WO2022166560A1 - 一种偶氮杯芳烃药用辅料及其用途 - Google Patents

一种偶氮杯芳烃药用辅料及其用途 Download PDF

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WO2022166560A1
WO2022166560A1 PCT/CN2022/071935 CN2022071935W WO2022166560A1 WO 2022166560 A1 WO2022166560 A1 WO 2022166560A1 CN 2022071935 W CN2022071935 W CN 2022071935W WO 2022166560 A1 WO2022166560 A1 WO 2022166560A1
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treatment
drug
azocalixarene
compound
formula
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French (fr)
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郭东升
王宏磊
岳宇昕
李华斌
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苏州隽德生物科技有限公司
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/69—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
    • A61K47/6949—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit inclusion complexes, e.g. clathrates, cavitates or fullerenes
    • 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
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/337—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having four-membered rings, e.g. taxol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/69—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A61P13/12—Drugs for disorders of the urinary system of the kidneys
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis

Definitions

  • the present application relates to the field of pharmacy, in particular to a pharmaceutical composition comprising an azocalixarene supramolecular compound as a hypoxia-responsive targeted pharmaceutical excipient, and a new use of the azocalixarene supramolecular compound in pharmacy.
  • Hypoxia is one of the important pathological and physiological features of many diseases, such as cancer (Nature Reviews Cancer, 2011, 11:393-410), myocardial infarction (N.Engl.J.Med., 2017,377:1240-1249) , stroke (Age and Aging, 2002, 31: 10-12), atherosclerosis (Curr. Opin. Lipidol. 2009, 20: 409-14), rheumatoid arthritis (J. Biomed Sci., 2016, 23:62), inflammatory bowel disease (J.Inflamm Res.2014, 7:113-120), chronic hypoxic lung disease (Lancet Respir Med.2016, 4:225), chronic kidney disease (Int.J.Mol. Sci. 2017, 18:950) et al.
  • hypoxia is also an effective target for these diseases.
  • a common response strategy is to covalently link drugs to hypoxia-responsive groups such as nitro, quinone, azo, etc. In a hypoxic environment, the reduction of the responsive group results in the release of the drug.
  • the covalent strategy faces the problems of complex molecular design, time-consuming and high cost of synthesis and separation, and changes in activity and toxicity caused by covalent attachment.
  • Calixarenes and their derivatives can form complexes with organic molecules or metal ions, so they are considered as the third-generation supramolecular host molecules with great development potential, and have been widely studied in the field of host-guest chemistry.
  • Calixarenes are a class of cyclic oligomers formed by the attachment of phenol units via methylene groups at the ortho position of the phenolic hydroxyl group. Techniques using amphiphilic sulfonated calixarene (see CN103550156A; CN104174026A) for drug delivery have been developed in the prior art.
  • Azocalixarene compounds are an important class of functional calixarene derivatives, which have been widely used in metal ion complexation (see CN108404854A; Lilin Lu et al., Analytica Chimica Acta 535 (2005): 183-187; Tae Hyun Kim et al. Human, Bull. Korean Chem. Soc. 2013, Vol. 34, No. 11: 3377-3380), small organic molecule binding (J.
  • CAC4A azocalixarene compound
  • the inventors have used the sulfonated azocalixarene host with accurate structure, fixed molecular weight, stable batch synthesis, easy derivatization and unique cavity bonding properties as a hypoxia-responsive drug-carrying molecule container platform, which can realize The release of an effective amount of targeted drugs in hypoxic regions of the body.
  • One object of the present invention is to provide a hypoxia-responsive targeted drug-loaded molecule container platform, thereby improving the effectiveness, targeting and safety of active drug molecule delivery.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising at least one pharmaceutically active substance and an azocalixarene compound of formula (I):
  • n is an integer from 4 to 8
  • M is independently selected from H, Na and K.
  • M is preferably Na.
  • n 4, 5 or 6.
