本申请作各种改动或修改，得到的等价形式同样落于本申请的保护范围。还应当理解，本领域技术人员在本申请提供的技术方案的基础上，通过合乎逻辑的分析、推理或者有限的试验得到的技术方案，均在本申请所附权利要求的保护范围内。因此，本申请的保护范围应以所附权利要求为准，说明书及附图可以用于解释权利要求的内容。

Claims (47)

  1. 一种载药单分子纳米聚合物，其包含多条聚氨基酸链，所述多条聚氨基酸链的链间通过多个二价连接基L Pt共价相连使所述多条聚氨基酸链构成非线性骨架，至少一条所述聚氨基酸链的端部连接有亲水性聚合物链；其中，所述二价连接基L Pt的线性骨架中包含铂原子，所述铂原子参与构成铂类药物单元，所述铂类药物单元为铂类药物活性成分或其前药的残基；
    可选地，所述聚氨基酸链的侧基接枝有第二药物单元；其中，所述第二药物单元为抗肿瘤药物活性成分或其前药的残基。
  2. 根据权利要求1所述载药单分子纳米聚合物，其中，所述第二药物单元每次出现，独立地通过响应性连接基L R连接至相应的氨基酸单元，所述响应性连接基L R能够在外界刺激下发生键断裂；
    优选地，所述聚氨基酸链由α氨基酸单元组成，所述聚氨基酸链的主链由-NH-C-C(＝O)-构成；所述第二药物单元每次出现，独立地连接到相应的α氨基酸单元的α碳。
  3. 根据权利要求1或2所述载药单分子纳米聚合物，其中，任一条所述聚氨基酸链的主链由多个
    Figure PCTCN2022108093-appb-100001
    所示结构通过-C(＝O)-NH-键依次键连而成，任一个所述
    Figure PCTCN2022108093-appb-100002
    中的U独立地为碳中心三价基团，任一个所示“*”端独立地连接到所述二价连接基L Pt，或者连接到氢原子或一价侧基R A；所述一价侧基R A中为含有所述第二药物单元的含药侧链，或者为不含药物单元的端基R 0；
    优选地，任一个所示“*”端独立地连接到所述二价连接基L Pt，或者连接到所述一价侧基R A；
    另优选地，任一个所示“*”端独立地连接到所述二价连接基L Pt，或者连接到含有所述第二药物单元的含药侧链。
  4. 根据权利要求3所述载药单分子纳米聚合物，其中，R 0每次出现，独立地选自如下任一种基团：C 1-6烷基、-L A-COOH、-L A-NH 2、-L A-OH、-L A-SH、-L A-CONH 2、-L A-咪唑基、-L A-NHC(＝NH)NH 2、-L A-苯基、-L A-吲哚基和-L A-S-C 1-3烷基；其中，任一个L A独立地选自C 1-6亚烷基，独立地优选为C 1-4亚烷基，进一步独立地优选为亚甲基、1,2-亚乙基、1,3-亚丙基或1,4-亚丁基；
    优选地，R 0每次出现，独立地选自如下任一种基团：-CH 3、-CH(CH 3) 2、-CH 2CH(CH 3) 2、 -CH(CH 3)CH 2CH 3、-CH 2CH 2SCH 3、
    Figure PCTCN2022108093-appb-100003
    -CH 2-OH、-CH(OH)CH 3、-CH 2SH、-CH 2CONH 2、-CH 2CH 2CONH 2、-CH 2CH 2CH 2NH 2及其离子形式、-CH 2CH 2CH 2CH 2NH 2及其离子形式、-CH 2CH 2CH 2NHC(＝NH)NH 2及其离子形式、
    Figure PCTCN2022108093-appb-100004
    及其离子形式、-CH 2COOH及其离子形式和-CH 2CH 2COOH或及离子形式。
  5. 根据权利要求3或4所述载药单分子纳米聚合物，其中，R 0每次出现，独立地为亲水性端基或疏水性端基。
  6. 根据权利要求3至5中任一项所述载药单分子纳米聚合物，其中，每一个
    Figure PCTCN2022108093-appb-100005
    中的U均为CH。
