WO2023051682A1 - 一种局部药物组合物、应用及试剂盒 - Google Patents

一种局部药物组合物、应用及试剂盒 Download PDF

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WO2023051682A1
WO2023051682A1 PCT/CN2022/122537 CN2022122537W WO2023051682A1 WO 2023051682 A1 WO2023051682 A1 WO 2023051682A1 CN 2022122537 W CN2022122537 W CN 2022122537W WO 2023051682 A1 WO2023051682 A1 WO 2023051682A1
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dextran
local
drug
pharmaceutical composition
synergistic
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PCT/CN2022/122537
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English (en)
French (fr)
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邹方霖
邹礼常
王建霞
钟晓丽
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成都夸常奥普医疗科技有限公司
夸常股份有限公司
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Priority claimed from PCT/CN2021/122134 external-priority patent/WO2023050297A1/zh
Application filed by 成都夸常奥普医疗科技有限公司, 夸常股份有限公司 filed Critical 成都夸常奥普医疗科技有限公司
Publication of WO2023051682A1 publication Critical patent/WO2023051682A1/zh

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
    • A61K31/716—Glucans
    • A61K31/721—Dextrans
    • 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
    • A61K47/02—Inorganic compounds
    • 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
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/16—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing nitrogen, e.g. nitro-, nitroso-, azo-compounds, nitriles, cyanates
    • A61K47/18—Amines; Amides; Ureas; Quaternary ammonium compounds; Amino acids; Oligopeptides having up to five amino acids
    • 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
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the invention relates to the technical field of pharmaceutical preparations, in particular to a new application of a dextran compound, a new pharmaceutical composition prepared according to the application, and a new treatment method using the composition.
  • solid tumors are a representative model of disease with localized lesions consisting of tissues containing tumor cells (tumor body and lymph nodes contaminated by tumor cells).
  • tumor cells tumor body and lymph nodes contaminated by tumor cells.
  • fibroblasts intercellular matrix, pipelines and other components in tumor tissues, which are sometimes called the microenvironment of tumor cells.
  • pancreatic cancer cells only account for about 30% of the tumor tissue. These microenvironments complicate the reaction kinetics between cytotoxic drugs and tumor cells, which may greatly reduce the drug efficacy.
  • synergistic or synergistic effect is widely accepted as "mutual enhancement of similar effects between synergists and common ingredients", and the local synergistic (or synergistic) effect between chemical drugs is that synergists and common ingredients
  • the principle of selection of synergists and co-components should then be such that their actual co-efficacy exceeds the expected co-efficacy.
  • the anticipation of the expected co-efficacy is obviously based on their respective chemical local effects, which are short-term drug effects similar to chemical ablation, rather than medium and long-term drug effects.
  • the choice of pharmacological conditions for local potentiators is obviously to maximize, at least effective local effects (tissue destruction, that is, similar to chemical ablation), rather than the opposite; Synergists or their combinations, rather than their non-synergistic combinations; and the choice of the preferred combination between the synergist and the co-ingredients should obviously be based on chemical ablation-like (short-term) drug effects, rather than mid- to long-term drug effects effect.
  • local synergy (or synergy) that meets the above technical schemes (local action strength of synergists, local synergists or their combination, short-term drug effect similar to chemical ablation) is called conventional local synergy (or synergism). effect), otherwise it is unconventional local synergy (or synergy).
  • the chemical drug sharing in reality is in line with the above expectations, that is, they are all conventional local synergistic technical solutions.
  • chemotherapeutic drugs 5-fluorouracil and cisplatin
  • the combination of intratumoral administration should be selected under the condition of administration concentration that is effective for each of them, generally maximizing the cytotoxic effect; conventional chemotherapy ineffective drugs—such as glucose and methylene blue
  • conventional chemotherapy ineffective drugs such as glucose and methylene blue
  • the intratumoral administration combination is selected under the administration concentration conditions of each being effective or maximizing chemical ablation (local effect).
  • topically acting drugs such as basic organic compounds and basic inorganic salts.
  • synergists are used to provide conventional local synergy rather than unconventional local synergy.
  • the problem to be solved by the present invention is how to make dextran have a new application besides being used as a blood volume expander: local shared effect reversal, that is, to convert a non-synergistic drug that does not have a synergistic effect with it into a local drug that does not have a synergistic effect with it.
  • the synergistic co-product components, or/and the co-products that make their medium- and long-term drug effects exhausted are converted into composition components with considerable mid- and long-term drug effects, providing an easy-to-prepare, almost no systemic side effects for the clinical needs of solid tumors Medium and long-term intratumoral administration composition and a medium and long-term treatment method. So far, there is no report on the function of dextran that can be used as a blood volume expander, let alone the use of this function.
  • the present invention aims to propose a new pharmacology of the dextran compound—reversal of shared intratumoral effects, and its new application.
  • dextran which can be used as a blood volume expander, is selected as a potentiator that minimizes rather than maximizes local effects, especially mid- and long-term local effects
  • its non-synergistic drugs, rather than its synergistic drugs are selected as its co-products Components, choose to convert these non-synergistic drugs into their synergistic co-components or/and inhibit the co-reversal effect of the mid-to-long-term drug efficacy failure of the co-products, rather than the direct synergistic effect on their co-operative drugs as co-pharmacology
  • the medium-to-long-term pharmaceutical composition prepared for intratumoral administration and the new medium-to-long-term treatment method thus achieve high efficiency and low toxicity, especially in the medium and long-term, so as to provide various options for clinical use.
  • polyvinylpyrrolidone commonly used as a blood volume expander
  • polyvinylpyrrolidone can, under certain specific conditions beyond conventional synergistic expectations (for example, minimize its local effects), actually enhance Synergistically acting non-synergistic drugs are reversed to their synergistic co-components.
  • dextran which can be used as an expander, can further reverse the mid- and long-term drug efficacy failure of the co-product.
  • the present invention provides a local pharmaceutical composition, its intratumoral administration form is the aqueous solution of dextran which can be used as a blood volume expander and its non-cooperative drugs, wherein the pharmacological concentration of the dextran is ⁇ 25%;
  • the synergistic drugs are respectively selected from basic organic compounds (drug group A) and basic inorganic salts (drug group B) that do not show local synergy with the expansion agent, and the basic organic compounds include tromethamine, At least one of triethanolamine, trishydroxymethylaminomethane, meglumine, arginine, and lysine, and the basic inorganic salt includes at least one of sodium bicarbonate, sodium carbonate, sodium lactate, and sodium acetate, so The pharmacological concentration of the various non-synergistic drugs is 4-40%.
  • the pharmacological concentration of the blood volume expander is 2-25%.
  • the pharmacological concentration of the blood volume expander is 2-4.5%.
  • concentration refers to weight/volume percentage concentration.
  • “Pharmacological concentration” refers to the intratumoral administration concentration required for a drug to provide a selected pharmacology (eg, co-reversal effects of dextran, co-effects of non-synergistic drugs that can be reversed by dextran).
  • the dextran is selected from dextran compounds that can be used as blood volume expanders, including at least one of dextran 10, dextran 20, dextran 40, and dextran 70.
  • the selection conditions of the basic organic compound (drug group A) and basic inorganic salt (drug group B) are: no synergistic effect occurs when the A or B is shared with the dextran, and the A and B There is a synergistic effect when B is shared with the dextran, and the synergistic effect includes short-term synergistic effect and/or mid- to long-term synergistic effect, and the synergistic effect includes drug effect synergy and/or safety synergy.
  • the pharmacological concentrations of the basic organic compound (drug group A) and the basic inorganic salt (drug group B) are: at least an intratumoral administration concentration that can provide considerable short-term local drug effects, wherein the basic organic compound (Drug A) has a pharmacological concentration of 5-20%, and when the basic organic compound is selected from arginine or acid-resistant acid, the pharmacological concentration of the basic organic compound is 10-25%; the alkali The pharmacological concentration of the active inorganic salt (drug B) is 3-15%.
  • the determining condition for the ratio of the basic organic compound (drug group A) to the basic inorganic salt (drug group B) is: the combination of A and B is a local synergistic combination (short-term synergy).
  • the concentration ratio of the combination of A and B (basic organic compound/basic inorganic salt) is (5-25%)/(3-15%).
  • the concentration ratio refers to the content ratio in the formed pharmaceutical composition.
  • the combination of the basic organic compound and the basic inorganic salt is selected from one of the following combinations: arginine/basic inorganic salt combination, the concentration ratio is (10-20%)/(2-15 %); arginine/tris/basic inorganic salt combination, the concentration ratio is (10-20%)/(2-15%)/(2-15%); lysine/basic Inorganic salt combination, the concentration ratio is (15-25%)/(2-15%); tromethamine/basic inorganic salt combination, the concentration ratio is (2-10%)/(2-15%); three Ethanolamine/basic inorganic salt combination, concentration ratio is (2-10%)/(2-15%); Trishydroxymethylaminomethane/basic inorganic salt combination, concentration ratio is (2-15%)/(2 -15%); meglumine/basic inorganic salt combination, the concentration ratio is (10-25%)/(2-15%).
  • cytotoxic drugs with a pharmacological concentration of 0.03-6%, or/and immunomodulators with a pharmacological concentration of 0.2-25% are also included.
  • the cytotoxic drugs refer to active ingredients that mainly target diseased cells or structures within diseased cells to achieve their drug effects, such as substances that have been shown to resist proliferation of cancer cells in cell experiments or in tumor-bearing animals, including: 1) destroying Drugs for the structure and function of DNA, such as alkylating agents - cyclophosphamide, carmustine, etc., metal platinum complexes - cisplatin, carboplatin, etc., DNA topoisomerase inhibitors - doxorubicin, topo Tecan, irinotecan, etc.; 2) Drugs embedded in DNA that interfere with RNA transcription, such as anti-tumor antibiotics-actinomycins, daunorubicin, doxorubicin, etc.; 3) Drugs that interfere with DNA synthesis, such as Pyrimidine antagonists - uracil derivatives, such as 5-fluorouracil, furofluorouracil, bisfururouracil, cytosine derivatives cytara
  • the chemotherapeutic drugs are selected from uracil derivatives, cyclophosphamides, gemcitabines, epirubicins, antitumor antibiotics, teniposide, metal platinum complexes, taxanes
  • uracil derivatives preferably, at least one of 5-fluorouracil, cyclophosphamide, ifosfamide, gemcitabine, epirubicin, antitumor antibiotics, teniposide, metal platinum complexes, paclitaxel A sort of.
  • the pharmacological concentration (w/v) of the chemotherapeutic drug in the local pharmaceutical composition is 0.1-5%, for example, the pharmacological concentration (w/v) of alkylating agents such as cyclophosphamide and carmustine is 0.5-5%, preferably 0.75-1.5%; the pharmacological concentration (w/v) of metal platinum complexes such as cisplatin and carboplatin is 0.1-0.25%; doxorubicin, topotecan, irinotecan The concentration (w/v) of such DNA topoisomerase inhibitors is 0.1-0.30%; The concentration (w/v) of antitumor antibiotics such as actinomycins, daunorubicin is 1-4%, preferably 1-2%; the concentration (w/v) of uracil derivatives such as 5-fluorouracil, furofluorouracil, and bisfururouracil, and cytosine derivatives such as cytarabine, cyclocyt
  • the immunomodulator includes at least one of BCG and components thereof, probiotics and components thereof, and at least one of immunomodulatory antibodies; the BCG and components thereof include BCG, inactivated BCG, BCG polysaccharide nucleic acid, etc., and the probiotics Bacteria and its components include inactivated probiotics, probiotic cell wall polysaccharides and their derivatives, probiotic ribonucleic acid, etc., and the immunomodulatory antibodies include plasma immunoglobulins, polyclonal immunoglobulins, and monoclonal immunoglobulins.
  • the monoclonal immunoglobulin includes antibody blockers against inhibitory receptors, such as blocking antibodies against CTLA-4 molecules and PD-1 molecules; antibody blockers against ligands of inhibitory receptors Antibodies, activating antibodies against stimulating molecules on the surface of immune response cells, such as anti-OX40 antibody, anti-CD137 antibody, anti-4-1BB antibody; neutralizing antibodies against immunosuppressive molecules in the local disease microenvironment, such as anti-TGF-p1 Antibody.
  • inhibitory receptors such as blocking antibodies against CTLA-4 molecules and PD-1 molecules
  • Antibodies antibody blockers against ligands of inhibitory receptors Antibodies, activating antibodies against stimulating molecules on the surface of immune response cells, such as anti-OX40 antibody, anti-CD137 antibody, anti-4-1BB antibody
  • neutralizing antibodies against immunosuppressive molecules in the local disease microenvironment such as anti-TGF-p1 Antibody.
  • BCG (or probiotic) component refers to a preparation derived from natural BCG (or probiotic) or its engineered bacteria (such as cell wall polysaccharide) or an engineering analog of the preparation (such as similar to cell wall Synthetic polysaccharides of polysaccharides or polysaccharides from other sources).
  • the present invention also provides an application of the local pharmaceutical composition in the preparation of a medicament for treating solid tumors, characterized in that the pharmaceutical composition is used as a local pharmaceutical composition for intratumoral administration; in the pharmaceutical composition
  • the dextran described in is used as the active ingredient (reversal agent) that can provide intratumoral common reversal effect, and the non-synergistic drug is used as the active ingredient (reversed agent) that provides intratumoral shared effect reversed, wherein the reversal refers to the Such a co-pharmacology of dextran that converts a drug with which it does not act synergistically (non-synergistic) (eg drug A or B) into a co-component with which it locally synergizes (short-term reversal), or/and
  • the co-product for example, the combination of drugs A and B) that has lost its mid- and long-term drug efficacy is transformed into a composition component that effectively inhibits mid- and long-term drug efficacy
  • the present invention also provides the application of dextran which can be used as a blood volume expander in the preparation of medicines for treating solid tumors, characterized in that the dextran is contained in a local pharmaceutical composition for intratumoral administration, in The dextran in the pharmaceutical composition is used as an active ingredient that can provide a common reversal effect in the tumor, wherein the reversal refers to such a common pharmacology of the dextran that it does not have a synergistic effect with it (not synergistic) drug (e.g. drug A or B) to a co-component with which it synergizes locally (short-term reversal), or/and a shift in said co-product (e.g. a combination of drugs A and B) that itself fails in the medium to long term It is a component of the composition that can effectively inhibit browning of medium and long-term drug efficacy (medium and long-term reversal).
  • a synergistic drug e.g. drug A or B
  • the common components are respectively selected from basic organic compounds and basic inorganic salts that share no local synergy with the dextran, and the basic organic compounds and basic inorganic salts each have a Pharmacological concentrations or concentration ratios at which the local effects of drugs are mutually reinforcing.
  • the term "reversal” refers to a co-pharmacology of a drug that converts a drug with which it does not act synergistically (non-synergistic) to a part of the co-component with which it synergizes (short-term reversal) , or/and transform the co-product that has lost its medium- and long-term drug efficacy into a composition component that is effectively inhibited from medium- and long-term drug effect browning (medium- and long-term reversal).
  • the drug is the reversal agent, and the non-synergistic drug or/and their combination is the reversed agent.
  • dextran is a reversal agent, and a basic organic compound, a basic inorganic salt, or a local synergistic combination of a basic organic compound and a basic inorganic salt is a reversed agent.
  • the present invention also provides a method for treating solid tumors, which includes the following steps: intratumorally administering a therapeutically effective amount of the above-mentioned drug to an individual in need thereof.
  • local administration refers to any administration method that directly delivers the drug to the target area (such as the tumor body), which is different from the blood transport to the target area, such as oral administration, intravenous injection, intraperitoneal injection, etc. Conventional methods of administration such as injections.
  • introduction refers to a kind of local administration whose target area is tumor tissue, including intratumoral administration through hemangioma and percutaneous intratumoral injection. Intratumoral administration of chemical drugs and their compositions is generally considered a local treatment.
  • local treatment is different from “systemic treatment”, the latter refers to the treatment of the whole body of the patient (such as the tumor body, the area connected with the tumor body, and tumor cells contained in other parts of the body), while the former refers to the treatment of the patient's local lesion.
  • the treatment of the local area (such as administration of local lesions and other diseased areas connected with it).
  • the term "local effect” is not a conventional effect (also known as absorption effect), which refers to a drug that is routinely administered (also known as absorption administration, such as oral, intravenous, intraperitoneal injection) and then absorbed into the blood and delivered to a highly specific The pharmacological effect produced by the drug target (such as the specific target of a specific cancer cell), while the former refers to the local area where the drug penetrates into the gap after local administration (direct delivery) to the diseased tissue (such as tumor cell microscopic environment) pharmacological effects.
  • the term "local chemical action” is not an absorption chemical action (eg cytotoxicity), the former and latter being a local action and an absorption effect produced by a chemical drug, respectively.