  • the pharmaceutically active substance used for targeted drug delivery using the drug-loaded molecule container platform of the present invention is selected from the group consisting of drugs for the treatment of one or more of the following diseases: cancer, myocardial infarction, stroke, atherosclerosis Sclerosis, rheumatoid arthritis, inflammatory bowel disease, chronic hypoxic lung disease, and chronic kidney disease.
  • the drug for treating cancer is selected from the group consisting of adriamycin, paclitaxel, camptothecin, hydroxycamptothecin, irinotecan, topotecan, belonotecan, oxaliplatin, clopidogrel, irinotecan Rodipine, hydroxychloroquine, lovastatin, tadalafil, tiratinib, lenvatinib, sorafenib, regorafenib, gefitinib, and methoxyestradiol.
  • the drug for the treatment of myocardial infarction is selected from: aspirin, ticagrelor, bisoprolol and clopidogrel.
  • the drug for the treatment of stroke is selected from the group consisting of: aspirin, heparin, alteplase, nimodipine, ropirazole, tirapazate, and elirodide.
  • Drugs for the treatment of atherosclerosis are selected from: lovastatin and ezetimibe.
  • the drug for the treatment of rheumatoid arthritis is selected from: methotrexate, tofacitinib, baricitinib, etanercept, abatacept, leflunomide, hydroxychloroquine and sulfasalazine.
  • Drugs for the treatment of inflammatory bowel disease selected from: mesalazine, azathioprine, mercaptopurine, methotrexate, infliximab, adalimumab, certolizumab, natalizumab, and thalizumab Spend.
  • Drugs for the treatment of chronic hypoxic pulmonary disease are selected from: tadalafil, riociguat, and treprostinil.
  • the drug for the treatment of chronic kidney disease is selected from the group consisting of roxadustat, dipyridamole, hydrochlorothiazide and vildarestat.
  • the molar ratio of the pharmaceutical active substance to the azocalixarene compound of formula (I) is 1:(0.8-5.0), preferably 1:(0.9-3.0), more preferably 1:(1.0-1.5), most preferably 1:1.
  • the present invention also relates to the use of the azocalixarene compound of formula (I) in the preparation of hypoxia-responsive targeted pharmaceutical excipients.
  • Azocalixarene compounds are a class of hypoxia-responsive targeted drug excipients, which can improve the solubility and stability of drug molecules and reduce their effects on normal cells.
  • Hypoxia is a typical feature of cancer, myocardial infarction, stroke, atherosclerosis, rheumatoid arthritis, inflammatory bowel disease, chronic hypoxic lung disease, chronic kidney disease and other diseases. In order to survive under hypoxic conditions, hypoxic cells often obtain energy through glycolysis, which is an effective way to resist hypoxia.
  • the present invention provides an azocalixarene compound of formula (I) for use or method for the treatment of one or more of the following diseases: cancer, myocardial infarction, stroke, atherosclerosis, rheumatoid joints inflammation, inflammatory bowel disease, chronic hypoxic lung disease and chronic kidney disease,
  • n is an integer from 4 to 8
  • M is independently selected from H, Na and K;
  • the use or method comprises mixing the azocalixarene compound as a pharmaceutical excipient with a pharmaceutically active substance for the treatment of the one or more diseases and administering to a patient in need thereof.
  • Figure 1 shows a schematic diagram of the hypoxia response of the PTX/SACnA host-guest inclusion complex.
  • Figure 2 shows the UV absorption curves of SAC4A before and after adding SDT.
  • Figure 3a shows photographs of clear solutions obtained after dissolving various concentrations of SAC5A and drug molecule PTX in PBS;
  • Figure 3b shows the phase solubility curve of SAC5A solubilized PTX.
  • Figures 4a-f show graphs of changes in ALT, AST, ALP, BUN, LDH and CRP in the blood of two groups of mice after administration of PBS and SAC5A.
  • FIG. 5 shows the H&E staining photographs of each organ in the two groups of mice after administration of PBS and SAC5A.