  7. 根据权利要求1至6中任一项所述载药单分子纳米聚合物，其中，一个分子中，所述二价连接基L Pt中的铂原子的数量相对于氨基酸单元总数量的百分比为10％～100％，优选为10％～90％，另优选为10％～80％，另优选为10％～60％，另优选为10％～50％，另优选为10％～40％，另优选为10％～30％，另优选为15％～25％，另优选为18％～22％，另优选为15％～80％，另优选为15％～60％，另优选为15％～50％，另优选为15％～40％，另优选为15％～30％。
  8. 根据权利要求1至7中任一项所述载药单分子纳米聚合物，其中，一个分子中，所述第二药物单元的数量与所述铂类药物单元的数量之比为(0～10):1，优选为(0～5):1，另优选为(0～3):1，另优选为(0～1):1，另优选为(0.5～10):1，另优选为(0.5～5):1，另优选为(0.5～3):1，另优选为(1～5):1，另优选为(1～3):1，另优选为(2～3):1。
  9. 根据权利要求1至8中任一项所述载药单分子纳米聚合物，其中，一个分子中，所述亲水性聚合物链的数量与所述铂类药物单元的数量之比为1:(2～100)，优选为1:(10～60)，另优选为1:(15～45)，另优选为1:(15～25)。
  10. 根据权利要求1-9中任一项所述载药单分子纳米聚合物，其包括式(I)所示的四价结构单元，式(III)所示的一价结构单元、可选的式(II)所示的二价结构单元以及可选的式(IV)所示的二价结构单元；
    Figure PCTCN2022108093-appb-100006
    Figure PCTCN2022108093-appb-100007
    式(I)每次出现，其中，U 1和U 2各自独立地为碳中心三价基团，D Pt为所述铂类药物单元；
    式(III)每次出现，其中，POL i为所述亲水性聚合物链；L 5独立地为二价连接基或无；Z 5独立地为-NH-或-C(＝O)-；
    式(II)每次出现，其中，U 3独立地为碳中心三价基团，L R独立地为响应性连接基，L 4独立地为二价连接基或无，D T为所述第二药物单元；其中，L R能够在外界刺激下发生键断裂；
    式(IV)每次出现，其中，U 6独立地为碳中心三价基团，R E独立地为H或R 0；其中，R 0为不含药物单元的端基；
    优选地，所述载药单分子纳米聚合物包括式(II)所示的二价结构单元以及式(IV)所示的二价结构单元中的至少一种；
    优选地，式(II)所示的二价结构单元以及式(IV)所示的二价结构单元中仅存在其中的一种；
    另优选地，所述载药单分子纳米聚合物不包括式(II)所示的二价结构单元；
    另优选地，所述载药单分子纳米聚合物不包括式(IV)所示的二价结构单元；
    另优选地，所述载药单分子纳米聚合物包括式(II)所示的二价结构单元以及式(IV)所示的二价结构单元；
    另优选地，所述聚氨基酸链由式(I)所示的四价结构单元和式(II)所示的二价结构单元组成；
    另优选地，所述聚氨基酸链由式(I)所示的四价结构单元和式(IV)所示的二价结构单元组成；
    另优选地，所述聚氨基酸链由式(I)所示的四价结构单元、式(II)所示的二价结构单元和式(IV)所示的二价结构单元组成。
  11. 根据权利要求10所述载药单分子纳米聚合物，其中，
    Figure PCTCN2022108093-appb-100008
    每次出现，独立地包含如下结构：
    Figure PCTCN2022108093-appb-100009
    其中，U 10独立地为三价烃基，独立地优选为三价烷基；更优选地，
    Figure PCTCN2022108093-appb-100010
    独立地为赖氨酸或鸟氨酸单元，独立地更优选为赖氨酸单元；
    Figure PCTCN2022108093-appb-100011
    每次出现，独立地包含如下结构：
    Figure PCTCN2022108093-appb-100012
    其中，U 20独立地为三价烃基，独立地优选为三价烷基；更优选地，