  • chemotherapeutic drugs chemotherapeutic drugs
  • cytotoxicity ineffective The increase in the local efficacy of chemotherapeutic drugs (cytotoxic drugs) that are effective with conventional medication does not exceed the maximum expected kinetics of cytotoxicity (for example, within 150% of their absorbed efficacy), indicating that their local chemical effects are still their cytotoxic effects; and
  • the improvement of local drug efficacy of chemical drugs that are ineffective by conventional drugs (cytotoxicity ineffective) can exceed the maximum expected kinetics of its absorption (such as cytotoxicity) (for example, more than 200% of the absorbed drug effect), indicating that the main role of its local chemical effect is
  • the drug target is not the specific target of a specific cancer cell when it is a tumor cytotoxic substance, but other tissue structures (such as protein deposits) other than the specific target.
  • the local action of these chemicals is apparently primarily a local chemical action as distinguished from cytotoxicity, ie tissue destruction.
  • chemical ablation traditionally refers to the local chemical action exhibited by classical chemical ablative agents (such as high-concentration ethanol, high-concentration hydrochloric acid or acetic acid, high-concentration sodium hydroxide), the drug effect of which is effective tissue destruction.
  • chemical-like ablation refers to other chemical drugs (also known as conventional ineffective drugs) that are ineffective (cytotoxic ineffective) by conventional drugs, such as dextran, basic organic compounds, and basic inorganic drugs that can be used as blood volume expanders in this application.
  • the local action or local chemical action shown by salt also has a drug effect of directly destroying the tissue, and the drug effect of tumor tissue necrosis can be displayed in a short period of time (such as within 5 days after administration), which is highly similar to chemical ablation. It is generally believed that like chemical ablation, its short-term efficacy will fail in the medium and long term, resulting in a high recurrence rate.
  • the term “topical (locally acting) drug” refers to a chemical substance and the conditions under which its local action is achieved.
  • drug with (not appreciable) local effect refers to a chemical drug that (fails to) meet the generally accepted criteria for appreciable drug effect in the industry. In animal experiments, “the local effect is appreciable (not appreciable)” means that the difference from the negative control is sufficiently (not) obvious, for example, the tumor inhibition rate is ⁇ 15% ( ⁇ 15%).
  • Drugs with considerable local effects such as glucose with an intratumoral concentration of ⁇ 30%, and drugs with insignificant local effects, such as dextran with an intratumoral concentration of ⁇ 30%.
  • the term "locally effective (ineffective) drug” refers to a chemical drug that can (fail) meet the generally accepted standard of drug effective effect in the industry. In animal experiments, "local effect is effective (ineffective)” means that the tumor inhibition rate is ⁇ 40% ( ⁇ 40%). Locally effective drugs such as ethanol with an intratumoral concentration of ⁇ 95%, while dextran with an intratumoral concentration of ⁇ 30% provide insignificant local effects, let alone effective.
  • the term "a drug with the maximum (minimum) local effect” refers to a drug whose local effect (eg, tumor inhibition rate) no longer increases (decreases) significantly with increasing (decreasing) concentration.
  • minimal local effect is clearly the local effect of the negative control, and “minimal local effect” is close to the local effect of the negative control.
  • a local effect is minimized when the insignificant local effect no longer significantly decreases with decreasing concentration (eg, tumor inhibition ⁇ 10%).
  • Drugs that maximize local effects such as dextran administered at an intratumoral concentration of ⁇ 35%, and drugs that minimize local effects, such as dextran administered at an intratumoral concentration of ⁇ 25%.
  • composition refers to a compound preparation comprising at least two active ingredients.
  • topical pharmaceutical composition refers to a combination formulation comprising fixed concentrations or concentration ratios of at least two active ingredients in a topically administered dosage form, wherein said combination formulation refers to a shared form in which said active ingredients can be thoroughly mixed into a target area the concentration or concentration ratio is the concentration or concentration ratio at which the active ingredient enters the target area; the topical dosage form is distinguished from the absorption dosage form, e.g., does not contain the latter as required to reduce the local effect at the site of administration For the purpose of inactive ingredients (such as isotonicity regulator sodium chloride, etc.).
  • inactive ingredients such as isotonicity regulator sodium chloride, etc.
  • the therapeutic effect of the treatment is mainly the intratumoral shared effect of the components of the drug, and its core pharmacology is the reversal effect of dextran, which makes the drug effect of the intratumoral shared effect short-term local treatment, or/and moderate long-term treatment.
  • the pharmacology of the short-term local treatment includes chemical-like ablation of local chemical synergy of one or more tumor tissues and optionally other chemotherapy, and the chemical-like ablation is independent of tumor cytotoxicity; the medium and long-term treatment
  • the pharmacology of the drug includes secondary immune effects and other secondary effects of local effects (or local synergistic effects) in tumor tissues.
  • the applicable patients for the treatment are selected from one or more of the following groups: immunosuppressed patients, patients who can be administered into local lesions, patients whose local lesions can be chemically ablated, patients whose local lesions can be administered Patients with secondary immunity.
  • the composition is a chemoablation-in situ immunotherapy-like drug.
  • immunotherapy is different from the term “immunoenhancement”.
  • the former refers to the immune effect (such as the immune effect of therapeutic vaccines, specific antibodies, etc.) Function, but still have auxiliary immune function (such as immune enhancer has the effect of improving the immune function of the body).
  • the local pharmaceutical composition is an injection for intratumoral administration.
  • the injection is a powder injection.
  • injection refers to a sterile preparation that contains the active ingredient and its vehicle and can be administered in vivo;
  • "powder injection” refers to an injection comprising a sterile dry powder and a vehicle, the sterile dry powder contains part or all of the active ingredient, and the vehicle Contains all liquid carriers.
  • concentration of the injection is usually the concentration of the preparation, and one of the characteristics of the injection for intratumoral administration is that the concentration of the preparation must be greater than or equal to its pharmacological concentration (usually much higher than the absorption pharmacological concentration).
  • the pharmacological concentration of the active ingredient in the injection for intratumoral administration is usually the concentration of the active ingredient in the liquid drug at the end point (such as needle hole, catheter outlet, etc.) of the local drug delivery device (syringe, puncture, injection catheter, etc.).
  • the concentration of the active ingredient is the concentration of the active ingredient in the mixture of a specified amount of sterile dry powder and a specified amount of vehicle (such as reconstitution solution or the pharmaceutically acceptable liquid carrier).
  • the "solid tumor” refers to tumors with tumor tissue, which can be due to any pathology (malignant and non-malignant) and tumors at any stage, according to tumor cell types, including epithelial cell tumors, sarcoma, lymphoma, germ cell tumors, etc.
  • Tumors, blastomas according to the organ or tissue where the tumor cells are concentrated, including brain tumors, skin tumors, bone tumors, muscle tumors, breast tumors, kidney tumors, liver tumors, lung tumors, gallbladder tumors, pancreatic tumors, and esophageal tumors , tumors of the bladder muscle, large intestine, small intestine, spleen, stomach, prostate, ovaries, or uterus.
  • the tumors include malignant tumors and non-malignant tumors
  • the malignant tumors include breast cancer, pancreatic cancer, thyroid cancer, nasopharyngeal cancer, prostate cancer, liver cancer, lung cancer, intestinal cancer, oral cancer, esophageal cancer, gastric cancer, laryngeal cancer, Testicular cancer, vaginal cancer, uterine cancer, ovarian cancer, malignant lymphoma, malignant brain tumor, etc.
  • the non-malignant tumors include breast tumors, pancreatic tumors, thyroid tumors, prostate tumors, liver tumors, lung tumors, intestinal tumors, oral tumors , Esophageal tumors, gastric tumors, nasopharyngeal tumors, laryngeal tumors, testicular tumors, vaginal tumors, uterine tumors, fallopian tube tumors, ovarian tumors, lymphomas, brain tumors, etc.
  • the solid tumor is selected from malignant tumors and non-malignant tumors suitable for intratumoral administration.
  • the conventional ineffective drugs and their combinations administered in the tumor can theoretically be used for any solid tumor, because they all contain the target of their local chemical action - tumor tissue, but currently in clinical practice, their use in anti-tumor drugs The proportion is less than 1%.
  • the main limitation is the many subjective and objective factors involved in the "suitable for intratumoral drug delivery".
  • the solid tumor is selected from solid tumors that are not suitable for chemical ablative agents or conventional local synergistic compositions due to easy recurrence, wherein the conventional local synergistic composition refers to a combination of locally acting drugs and their synergists or synergists Formed topical pharmaceutical compositions such as said co-administrations, or combinations of dextran analogues such as glucose with these co-examinations.
  • the above tumors suitable for intratumoral administration includes at least a very small number of tumors that are prone to relapse after chemical ablation or chemical-like ablation.
  • the solid tumor has at least one tumor size greater than 6 cm, or greater than 10 cm.
  • the present invention also provides a pharmaceutical kit, which is characterized by comprising a container comprising the following independently packaged formulations: comprising the above topical pharmaceutical composition; and a formulation comprising a vehicle.
  • a container comprising the following independently packaged formulations: comprising the above topical pharmaceutical composition; and a formulation comprising a vehicle.
  • Such containers may include ampoules, vials, and the like.
  • the pharmaceutical kit further includes instructions or labels.
  • the ratio of the administration amount of the local pharmaceutical composition to the volume of the target area in the local lesion is >0.1, 0.15-1.5, preferably 0.23-1.5 or 0.5-1.5.
  • the dosage of the local pharmaceutical composition is ⁇ 1ml, or the dosage inside the local lesion is 10-150ml or/and the dosage outside the local lesion is 1.5-50ml.
  • the topical pharmaceutical composition of the present invention has the following advantages:
  • the local pharmaceutical composition has good medium and long-term curative effect and has almost non-toxic systemic drug safety; compared with molecular targeted drugs, it does not require harsh indication screening, and is suitable for rapid Growing tumors, large tumors, and hypovascular tumors show great potential; compared with chemical ablative agents or combined chemoablative agents (including topical pharmaceutical compositions of locally acting drugs and their synergists or synergistic combinations), The local irritation is smaller, the specificity is strong, and the medium and long-term curative effect is higher; (2) the local pharmaceutical composition described in the application does not have the problem of drug resistance, the preparation method is simple, the cost is low, and it benefits a wide range of groups.
  • Fig. 1 is the histopathological HE staining examination result of the sample of embodiment 6 of the present invention
  • Fig. 2 is the TUNEL inspection result of the 7 part samples of the embodiment of the present invention.
  • Fig. 3 is the appearance diagram of the tumor before and after the experiment in Example 10 of the present invention.
  • the above includes any dextran having an average molecular weight between 9,000 and 76,000.
  • mice All SPF (Specific Pathogen Free, no specific pathogen) grade animals purchased from professional laboratory animal companies, unless otherwise specified, wherein mice (BALB/c) and nude mice (BALB /c Nude) were healthy females aged 6-8 weeks, weighing 17.5-20.5g. Nude mice are mutant mice with congenital thymus defects, lack of immune response, and are often used for pure chemotherapy of tumors, such as local chemical effects (tissue destruction or chemical ablation).
  • SPF Specific Pathogen Free, no specific pathogen
  • Animal modeling of subcutaneous transplantation of tumor cells to generate tumors is a known technology. When the tumor grows to the required volume (such as 50-500mm3 in mice), the modeling is successful. After modeling, the animals are divided into several groups. Six rats in each group were regularly observed and measured for general state, body weight, food intake, animal graft-versus-host disease, tumor volume, tumor weight, and survival time.
  • Tumor volume (V), tumor proliferation rate (R), and tumor inhibition rate (r) were calculated according to the following formulas:
  • Tumor volume V 0.5 ⁇ a ⁇ b 2 , where a and b represent the length and width of the tumor respectively;
  • Tumor proliferation rate R TV/CV ⁇ 100, wherein TV and CV are the tumor volumes of the study group and the negative control group respectively;
  • Tumor inhibition rate (r) 100% - R, where R is the tumor proliferation rate.
  • the efficacy of drug i is recorded as Ei, which can be represented by ri.
  • the pharmacology of a drug can be studied through its efficacy, especially the efficacy of the same drug in different regimens. For example, when the drug i has little difference in efficacy between schemes X and Y (such as Ei X /Ei Y ⁇ 200%), it indicates that the pharmacology of the medicine is the same; When the difference in drug efficacy is large (such as Ei X /Ei Y > 200%), it means that the pharmacology of drug i in program X greatly exceeds the expected range of its pharmacological kinetics in program Y, and it is likely to involve a new drug that is different from program Y.
  • Positive controls for chemotherapy include classic cytotoxic drugs (such as 0.5-1% 5-fluorouracil, whose short-term tumor inhibition rate is ⁇ 30% under the conditions of the following examples) and classic chemical ablative agents (such as 75-99% ethanol, Its short-term tumor inhibition rate under the conditions of the following examples is ⁇ 15%).
  • classic cytotoxic drugs such as 0.5-1% 5-fluorouracil, whose short-term tumor inhibition rate is ⁇ 30% under the conditions of the following examples
  • classic chemical ablative agents such as 75-99% ethanol, Its short-term tumor inhibition rate under the conditions of the following examples is ⁇ 15%).
  • copharmacology refers to the mechanism of interaction between the different components of a pharmaceutical composition (eg, a potentiator and its co-pharmaceutics).
  • Drug sharing pharmacology is highly uncertain. The same shared components can form different combinations under different conditions, such as research combination 1 and control combination 2 .
  • the drug effect of research combination 1 such as tumor inhibition rate r 1
  • the drug effect of control combination 2 such as tumor inhibition rate r 2
  • the expected shared drug effect of the latter is defined as Defined as the additive effect of co-administered drugs that does not interfere with each other.
  • the "synergistic effect” then refers to the actual interaction resulting from drug co-administration that is mutually reinforcing
  • short-term synergistic effect is different from the term “medium- and long-term synergistic effect”.
  • the former refers to the synergistic effect shown based on short-term (eg, within 5 days after the first administration) drug effects (such as r 3d1 , r 3d2 ), and the latter refers to the synergistic effect based on The synergistic effect of drug effects (r 21d1 , r 21d2 ) in the medium and long term (eg after the 14th day after the first administration).
  • local synergy is distinguished from the term “non-local synergy”, which refers to synergistic effects based on locally acting pharmacological effects (usually short-term pharmacological effects such as r 3d as mentioned above), which refer to non-locally acting pharmacological effects (Medium and long-term drug effects such as r 21d ) synergistic effect. It is generally believed that the local synergistic effect observed by the combination of chemical drugs on the nude mouse model is mainly the mutual enhancement between the local chemical effects of the compounds, which is obviously a short-term synergistic effect.
  • the acceptable error level between batches is ⁇ 20%, and the acceptable error level within batches is ⁇ 15%, and the difference between groups within the error range is obviously not obvious (or not appreciable).
  • the above r 1 and r 2 may both come from the medium and long-term drug effects, both from the short-term drug effects, or from the medium- and long-term drug effects and the expected (control) short-term drug effects respectively.
  • the short-term drug effect produced by intratumoral administration on local lesions is actually the drug effect of local interactions
  • the medium and long-term drug effect produced by intratumoral administration on local lesions is a drug effect that involves a complex interaction between local effects and secondary immune responses.
  • the shared toxicological effects of drugs I and II are the same as their pharmacological effects, and the toxicological effects (toxic effects) of the research combination and the control combination, such as systemic safety and local irritation, can be used for comparative research, wherein the control combination
  • the safety expectation of shared use is the theoretical simple additive expectation of toxicological effects, that is, the shared toxic effect is not less than the maximum toxic effect of the two drugs.
  • the shared toxicological effect of the drug I/II composition is judged by the following method: if the actual toxic effect of the composition is consistent with the expected toxic effect of theoretical simple addition, then the sharing of the composition is a toxicological additive effect; If the actual toxic effect is lower than the expected toxic effect of theoretical simple addition (for example, the shared toxic effect is less than the maximum toxic effect of the two drugs), the sharing of the composition is an obvious toxicological synergy; if the actual shared toxic effect of the composition is not If it is less than the maximum toxic effect of the two drugs, greater than the expected combined toxic effect of the theoretical simple addition, and not less than the maximum toxic effect of the two drugs, then the combination of the composition is an obvious toxicological antagonism.
  • composition preparation is prepared with water for injection according to the composition components, pharmacological concentration and concentration ratio.
  • the preparation method of related solutions is the prior art, and will not be repeated here.
  • the prepared liquid preparation can be freeze-dried to obtain the freeze-dried powder for injection.
  • the freeze-drying adopts the following process: keep the pre-freezing temperature -45°C for 4 hours, the temperature increase rate is 0.1°C/min and rise to -15°C for at least 10 hours for sublimation; Keep for 6 hours for desorption and drying, subpackage (such as 7.5ml/bottle), and cap.
  • the aseptic medium is pumped into the bottle, mixed evenly and can be used as an injection medicine, such as 1.5% broken components of Saccharomyces cerevisiae/20% amino acid.
  • each of the four compositions contains 20% expanders, 20% arginine and 5.8% sodium carbonate, wherein the expanders are: dextran 40, dextran 70, carboxyethyl starch 200, and polyvinylpyrrolidone k70.