  • Figure 6a shows lymphocyte levels in mice treated with PBS, PTX (5.00 mg/kg), SAC5A (15.61 mg/kg), SAC5A/PTX (PTX: 5.00 mg/kg, SAC5A: 10.93 mg/kg);
  • Figure 6b shows leukocyte levels in mice treated with PBS, PTX (5.00 mg/kg), SAC5A (15.61 mg/kg), SAC5A/PTX (PTX: 5.00 mg/kg, SAC5A: 10.93 mg/kg).
  • Figure 7 shows PBS, SAC5A/PTX (PTX: 5.00 mg/kg, SAC5A: 10.93 mg/kg), SAC5A/PTX (PTX: 2.50 mg/kg, SAC5A: 5.46 mg/kg), PTX (5.00 mg/kg) , Changes in tumor volume of mice treated with Lipo-PTX (5.00 mg/kg), HSA-PTX (5.00 mg/kg), and SAC5A (10.93 mg/kg).
  • Figure 8 shows PBS, SAC5A/PTX (PTX: 5.00 mg/kg, SAC5A: 10.93 mg/kg), SAC5A/PTX (PTX: 2.50 mg/kg, SAC5A: 5.46 mg/kg), PTX (5.00 mg/kg) , Changes in body weight of tumor-bearing mice treated with Lipo-PTX (5.00 mg/kg), HSA-PTX (5.00 mg/kg), and SAC5A (10.93 mg/kg).
  • Figure 9 shows PBS, SAC5A/PTX (PTX: 5.00 mg/kg, SAC5A: 10.93 mg/kg), SAC5A/PTX (PTX: 2.50 mg/kg, SAC5A: 5.46 mg/kg), PTX (5.00 mg/kg) ), Lipo-PTX (5.00 mg/kg), HSA-PTX (5.00 mg/kg), SAC5A (10.93 mg/kg) treatment of tumor-bearing mice survival rate.
  • Figure 10 shows confocal micrographs of Ki67, TUNLE and H&E staining of mouse tumors in each group.
  • the present invention employs conventional methods of mass spectrometry, NMR, HPLC, protein chemistry, biochemistry, recombinant DNA techniques and pharmacology, which are within the skill in the art.
  • mass spectrometry NMR, HPLC, protein chemistry, biochemistry, recombinant DNA techniques and pharmacology
  • specific definitions are provided, the nomenclature and laboratory procedures and techniques associated with the chemistry of analytical chemistry, synthetic organic chemistry, and medical and medicinal chemistry described herein are known to those skilled in the art.
  • the foregoing techniques and procedures can be performed by conventional methods well known in the art and described in various general and more specific documents, which are cited and discussed in this specification.
  • Azocalixarene compound of the present invention and drug-carrying use thereof
  • the present invention provides a hypoxia-responsive targeted drug-carrying molecule container platform, which is an azocalixarene compound of formula (I):
  • n is an integer from 4 to 8
  • M is independently selected from H, Na and K.
  • M is preferably Na.
  • n 4, 5 or 6.
  • the sulfonated azocalixarene compound as the hypoxia-responsive targeting drug-carrying molecule container platform of the present invention is selected from the following compounds:
  • the sulfonated azocalixarene compounds of the present invention can form host-guest inclusion complexes with active pharmaceutical substances through non-covalent interactions such as hydrogen bonds, electrostatics, hydrophobicity, etc., and the bonding constant is 10 3 or more, more preferably 10 4 or more, and then More preferably 10 5 or more, particularly preferably 10 6 or more, and most preferably 10 7 or more.
  • the azo bond of the sulfonated azocalixarene compound of the present invention is reduced and broken under the action of the overexpressed azoreductase, so that the active substance of the drug is released from the prototype, achieving The targeting of the drug is reduced, the dosage is reduced, and the safety is improved.
  • the sulfonated azocalixarene compound of the present invention can form a stable host-guest non-covalent binding structure with various small-molecule pharmaceutical active substances, and target the pharmaceutical active substances to be released in the hypoxic region of the body, it can be used as Hypoxia-responsive targeted drug-loading molecular container platforms or drug excipients are used in the pharmaceutical field.