    Figure PCTCN2022108093-appb-100013
    独立地为赖氨酸或鸟氨酸单元，独立地更优选为赖氨酸单元；
    另优选地，一个分子中的U 1和U 2均相同；
    另优选地，一个分子中的U 10和U 20均相同。
  12. 根据权利要求10或11所述载药单分子纳米聚合物，其中，D Pt与相邻基团形成如下结构的主链：-C(＝O)-O-Pt-O-C(＝O)-或-C(＝O)-NH-O-Pt-O-NH-C(＝O)-。
  13. 根据权利要求10至12中任一项所述载药单分子纳米聚合物，其中，式(I)每次出现，独立地具有式(I-1)所示结构：
    Figure PCTCN2022108093-appb-100014
    其中，U 10和U 20分别独立地如权利要求11所定义；
    Z 11和Z 21各自独立地为无、-C(＝O)-或-C(＝O)-O-*，其中的“*”指向D Pt；
    R 11和R 21各自独立地为C 1-6亚烃基，各自独立地优选为C 1-6亚烷基，各自独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基、1,4-亚丁基、1,5-亚戊基或1,6-亚己基，各自独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，另各自独立地优选为1,2-亚乙基或1,3-亚丙基，另各自独立地优选为1,2-亚乙基；
    X 11和X 21各自独立地为-C(＝O)-O-*或-C(＝O)-NH-O-*，可以各自独立地优选为-C(＝O)-O-*，其中的“*”指向D Pt。
  14. 根据权利要求10至13中任一项所述载药单分子纳米聚合物，其中，D Pt每次出现，独立地选自顺铂、卡铂、奈达铂、奥沙利铂和洛铂中任一种的残基。
  15. 根据权利要求10至14中任一项所述载药单分子纳米聚合物，其中，式(I)每次出现，具有相同结构。
  16. 根据权利要求10至15中任一项所述载药单分子纳米聚合物，其中，
    Figure PCTCN2022108093-appb-100015
    每次出现，独立地包含如下结构：
    Figure PCTCN2022108093-appb-100016
    其中，U 30独立地为三 价烃基，独立地优选为三价烷基；更优选地，
    Figure PCTCN2022108093-appb-100017
    独立地为赖氨酸或鸟氨酸单元，独立地更优选为赖氨酸单元；
    另优选地，一个分子中的U 3均相同；
    另优选地，一个分子中的U 30均相同。
  17. 根据权利要求10至16中任一项所述载药单分子纳米聚合物，其中，L R每次出现，独立地包含能够在如下至少一种条件下发生断裂的连接基：胞内还原条件、活性氧条件、pH条件、酶解条件和水解条件；
    优选地，
    所述pH条件满足pH值小于6.8，进一步优选pH为4.0～6.8；
    所述酶解条件选自如下的一种或多种酶：MMP-2酶和偶氮还原酶；
    所述水解条件为酸性水解条件或碱性水解条件。
  18. 根据权利要求17所述载药单分子纳米聚合物，其中，L R每次出现，独立地包含如下(a)组、(b)组、(c)组、(d)组和(e)组中的一种或多种连接基；
    (a)组：-S-S-；
    (b)组：草酸酯基、硼酸酯基、酮缩硫醇基、硫醚基、单硒基、二硒基、二价碲基、噻唑啉酮基、硼酸基和3～7元脯氨酸低聚链；
    (c)组：缩醛基和腙键；
    (d)组：GPLGVRG肽段和偶氮基；
    (e)组：-C(＝O)-O-和-O-C(＝O)-；
    优选地，L R每次出现，独立地包含如下的一种或多种连接基：-S-S-、草酸酯基、芳基硼酸酯基、缩醛基、腙键、GPLGVRG肽段、偶氮基、-C(＝O)-O-和-O-C(＝O)-；另优选地，所述芳基硼酸酯基为苯基硼酸酯基。