  • the aqueous solution of the composition prepared by the conventional method is filtered with the same filter (pore size 0.2um) at room temperature according to the conventional sterilizing filtration method, and the dextran composition can be sterilized and filtered through the filter, and then the two compositions are due to pressure when passing through the filter. Too high Sterile filtration not done.
  • each of the 4 compositions contained X% dextran 70, 20% arginine and 5.8% sodium carbonate, wherein said X% was: 2%, 4.5%, 10%, 25%, 30%, respectively.
  • the aqueous solution of the composition prepared according to the conventional method is filtered with the same filter (pore size 0.2um) according to the conventional sterilization filtration method at room temperature, and the first 4 compositions with a dextran concentration of 2-25% can be sterilized by the filter.
  • the sterilizing filtration was not completed because the pressure was too high. It could only be diluted, filtered and then concentrated, which increased the risk of bacterial contamination.
  • the preferred cleanliness of the manufacturing process for the pharmaceutical preparation of dextran and different combinations was compared.
  • the prescriptions of the two compositions were: 20% arginine/5.8% sodium carbonate/4.5% dextran 70, 1% methylene blue/5.8% sodium carbonate/4.5% dextran 70, respectively.
  • the first composition did not have any obvious contamination; while the second composition was stained on any utensils it was connected to and was difficult to clean, especially causing freeze-drying machines Pollution will greatly increase the preparation cost.
  • the production batch is the active ingredient of each batch of 5000g dry weight, including arginine (all L-arginine in this application), sodium carbonate and dextran 70, and the ratio of each component is 3.5g arginine/1g sodium carbonate/ 0.6g of dextran 70
  • the pharmaceutical dosage form is freeze-dried dosage form, when the drug is used, it is prepared with an appropriate amount of water for injection into an aqueous solution of 2-4.3% dextran 70, 11.7-25% arginine, and 3.3-7.1% sodium carbonate for intratumoral administration.
  • Tumor-bearing nude mice are widely used in drug research of chemotherapy rather than immunotherapy for patients with solid tumors.
  • Nude mice were used as experimental objects, and 2.5 ⁇ 10 6 mouse liver cancer Hepa1-6 cells/mouse were injected into the right axilla
  • the transplanted tumor was modeled subcutaneously.
  • the average volume of the tumor in nude mice successfully modeled was 75.7mm 3 .
  • the model animals were randomly divided into 12 groups, and the components of each group in Table 2 were injected into the tumor, and each group was administered twice.
  • the medication interval is 2 days, and the volume of each injection is 50 ⁇ l/monkey.
  • the local lesion volume (V) was measured, and the tumor inhibition rate (r) was calculated according to the negative control group.
  • the tumor inhibition rate (12.3%) of the expansion agent A 2 in group 3 is less than 1/2 (25.7%) of the control A 1 with considerable local effect, which does not provide considerable local effect.
  • the drug efficacy of groups 10 and 11 is lower than that of groups 7 and 8, which is in line with the expectation of conventional local synergism: co-administered with the same drug (B or C), the drug with weak local effect (A 2 ) has a low co-pharmaceutical effect Drugs with strong local effects (A 1 ).
  • a 2 and A 1 were combined with the same co-products (B and C) respectively, the efficacy of group 12 was higher than that of group 9, which was contrary to the aforementioned conventional synergistic expectations.
  • the actual/expected ratio q of the co-drug effect of group 12 is >1.00, which is contrary to the aforementioned conventional synergistic expectations, that is, unconventional use of non-synergistic drugs (B, C) Reversion to the active ingredient of the synergistic co-occurrence (B/C).
  • the reversal effect of A 2 but not of A 1 should be the pharmacological basis for the pharmacological effect of A 2 /B/C to be greater than that of A 1 /B/C, which also shows that the reversal effect is drug-selective.
  • compositions of A 2 and A 1 and the same combination B/C are not only completely opposite in the above shared pharmacology (reversal effect vs no reversal effect), but also very different in shared toxicology.
  • Moderate local irritation was observed with B/C in group 6 (33% of the animals observed a high level of struggle) and local irritation was observed with A 1 /B/C in group 9
  • Significantly strengthened (100% of the animals struggled with high intensity) while group 12 observed obvious antagonism of the local stimulation phenomenon (no animals struggled with high intensity) when A 2 /B/C was administered.
  • the number of animals with significant local leakage was more than 50% less in group 12 (A 2 /B/C) when administered compared to group 9 (A 1 /B/C).
  • cohort 12 ( A2 /B/C) showed a safety synergy that significantly improved the local safety of B/C.
  • the modeled animals are nude mice, and the modeled cells (2.5 ⁇ 10 6 cells/only) are human pancreatic cancer cells (PANC-1).
  • the modeling method is the same as that in Example 2, and will not be repeated here.
  • the average tumor volume of the mice was 81.3mm 3 .
  • the model animals were randomly divided into 38 groups, and the components of each group were injected into the tumor according to Table 3. Each group was treated twice with an interval of 3 days, and the volume of each injection was 50 ⁇ l /Only. On the 3rd day after the second administration, the local lesion volume (V) was measured, and the tumor inhibition rate (r) was calculated according to the negative control group.
  • V local lesion volume
  • r tumor inhibition rate
  • the actual/expected ratio q of the co-efficacy of groups 25-28 is all less than 1.00, and the glyconutrient (A1) does not show its inconsistency drug (B, C) and its combination (B/C) Reverses the activity of its synergistic counterparts.
  • the structures of the blood volume expanders in this example are highly similar to glucose, and they probably have the same shared pharmacology under the same conditions. The q ⁇ 1.00 for group 29 seems to confirm this.
  • the B/C in the composition of group 29 is the combination of q ⁇ 1.00 in group 19
  • the B/C in the composition of group 30-37 is the combination of q>1.00 in group 20-28, respectively Combination, indicating that not any combination of non-synergistic drugs, only when the amount ratio between them makes their local effects mutually enhance the combination of non-synergistic drugs can be reversed by dilators, or that the reversal effect has non-synergistic Drug combination selectivity.
  • dextran 10, dextran 20, and dextran 70 replaced dextran 40 in this experiment, tromethamine, triethanolamine, tris, meglumine, lysine, etc.
  • dextran with an intratumoral administration concentration of ⁇ 25% can be used as an active ingredient (reversal agent) that provides a reversal of the shared effect
  • a basic organic compound or a basic inorganic salt that does not produce a local synergistic effect with the dextran can be used as its Non-synergistic drugs (reversed agents)
  • the synergistic combination of basic organic compounds and basic inorganic salts can be used as a synergistic co-share of dextran, and dextran can transform its non-synergistic drugs into synergistic co-components to provide synergistic effects ; and/or reduce the local irritation of the joint, providing a synergistic effect on safety.
  • the modeling animals are nude mice, and the modeling cells (2 ⁇ 10 6 cells/mouse) are mouse malignant sarcoma cells (S180 cells).
  • the modeling method is the same as in Example 2, and will not be repeated here.
  • the average volume of the tumor in the successfully modeled nude mice was 74.9mm 3 , and the model animals were randomly divided into 22 groups, and each group except the 16-18 group was treated with A, conventional administration (tail vein injection); B, tumor Intra-injection was administered in two ways, and each single group was administered twice, with an interval of 3 days, and the volume of each injection was 50 ⁇ l per mouse.
  • the local lesion volume (V) was measured, and the tumor inhibition rate (r A , r B ) was calculated according to the negative control group of series A and B.
  • the results are shown in Table 5.
  • the pharmaceutical compositions used in each group were prepared according to the conventional aqueous solution preparation method or the preparation method in Example 1.
  • the administration method of group 16-17 in series A is: conventional administration of blood volume expansion agent + intratumoral injection of common ingredient composition
  • the administration method in series B is intratumoral injection of blood volume expander + common ingredient composition Compositions are administered conventionally.
  • the administration method of group 18 in series A is intratumoral sequential injection of blood volume expansion agent and common component composition
  • the administration mode of series B is intratumoral sequential injection of common component composition and blood volume expansion agent.
  • Table 5 The tumor inhibition rate data of different drug combinations and administration methods
  • the drug effect produced by conventional administration reflects the systemic pharmacological effects of the drug (such as individual nutrition enhancement or overall imbalance, tumor cytotoxicity, immune enhancement, sustained release in the blood, etc.)
  • the drug effect produced by intratumoral administration reflects local effects such as local chemical damage caused by the drug directly entering the local lesion.
  • the efficacy of chemical drugs is mainly short-term.
  • the efficacy of chemotherapy is concerned, if the efficacy of the drug is considerable after conventional administration (r A is greater than 15%) and the efficacy of intratumoral administration can also be improved based on kinetic conditions (such as 100% ⁇ r B /r A ⁇ 200%), it shows that its pharmacology is cytotoxicity (such as group 02); if the drug effect is not appreciable through conventional administration (r A is less than 15%), the drug effect of intratumoral administration is considerable (r B > 15%) and r B /r A > 200%, it means that its pharmacology is local tissue destruction independent of cytotoxicity, which is also referred to as local effect in the application of the present invention (such as group 03).
  • compositions in groups 16, 18 and group 12, group 17 and group 15 have the same components and component ratios, the only difference is that the dosage forms are different (non- topical dosage form and topical dosage form where the active ingredients must be administered in admixture); unexpectedly, different dosage forms showed completely different co-efficacy, indicating that dextran can only be used under the condition of intratumoral co-administration. Showing its shared reversal effect.
  • necrotic areas were of different sizes, mostly in the form of flakes or bands , distributed in the edge and central area of tumor tissue.
  • the number of necrotic tumor cells in the necrotic area is large, a large number of tumor cell nuclei are condensed and deeply stained, and a large number of nuclear fragments, which further confirm the special pharmacological effects of the compositions of groups 12 and 15 after intratumoral injection.
  • Example 5 Study of local action characteristics required for reversal
  • the modeling animals are nude mice, and the modeling cells (1 ⁇ 10 6 cells/mouse) are mouse breast cancer cells (4T1 cells).
  • the modeling method is the same as in Example 2, and will not be repeated here.
  • the average tumor volume of nude mice successfully modeled was 84.1 mm 3 , and the model animals were randomly divided into 26 groups.
  • Intratumoral injection was performed according to the components in Table 6.
  • the pharmaceutical compositions used in each group were prepared according to the conventional aqueous solution preparation method or the preparation method in Example 1. Each group was administered twice, with an interval of 3 days, and the volume of each injection was 50 ⁇ l per mouse. On the 3rd day after the second administration, the local lesion volume (V) was measured, and the tumor inhibition rate (r) was calculated according to the negative control group.
  • group 25 (dilator/combination of 3 non-synergistic drugs) had an actual/expected ratio q greater than 1.00 for co-efficacy, also showing local synergy.
  • the shared effect of at least 2 non-synergistic drugs is a partial synergistic effect.
  • Examples 2-4 basic organic compounds and basic inorganic salts are non-synergistic drugs of dilators with minimized local effects, respectively.
  • the modeled animals are mice, and the modeled cells (0.5 ⁇ 10 5 cells/mouse) are mouse breast cancer cells (4T1 cells).
  • the modeling method is the same as in Example 2, and will not be repeated here.
  • the average tumor volume of the successfully modeled mice was 107.3mm 3 .
  • the model animals were randomly divided into 23 groups, and each group was given intratumoral injection. The injection volume is 100 ⁇ l/only.
  • Each composition was prepared according to the conventional aqueous solution preparation method or the preparation method in Example 1, wherein the compositions used in Groups 15-17 respectively formed Compositions 1-4 for common use.
  • the local lesion volume (V) was measured on the 3rd and 21st day after the second administration, and the drug efficacy-tumor inhibition rate (r 3d , r 21d ) was calculated according to the negative control group.
  • the results are shown in Table 7. According to the drug effects r 3d and r 21d , the actual/expected ratio q 3d of the short-term drug effect and the actual/expected ratio q 21d of the medium and long-term drug effect can be calculated respectively,
  • the medium and long-term drug effects of dextran are as unsatisfactory as its short-term drug effects, and the long-term local effects are minimized as its short-term local effects;
  • arginine, sodium bicarbonate (group 4 , 7)
  • methylene blue (group 5)
  • their mid- and long-term drug efficacy declines more obviously, all of which are exhausted to almost nothing (r 3d ⁇ 5%).
  • the dextran 10, dextran 20, dextran 40, and dextran 70 of intratumoral administration concentration of 2-25% all show the result consistent with group 1 in the above table
  • the following intratumoral administration concentration (2 -25%) of the basic organic compounds showed results consistent with group 4 in the table above: 2-10% tromethamine, 2-15% triethanolamine, 2-15% tris, 10-25% meglumine, 10-20% arginine, 15-25% lysine
  • the following intratumoral administration concentrations (2-15%) of basic inorganic salts all show the same group as the above table 7
  • Consistent results 2-15% sodium carbonate, 5-15% sodium lactate, 5-15% sodium acetate, 5-15% sodium bicarbonate, ie dextran, basic organic compound, basic inorganic salt at the stated experimental concentrations At most, they can show a certain short-term drug effect, and their mid- and long-term drug effects are all reduced to insignificant, showing that they are all drugs with minimal mid- and long-term local effects, and the
  • the mid- and long-term drug efficacy was significantly higher than that of the shared substances (groups 15-17).
  • the mid- and long-term synergies of dextran to its co-products are much higher than the short-term synergies (i.e. rcomposition 21d / co-occurrence r 21d > 150% ⁇ rcomposition 3d / co-production r 3d ), showing that the long-term reversal effect Significantly higher than the short-term reversal effect.
  • necrotic areas vary in size, mostly in the form of flakes or bands; scattered in the edge and central area of the tumor tissue, the distribution is relatively uniform, and the area of necrosis accounts for about 10% of the tumor tissue.
  • necrotic tumor cells were seen in the necrotic area, and the stroma was widened.
  • necrotic tumor cells in the necrotic area The number of necrotic tumor cells in the necrotic area is large, and the nuclei of a large number of tumor cells are condensed and deeply stained, and a large number of nuclei are fragmented; there are more immune cells, neutrophils, and lymphocytes than normal in the area near the tumor.
  • the medium and long-term efficacy may be related to the effect of secondary immunotherapy, which needs to be confirmed by follow-up studies.
  • dextran can provide medium- and long-term reversal effects that are significantly higher than its short-term reversal effects, reverse its non-synergistic drugs into medium- and long-term synergistic co-components, or lower the mid- and long-term efficacy of the co-products themselves.
  • the drug effect eg r 21d /r 3d ⁇ 50%) is reversed to a higher drug effect with less debilitating mid- and long-term drug effect (eg r 21d /r 3d >50%), showing a new pharmacological significance.
  • Embodiment 7 Medium and long-term reversal agent (composition) research
  • Example 2-6 A comparative study was carried out on the drugs that can be used as blood volume expansion agents in Examples 2-6, so as to optimize the reversal agent with the best mid- and long-term common reversal effect.
  • the modeled animals were mice, and the modeled cells were mouse synovial sarcoma cells (ZM755 cells, 3 ⁇ 10 6 cells/mouse). The modeling method was the same as in Example 2, and will not be repeated here.
  • the average tumor volume of the successfully modeled mice was 94.2mm 3 , and the model animals were randomly divided into 11 groups, each group was given medicine twice, and the medicine interval was 3 days, except for group 7, the amount of intratumoral injection 100 ⁇ l/mouse, while the intratumoral injection volume of group 7 was 150 ⁇ l/mouse.
  • the local lesion volume (V) was measured on the 21st day after the second administration, and the tumor inhibition rate (r 21d ) was calculated according to the negative control group.
  • each composition is prepared according to the conventional aqueous solution preparation method or the preparation method in Example 1.
  • the actual/expected ratio q 21d of medium and long-term drug effects can be calculated according to the drug effect r 21d on the 21st, and the drug effect data are listed in the following table.
  • the r 21d of groups 1-5 were all ⁇ 5%, indicating that the three dilators and their two non-synergistic drugs were all drugs with minimal mid-term local effects; the r 21d of groups 6-7 were all ⁇ 5%, indicating that the combination of 2 non-synergistic drugs (at the same dose and increasing dose) does not provide mid-term synergistic effect, and the intratumoral co-drugs of different dilators in groups 9-10 and the non-synergistic drug combination
  • the actual/anticipated ratio q 21d of the effect was both >1, indicating that these dilators all have medium- and long-term communal reversal effects.
  • the histopathological HE staining results of the corresponding tumor samples of r 21d in group 8 are consistent with the results of the representative tumor samples of group 18 in Example 6, and will not be repeated here.
  • the results of the TUNEL examination (entrusted to Chengdu Yonkers Biotechnology Co., Ltd.
  • Embodiment 8 Medium and long-term reversal agent (pharmacological conditions) research
  • the modeled animals were mice, and the modeled cells were mouse breast cancer cells (4T1 cells, 0.5 ⁇ 10 5 cells/mouse), and the modeling method was the same as that in Example 2, which will not be repeated here.
  • the average tumor volume of the successfully modeled mice was 107.3mm3.
  • the local lesion volume (V) was measured on the 21st day after the second administration, and the drug efficacy-tumor inhibition rate (r21d) was calculated according to the negative control group.