  • compositions comprising at least one pharmaceutically active substance and an azocalixarene compound of formula (I).
  • the pharmaceutical compositions of the present invention may also include other pharmaceutically acceptable carriers or excipients, and other therapeutic agents.
  • the pharmaceutically active substance used for targeted drug delivery using the drug-loaded molecule container platform of the present invention is selected from the group consisting of drugs for the treatment of one or more of the following diseases: cancer, myocardial infarction, stroke, atherosclerosis Sclerosis, rheumatoid arthritis, inflammatory bowel disease, chronic hypoxic lung disease, and chronic kidney disease.
  • the preferred drug for treating cancer is selected from: doxorubicin, paclitaxel, camptothecin, hydroxycamptothecin, irinotecan, topotecan, belonotecan, oxaliplatin, clopidogrel, Elirodib, hydroxychloroquine, lovastatin, tadalafil, tiratinib, lenvatinib, sorafenib, regorafenib, gefitinib, and methoxyestradiol.
  • Preferred drugs for the treatment of myocardial infarction are selected from the group consisting of aspirin, ticagrelor, bisoprolol and clopidogrel.
  • Preferred drugs for the treatment of stroke are selected from the group consisting of aspirin, heparin, alteplase, nimodipine, ropirazole, tirapazate and elirodide.
  • Preferred drugs for the treatment of atherosclerosis are selected from: lovastatin and ezetimibe.
  • Preferred drugs for the treatment of rheumatoid arthritis are selected from: methotrexate, tofacitinib, baricitinib, etanercept, abatacept, leflunomide, hydroxychloroquine and sulfasalazine.
  • Preferred drugs for the treatment of inflammatory bowel disease are selected from: mesalazine, azathioprine, mercaptopurine, methotrexate, infliximab, adalimumab, certolizumab, natalizumab and Salidor.
  • Preferred drugs for the treatment of chronic hypoxic pulmonary disease are selected from the group consisting of: tadalafil, riociguat and treprostinil.
  • Preferred drugs for the treatment of chronic kidney disease are selected from the group consisting of: roxadustat, dipyridamole, hydrochlorothiazide and vildarestat.
  • the at least one pharmaceutically active substance and the sulfonated azocalixarene compound of formula (I) of the present invention are administered to the patient in a molar ratio of 1:(0.8-5.0), preferably 1:(0.9-3.0 ), more preferably in a molar ratio of 1:(1.0-1.5), most preferably in a molar ratio of about 1:1.
  • the medicament comprising the pharmaceutical excipient and pharmaceutically active substance of the present invention can be administered to a patient by at least one of conventional injection, oral, inhalation, rectal and transdermal administration.
  • Other applicable modes of administration include, but are not limited to, auricular administration, intra-articular administration, intramuscular administration, intrathecal administration, in vitro administration, buccal administration, intrabronchial administration, and conjunctival administration.
  • the amount of a given drug depends on factors such as the particular dosing regimen, the type of disease or condition and its severity, the subject in need of treatment or uniqueness of the host (eg body weight). However, depending on the particular surrounding circumstances, including, for example, the particular drug employed, the route of administration, the condition being treated, and the subject or host being treated, the dosage administered can be routinely determined by methods known in the art. Generally, for adult therapeutic use, the administered dose is typically in the range of 0.02-5000 mg/day, eg, about 1-1500 mg/day.
  • the desired dose may conveniently be presented as a single dose, or as divided doses administered simultaneously (or over a short period of time) or at appropriate intervals, eg, two, three, four or more divided doses per day. It will be understood by those skilled in the art that although the above dosage range is given, the specific effective amount can be appropriately adjusted according to the patient's condition and in conjunction with the physician's diagnosis.
  • the sulfonated azocalixarene compound of the present invention has the advantage as a drug-loading molecule container platform: it has high solubility in solution, especially in aqueous solution, In addition, it has additional solubilizing effect for poorly soluble drugs, so when used as a drug delivery platform, it will not limit the administration concentration of active drugs due to the limitation of solubility, which can provide a higher degree of freedom for the choice of administration concentration.