  19. 根据权利要求10至18中任一项所述载药单分子纳米聚合物，其中，-L 4-D T每次出现，独立地包括Z 4-D T，其中，Z 4每次出现，独立地为化学键或者选自如下任一种基团：-C(＝O)-、-O-、-S-、-O-C(＝O)-*、-NH-C(＝O)-*、和-NH-，其中，“*”所示端指向D T；
    优选地，-L 4-D T每次出现，其结构独立为-R 32-Z 4-D T，其中，R 32每次出现，独立地为C 1-6亚烃基，独立地优选为C 1-6亚烷基，独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基、1,4-亚丁基、1,5-亚戊基或1,6-亚己基，独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，另独立地优选为1,2-亚乙基或1,3-亚丙基，另独立地优选为1,2-亚乙基；
    更优选地，D T与Z 4独立地形成如下任一种连接基：-C(＝O)-O-、-O-C(＝O)-、-O-C(＝O)-O-、-O-C(＝O)-NH-、-NH-C(＝O)-O-、-C(＝O)-NH-和-NH-C(＝O)-；更优选地，D T与Z 4独立地形成如下任一种连接基：-C(＝O)-O-、-O-C(＝O)-、-O-C(＝O)-O-、-O-C(＝O)-NH-和-NH-C(＝O)-O-；另优选地，D T与Z 4独立地形成如下连接基：-O-C(＝O)-O-。
  20. 根据权利要求10至19中任一项所述载药单分子纳米聚合物，其中，式(II)每次出现，具有式(II-1)所示结构：
    Figure PCTCN2022108093-appb-100018
    其中，U 30每次出现，独立地如权利要求17所定义；
    R 32和Z 4每次出现，分别独立地如权利要求20所定义；
    Z 3每次出现，独立地为无、-C(＝O)-或-C(＝O)-O-*，独立地优选为-C(＝O)-或-C(＝O)-O-*，另独立地优选为-C(＝O)-O-*，另独立地优选为-C(＝O)-，其中的“*”指向R 31；
    R 31每次出现，独立地为C 1-6亚烃基，独立地优选为C 1-6亚烷基，独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基、1,4-亚丁基、1,5-亚戊基或1,6-亚己基，独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，另独立地优选为1,2-亚乙基或1,3-亚丙基，另独立地优选为1,2-亚乙基。
  21. 根据权利要求10至20中任一项所述载药单分子纳米聚合物，其中，D T每次出现，独立地选自喜树碱类化合物、瑞喹莫德和紫杉醇中任一种的残基；
    优选地，所述喜树碱类化合物包括喜树碱及其衍生物或类似物；
    更优选地，所述喜树碱类化合物包括依立替康、拓扑替康、卢比替康、吉咪替康、9-氨基喜树碱、9-硝基喜树碱和7-乙基-10-羟基喜树碱。
  22. 根据权利要求10至21中任一项所述载药单分子纳米聚合物，其中，式(II)每次出现，具有相同结构。
  23. 根据权利要求10至22中任一项所述载药单分子纳米聚合物，其中，Z 5每次出现，独立地为-NH-、-C(＝O)-或*-O-C(＝O)-，其中的“*”指向L 5。
  24. 根据权利要求10至23中任一项所述载药单分子纳米聚合物，其中，L 5每次出现，独立地为C 1-6亚烃基，独立地优选为C 1-6亚烷基，独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基、1,4-亚丁基、1,5-亚戊基或1,6-亚己基，独立地更优选为亚甲基、1,2-亚乙基、1,3-亚丙基或1,4-亚丁基，另独立地更优选为1,2-亚乙基或1,3-亚丙基；
    另优选地，式(III)每次出现，具有相同的L 5和Z 5。