  • each composition is prepared according to the conventional aqueous solution preparation method or the preparation method in Example 1.
  • According to the drug effect r21d on the 21st calculate the actual/expected ratio q21d of the medium and long-term drug effect, and the following table lists the drug effect data.
  • the r21d of groups 1-8 are all ⁇ 5%, indicating that 2-25% dextran 40 and its 2 non-synergistic drugs are all drugs with minimal mid-term local effects, and the combination of 2 non-synergistic drugs Does not provide mid-term synergy.
  • the actual/expected ratio q21d of intratumoral co-drug efficacy of different dilators in groups 9-13 and the non-synergistic drug combination was all >1, indicating that the concentrations of these dilators all have medium- and long-term co-reversal effects.
  • the administration dosage of 25% dextran is all lower than its prescribed dosage when it is used as intravenous injection, and administration composition (group 9) and No systemic adverse reactions were seen in the test animals of each corresponding constituent component (group 1, 6-8), indicating that the composition did not produce significantly higher systemic adverse reactions than its respective constituent constituents at this dosage. Toxicity, but produced a significantly higher efficacy.
  • mouse melanoma cells B16F1 cells, 3 ⁇ 106 cells/only
  • mouse liver cancer cells HepG2 cells, 3 ⁇ 106 cells/only
  • mouse rectal cancer cells C26 cells, 3 ⁇ 106 cells/only
  • dextran 10, dextran 20, and dextran 70 to replace dextran 40 in this experiment were also obtained.
  • the experimental results are consistent, and the experimental results will not be repeated here.
  • Dextran at an intratumoral administration concentration of 2-25% thus becomes the preferred technical solution of the present application for active ingredients that can provide intratumoral co-action reversal, especially mid- to long-term reversal.
  • Example 9 Medium and long-term non-synergistic drugs and their combination optimization
  • the average tumor volume of the successfully modeled mice was 103.1mm 3 .
  • the model animals were randomly divided into 27 groups, and each single group was administered twice with an interval of 3 days, and the intratumoral injection volume was 100 ⁇ l per mouse.
  • the local lesion volume (V) was measured on the 21st day after the second administration, and the drug efficacy-tumor inhibition rate (r 21d ) was calculated based on the negative control group.
  • each composition is prepared according to the conventional aqueous solution preparation method or the preparation method in Example 1. According to the drug effect r 21d on the 21st, calculate the actual/expected ratio q 21d of the medium and long-term drug effect, and the following table lists the drug effect data.
  • the r 21d of groups 1-10 are all ⁇ 5%, which is consistent with the results of the single-drug group in Example 6.
  • the dextran 10, dextran 20, dextran 40, and dextran 70 of intratumoral administration concentration of 2-25% all show the result consistent with group 1 in the above table
  • the following intratumoral administration concentration (2 -25%) of the basic organic compounds showed consistent results with Groups 2-5 in the table above: 2-10% tromethamine, 2-15% triethanolamine, 2-15% trishydroxymethylamino Methane, 10-25% meglumine, 10-20% arginine, 15-25% lysine
  • the following intratumoral administration concentrations (2-15%) of basic inorganic salts all show the same as in the above table Consistent results for groups 6-10: 2-15% Sodium Carbonate, 5-15% Sodium Lactate, 5-15% Sodium Acetate, 5-15% Sodium Bicarbonate, ie the stated experimental concentrations of dextran, basic organic compound, Basic inorganic
  • group 20-25 has the highest mid- and long-term drug efficacy, showing that dextran has the strongest mid- and long-term co-reversal effect on the combination of basic organic compounds and basic inorganic salts with minimal mid- and long-term local effects, so It becomes the preferred technical scheme of the combination of non-synergistic drugs of dextran of the present application.
  • the prescription of the pharmaceutical composition is: 200g dextran 40, 200g arginine, and 50g sodium carbonate are dissolved in an appropriate amount of water for injection to prepare a 1L aqueous solution, which is sterilized and filtered through a 0.2um filter and then divided into 30ml/bottles and stored in a refrigerator at 4°C .
  • the administration was percutaneous intratumoral injection, 25ml each time, 3 times in total, and the interval between each time was 14 days.
  • the first administration will be on October 4, 2021 ( Figure 3-2), the tumor will start to shrink significantly on November 7, 2021 ( Figure 3-3), and the tumor will completely disappear on March 28, 2022 ( Figure 3 -4), and the weight of the pig increased to 52 kg.
  • the preferred indication of the present application is the local treatment of large tumors, which according to common knowledge is those whose largest dimension is larger than 6 cm, preferably larger than 7 cm or even larger than 10 cm.
  • Example 11 Further synergistic research on mid- and long-term reversal effects
  • the modeled animals were mice, and the modeled cells were mouse melanoma cells (B16F1 cells, 3 ⁇ 10 6 cells per mouse).
  • the modeling method was the same as that in Example 2, and details will not be repeated here.
  • the average tumor volume of the successfully modeled mice was 98.3 mm 3 .
  • the model animals were randomly divided into 25 groups, and each single group was administered twice with an interval of 3 days, and the intratumoral injection volume was 100 ⁇ l per mouse.
  • the r 21d of groups 18-24 are all > 15%, all showing considerable medium and long-term local effects, and the actual/expected ratio q 21d of their shared intratumoral drug effects are all > 1, showing that the expansion agent
  • These non-synergistic drug combinations (groups 11-17) all have medium and long-term shared reversal effects.
  • the r 21d of groups 20-24 were all higher than those of groups 18-19, which provided higher medium and long-term efficacy for patients who could accept higher side effects (cytotoxic drug toxicity).
  • the specific compound is preferably selected from dextran including dextran 10, dextran 20, dextran 40, and dextran 70; its specific role is selected from, rather than its sugar nutrient analogues (such as Glucose); its specific effect is selected from ineffective, preferably not appreciable, more preferably minimized local effect, rather than at least appreciable, preferably effective, more preferably maximized local effect selected according to conventional synergist requirements; its specific composition conditions Medium and low intratumoral administration concentration to meet the specific effect: 2-25%, 2-20%, 2-10%, or 2-4.5%, rather than the high intratumoral administration required by conventional synergists concentration;
  • the above-mentioned reversed agent of dextran can only select a specific compound, its specific effect, and its specific composition conditions.
  • the specific compound is preferably selected from basic organic compounds and basic inorganic salts, rather than its basic analogues such as metal sodium hydroxide; its specific role is as the dextran non-synergistic drugs, rather than synergistic drugs required by conventional synergists; its specific compositional conditions are the compositional conditions required for partial synergy between the basic organic compound and basic inorganic salts, rather than conventional synergistic The compositional conditions required for local synergy between the desired basic organic compound or basic inorganic salt and dextran.