  • the reactions can be used sequentially to provide the compounds described herein; or they can be used to synthesize fragments that are subsequently added by methods described herein and/or methods known in the art.
  • reaction product can be isolated and purified using conventional techniques including, but not limited to, filtration, distillation, crystallization, chromatography, and the like. These products can be characterized using conventional methods, including physical constants and spectral data.
  • the general synthetic route of the azocalixarene compound of the present invention is as follows.
  • CAC4A was additionally synthesized.
  • Example 1 Determination of the bonding constant between sulfonated azocalixarene compounds and drug molecules
  • Test method Fluorescence titration.
  • Test tools Quartz cuvette is used as the sample cell for the test, the test optical path is 10mm, the instrument model is Varian Cary Eclipse, and the temperature control device of the cuvette is equipped with the model Cary Single-cuvette Peltier.
  • Rhodamine B was purchased from Shanghai McLean Biochemical Technology Co., Ltd.
  • Paclitaxel (PTX) was purchased from Melan Bio, and other active drugs were commercially available products.
  • PBS phosphate buffer solution
  • the paclitaxel molecular solution was added into the fluorescence cell in a predetermined volume, and the change of fluorescence intensity was recorded.
  • the volume of paclitaxel added was less than 0.8% of the total volume.
  • the fluorescence titration data were fitted by the host-guest 1:1 competitive bonding model.
  • the excitation wavelength was 554 nm, and the binding constant Ka of the host-guest inclusion was determined. The results are shown in Table 1 below.
  • PBS phosphate buffer solution
  • the azocalixarene compound of the present invention has strong binding strength with active drug molecules, and the binding constant reaches 10 3 or more, preferably 10 4 or more, more preferably 10 5 or more, still more preferably 10 6 or more , particularly preferably 10 7 or more. Therefore, the sulfonated azocalixarene compounds of the present invention can form stable host-guest non-covalent bonds with active drug molecules.
  • Test method UV-Vis spectroscopy.
  • Test tool Japan Shimadzu UV-3600 UV-Vis spectrophotometer, equipped with a temperature control module (model: PTC-348WI), the test sample uses Shimadzu's own quartz cuvette, with an optical path of 10mm.
  • the fluorescence instrument model is Varian Cary Eclipse, with a cuvette temperature control device model Cary Single-cuvette Peltier.
  • the test uses a quartz cuvette as the sample cell, and the test optical path is 10mm.
  • PBS phosphate buffered saline
  • the test results are shown in Figure 2.
  • Figure 2 shows the UV absorption curves of SAC4A before and after adding SDT. It can be seen from Figure 2 that after the addition of SDT, the UV absorption corresponding to the azo bond in SAC4A gradually decreased with time, indicating that SAC4A can be reduced by SDT and has hypoxia responsiveness.
  • Test method UV-Vis spectroscopy.
  • Test tool Japan Shimadzu UV-3600 UV-Vis spectrophotometer, equipped with a temperature control module (model: PTC-348WI), the test sample uses Shimadzu's own quartz cuvette, with an optical path of 10mm.
  • the fluorescence instrument model is Varian Cary Eclipse, with a cuvette temperature control device model Cary Single-cuvette Peltier.
  • the test uses a quartz cuvette as the sample cell, and the test optical path is 10mm.
  • SAC5A As an example, first test the UV standard curve of SAC5A, configure a series of standard solutions of SAC5A, the concentrations are: 1 ⁇ M, 2 ⁇ M, 4 ⁇ M, 6 ⁇ M, 8 ⁇ M, and obtain the standard curve.
  • PBS phosphate buffered saline
  • the solubility of the sulfonated azocalixarene compounds of the present invention is significantly improved. Therefore, when the sulfonated azocalixarene compound of the present invention is used as a drug delivery platform, the administration concentration of the active drug will not be restricted due to the limitation of solubility, and can provide a higher degree of freedom for the choice of administration concentration.