  25. 根据权利要求10至24中任一项所述载药单分子纳米聚合物，其中，POL i每次出现，独立地包含如下任一种亲水聚合物链：聚乙二醇、聚(丙二醇)、乙二醇和丙二醇的共聚物、聚(乙氧基化多元醇)、聚(烯醇)、聚(乙烯基吡咯烷酮)、聚(羟烷基甲基丙烯酰胺)、聚(羟烷基甲基丙烯酸酯)、聚(糖)、聚(α-羟基酸)、聚(乙烯醇)、聚膦腈、聚噁唑啉、聚(N-丙烯酰基吗啉)以及前述聚合物链的任意组合；
    优选地，所述亲水聚合物链的分子量选自50Da～100kDa，另优选为100Da～80kDa，另优选为500Da～50kDa，另优选为500Da～10kDa，另优选为500Da～8000Da，另优选为500Da～6000Da，另优选为500Da～5000Da，另优选为1000Da～50kDa，另优选为1000Da～10kDa，另优选为1000Da～8000Da，另优选为1000Da～6000Da，另优选为1000Da～5000Da；
    另优选地，POL i每次出现，独立地包含聚乙二醇链段；另优选地，所述聚乙二醇链段为mPEG，另优选地，所述聚乙二醇链段的分子量选自50Da～100kDa，另优选为100Da～80kDa，另优选为500Da～50kDa，另优选为500Da～10kDa，另优选为500Da～8000Da，另优选为500Da～6000Da，另优选为500Da～5000Da，另优选为1000Da～50kDa，另优选为1000Da～10kDa，另优选为1000Da～8000Da，另优选为1000Da～6000Da，另优选为2000Da～6000Da，另优选为4000Da～6000Da，另优选为1000Da～5000Da，另优选为约500Da、约600Da、约800Da、约1000Da、约1100Da、约1200Da、约1500Da、约1600Da、约2000Da、约2200Da、约2500Da、约3000Da、约3500Da、约4000Da、约4400Da、约4500Da、约5000Da、约5500Da、约6000Da、约6500Da、约7000Da、约8000Da、约9000Da、约10kDa、约12kDa、约15kDa、约20kDa、约25kDa、约30kDa、约35kDa、约40kDa或约40kDa，其中的“约”表示±10％；
    上述任一种“分子量”独立地表示重均分子量或数均分子量。
  26. 根据权利要求10至25中任一项所述载药单分子纳米聚合物，其中，式(IV)每次出现，具有式(IV-1)所示结构：
    Figure PCTCN2022108093-appb-100019
    其中，R E每次出现，独立地为氢原子或R 0，其中R 0为不含药物单元的端基；
    优选地，R E每次出现，独立地为R 0；
    另优选地，R 0如权利要求5或6所定义。
  27. 根据权利要求10至26中任一项所述载药单分子纳米聚合物，其中，式(IV)每次出现，具有相同结构。
  28. 根据权利要求10至27中任一项所述载药单分子纳米聚合物，其中，所述载药单分子纳米聚合物包括式(I-2)所示的四价结构单元，式(III-2)所示的一价结构单元、可选的式(II-2)所示的二价结构单元以及可选的式(IV-1)所示的二价结构单元；
    Figure PCTCN2022108093-appb-100020
    Figure PCTCN2022108093-appb-100021
    优选地，
    n11和n21各自独立地为3或4，n12和n22各自独立地为1、2、3、4或5；
    n31独立地为3或4，n32独立地为2、3或4，n33独立地为2、3或4；
    n51独立地为1、2、3或4；