  • the basic organic compound is selected from at least one of tromethamine, triethanolamine, trishydroxymethylaminomethane, meglumine, arginine, and lysine
  • the basic inorganic salt is selected from the group consisting of At least one of sodium bicarbonate, sodium carbonate, sodium lactate, sodium acetate
  • the composition condition between the non-synergistic drugs that can be reversed by the synergist is: the concentration between the basic organic compound and the basic inorganic salt The ratio is (3-25%)/(4-25%).
  • the dextran/basic organic compound/basic inorganic salt composition can also optionally add one or more of the following groups: methylene blue and its analogs, immunomodulators, cytotoxic drugs, such as The following combination: the concentration ratio (W basic organic compound /W basic inorganic salt / methylene blue and its analogs ) is (5-25%)/(3-15%)/(0.35-5%) basic Organic compound/basic inorganic salt/methylene blue and its analog combination, concentration ratio (W basic organic compound /W basic inorganic salt / immune regulator ) is (5-25%)/(3-15%) / (1-15%) basic organic compound/basic inorganic salt/immunomodulator combination, concentration ratio (W basic organic compound /W basic inorganic salt / cytotoxic drug ) is (5-25%)/ (3-15%)/(0.1-10%) combination of basic organic compound/basic inorganic salt/cytotoxic drug.
  • the concentration ratio (W basic organic compound /W basic inorganic salt / methylene blue and its analogs ) is

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Abstract

本发明公开了作为血容量扩张剂的右旋糖酐的新用途。所述右旋糖酐在特定条件下显示出针对特定药物的共用逆转作用，从而赋予其新的成药性。本发明还公开了一种根据该用途的瘤内给药组合物及治疗方法。本发明所述的应用、瘤内给药组合物以及方法的中长期疗效更佳，对快速生长瘤体、大瘤体和乏血供瘤体显示出巨大的潜力。

Description

一种局部药物组合物、应用及试剂盒 技术领域
本发明涉及药物制剂技术领域，特别涉及右旋糖酐化合物的一种新用途，以及根据该用途成药的一种新药物组合物和应用该组合物的一种新治疗方法。
背景技术
从病变组织的角度看，实体肿瘤是局部病变疾病的一个代表性模型，其局部病变为包含肿瘤细胞的组织(瘤体和被肿瘤细胞污染的淋巴结)。除肿瘤细胞，瘤体组织中往往还存在成纤维细胞、细胞间质、管道等组成，有时也被称作肿瘤细胞的微环境。以胰腺癌瘤体为例，胰腺癌细胞在瘤体组织中仅占约30％，这些微环境使细胞毒药物与肿瘤细胞之间的反应动力学十分复杂，可能大大降低药效。例如，研究者在体外反应器中发现针对某特定肿瘤细胞产生细胞毒作用的物质针对某特定肿瘤细胞产生细胞毒作用的物质很多，但可用于临床的细胞毒药物却少之又少。老药新用及其共用于是成为一个重要研究方向。
药物共用研究的目标为协同作用。协同(或增效)作用被广泛接受的定义为“增效剂与共用成分之间类似作用的相互增强”，化学药物之间的局部协同(或增效)作用便是增效剂与共用成分之间各自化学局部作用(组织破坏作用)的互相增强，增效剂与共用成分的选择原则于是应当是使它们的实际共用药效超过预期共用药效。预期共用药效的预期显然基于它们各自化学局部作用药效，均为类化学消融的短期药物效应,而非中长期药物效应。具体而言，局部增效剂的药理条件的选择显然为最大化、至少为有效的局部作用(组织破坏作用，即类化学消融)，而非相反；其瘤内共用成分的选择显然为其局部协同物或它们的组合，而非其不协同药物的组合；而所述增效剂与共用成分之间的优选组合的选择显然应当基于类化学消融的(短期)药物效应，而非中长期药物效应。在本申请中，符合上述技术方案(增效剂局部作用强度、局部协同物或它们的组合、类化学消融的短期药效)的局部协同(或增效)被称作常规局部协同(或增效)，否则便是非常规局部协同(或增效)。
现实中的化学药物共用合乎上述预期，即均为常规局部协同的技术方案。例如化疗药物－5-氟尿嘧啶与顺铂之间，选择在各自均有效、一般为最大化细胞毒作用的给药浓度条件下进行瘤内给药组合；常规化疗无效药物－例如葡萄糖和亚甲蓝之间，选择各自均为有效或最大化类化学消融(局部作用)的给药浓度条件下进行瘤内给药组合。类似的其它局部作用药物(例如碱性有机化合物和碱性无机盐)，也是如此。在这些技术方案中，增效剂的用途均为提供常规局部协同、而不是非常规局部协同。这些常规局部药物组合物虽然可以为患者提供增效的瘤组织破坏药效，但普遍显示高复发率，反应其协同药效在中长期完全衰竭，远远满足不了临床上对中长期疗效的需要，这也是这类组合物乏人问津的原因之一。
本发明要解决的问题，便是如何使得右旋糖酐除了用作血容量扩张剂之外还具有新的用途：局部共用作用逆转，即将本来与之并不发生协同作用的不协同药物转变为与之局部协同的共用物成分、或/和 使得本身中长期药效衰竭的所述共用物转变为中长期药效可观的组合物成分，为实体肿瘤临床需要提供一种容易制备、几无全身性副作用的中长效瘤内给药组合物及一种中长效治疗方法。迄今仍无可用作血容量扩张剂的右旋糖酐该项功能的报道，更谈不上该项功能的用途。
发明内容
有鉴于此，本发明旨在提出右旋糖酐化合物的一种新药理-瘤内共用作用逆转，及其新用途。具体而言，选择局部作用尤其是中长期局部作用最小化、而非最大化的可用作血容量扩张剂的右旋糖酐作为增效剂，选择其非协同药物、而非其协同药物作为其共用物组分，选择将这些非协同药物转化为其协同共用物组分或/和抑制所述共用物的中长期药效衰竭的共用逆转作用、而非对其协同药物的直接协同作用作为共用药理，从而制备的用于瘤内给药的中长效药物组合物和提供的新的中长效治疗方法，实现高效低毒，尤其是中长期内，以为临床提供供各种选择。
本申请的发明人意外地发现，通常用作血容量扩张剂的聚乙烯吡咯烷酮在超出常规增效预期的某些特定条件下(例如其局部作用最小化)，居然可使本来与之共用并无协同作用的不协同药物逆转为其协同共用物成分。进一步的研究更发现，可作为扩张剂的右旋糖酐居然还能进一步逆转所述共用物的中长期药效衰竭。
本发明提供了一种局部药物组合物，其瘤内给药形式为可用作血容量扩张剂的右旋糖酐及其不协同药物的水溶液，其中所述右旋糖酐的药理浓度为≦25％；所述不协同药物分别选自与所述扩张剂共用未显示出局部协同作用的碱性有机化合物(药物组A)和碱性无机盐(药物组B)，所述碱性有机化合物包括氨丁三醇、三乙醇胺、三羟甲基氨基甲烷、葡甲胺、精氨酸、赖氨酸的至少一种，所述碱性无机盐包括碳酸氢钠、碳酸钠、乳酸钠、乙酸钠的至少一种，所述多种不协同药物的药理浓度为4-40％。
优选的，所述血容量扩张剂的药理浓度为2-25％。
优选的，所述血容量扩张剂的药理浓度为2-4.5％。
在本申请中，除另有说明外，“浓度”是指重量/体积百分比浓度。＂药理浓度＂是指使得药物提供选定药理(例如右旋糖酐的共用逆转作用、不协同药物可被右旋糖酐逆转的共用作用)所需的瘤内给药浓度。
优选的，所述右旋糖酐选自可用作血容量扩张剂的右旋糖酐类化合物，包括右旋糖酐10、右旋糖酐20、右旋糖酐40、右旋糖酐70中的至少一种。
优选的，所述碱性有机化合物(药物组A)和碱性无机盐(药物组B)的选择条件为：所述A或B与所述右旋糖酐共用时不发生协同作用、而所述A和B与所述右旋糖酐共用时存在协同作用，以及所述协同作用包括短期协同作用和/或中长期协同作用，所述协同作用包括药效协同和/或安全性协同。
优选的，碱性有机化合物(药物组A)和碱性无机盐(药物组B)的药理浓度分别为：至少可提供可观短期局部药效的瘤内给药浓度，其中所述碱性有机化合物(药物A)的药理浓度为5－20％,以及当所述碱性有机化合物选自精氨酸或耐氨酸时,所述碱性有机化合物的药理浓度为10-25％；所述碱性无机盐(药物B)的药理浓度为3-15％。
优选的，所述碱性有机化合物(药物组A)和碱性无机盐(药物组B)的量比决定条件为：所述A和B的形成的组合为局部协同组合(短期协同)。
优选的，所述A、B组合(碱性有机化合物/碱性无机盐)的浓度比为(5-25％)/(3-15％)。所述浓度比是指在形成的药物组合物中的含量比例。
优选的，所述碱性有机化合物和所述碱性无机盐的组合选自以下组合之一种：精氨酸/碱性无机盐组合，浓度比为(10-20％)/(2-15％)；精氨酸/三羟甲基氨基甲烷/碱性无机盐组合，浓度比为(10-20％)/(2-15％)/(2-15％)；赖氨酸/碱性无机盐组合，浓度比为(15-25％)/(2-15％)；氨丁三醇/碱性无机盐组合，浓度比为(2-10％)/(2-15％)；三乙醇胺/碱性无机盐组合，浓度比为(2-10％)/(2-15％)；三羟甲基氨基甲烷/碱性无机盐组合，浓度比为(2-15％)/(2-15％)；葡甲胺/碱性无机盐组合，浓度比为(10-25％)/(2-15％)。
任选的，还包括药理浓度为0.03-6％的细胞毒药物、或/和药理浓度为0.2-25％的免疫调节剂。
所述细胞毒药物是指主要以病变细胞或病变细胞内结构为靶实现其药物效应的活性成分，例如在细胞实验或在荷瘤动物中显示出抗癌细胞增殖的物质，包括：1)破坏DNA结构和功能的药物，如烷化剂-环磷酰胺、卡莫司汀等、金属铂络合物-顺铂、卡铂等、DNA拓扑异构酶抑制剂-多柔比星类、拓扑替康、伊立替康等；2)嵌入DNA中干扰转录RNA的药物，如抗肿瘤抗生素-放线菌素类、柔红霉素、多柔比星等；3)干扰DNA合成的药物，如嘧啶拮抗物-尿嘧啶衍生物，如5-氟尿嘧啶、呋氟尿嘧啶、双呋氟尿嘧啶，胞嘧啶衍生物阿糖胞苷、环胞苷、5-氮杂胞苷等、嘌呤拮抗物-如溶癌呤、硫鸟嘌呤等、叶酸拮抗物-如甲氨蝶呤等、4)影响蛋白质合成的药物，如秋水仙碱类、长春碱类、紫杉烷类-如紫杉醇、多西紫杉等。
优选的，所述化疗药物选自尿嘧啶衍生物类、环磷酰胺类、吉西他滨类、表柔比星类、抗肿瘤抗生素类、替尼泊苷、金属铂络合物、紫杉烷类中的一种或多种：优选的，包括5-氟尿嘧啶、环磷酰胺、异环磷酰胺、吉西他滨、表柔比星、抗肿瘤抗生素、替尼泊苷、金属铂络合物、紫杉醇中的至少一种。
所述化疗药物在所述局部药物组合物中的药理浓度为(w/v)为0.1-5％，例如环磷酰胺、卡莫司汀等烷化剂的药理浓度为(w/v)为0.5-5％、优选为0.75-1.5％；顺铂、卡铂等金属铂络合物的药理浓度(w/v)为0.1-0.25％；多柔比星类、拓扑替康、伊立替康等DNA拓扑异构酶抑制剂的浓度(w/v)为0.1-0.30％；放线菌素类、柔红霉素等抗肿瘤抗生素的浓度(w/v)为1-4％、优选为1-2％；5-氟尿嘧啶、呋氟尿嘧啶、双呋氟尿嘧啶等尿嘧啶衍生物以及阿糖胞苷、环胞苷、5-氮杂胞苷等胞嘧啶衍生物的浓度(w/v)浓度为0.25-2％、优选为0.5-1.5％。
所述免疫调节剂包括卡介苗及其组分、益生菌及其组分、免疫调节抗体中的至少一种；所述卡介苗及其组分包括卡介苗、灭活卡介苗、卡介苗多糖核酸等，所述益生菌及其组分包括灭活益生菌、益生菌胞壁聚多糖及其衍生物、益生菌核糖核酸等，所述免疫调节抗体包括血浆免疫球蛋白、多克隆免疫球蛋白、单克隆免疫球蛋白等，所述单克隆免疫球蛋白包括针对抑制性受体的抗体阻断剂，如针对CTLA-4分子和PD-1分子的阻断性抗；针对抑制性受体的配体的抗体阻断剂、针对免疫反应细胞表面刺激分子的激活性抗体，如抗OX40抗体、抗CD137抗体、抗4-1BB抗体；针对局部病变疾病微环境中免疫抑制性分子的中和抗体，如抗TGF-p1抗体。术语“卡介苗(或益生菌)组分”是指源于天然卡介苗(或益生菌)或其工程菌的制备物(例如胞壁聚多糖)或该制备物的工程类似物(例如类似于胞壁聚多糖的合成聚多糖或其它来源聚多糖)。
本发明还提供了所述局部药物组合物在制备治疗实体肿瘤的药物中的一种应用，其特征在于，所述药物组合物作为瘤内给药的局部药物组合物；在所述药物组合物中所述右旋糖酐作为可提供瘤内共用逆转作用的活性成分(逆转剂)，所述不协同药物作为提供瘤内共用作用被逆转的活性成分(被逆转剂)，，其中所述逆转是指所述右旋糖酐的这样一种共用药理，其使得本来与之并不发生协同作用的(不协同)药物(例如药物A或B)转变为与之局部协同的共用物成分(短期逆转)、或/和使得本身中长期药效衰竭的所述共用物(例如药物A与B的组合)转变为中长期药效衰褐得到有效抑制的组合物成分(中长期逆转)。
本发明还提供了可用作血容量扩张剂的右旋糖酐在制备用于治疗实体肿瘤的药物中的应用，其特征在于，所述右旋糖酐包含在用于瘤内给药的局部药物组合物中，在所述药物组合物中所述右旋糖酐作为可提供瘤内共用逆转作用的活性成分，其中所述逆转是指所述右旋糖酐的这样一种共用药理，其使得本来与之并不发生协同作用的(不协同)药物(例如药物A或B)转变为与之局部协同的共用物成分(短期逆转)、或/和使得本身中长期药效衰竭的所述共用物(例如药物A与B的组合)转变为中长期药效衰褐得到有效抑制的组合物成分(中长期逆转)。
优选的，所述共用成分分别选自与所述右旋糖酐共用未显示出局部协同作用的碱性有机化合物和碱性无机盐，且所述碱性有机化合物和碱性无机盐各自具有使得它们之间的局部作用互相增强的药理浓度或浓度比。
在本申请范围内，术语＂逆转＂是指药物的这样一种共用药理，其使得本来与之并不发生协同作用的(不协同)药物转变为与之局部协同的共用物成分(短期逆转)、或/和使得本身中长期药效衰竭的所述共用物转变为中长期药效衰褐得到有效抑制的组合物成分(中长期逆转)。该药物即为逆转剂，该不协同药物或/和它们组合而成的共用物即为被逆转剂。在本申请中，右旋糖酐为逆转剂，碱性有机化合物、碱性无机盐、或碱性有机化合物与碱性无机盐的局部协同组合为被逆转剂。
本发明还提供了一种治疗实体肿瘤的方法，其包括下述步骤：向有此需要的个体的瘤内给药治疗有效量的上述药物。
术语“局部给药”(又称介入给药)是指药物直接递送至靶区内(如瘤体内)的任何给药方式，区别于经血液输运至靶区，如口服、静脉注射、腹腔注射等常规给药方式。术语“瘤内给药”是一种靶区为瘤体组织的局部给药，包括经血管瘤内给药和经皮瘤内注射。通常认为瘤内给药化学药物及其组合物为一种局部治疗。术语“局部治疗”区别于“全身治疗”，后者是指针对患者全身(如瘤体、瘤体联通的区域、体内其它部分所含肿瘤细胞)的治疗，而前者是指针对患者局部病变所在的局部区域(如给药局部病变及与其联通的其它病变区域)的治疗。
术语“局部作用”并非常规作用(又称吸收作用)，后者是指药物常规用药(又称吸收给药，如口服、静脉注射、腹腔注射)后经血液吸收以血药形式递送至高度特异的药靶(如特定癌细胞的特定靶点)所产生的药理作用，而前者是指药物局部给药(直接递送)至病变组织后在其间隙渗透所及的局部范围内(如肿瘤细胞微环境)产生的药理作用。术语“局部化学作用”并非吸收化学作用(例如细胞毒作用)，前者和后者分别为由化学药物产生的局部作用和吸收作用。常规用药有效的化疗药物(细胞毒药物)的局部药效提高不超过细胞毒作用动力学最大预期(例如其吸收药效的150％以内)，说明其局部化学 作用仍为其细胞毒作用；而常规用药无效(细胞毒无效)的化学药物的局部药效提高则可以超过其吸收作用(例如细胞毒作用)动力学最大预期(例如吸收药效的200％以上)，说明其局部化学作用的主要药靶并非其作为肿瘤细胞毒物质时的特定癌细胞的特定靶点，而是特定靶点之外的其它组织结构(例如蛋白质沉淀)。这些化学药物的局部作用显然主要为区别于细胞毒的局部化学作用，即组织破坏。
术语“化学消融”习惯上是指经典化学消融剂(例如高浓度乙醇、高浓度盐酸或乙酸、高浓度氢氧化钠)所显示的局部化学作用，其药物效应为有效的组织破坏。术语“类化学消融”是指常规用药无效(细胞毒无效)的其它化学药物(又称常规无效药物，例如本申请中的可用作血容量扩张剂的右旋糖酐、碱性有机化合物、碱性无机盐)所显示的局部作用或局部化学作用，其药物效应亦为直接破坏组织，短期内(如给药后5日以内)便可显示瘤体组织坏死的药效，与化学消融高度类似。通常认为类化学消融如同化学消融其短期药效均在中长期出现药效衰竭，结果为复发率高。