    p独立地为正整数，优选为小于等于2500的正整数，另优选为小于等于2000的正整数，另优选为小于等于1500的正整数，另优选为小于等于1000的正整数，另优选为小于等于800的正整数，另优选为小于等于600的正整数，另优选为小于等于500的正整数，另优选为小于等于400的正整数，另优选为小于等于300的正整数，另优选为小于等于250的正整数，另优选为小于等于200的正整数，另优选为选自2～2500的整数，另优选为选自3～2000的整数，另优选为选自5～1500的整数，另优选为选自5～1000的整数，另优选为选自5～800的整数，另优选为选自5～600的整数，另优选为选自5～500的整数，另优选为选自5～400的整数，另优选为选自5～300的整数，另优选为选自5～250的整数，另优选为选自5～200的整数，另优选为选自5～1500的整数，另优选为选自5～1000的整数，另优选为选自5～800的整数，另优选为选自10～600的整数，另优选为选自10～500的整数，另优选为选自10～400的整数，另优选为选自10～300的整数，另优选为选自10～250的整数，另优选为选自10～200的整数，另优选为选自20～600的整数，另优选为选自20～500的整数，另优选为选自20～400的整数，另优选为选自20～300的整数，另优选为选自20～250的整数，另优选为选自20～200的整数；另优选为选自50～500的整数，另优选为选自50～400的整数，另优选为选自50～300的整数，另优选为选自50～250的整数，另优选为选自50～200的整数；另优选为选自100～500的整数，另优选为选自100～400的整数，另优选为选自100～300的整数，另优选为选自100～250的整数，另优选为选自100～200的整数，另优选为选自100～150的整数；
    式(IV-1)所示的二价结构单元如权利要求26所定义。
  29. 根据权利要求28所述载药单分子纳米聚合物，其中，
    n11和n21各自独立地为4，n12和n22各自独立地为4；
    n31独立地为4，n32独立地为2，n33独立地为2；
    n51独立地为2、3或4；
    优选地，L R为-S-S-，Z 5为-NH-。
  30. 根据权利要求29所述载药单分子纳米聚合物，其中，D Pt为顺铂、奥沙利铂或
    Figure PCTCN2022108093-appb-100022
    的残基，D T为喜树碱、紫杉醇或瑞喹莫德的残基。
  31. 根据权利要求1～30中任一项所述载药单分子纳米聚合物，其中，所述载药单分子纳米聚合物的重均分子量选自200kDa～5000kDa，优选为500kDa～5000kDa，另优选为500kDa～4000kDa，另优选为500kDa～3000kDa，另优选为500kDa～2500kDa，另优选为500kDa～2000kDa，另优选为500kDa～1500kDa，另优选为600kDa～1500kDa，另优选为800kDa～1200kDa；
    优选地，一个分子中的铂原子数量大于40，优选大于50，另优选选自50～5000，进一步可选自50～4000，进一步可选自50～2000，进一步可选自50～1500，进一步可选自50～1000，进一步可选自50～500，另优选选自60～2000，另优选选自60～1500，另优选选自60～1000，另优选选自60～500，另优选选自80～2000，另优选选自80～1500，另优选选自80～1000，另优选选自80～500，另优选选自100～2000，另优选选自100～1500，另优选选自100～1000，另优选选自100～500，另优选选自150～2000，另优选选自150～1500，另优选选自150～1000，另优选选自150～500，另优选选自200～2000，另优选选自200～1500，另优选选自200～1000，另优选选自200～500，另优选选自250～2000，另优选选自250～1500，另优选选自250～1000，另优选选自250～500，另优选选自300～400。
  32. 一种载药单分子纳米聚合物的制备方法，其包括如下步骤：将结构如式(I-3)所示的含铂化合物、结构如式(III-3)所示的单官能化亲水聚合物、可选的结构如式(II-3)所示的药物化合物以及可选的如式(IV-3)所示化合物在有机溶剂中混合，进行开环聚合反应；
    Figure PCTCN2022108093-appb-100023
    其中，
    P E为R E或被保护的R E，在所述开环聚合反应中不具备反应性；
    U 1、U 2、D Pt、U 3、L R、L 4、D T、L 5和R E分别如权利要求10至30中任一项所定义；
    mPEG为甲氧基聚乙二醇链段；
    F 5为-NH 2、-COOH、
    Figure PCTCN2022108093-appb-100024
    优选为-NH 2；
    进一步优选地，所述开环聚合反应于无水条件下进行；
    更进一步优选地，开环聚合反应温度为15～40℃，更优选地，开环聚合反应时间为24～96h。
  33. 一种双药单分子纳米聚合物前药，其包含与亲水性聚合物连接的聚氨基酸，其中在该聚氨基酸的重复单元的α碳上键合了铂类药物活性成分的前药部分和抗肿瘤药物活性成分的前药部分；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
  34. 根据权利要求33的双药单分子纳米聚合物前药，其中所述的亲水性聚合物选自如下任一种：聚乙二醇、聚(丙二醇)、乙二醇和丙二醇的共聚物、聚(乙氧基化多元醇)、聚(烯醇)、聚(乙烯基吡咯烷酮)、聚(羟烷基甲基丙烯酰胺)、聚(羟烷基甲基丙烯酸酯)、聚(糖)、聚(α-羟基酸)、聚(乙烯醇)、聚膦腈、聚噁唑啉、聚(N-丙烯酰基吗啉)以及这些物质的任意组合。
  35. 根据权利要求33或34的双药单分子纳米聚合物前药，其中所述的铂类药物选自顺铂、卡铂、奈达铂、奥沙利铂和洛铂中的任一种。
  36. 根据权利要求34至35任意一项的双药单分子纳米聚合物前药，其中所述分子中含有游离羟基、游离氨基或两者的组合的抗肿瘤药物活性成分是喜树碱类化合物、瑞喹莫德和紫杉醇中的一种或多种；优选地，所述喜树碱类化合物包括喜树碱及其衍生物或类似物，更优选地，所述喜树碱类化合物包括依立替康、拓扑替康、卢比替康、吉咪替康、9-氨基喜树碱、9-硝基喜树碱和7-乙基-10-羟基喜树碱。
  37. 一种制备权利要求33至36任意一项的双药单分子纳米聚合物前药的方法，该方法包括如下步骤：
    (1)在合适反应条件下，合成抗肿瘤药物活性成分的单NCA单体，优选地，所述抗肿瘤药物活性成分的分子结构中含有游离羟基或游离氨基；
    (2)在合适反应条件下，合成铂类药物活性成分的双NCA单体，
    (3)在合适反应条件下，使步骤(1)和步骤(2)得到的单体与具有末端氨基的亲水性聚合物反应，得到所述的双药单分子纳米聚合物前药，以及
    (4)分离所得到的双药单分子纳米聚合物前药。
  38. 一种载药单分子纳米聚合物胶束，其组成选自如下任一种：权利要求1～31中任一项所述载药单分子纳米聚合物，权利要求32所述的制备方法制备得到的载药单分子纳米聚合物，权利要求33至36中任一项的双药单分子纳米聚合物前药，和权利要求37的制备方法制得的双药单分子纳米聚合物前药；所述载药单分子纳米聚合物胶束具有核壳结构，外壳结构为亲水性聚合物链形成的亲水层，所包载的药物单元位于内核中。
  39. 根据权利要求38所述载药单分子纳米聚合物胶束，其中，25℃时水中的胶束粒径选自10～120nm，优选为10～110nm，另优选为10～100nm，另优选为10～80nm，另优选为10～50nm，另优选为10～40nm，另优选为10～30nm，另优选为15～120nm，优选为15～110nm，另优选为15～100nm，另优选为15～80nm，另优选为15～50nm，另优选为15～40nm，另优选为15～30nm，另优选为20～120nm，优选为20～110nm，另优选为20～100nm，另优选为20～80nm，另优选为20～70nm，另优选为20～50nm，另优 选为20～40nm，另优选为25～120nm，优选为25～110nm，另优选为25～100nm，另优选为25～80nm，另优选为25～50nm，另优选为25～40nm，另优选为25～35nm，另优选为30～35nm；
    进一步地，所述载药单分子纳米聚合物胶束的平均直径选自15～50nm，优选为15～40nm，另优选为20～40nm，另优选为25～35nm。