术语“局部(局部作用)药物”是指化学物质及其局部作用实现的条件。术语“局部作用(不可观)可观的药物”是指(未能)能达到业内普遍接受的药物可观效应标准的化学药物。在动物实验中，＂局部作用可观(不可观)＂则为与阴性对照物的差异足够(不足够)明显，例如抑瘤率为≧15％(＜15％)。局部作用可观的药物例如瘤内给药浓度为≧30％的葡萄糖，局部作用不可观的药物例如瘤内给药浓度为＜30％的右旋糖酐。术语“局部作用有效(无效)的药物”是指能(未能)达到业内普遍接受的药物有效效应标准的化学药物。在动物实验中，＂局部作用有效(无效)＂则为抑瘤率≧40％(＜40％)。局部作用有效的药物例如瘤内给药浓度为≧95％的乙醇，而瘤内给药浓度为＜30％的右旋糖酐提供的局部作用不可观、更说不上有效。术语“局部作用最大(最小)化的药物”是指其局部作用药效(例如抑瘤率)不再随浓度增大(減小)而明显增大(減小)的药物。在动物实验中，＂最小局部作用＂显然为阴性对照物的局部作用，＂最小化局部作用＂则为接近阴性对照物的局部作用。当不可观局部作用不再随浓度減小而明显減小时(例如抑瘤率为＜10％)，即为最小化局部作用。局部作用最大化的药物，例如瘤内给药浓度为≧35％的右旋糖酐，局部作用最小化的药物例如瘤内给药浓度为≦25％的右旋糖酐。
术语“药物组合物”是指包含至少两种活性成分的复方制剂。术语“局部药物组合物”是指包含至少两种活性成分的固定浓度或浓度比的局部给药剂型的复方制剂，其中所述复方制剂是指所述活性成分可充分混合进入靶区的共用形式；所述浓度或浓度比为所述活性成分进入靶区的浓度或浓度比；所述局部给药剂型区别于吸收给药剂型，例如不包含后者所需的以减小给药处局部作用为目的非活性成分(例如等渗调节剂氯化钠等)。
优选的，所述治疗的治疗作用主要为药物各成分瘤内共用作用，其核心药理为右旋糖酐的逆转作用，该逆转作用使得所述瘤内共用作用的药物效应为短期局部治疗、或/和中长期治疗。其中所述短期局部治疗的药理包括一处或多处肿瘤组织的局部化学协同作用的类化学消融和任选存在的其它化疗，所述类化学消融独立于肿瘤细胞毒作用；所述中长期治疗的药理包括肿瘤组织内的局部作用(或局部协同作用)的次生免疫作用和其它次生作用。在一个实施方案中，所述治疗的适用患者选自包括以下组之一种或多种：免疫抑制患者、可局部病变内给药的患者、局部病变组织可类化学消融的患者、局部病变可产生次生免疫作用的患者。在一个实施方案中，所述组合物为类化学消融-原位免疫治疗药物。
术语“免疫治疗”区别于术语“免疫增强”，前者是指其单独使用便可达到治疗作用的免疫作用(如治疗疫苗、特异性抗体等的免疫作用)，而后者是指单独使用不能达到治疗作用、但仍有辅助作用的免疫作用(如免疫增强剂具有提高机体免疫功能的作用)。
优选的，所述局部药物组合物为瘤内给药注射剂。
优选的，所述注射剂为粉针剂。
所述“注射剂”指含活性成分及其溶媒并可供体内给药的无菌制剂；＂粉针剂＂是指包含无菌干粉和溶媒的注射剂，无菌干粉中包含部分或全部活性成分，溶媒中包含全部液体载体。注射剂浓度通常为制剂浓度，而瘤内给药注射剂的特征之一为其制剂浓度必须大于或等于其药理浓度(通常远高于吸收药理浓度)。瘤内给药注射剂中所述活性成分的药理浓度通常是局部给药器械(注射器、穿刺器、注入导管等)终点(如针孔、导管出口等)的液体药物中的活性成分浓度。对注射用粉针剂而言，所述活性成分的浓度即为规定量的无菌干粉和规定量的溶媒的混合物(如复溶液或所述药物学可接受的液体载体)中的活性成分浓度。
所述“实体肿瘤”指具有瘤体组织的肿瘤，其可以是由于任何病理(恶性和非恶性)和处于任何阶段的肿瘤，按照肿瘤细胞类型，包括上皮细胞肿瘤、肉瘤、淋巴瘤、生殖细胞肿瘤、胚细胞瘤；按照肿瘤细胞集中区所在的器官或组织，包括脑肿瘤、皮肤肿瘤、骨肿瘤、肌肉肿瘤、乳腺肿瘤、肾肿瘤、肝肿瘤、肺肿瘤、胆囊肿瘤、胰腺肿瘤、食道肿瘤、膀肌肿瘤、大肠肿瘤、小肠肿瘤、脾肿瘤、胃肿瘤、前列腺肿瘤、卵巢肿瘤或子宫肿瘤。所述肿瘤包括恶性肿瘤和非恶性肿瘤，所述恶性肿瘤包括乳腺癌、胰腺癌、甲状腺癌、鼻咽癌、前列腺癌、肝癌、肺癌、肠癌、口腔癌、食道癌、胃癌、喉癌、睾丸癌、阴道癌、子宫癌、卵巢癌、恶性淋巴瘤、恶性脑瘤等，所述非恶性肿瘤包括乳腺瘤、胰腺瘤、甲状腺瘤、前列腺瘤、肝瘤、肺瘤、肠瘤、口腔瘤、食道瘤、胃瘤、鼻咽瘤、喉瘤、睾丸瘤、阴道瘤、子宫瘤、输卵管瘤、卵巢瘤、淋巴瘤、脑瘤等。
优选的，所述实体肿瘤选自适于瘤内给药的恶性肿瘤和非恶性肿瘤。尽管瘤内给药的常规无效药物及其组合在理论上可用于任何实体肿瘤，因为它们都含有其局部化学作用的药靶－瘤体组织，但目前在临床上，其在抗肿瘤药物中的占比还不到1％，主要限定便是该＂适于瘤内给药＂涉及的众多主客观因素。
优选的，所述实体肿瘤选自因容易复发而不适用化学消融剂或常规局部协同组合物的实体肿瘤，其中所述常规局部协同组合物是指由局部作用药物与其协同物或协同物的组合形成的局部药物组合物，例如所述共用物、或右旋糖酐类似物例如葡萄糖与这些共用物的组合物。以上每种适于瘤内给药的肿瘤中，均包含至少极个别经化学消融或类化学消融后容易复发的瘤体。
优选的，所述实体肿瘤具有至少一个瘤体尺寸为大于6cm、或大于10cm。
本发明还提供了一种药物试剂盒，其特征在于，包括包含以下独立包装制剂的容器：含有上述的局部药物组合物；含有溶媒的制剂。所述容器可以包括安瓿、小玻璃瓶等。优选的，所述药物试剂盒还包括说明书或标签。在施用时，所述局部药物组合物的施用量与所述局部病变内靶区体积之比＞0.1、0.15-1.5、优选为0.23-1.5或0.5-1.5。亦或者根据具体情况，所述局部药物组合物的施用量为≥1ml，或局部病变内的施用量为10-150ml或/和局部病变外的施用量为1.5-50ml。
相对于现有技术，本发明所述的局部药物组合物具有以下优势：
(1)与细胞毒药物相比，所述局部药物组合物的中长期疗效佳且具有几乎无毒的全身用药安全性；与分子靶向药物相比，不需要苛刻的适应症筛选，对快速生长瘤体、大瘤体和乏血供瘤体显示出巨大的潜力；与化学消融剂或组合类化学消融剂(包括局部作用药物与其协同物或协同物组合的局部药物组合物)相比，局部刺激性更小，特异性强且具有更高的中长期疗效；(2)本申请所述的局部药物组合物不存在耐药性问题，制备方法简单、成本低廉，惠及群体广泛。
附图说明
图1为本发明实施例6部分样品的组织病理学HE染色检查结果；
图2为本发明实施例7部分样品的TUNEL检查结果；
图3为本发明实施例10在实验前后的肿瘤外观图；
具体实施方式
需要说明的是，在不冲突的情况下，本发明中的实施例及实施例中的特征可以相互组合。下面将结合实施例来详细说明本发明。本申请所使用的高分子血容量扩张剂等制剂均可从商业途径得到，例如表1。
表1实施例中使用的部分右旋糖酐
品名 平均分子量 CAS号 供应商
右旋糖酐10 9000～11000 9004-54-0 山东金洋药业有限公司
右旋糖酐20 16000～24000 9004-54-0 山东金洋药业有限公司
右旋糖酐40 32000～42000 9004-54-0 山东金洋药业有限公司
右旋糖酐70 64000～76000 9004-54-0 山东金洋药业有限公司
以上包括平均分子量在9000－76000之间的任何右旋糖酐。
以下实施例中所用的实验动物均为从专业实验动物公司购入的SPF(Specific Pathogen Free，无特定病原体)级动物，除非另有说明，其中小鼠(BALB/c)和裸小鼠(BALB/c Nude)均为6-8周龄健康雌性、体重17.5-20.5g。裸鼠为先天性胸腺缺陷的突变小鼠，缺乏免疫反应，常用作肿瘤的纯化疗例如局部化学作用(组织破坏或化学消融)的研究。
皮下移植肿瘤细胞生成瘤体的动物建模为现有公知技术，当瘤体长至所需体积(如小鼠荷瘤为50-500mm3)为建模成功，建模后将动物分成若干组，每组6只，定期观察测定一般状态、体重、摄食量、动物移植物抗宿主病、瘤体体积、瘤重、生存时间等。
瘤体体积(V)、瘤体增殖率(R)、抑瘤率(r)分别按照下述公式计算：
瘤体体积V＝0.5×a×b 2，其中a、b分别代表瘤体的长、宽；
瘤体增殖率R＝TV/CV×100，其中TV和CV分别为研究组和阴性对照组的瘤体体积；
抑瘤率(r)＝100％-R，其中R为瘤体增殖率。
在下述实施例中，药物i的药效记为Ei，可以用ri表示。药物的药理可通过药效，尤其是同一药物在不同方案中的药效来研究。如当药物i在方案X、Y之间的药效差异不大时(如Ei X/Ei Y＜200％)，其说明药物的药理是相同的；当药物i在方案X、Y之间的药效差异大时(如Ei X/Ei Y＞200％)，说明药物i在方案X中的药理大大超过了其在方案Y中药理的动力学预期范围，很可能涉及与方案Y不同的新 药理。若两个药物显示出显然不同的Ei X/Ei Y关系，则很可能涉及不同的药理；若两个药物显示出相似的Ei X/Ei Y关系，则它们很可能涉及相同的药理，至少涉及相似的药理，如类化学消融药理和化学消融药理。
在以下实施例中，实验结果(例如瘤重、瘤体积)采用均数±标准差(x±s)表示，两个实验动物组之间的差别采用统计学软件SPSS13.0或SPSS19.0进行显著性检验，检验选用统计量t来进行，检验水准α＝0.05，P＜0.05表示差异有统计学意义，P＞0.05则无统计学意义。
化疗的阳性对照包括经典的细胞毒药物(如0.5-1％5-氟尿嘧啶，其在下述实施例条件下的短期抑瘤率为≥30％)和经典化学消融剂(如75-99％乙醇，其在下述实施例条件下的短期抑瘤率为≥15％)。
术语“共用药理”是指药物组合物中不同组分(例如增效剂与其共用物)之间的相互作用机理。药物共用药理具有高度不确定性。相同共用组分在不同条件下可以形成不同组合，例如研究组合 1和对照组合 2。业内通常将研究组合 1的药效例如抑瘤率r 1定义为实际共用药效；将对照组合 2的药效例如抑瘤率r 2定义为预期共用药效；并将后者的预期共用药理定义为共用药物互不干扰的加和作用。从而，依据实际共用药效/预期共用药效比(本申请中简称实际/预期比)q(q＝r 1/r 2)可判断共用药理：当q＝1，显示实际共用药效(及实际相互作用)合符预期药效(及预期相互作用)，判断实际共用药理也为加和作用；当q＜1，显示实际共用药效(及实际相互作用)不及预期药效(及预期相互作用)，判断共用药理为共用药物作用彼此减弱的拮抗作用；当q＞1，显示实际共用药效(及实际相互作用)超预期药效(及预期相互作用)，判断共用药理为共用药物作用互相增强的协同作用。所述“协同作用”于是是指药物共用产生的较之预期相互作用而言互相增强的实际相互作用，具体为q＝r 1/r 2＞1。
术语“短期协同作用”区别于术语“中长期协同作用”，前者指根据短期(如未次给药后5日以内)药效(例如r 3d1、r 3d2)显示的协同作用，后者指根据中长期(如未次给药的第14天后)药效(r 21d1、r 21d2)的协同作用。术语“局部协同作用”区别于术语“非局部协同作用”，前者指根据局部作用药效(如前所述通常为短期药效例如r 3d)的协同作用，后者指根据非局部作用药效(中长期药效例如r 21d)的协同作用。通常认为，化学药物组合在裸鼠模型上观察到的局部协同作用主要为化合物局部化学作用之间的相互增强，其显然为一种短期协同作用。
由于组间差异判断涉及到实验误差问题，尤其是动物实验。通常批间可以接受的误差水平为±20％，批内可以接受的误差水平为≦15％，而在误差范围内的组间区别显然为不明显(或不可观)。当研究组合 1与阴性对照的区别明显例如r 1＞15％(不明显例如r 1≦15％)，则能(不能)根据实际/预期比q＝r 1/r 2判断，不能据此判断时说明并无明显共用作用、包括协同作用。
对照组合 2的加和药效r 2的假设很多，如Burgi法(Burgi Y.Pharmacology；Drug actions and reactions.Cancer res.1978，38(2)，284-285)，对Burgi法改进的金正均法(金正均，等概率和曲线和“Q50”，上海第二医学院学报；1981，1，75-86)。按照金正均法，假设r I和r II分别为药物I和II的单药实际药效，则r 2＝r I+r II-r I×r II为两个药物通过互不干扰的加和作用产生的理论预期药效。
更具体而言，上述r 1、r 2可以都来自中长期药效，都来自短期药效，或分别来自中长期药效和预期(对照)短期药效。瘤内给药在局部病变上产生的短期药效(例如瘤体组织坏死而使其生长受到抑制) 实际上是局部相互作用的药效，瘤内给药在局部病变上产生的中长期药效(例如瘤体坏死减小)则是涉及到局部作用与次生免疫反应之间复杂的相互作用的药效。
通常认为，药物I和II共用的毒理作用如同其药理作用一样，可以用研究组合和对照组合的毒理效应(毒效)例如全身性安全性和局部刺激反应来比较研究，其中对照组合的共用安全性预期为毒理作用的理论单纯相加预期，即共用毒效不小于两药物中的最大毒效。药物I/II组合物的共用毒理作用采用如下方法判断：若组合物实际毒效与理论单纯相加预期的毒效一致，则该组合物的共用为毒理加和作用；若组合物的实际毒效不及理论单纯相加预期的毒效(例如共用毒效小于两药物中的最大毒效)，则该组合物的共用为明显的毒理协同作用；若组合物的实际共用毒效不小于两药物中的最大毒效大于理论单纯相加预期的共用毒效不小于两药物中的最大毒效，则该组合物的共用为明显的毒理拮抗作用。
实施例1
用注射用水按组合物组分、药理浓度和浓度比例制备各组合物制剂，相关溶液的配制方法为现有技术，在此不进行赘述。
制备的液体制剂经冷冻干燥后可获得注射用冻干粉。作为本发明的一个示例，所述冷冻干燥采用下述工艺：在预冻温度-45℃保持4小时，升温速率为0.1℃/分钟且升至-15℃时至少保持10小时进行升华；30℃保持6h进行解吸附干燥，分装(如7.5ml/瓶)、封盖。使用时将无菌溶媒抽入瓶中，混匀即可用作注射药物，如1.5％酿酒酵母菌破碎组分/20％氨基酸。
为进行本发明的组合物的新药申报工作中的制备方案优选，进一步进行了以下制备实验。
在一个实验中，比较了包含相同浓度不同扩张剂的不同组合物成药所必须的可除菌过滤性。4个组合物各包含20％扩张剂、20％精氨酸和5.8％碳酸钠，其中所述扩张剂分别为：右旋糖酐40、右旋糖酐70、羧乙基淀粉200、聚乙烯吡咯烷酮k70。按常规方法配制的组合物水溶液分别在室温按常规除菌过滤方法以相同滤器(孔径0.2um)进行过滤，右旋糖酐组合物均可通过滤器进行除菌过滤，而后2个组合物通过滤器时因压力过高未完成除菌过滤。这些结果说明，右旋糖酐用于组合物的制备具有不同而更容易进行的工艺条件。
在一个实验中，比较了包含相同不同浓度(X％)右旋糖酐的不同组合物成药所必须的可除菌过滤性。4个组合物各包含X％右旋糖酐70、20％精氨酸和5.8％碳酸钠，其中所述X％分别为：2％、4.5％、10％、25％、30％。按常规方法配制的组合物水溶液分别在室温按常规除菌过滤方法以相同滤器(孔径0.2um)进行过滤，右旋糖酐浓度为2-25％的前4个组合物均可通过滤器进行除菌过滤，而最后1个组合物通过滤器时因压力过高未完成除菌过滤，其只能稀释后过滤然后浓缩，又增大了染菌风险。这些结果说明，2-25％右旋糖酐用于组合物的制备具有不同而更容易进行的工艺条件。
在一个实验中，比较了右旋糖酐与不同共用物的组合物成药所优选的生产过程洁净性。2个组合物的处方分别为：20％精氨酸/5.8％碳酸钠/4.5％右旋糖酐70、1％亚甲蓝/5.8％碳酸钠/4.5％右旋糖酐70。在称样至药物冻干的整个生产过程中，第1个组合物均无任何明显污染；而第2个组合物均着色于其所接融的任何器具且难以清洗，尤其是造成冻干机污染，会大大提高制备成本。这些结果说明，较之亚甲蓝，碱性有机化合物和碱性无机盐作为右旋糖酐不协同药物的使用使得组合物的制备具有不同而更容易进行的工艺条件。
(2)根据以上实验的结果，委托杭州澳亚生物技术股份有限公司进行了将本申请的组合物用以申报新药的GMP生产。生产批量为每批5000g干重的活性成分，包括精氨酸(本申请中均为L-精氨酸)、碳酸钠和右旋糖酐70，各成分量比为3.5g精氨酸/1g碳酸钠/0.6g右旋糖酐70，药物剂型为冻干剂型，药物使用时以适量注射用水配制为2-4.3％右旋糖酐70、11.7-25％精氨酸、3.3-7.1％碳酸钠的水溶液供瘤内给药。
实施例2扩张剂短期逆转作用的发现
荷瘤裸小鼠被广泛用于实体肿瘤患者化疗而非免疫治疗的药物研究，以裸小鼠为实验对象，将2.5×10 6个小鼠肝癌Hepa1-6细胞/只注射入右侧腋部皮下进行移植瘤建模，建模成功裸小鼠的瘤体平均体积为75.7mm 3，将模型动物随机分为12组，按表2各组成分进行瘤内注射，每组均用药2次，用药间隔为2日，每次注射量50μl/只。第二次用药后第3日，测局部病变体积(V)，并根据阴性对照组计算抑瘤率(r)。
表2不同组别的抑瘤率数据
Figure PCTCN2022122537-appb-000001
由上表可知，组3中扩张剂A 2抑瘤率(12.3％)还不到可观局部作用对照物A 1的1/2(25.7％)，其并未提供可观局部作用。组别10、11的药效低于组别7、8，合乎常规局部增效的预期：与相同共用 物(B或C)共用，局部作用较弱的药物(A 2)的共用药效低于局部作用较强的药物(A 1)。然而，当A 2和A 1分别与相同共用物(B和C)形成组合物后，组别12的药效高于组别9，这有悖于前述常规增效预期。
由上表可知，组别7、8、10、11的共用药效的实际/预期比q均为＜1.00，说明单药B或C均是A 1和A 2的不协同药物。尽管组6(B/C)的共用药效的实际/预期比q＞1.00，组别9(A 1/B/C)的共用药效的实际/预期比q仍然为＜1.00，这合乎常规增效预期：增效剂只能协同其协同药物或协同药物的组合。然而，组别12(A 2/B/C)的共用药效的实际/预期比q却为＞1.00，这有悖于前述常规增效预期，即非常规地将其不协同药物(B、C)逆转为协同共用物(B/C)的活性成分。A 2具有、而A 1不具有的该逆转作用应为A 2/B/C药效大于A 1/B/C的药理基础，这也说明该逆转作用具有药物选择性。
A 2和A 1和相同组合B/C的不同组合物不仅在上述共用药理上完全相反(逆转作用vs无逆转作用)，而且在共用毒理上也大不相同。组别6中给药B/C时观察到的局部刺激作用现象为中等(33％动物观察到较高强度挣扎)，组别9中给药A 1/B/C过程中观察到局部刺激现象明显加强(100％动物均出现高强度挣扎)，而组别12在给药A 2/B/C时观察到局部刺激现象明显拮抗(无动物出现较高强度挣扎)。此外，与组别9(A 1/B/C)相比，组别12(A 2/B/C)给药时出现明显局部泄漏的动物数目要少50％以上。总之，组别12(A 2/B/C)显示出明显改善B/C局部安全性的安全性协同作用。
上述意外发现为类似扩张剂的新活性研究提供了新的思路。
实施例3：逆转作用所需共用成分特征研究
建模动物为裸小鼠，建模细胞(2.5×10 6个细胞/只)为人胰腺癌细胞(PANC-1)，建模方法同实施例2，在此不进行赘述，建模成功的裸小鼠的瘤体平均体积为81.3mm 3，将模型动物随机分为38组，按表3各组成分进行瘤内注射，每组均用药2次，用药间隔为3日，每次注射量50μl/只。第二次用药后第3日，测局部病变体积(V)，并根据阴性对照组计算抑瘤率(r)。
表3不同组合的抑瘤率数据
Figure PCTCN2022122537-appb-000002
Figure PCTCN2022122537-appb-000003
在本发明申请中，以葡萄糖作为糖营养素的代表，30％-50％糖营养素可提供可观局部作用。在上表中，组别1-3的r均为＜8％，还不到有效r标准(r≥40％)的20％，与阴性对照物(局部作用最小)非常接近，A1、A2和A3均为局部作用最小化的药物。在其它类似于组别2的实验中，(25％)右旋糖酐10、(20％)右旋糖酐20、(10％)右旋糖酐70等其它右旋糖酐代替本实验中的(15％)右旋糖酐40，也分别获得与本实验一致的结果，即瘤内给药浓度≦25％的右旋糖酐只能提供最小化的局部作用，实验结果不再冗述。
在上表中，组别10-18的共用药效的实际/预期比q均为＜1.00，均未显示局部协同作用，说明其中的共用药物(Bi或Ci)皆为上述局部作用最小化药物(A1、A2、A3)的不协同药物。在其它类似于组别14-15的实验中，右旋糖酐10、右旋糖酐20、右旋糖酐70等其它右旋糖酐代替本实验中的右旋糖酐40，氨丁三醇、三乙醇胺、三羟甲基氨基甲烷、葡甲胺、赖氨酸等碱性有机化合物代替本实验中的精氨酸，碳酸钠、乳酸钠、乙酸钠等碱性无机盐代替本实验中的碳酸氢钠，也分别获得与本实验相类似 的结果，可合理推及碱性有机化合物、碱性无机盐为局部作用最小化的右旋糖酐的局部不协同药物，实验结果不再冗述。
在上表中，组别25-28的共用药效的实际/预期比q均小于1.00，糖营养素(A1)并未显示将其不协同药物(B、C)及其组合(B/C)逆转为其协同共用物的活性。本实施例中的血容量扩张剂(右旋糖酐40和羟乙基淀粉)的结构与葡萄糖高度相似，在相同条件下很可能具有相同的共用药理。组别29的q＜1.00，似乎对此加以了证实。然而，组别30-37的共用药效的实际/预期比q均为＞1.00，血容量扩张剂于是显示共用逆转药理，即将其不协同药物的组合(B/C)逆转为其协同共用物，从而其不协同药物(B、C)逆转为其协同共用物的活性成分。此外，组别29的组合物中的B/C为组别19中q＜1.00的组合，而组别30-37的组合物中的B/C分别为组别20-28中q＞1.00的组合，说明并非任何组合中的不协同药物、只有当它们之间的量比使得它们之间的局部作用互相增强的组合中的不协同药物才能被扩张剂逆转，或者说明该逆转作用具有不协同药物组合选择性。在其它类似于组别30的实验中，右旋糖酐10、右旋糖酐20、右旋糖酐70代替本实验中的右旋糖酐40，氨丁三醇、三乙醇胺、三羟甲基氨基甲烷、葡甲胺、赖氨酸等碱性有机化合物代替本实验中的精氨酸，碳酸钠、乳酸钠、乙酸钠等碱性无机盐代替本实验中的碳酸氢钠，也分别获得与本实验一致的结果，即右旋糖酐可逆转选自碱性有机化合物和碱性无机盐的不协同药物的组合(它们之间协同)，实验结果不再冗述。
此外，组别20-24(不协同药物的组合)给药时观察到33％以上动物出现较高强度挣扎，说明局部刺激作用现象较强；组别25-28(糖营养素/不协同药物组合)给药时实验动物刺激反应与组别20-24相当(39％以上动物出现较高强度挣扎)。与组别25-28相比，组别30-37给药时实验动物刺激反应明显更弱(35％以上动物出现较高强度挣扎)，且给药时出现明显局部泄漏的动物数目要少50％以上。
综上，瘤内给药浓度为≤25％的右旋糖酐可作为提供共用作用逆转的活性成分(逆转剂)，与所述右旋糖酐不产生局部协同作用的碱性有机化合物或碱性无机盐可作为其不协同药物(被逆转剂)，而碱性有机化合物和碱性无机盐的协同组合可作为右旋糖酐的协同共用物，右旋糖酐从而將其不协同药物转变为协同共用物组分，提供药效协同作用；和/或使该共用物的局部刺激性下降，提供安全性协同作用。
实施例4逆转作用所需共用方式特征研究
建模动物为裸小鼠，建模细胞(2×10 6个细胞/只)为小鼠恶性肉瘤细胞(S180细胞)，建模方法同实施例2，在此不进行赘述。建模成功裸小鼠的瘤体平均体积为74.9mm 3，将模型动物随机分为22组，除16-18组之外每组分别通过A、常规给药(尾静脉注射)；B、瘤内注射两种方式给药，每个单组均用药2次，用药间隔为3日，每次注射量50μl/只。第二次用药后第3日，测局部病变体积(V)，并根据系列A、B的阴性对照组计算抑瘤率(r A、r B)，结果见表5。
各组别所用的药物组合物均按常规的水溶液制备方法或实施例1中的制备方法制备。组别16-17在系列A中给药方式为：血容量扩张剂采用常规给药+共用成分组合物采用瘤内注射，系列B中给药方式为血容量扩张剂采用瘤内注射+共用成分组合物采用常规给药。组别18在系列A中给药方式为瘤内序贯注射血容量扩张剂和共用成分组合物，系列B中的给药方式为瘤内序贯注射共用成分组合物和血容量扩张剂。
表5不同药物组合、给药方式的抑瘤率数据
Figure PCTCN2022122537-appb-000004
众所周知，常规给药(如腹腔注射、静脉注射等)产生的药效反映了该药物的全身性药理作用(如营养个别增强或总体不平衡、肿瘤细胞毒、免疫增强、血液内缓释等)，而瘤内给药产生的药效则反映了该药物直接进入局部病变部位引发的局部化学损伤等局部作用。在实体肿瘤治疗中，化学药物的药效 主要是短期药效。就化疗药效而言，如果药物经常规给药的药效可观(r A大于15％)且瘤内给药的药效还可基于动力学条件提高(如100％＜r B/r A＜200％)，则说明其药理为细胞毒作用(如组别02)；如果药物经常规给药的药效不可观(r A小于15％)，瘤内给药的药效可观(r B＞15％)且r B/r A＞200％，则说明其药理为独立于细胞毒作用的局部组织破坏作用，在本发明申请中亦简称为局部作用(如组别03)。
由表5可知，组别5-7的r B/r A＞200％，瘤内给药显示出不同于常规给药全身性药理的局部化学作用。这些组合物的局部共用药效的实际/预期比q均为＜1.00，说明其中的单药(精氨酸、精氨酸盐酸盐、碳酸氢钠)均为扩张剂的不协同药物。在其它类似于组别5-7的瘤内给药的实验中，右旋糖酐10、右旋糖酐20、右旋糖酐70等其它右旋糖酐代替本实验中的右旋糖酐40，氨丁三醇、三乙醇胺、三羟甲基氨基甲烷、葡甲胺、赖氨酸等碱性有机化合物代替本实验中的精氨酸，碳酸钠、乳酸钠、乙酸钠等碱性无机盐代替本实验中的碳酸氢钠，以及赖氨酸盐酸盐等碱性有机化合物酸化盐代替本实验中的精氨酸盐酸盐，也分别获得与本实验一致的结果，即碱性有机化合物、碱性无机盐、碱性有机化合物酸化盐均为右旋糖酐的局部不协同药物，实验结果不再冗述。
由表5可知，在组别12-13中，不协同药物的组合的成分近似，却只有组别12中的组合物(包含组别8中的不协同药物的组合)的局部共用药效的实际/预期比q大于1.00，还是说明扩张剂只能对某些特定种类、而非所有不协同药物提供共用逆转作用，其逆转作用具有选择性。在其它类似于组别12-13的瘤内给药的实验中，右旋糖酐10、右旋糖酐20、右旋糖酐70等其它右旋糖酐代替本实验中的右旋糖酐40，氨丁三醇、三乙醇胺、三羟甲基氨基甲烷、葡甲胺、赖氨酸等碱性有机化合物代替本实验中的精氨酸，碳酸钠、乳酸钠、乙酸钠等碱性无机盐代替本实验中的碳酸氢钠，以及赖氨酸盐酸盐等碱性有机化合物酸化盐代替本实验中的精氨酸盐酸盐.也分别获得与本实验一致的结果，实验结果不再冗述。
由表5可知，组别16、18与组别12、组别17与组别15中的组合物具有相同的组分和组分量比，唯一区别在于剂型不同(活性成分可分别给药的非局部给药剂型和活性成分须混合给药的局部给药剂型)；出乎意料的是，不同的剂型显示出完全不同的共用药效，说明右旋糖酐只能在瘤内联合给药的条件下才能显示其共用逆转作用。
组织病理学结果显示，组别12、15中瘤内给药与常规给药相比，实验动物的肿瘤样本存在较多局灶性坏死区域，坏死区域大小不等，多呈片状或带状，分布于肿瘤组织的边缘和中央区。坏死区内坏死肿瘤细胞数量较多，大量肿瘤细胞胞核浓缩深染，大量细胞核碎片，而这进一步证实组别12、15的组合物经瘤内注射后显示出的特别药理作用。
实施例5：逆转作用所需局部作用特征研究
建模动物为裸小鼠，建模细胞(1×10 6个细胞/只)为小鼠乳腺癌细胞(4T1细胞)，建模方法同实施例2，在此不进行赘述。建模成功裸小鼠的瘤体平均体积为84.1mm 3，将模型动物随机分为26组。按表6各组成分进行瘤内注射。各组别所用的药物组合物均按常规的水溶液制备方法或实施例1中的制备方法制备。每组均用药2次，用药间隔为3日，每次注射量50μl/只。第二次用药后第3日，测局部病变体积(V)，并根据阴性对照组计算抑瘤率(r)。
表6不同组合的抑瘤率数据
Figure PCTCN2022122537-appb-000005
在上表中，组别1-3中单糖的局部作用强度(药效r)随给药浓度增加而明显增加；而组别4-6中的右旋糖酐的局部作用强度(药效r)并不随给药浓度增加而明显增加。在其它类似于组别4-6的实验中，右旋糖酐10(给药浓度2-25％)、右旋糖酐70(给药浓度2-15％)等其它右旋糖酐代替本实验中的右旋糖酐40，也分别获得与本实验一致的结果，实验结果不再冗述。这些结果进一步证实了实施例3的结果：瘤内给药浓度为2-25％右旋糖酐为局部作用最小化的药物。
在上表中，组别11-13的共用药效随糖营养素瘤内给药浓度增加而增加，而组别14-16的共用药效并未随扩张剂给药浓度增加而明显增加。其中，组别11的实际/预期比q＞1.00，而组别12-16的实际/预期比q＜1.00，合乎常规协同预期和临床现实：作用较强的药物方能用于产生协同作用。然而，组别20－22的共用药效的实际/预期比q均小于1.00，而组别23、24的共用药效的实际/预期比q大于1.00。这些比较研究的结果进一步说明：局部作用较小(瘤内给药浓度为2-25％右旋糖酐)、而非较大 (30％右旋糖酐)的右旋糖酐更适合作为共用逆转剂。
此外，组别25(扩张剂/3个不协同药物的组合)的共用药效的实际/预期比q大于1.00，也显示出局部协同作用。在该3个不协同药物的组合中，至少2个不协同药物的共用作用为局部协同作用。而组别14－16的结果进一步证实了实施例2-4的结果：碱性有机化合物和碱性无机盐分别是局部作用最小化的扩张剂的不协同药物。
实施例6血容量扩张剂中长期逆转作用的发现
建模动物为小鼠，建模细胞(0.5×10 5个细胞/只)为小鼠乳腺癌细胞(4T1细胞)，建模方法同实施例2，在此不进行赘述。建模成功小鼠的瘤体平均体积为107.3mm 3，将模型动物随机分为23组，每组经瘤内注射给药，每个单组均用药2次，用药间隔为3日，每次注射量100μl/只。各组合物均按常规的水溶液制备方法或实施例1中的制备方法制备，其中组别15-17中所用组合物分别形成供共用的组合物1-4。第二次用药后在第3日、第21日测局部病变体积(V)，并根据阴性对照组计算药效-抑瘤率(r 3d、r 21d)，结果见表7。根据药效r 3d和r 21d可分别计算短期药效的实际/预期比q 3d和中长期药效的实际/预期比q 21d，
表7不同组别用药后的短效、长效数据
Figure PCTCN2022122537-appb-000006
Figure PCTCN2022122537-appb-000007
通常认为，各化学药物(例如右旋糖酐、碱性有机化合物和碱性无机盐)的局部作用主要是对其弥散所及瘤体组织的化学破坏作用，其药物效应在短期内(例如3-10日内)就应当完全显示出来(体现为抑瘤率最高)，而后随时间明显衰减(例如21日抑瘤率明显下降)。由表7可知，组别1-8中各药物的结果均合乎该常规预期。其中右旋糖酐(组别2)的中长期药效和它的短期药效一样不可观，其中长期局部作用和它的短效局部作用一样均为最小化；精氨酸、碳酸氢钠(组别4、7)与亚甲蓝(组别5)相比，它们的中长期药效的下降更明显，均为衰竭至几近于无(r 3d＜5％)。在其它类似实验中，瘤内给药浓度为2-25％的右旋糖酐10、右旋糖酐20、右旋糖酐40、右旋糖酐70均显示出与上表中组别1一致的结果，以下瘤内给药浓度(2-25％)的碱性有机化合物均显示出与上表中组别4一致的结果：2-10％氨丁三醇、2-15％三乙醇胺、2-15％三羟甲基氨基甲烷、10-25％葡甲胺、10-20％精氨酸、15-25％赖氨酸，以下瘤内给药浓度(2-15％)的碱性无机盐均显示出与上表中组别7一致的结果：2-15％碳酸钠、5-15％乳酸钠、5-15％乙酸钠、5-15％碳酸氢钠，即所述实验浓度的右旋糖酐、碱性有机化合物、碱性无机盐至多能显示一定的短期药效，它们的中长期药效均下降至不可观，显示它们均为中长期局部作用最小化的药物，实验结果不再冗述。
由表7可知，组别9-13的短期实际/预期比q 3d均为＜1.00，合乎常规局部协同预期：中长期局部作用均为最小化的药物之间不能观察到局部作用的互相增强，结果为这些常规局部组合物的中长期共用药效均衰竭至不可观。在其它类似实验中，上述右旋糖酐分别与所述碱性有机化合物或碱性无机盐的组合物显示与上表中组别9或12一致的结果，实验结果不再冗述。这些结果说明上述实施例2-5中扩张剂的短期不协同药物显然也是其中长期不协同药物。
由表7可知，组别14-17的短期实际/预期比q 3d均为＞1.00、且短期药效为有效，而它们的中长期药效衰竭(r 21d/r 3d＜50％)。这些结果合乎上述常规预期。
由表7可知，组别22的中长期共用药效衰竭至不可观，说明糖营养素不能将其不协同药物(精氨酸、亚甲蓝、5-氟尿嘧啶)的组合(10％精氨酸/1％亚甲蓝/1％5-氟尿嘧啶)逆转为其中长期协同共用物，这合乎以上常规局部协同预期。完全超过该预期，组别18－21的中长期共用药效实际/预期比q 21d＞1，中长期药效衰竭得到明显抑制(即r 21d/r 3d＞50％)，结果为组合物的中长期药效明显高于共用物 (组别15－17)。实质上，右旋糖酐对其共用物的中长期增效远高于短期增效(即r 组合物21d/ 共用物r 21d＞150％×r 组合物3d/ 共用物r 3d)，显示其中长期逆转作用明显高于短期逆转作用。在其它类似实验(包括例如以下实施例7)中，上述中长期局部作用最小化的右旋糖酐和上述碱性有机化合物和碱性无机盐的组合(与组别15中不协同药物组合共用性质类似：q 3d＞1和q 21d＜1)的组合物可获得与组别18-20类似的结果。
r 21d相应瘤体(组别18的瘤体样本代表)的组织病理学HE染色检查(委托成都扬克斯生物科技有限公司按公知技术方法进行)结果(图1)显示，瘤体中见较多局灶性坏死区域；坏死区域大小不等，多呈片状或带状；散在分布于肿瘤组织的边缘和中央区，分布较均匀，坏死区面积约占肿瘤组织的10％。坏死区域内见大量坏死肿瘤细胞，间质增宽。坏死区内坏死肿瘤细胞数量较多，大量肿瘤细胞胞核浓缩深染，大量细胞核碎片；靠近瘤周的区域分布有较正常情况过多的免疫细胞、中性粒细胞和淋巴细胞。
因此，推测中长期疗效可能和次生免疫治疗作用有关，这需要后续研究来证实。
此外，与实施例3的结果类似，组别18-21的局部刺激作用现象明显弱于组别15-17和22。
这些结果说明，右旋糖酐可以提供明显高于其短期逆转作用的中长期逆转作用，将其不协同药物逆转为中长期协同共用物组分，或将该共用物本身的中长期药效衰竭的较低药效(例如r 21d/r 3d＜50％)逆转为中长期药效不那么衰竭(例如r 21d/r 3d＞50％)的较高药效，显示出全新的药理意义。
实施例7：中长期逆转剂(组合物)研究
对实施例2-6中的可用作血容量扩张剂的药物进行比较研究，以优选具有最佳中长期共用逆转作用的逆转剂。在一个实验中，建模动物为小鼠，建模细胞为小鼠滑膜肉瘤细胞(ZM755细胞，3×10 6个细胞/只)，建模方法同实施例2，在此不进行赘述。建模成功小鼠的瘤体平均体积为94.2mm 3，将模型动物随机分为11组，每个组别均用药2次，用药间隔为3日，除组别7外每次瘤内注射量100μl/只，而组别7每次瘤内注射量150μl/只。第二次用药后在第21日测局部病变体积(V)，并根据阴性对照组计算抑瘤率(r 21d)。其中各组合物均按常规的水溶液制备方法或实施例1中的制备方法制备。可根据21日的药效r 21d计算中长期药效的实际/预期比q 21d，下表列出药效数据。
表8不同组别用药后的短效、长效数据
Figure PCTCN2022122537-appb-000008
Figure PCTCN2022122537-appb-000009