  40. 根据权利要求38或39所述载药单分子纳米聚合物胶束，其中，25℃时水中的胶束内核半径为5～50nm，优选为5～45nm，另优选为5～40nm，另优选为5～35nm，另优选为5～30nm，另优选为5～25nm，另优选为5～20nm，另优选为5～15nm，另优选为5～10nm，另优选为10～50nm，另优选为10～45nm，另优选为10～40nm，另优选为10～35nm，另优选为10～30nm，另优选为10～25nm，另优选为10～20nm，另优选为6～8nm；
    25℃时水中的胶束外壳厚度为5～40nm，优选为5～35nm，另优选为5～30nm，另优选为5～25nm，另优选为5～20nm，另优选为5～15nm，另优选为10～40nm，另优选为10～35nm，另优选为10～30nm，另优选为10～25nm，另优选为10～20nm，另优选为8～12nm。
  41. 一种药物递送系统，它包含载药单分子纳米聚合物胶束，该载药单分子纳米聚合物胶束包含权利要求1～31中任一项所述载药单分子纳米聚合物或权利要求37所述的制备方法制备得到的载药单分子纳米聚合物；
    优选地，
    所述亲水性聚合物链位于所述载药单分子纳米聚合物胶束的外壳；
    所述铂类药物单元以及所述第二药物单元均位于所述载药单分子纳米聚合物胶束的内核。
  42. 一种药物递送系统，它包含双药单分子纳米聚合物胶束，该双药单分子纳米聚合物胶束包含与亲水性聚合物连接的聚氨基酸，其中在该聚氨基酸的重复单元的α碳上键合了铂类药物活性成分的前药部分和抗肿瘤药物活性成分的前药部分；优选地，所述抗肿瘤药物活性成分的分子中含有游离羟基、游离氨基或两者的组合。
  43. 根据权利要求42的药物递送系统，其中所述的亲水性聚合物选自聚乙二醇、聚(丙二醇)、乙二醇和丙二醇的共聚物、聚(乙氧基化多元醇)、聚(烯醇)、聚(乙烯基吡咯烷酮)、聚(羟烷基甲基丙烯酰胺)、聚(羟烷基甲基丙烯酸酯)、聚(糖)、聚(α-羟基酸)、聚(乙烯醇)、聚膦腈、聚噁唑啉、聚(N-丙烯酰基吗啉)以及这些物质的任意组合。
  44. 根据权利要求42或43的药物递送系统，其中所述的铂类药物选自顺铂、卡铂、奈达铂、奥沙利铂和洛铂中的一种或多种。
  45. 根据权利要求42至44任意一项的药物递送系统，其中所述分子中含有游离羟基、游离氨基或两者的组合的抗肿瘤药物活性成分选自喜树碱类化合物、瑞喹莫德和紫杉醇中的一种或多种；优选地，所述喜树碱类化合物包括喜树碱及其衍生物或类似物，更优选地，所述喜树碱类化合物包括依立替康、拓扑替康、卢比替康、吉咪替康、9-氨基喜树碱、9-硝基喜树碱和7-乙基-10-羟基喜树碱。
  46. 铂类药物活性成分的双NCA单体和抗肿瘤药物活性成分的单NCA单体在制备单分子纳米聚合物前药或药物递送系统中的用途；优选地，所述抗肿瘤药物活性成分 的分子结构中含有游离羟基或游离氨基。
  47. 权利要求1至31任意一项所述载药单分子纳米聚合物，权利要求32所述的制备方法制备得到的载药单分子纳米聚合物，权利要求33至36任意一项的双药单分子纳米聚合物前药，权利要求37的制备方法制得的双药单分子纳米聚合物前药，权利要求38～40中任一项所述载药单分子纳米聚合物胶束，或权利要求41至45任意一项所述的药物递送系统在制备用于治疗肿瘤疾病的药物中的用途。
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CN116271074B (zh) * 2023-05-11 2023-08-25 东曜药业有限公司 一种具有双重治疗机制的肿瘤靶向治疗药物
CN116459215A (zh) * 2023-06-02 2023-07-21 辽宁大学 一种刺激响应型双药共递送胶束及其制备方法和应用
CN117126095A (zh) * 2023-08-18 2023-11-28 中国科学技术大学 含单一分子量聚乙二醇链段的可释放性功能化合物、包含其的纳米粒子及制备方法

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