在上表中，组别1-5的r 21d均为＜5％，说明三个扩张剂及其2个不协同药物均为中期局部作用最小化的药物；组别6－7的r 21d均为＜5％，说明2个不协同药物的组合(在等剂量及提高剂量下)并不提供中期协同作用，而组别9-10的不同扩张剂与该不协同药物组合的瘤内共用药效实际/预期比q 21d均为＞1，显示这些扩张剂均有中长期共用逆转作用。此外，在本实验的给药剂量(约5ml/kg/次)下，各组分(组别1-10)的给药剂量均低于它们各自用作静脉注射时的规定剂量，且给药组合物(组别8-10)和各相应构成组分(组别1-6)的试验动物均未见任何全身性不良反应，说明在该给药剂量下组合物并未产生明显高于其各相应构成组分的系统性毒性，却产生了明显更高的药效。比较而言，组别8的中长期药效最高，显示右旋糖酐具有最强的中长期共用逆转作用。在其它类似实验中，右旋糖酐10、右旋糖酐20、右旋糖酐70代替本实验中的右旋糖酐40也得到与本实验一致的结果，可用作血容量扩张剂的右旋糖酐于是成为本申请的可提供瘤内共用作用逆转的活性成分的优选技术方案。
组别8的r 21d相应瘤体样本代表的组织病理学HE染色检查结果与实施例6中组别18的肿瘤样本代表的结果一致，不再冗述。与r 21d相应的瘤体(组别8的瘤体样本代表)的TUNEL检查(委托成都扬克斯生物科技有限公司按公知技术方法进行)结果显示(图2)，在阳性细胞为细胞核呈棕黄色的细胞时，肿瘤组织坏死区见大量TUNEL阳性细胞，较大坏死区TUNEL阳性细胞密集分布，较正常肿瘤区域偶见散在的TUNEL阳性细胞，进一步证实其中长期疗效。
实施例8：中长期逆转剂(药理条件)研究
对上述实验优选的血容量扩张剂的药理浓度进行比较研究，以确定中长期共用逆转作用的增效浓度范围。在一个实验中，建模动物为小鼠，建模细胞为小鼠乳腺癌细胞(4T1细胞，0.5×105个细胞/只)，建模方法同实施例2，在此不进行赘述。建模成功小鼠的瘤体平均体积为107.3mm3，将模型动物随机分为14组，每个单组均用药2次，用药间隔为3日，每次瘤内注射量100μl/只。第二次用药后在第21日测局部病变体积(V)，并根据阴性对照组计算药效-抑瘤率(r21d)。其中各组合物均按常规的水溶液制备方法或实施例1中的制备方法制备。根据21日的药效r21d分别计算中长期药效的实际/预期比q21d，下表列出药效数据。
表9不同组别用药后的短效、长效数据
Figure PCTCN2022122537-appb-000010
Figure PCTCN2022122537-appb-000011
在上表中，组别1-8的r21d均为＜5％，说明2-25％右旋糖酐40及其2个不协同药物均为中期局部作用最小化的药物，且2个不协同药物的组合并不提供中期协同作用。而组别9-13的不同扩张剂与该不协同药物组合的瘤内共用药效实际/预期比q21d均为＞1，显示这些扩张剂浓度均有中长期共用逆转作用。此外，在本实验的给药剂量(约5ml/kg/次)下，25％右旋糖酐的给药剂量均低于其用作静脉注射时的规定剂量，且给药组合物(组别9)和各相应构成组分(组别1、6-8)的试验动物均未见任何全身性不良反应，说明在该给药剂量下组合物并未产生明显高于其各相应构成组分的系统性毒性，却产生了明显更高的药效。在其它类似实验中，小鼠黑色素瘤细胞(B16F1细胞，3×106个细胞/只)、小鼠肝癌细胞(HepG2细胞，3×106个细胞/只)、小鼠直肠癌细胞(CT26细胞，3×106个细胞/只)代替本实验中的小鼠乳腺癌细胞(4T1细胞，0.5×105个细胞/只)，右旋糖酐10、右旋糖酐20、右旋糖酐70代替本实验中的右旋糖酐40也得到与本实验一致的结果，实验结果在此不进行赘述。2-25％瘤内给药浓度的右旋糖酐于是成为本申请的可提供瘤内共用作用逆转、尤其是中长期逆转的活性成分的优选技术方案。
实施例9：中长期不协同药物及其组合优选
考虑到亚甲蓝在制备过程中易污染设备(实施例1)、在注射时公认的安全剂量(中国药典)较小(≦10mg/kg)，以下试验从实施例2-6中的无协同药物选择了在制备过程中不污染、以及可注射剂量较高的其它药物进行进一步研究，以确定实验例2－6中的不协同药物的优选方案。在一个实验中，建模动物为小鼠，建模细胞为小鼠乳腺癌细胞(4T1细胞，0.5×10 5个细胞/只)，建模方法同实施例2，在此不进行赘述。建模成功小鼠的瘤体平均体积为103.1mm 3，将模型动物随机分为27组，每个单组均用药2次，用药间隔为3日，每次瘤内注射量100μl/只。第二次用药后在第21日测局部病变体积(V)，并根据阴性对照组计算药效-抑瘤率(r 21d)。其中各组合物均按常规的水溶液制备方法或实施例1中的制备方法制备。根据21日的药效r 21d分别计算中长期药效的实际/预期比q 21d，下表列出药效数据。
表10不同组别用药后的短效、长效数据
Figure PCTCN2022122537-appb-000012
在上表中，组别1-10的r 21d均为＜5％，与实施例6中的单药组别的结果一致。在其它类似实验中，瘤内给药浓度为2-25％的右旋糖酐10、右旋糖酐20、右旋糖酐40、右旋糖酐70均显示出与上表中组别1一致的结果，以下瘤内给药浓度(2-25％)的碱性有机化合物均显示出与上表中组别2-5一致的结果：2-10％氨丁三醇、2-15％三乙醇胺、2-15％三羟甲基氨基甲烷、10-25％葡甲胺、10-20％精氨酸、15-25％赖氨酸，以下瘤内给药浓度(2-15％)的碱性无机盐均显示出与上表中组别6-10一致的结果：2-15％碳酸钠、5-15％乳酸钠、5-15％乙酸钠、5-15％碳酸氢钠，即所述实验浓度的右旋糖酐、碱性有机化合物、碱性无机盐至多能显示一定的短期药效，它们的中长期药效均下降至不可观，显示它们均为中长期局部作用最小化的药物，实验结果不再冗述。
在上表中，组别11-16、19的q 21d均为＜1，与实施例6中的不协同药物组合组别的结果一致，进一步说明上述中长期局部作用最小化的不协同药物之间不会产生可观的中长期局部协同作用，既使会产生短期局部协同作用。
在上表中，组别20-25的扩张剂与不同不协同药物组合的瘤内共用药效实际/预期比q 21d均为＞1，显示扩张剂对这些不协同药物组合均有中长期共用逆转作用。此外，在本实验的给药剂量(约5ml/kg/次)下，各组分(组别2-19)的给药剂量均低于它们各自用作静脉注射时的规定剂量，且给药组合物(组别20-25)和各相应构成组分(组别1-7、17-18)的试验动物均未见任何全身性不良反应，说明在该给药剂量下组合物并未产生明显高于其各相应构成组分的系统性毒性，却产生了明显更高的药效。比较而言， 组别20-25的中长期药效最高，显示右旋糖酐对所述中长期局部作用最小化的碱性有机化合物和碱性无机盐的组合具有最强的中长期共用逆转作用，于是成为本申请的右旋糖酐的不协同药物的组合的优选技术方案。
实施例10：大瘤体研究
对上述实验优选的血容量扩张剂和不协同药物及其组合进行大动物研究，以确定本申请优选的肿瘤适应症。在一个实验中，通过成都达硕动物试验公司获得病猪一只，体重35公斤，其下腹部皮下有一巨型结节(图3-1，长度14cm、宽度6cm)，经该公司兽医诊断为纤维瘤，除手术外无任何现有技术药物可以治疗，同意用本公司生产的药物组合物治疗。药物组合物的处方为：200g右旋糖酐40、200g精氨酸、50g碳酸钠溶于适量注射用水中配为1L水溶液，经0.2um滤器除菌过滤后分装为30ml/瓶放置于4℃冰箱贮存。给药均为经皮瘤内注射，每次给药25ml，共计3次，每次之间的间隔时间为14天。第一次给药为2021年10月4日(图3-2)，2021年11月7日瘤体开始明显缩小(图3-3)，2022年3月28日瘤体完全消失(图3-4)，且猪的重量增至52公斤。
该结果说明，本申请优选的适应症为大瘤体的局部治疗，所述大瘤体按公识为最大尺寸大于6cm者、优选为大于7cm甚至大于10cm者。
实施例11：中长期逆转作用的进一步增效研究
对上述实验优选的血容量扩张剂和不协同药物及其组合进行进一步的中长期增效。在一个实验中，建模动物为小鼠，建模细胞为小鼠黑色素瘤细胞(B16F1细胞，3×10 6个细胞/只)，建模方法同实施例2，在此不进行赘述。建模成功小鼠的瘤体平均体积为98.3mm 3，将模型动物随机分为25组，每个单组均用药2次，用药间隔为3日，每次瘤内注射量100μl/只。第二次用药后在第21日测局部病变体积(V)，并根据阴性对照组计算药效-抑瘤率(r 21d)，结果见表11。其中各组合物均按常规的水溶液制备方法或实施例1中的制备方法制备。
表11不同组别用药后的中长效数据
Figure PCTCN2022122537-appb-000013
Figure PCTCN2022122537-appb-000014
在上表中，组别2-10的r 21d均为＜10％，均未显示出可观的中长期局部作用，其中的组别6-10也都未显示出中长期协同作用，所用4种细胞毒药物均为右旋糖酐的不协同药物。
在上表中，组别11-17的r 21d均为＜10％，均未显示出可观的中长期局部作用，也都未显示出中长期协同作用。
在上表中，组别18-24的r 21d均为＞15％，均显示出可观的中长期局部作用，它们的瘤内共用药效实际/预期比q 21d均为＞1，显示扩张剂对这些不协同药物组合(组别11-17)均有中长期共用逆转作用。其中，组别20-24的r 21d均高于组别18-19，这为可接受更高副作用(细胞毒药物毒性)的患者提供了更高的中长期疗效。
以上实施例6-11的结果的共用药理超过现有技术中协同作用定义(共用使得相同作用彼此增强)的药理预期：并不产生中长期药效的右旋糖酐(r 21d＜15％)在本申请技术方案中可提供一种先前未经发现却很有用的药理功能-即将与之本来并不发生中长期协同作用的药物(不协同药物)逆转为可与之产生协同作用的共用物成分，或/和将本来中长期药效衰竭的该共用物逆转为中长期药效不那么衰竭的药物。右旋糖酐中长期逆转作用的技术方案与其短期逆转作用的技术方案一致，包括以下特征：
1、逆转剂只能选择特定化合物、其特定作用、及其特定组成条件。经本申请相关实施例及其它类似实验综合，特定化合物优选为选自包括右旋糖酐10、右旋糖酐20、右旋糖酐40、右旋糖酐70在内的右旋糖酐；其特定作用选自，而非其糖营养素类似物(例如葡萄糖)；其特定作用选自无效、优选不可观、更优选为最小化局部作用，而非为按常规协同物要求选择的至少可观、优选有效、更优选为最大化局部作用；其特定组成条件为满足所述特定作用的中低瘤内给药浓度：2-25％、2-20％、2-10％、或2-4.5％，而非为按常规协同物要求的高瘤内给药浓度；
2)、上述右旋糖酐的被逆转剂只能选择特定化合物、其特定作用、及其特定组成条件。经本申请相关实施例及其它类似实验综合，特定化合物优选为选自碱性有机化合物和碱性无机盐，而非其碱性类似物例如金属钠氢氧化物；其特定作用为作为所述右旋糖酐的不协同药物，而不是按常规协同物要求的协同药物；其特定组成条件为所述碱性有机化合物和碱性无机盐之间产生局部协同作用所需的组成条件，而不是按常规协同物要求的碱性有机化合物或碱性无机盐与右旋糖酐之间产生局部协同作用所需的组成条件。其中所述碱性有机化合物选自包括氨丁三醇、三乙醇胺、三羟甲基氨基甲烷、葡甲胺、精氨酸、赖氨酸之至少一种，所述碱性无机盐选自包括碳酸氢钠、碳酸钠、乳酸钠、乙酸钠之至少一种；所述可被增效剂逆转的不协同药物之间的组成条件为：所述碱性有机化合物和碱性无机盐之间的浓度比为(3-25％)/(4-25％)。
3)、上述特征1、2在瘤内给药而非常规的吸收给药中的相互作用(逆转－被逆转)。
此外，其中所述右旋糖酐/碱性有机化合物/碱性无机盐组合物还可以任选地加入以下组之一种或多种：亚甲蓝及其类似物、免疫调节剂、细胞毒药物，例如以下组合：浓度比(W 碱性有机化合物/W 碱性无机盐/ 亚甲蓝及其类似物) 为(5-25％)/(3-15％)/(0.35-5％)的碱性有机化合物/碱性无机盐/亚甲蓝及其类似物组合、浓度比(W 碱性有机 化合物/W 碱性无机盐/ 免疫调节剂)为(5-25％)/(3-15％)/(1-15％)的碱性有机化合物/碱性无机盐/免疫调节剂组合、浓度比(W 碱性有机化合物/W 碱性无机盐/ 细胞毒药物)为(5-25％)/(3-15％)/(0.1-10％)的碱性有机化合物/碱性无机盐/细胞毒药物组合。
以上所述仅为本发明的较佳实施例而已，并不用以限制本发明，凡在本发明的精神和原则之内，所作的任何修改、等同替换、改进等，均应包含在本发明的保护范围之内。本申请中所引用的各参考文献(包括所有专利、专利申请、期刊文章、书籍及任何其它公开)均以其整体援引加入本文。

Claims (17)

  1. 一种用于治疗实体肿瘤的局部药物组合物，其特征在于，瘤内给药形式为可用作血容量扩张剂的右旋糖酐及其不协同药物的水溶液，其中所述右旋糖酐、不协同药物的药理浓度分别为≤25％、4-40％；所述不协同药物分别选自与所述右旋糖酐共用未显示出局部协同作用的碱性有机化合物和碱性无机盐，所述碱性有机化合物包括氨丁三醇、三乙醇胺、三羟甲基氨基甲烷、葡甲胺、精氨酸、赖氨酸中的至少一种，所述碱性无机盐包括碳酸氢钠、碳酸钠、乳酸钠、乙酸钠中的至少一种。
  2. 根据权利要求1所述的局部药物组合物，其特征在于，所述右旋糖酐的药理浓度为2-25％。
  3. 根据权利要求2所述的局部药物组合物，其特征在于，所述右旋糖酐的药理浓度为2-4.5％。
  4. 根据权利要求1-3之一所述的局部药物组合物，其特征在于，所述右旋糖酐选自右旋糖酐10、右旋糖酐20、右旋糖酐40、右旋糖酐70中的至少一种。
  5. 根据权利要求1所述的局部药物组合物，其特征在于，所述碱性有机化合物和碱性无机盐的药理浓度比为(5-25％)/(3-15％)。
  6. 根据权利要求5所述的局部药物组合物，其特征在于，所述碱性有机化合物选自精氨酸或耐氨酸时，所述碱性有机化合物的药理浓度为10-25％。
  7. 根据权利要求5所述的局部药物组合物，其特征在于，所述碱性有机化合物和所述碱性无机盐的组合选自以下组合之一种：精氨酸/碱性无机盐组合，浓度比为(10-20％)/(2-15％)；精氨酸/三羟甲基氨基甲烷/碱性无机盐组合，浓度比为(10-20％)/(2-15％)/(2-15％)；赖氨酸/碱性无机盐组合，浓度比为(15-25％)/(2-15％)；氨丁三醇/碱性无机盐组合，浓度比为(2-10％)/(2-15％)；三乙醇胺/碱性无机盐组合，浓度比为(2-10％)/(2-15％)；三羟甲基氨基甲烷/碱性无机盐组合，浓度比为(2-15％)/(2-15％)；葡甲胺/碱性无机盐组合，浓度比为(10-25％)/(2-15％)。
  8. 根据权利要求1所述的局部药物组合物，其特征在于，所述局部药物组合物还包括药理浓度为0.1-5％的细胞毒药物和/或药理浓度为0.2-25％的免疫调节剂。
  9. 根据权利要求1所述的局部药物组合物，其特征在于，所述局部药物组合物为瘤内给药注射剂。
  10. 权利要求1-9任意一项所述的局部药物组合物在制备治疗实体肿瘤的药物中的应用，其特征在于，所述局部药物组合物通过瘤内注射给药，所述右旋糖酐作为可提供共用作用逆转、而非提供与各所述共用成分的局部协同作用的活性成分。
  11. 可用作血容量扩张剂的右旋糖酐在制备用于治疗实体肿瘤的药物中的应用，其特征在于，所述右旋糖酐包含在用于瘤内给药的局部药物组合物中，在所述局部药物组合物中所述右旋糖酐作为可提供共用作用逆转、而非提供与各个共用成分的局部协同作用的活性成分。
  12. 根据权利要求11所述的应用，其特征在于，所述共用成分分别选自与所述右旋糖酐共用未显示出局部协同作用的碱性有机化合物和碱性无机盐，且所述碱性有机化合物和碱性无机盐各自具有使得它们之间的局部作用互相增强的药理浓度或浓度比。
  13. 根据权利要求11所述的应用，其特征在于，所述药物组合物为权利要求1-8任意一项所述的局部药物组合物。
  14. 根据权利要求10-13之一所述的应用，其特征在于，所述实体肿瘤选自适于瘤内给药的恶性肿瘤和良性肿瘤，所述恶性肿瘤包括乳腺癌、胰腺癌、甲状腺癌、鼻咽癌、前列腺癌、肝癌、肺癌、肠癌、口腔癌、食道癌、胃癌、喉癌、睾丸癌、阴道癌、子宫癌、卵巢癌、恶性淋巴瘤、肉瘤，所述良性肿瘤包括甲状腺瘤、脂肪瘤、纤维瘤、囊肿、息肉。
  15. 根据权利要求14所述的应用，其特征在于，所述实体肿瘤选自因容易复发而不适用化学消融剂或常规局部协同组合物的实体肿瘤，其中所述常规局部协同组合物是指由局部作用药物与其协同物或协同物的组合形成的局部药物组合物。
  16. 根据权利要求15所述的应用，其特征在于，所述实体肿瘤为尺寸为大于6cm、或大于10cm的超大瘤体。
  17. 一种治疗实体肿瘤的方法，其包括给药权利要求10-13任意一项应用中所述治疗实体肿瘤的药物。
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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110870869A (zh) * 2018-08-31 2020-03-10 成都夸常奥普医疗科技有限公司 包含糖类营养素和常规无效化合物的药物组合物及其应用
WO2021164706A1 (zh) * 2020-02-18 2021-08-26 成都夸常奥普医疗科技有限公司 包含亚甲蓝类染料的药物组合物及其应用
WO2022068918A1 (zh) * 2020-09-30 2022-04-07 成都夸常奥普医疗科技有限公司 包含酸碱中和组合的药物组合物及其应用

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110870869A (zh) * 2018-08-31 2020-03-10 成都夸常奥普医疗科技有限公司 包含糖类营养素和常规无效化合物的药物组合物及其应用
WO2021164706A1 (zh) * 2020-02-18 2021-08-26 成都夸常奥普医疗科技有限公司 包含亚甲蓝类染料的药物组合物及其应用
WO2022068918A1 (zh) * 2020-09-30 2022-04-07 成都夸常奥普医疗科技有限公司 包含酸碱中和组合的药物组合物及其应用

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