  • chemotherapeutic drug camptothecin its commonly administered intravenous concentration (100 microliters per 10 grams of mice) is 1.44-2.87 mM.
  • the molar concentration ratio of the azocalixarene compound combined with the chemotherapeutic drug as a drug carrier is at least 1:1, and the solubility of the carboxyl azocalixarene CAC4A itself limits the administration dose of the active compound, so the sulfonated azocalix of the present invention is selected.
  • Aromatic compounds have significant advantages.
  • Example 4 Solubilization experiment of active drug molecules by sulfonated azocalixarene compounds
  • PTX solids and SAC5A solids were weighed at the same molar concentration and ground in a mortar for 15 minutes, and then added quantitative PBS (10 mM, pH 7.4) to dissolve to obtain a clear solution.
  • PBS 10 mM, pH 7.4
  • Liquid phase conditions were: Waters 2695 HPLC (Waters, Alliance 2695, USA) at 37°C at Chromatographic separation was performed on a chromatographic column (4.6 mm x 250 mm, 5 ⁇ m) with isocratic elution with water (A) and methanol (B).
  • the flow rate of the mobile phase was set to 1.0 mL min -1 , the injection volume was 10 ⁇ L, and the detection wavelength was 232 nm.
  • the sulfonated azocalixarene compound SAC5A of the present invention has a significant solubilizing effect on poorly water-soluble drugs, so it can solubilize poorly water-soluble drug molecules, improve drug utilization and safety, and can be used for water-soluble drugs. Then, the side effects are reduced and the therapeutic effect is enhanced through the strong inclusion effect.
  • mice Female Balb/c, 6-8 week old mice (purchased from Beijing Speifu Company) were selected and randomly divided into PBS and SAC5A groups, with 3 mice in each group, and each mouse weighed about 20 grams.
  • the drug was administered by tail vein injection, and the dose was 1 mM, 200 ⁇ L each time, and the drug dose of SAC5A was 15.61 mg/kg, four times every other day.
  • the safety experiment was determined according to the kit provided by Nanjing Jiancheng Bioengineering Institute. Blood biochemical test results are shown in Figures 4a-4f.
  • Alkaline phosphatase (ALP) alkaline phosphatase
  • Lactate dehydrogenase lactate dehydrogenase
  • CRP C-reactive protein
  • mice were dissected out of the heart, liver, spleen, lung and kidney. Each organ tissue was fixed in 4% paraformaldehyde (sigma) for 24 hours, and then sent to Tianjin Yishengyuan Biotechnology Co., Ltd. for paraffin section and H&E staining experiments. Microscopic analysis was subsequently performed.
  • the Lymph and WBC indexes in the PTX group were higher than those in the PBS group, indicating that free PTX could trigger an inflammatory response, while the SAC5A and PTX/SAC5A indexes were not significantly different from those in the PBS group, proving their higher safety.
  • 106 4T1 cells were injected subcutaneously into the armpits of 6-8 week old female Balb/C mice and allowed to grow to a tumor size of 100 mm3 .
  • mice were randomly divided into PBS, SAC5A (10.93 mg/kg), PTX (5.00 mg/kg), SAC5A/PTX (PTX: 5.00 mg/kg, SAC5A: 10.93 mg/kg), SAC5A/PTX (PTX: 2.50 mg/kg, SAC5A: 5.46mg/kg), Lipo-PTX (Liposu (paclitaxel liposome for injection), purchased from Nanjing Luye Pharmaceutical Co., Ltd., 5.00mg/kg, calculated as PTX), HSA-PTX ( Paclitaxel for injection (albumin-bound type), purchased from Shijiazhuang Pharmaceutical Group Ouyi Pharmaceutical Co., Ltd., 5.00 mg/kg, calculated as PTX) seven groups, 6 mice in each group.
  • Lipo-PTX Liposu (paclitaxel liposome for injection)
  • HSA-PTX Paclitaxel for injection (albumin-bound type), purchased from Shijiazhuang Pharmaceutical Group Ouyi Pharmaceutical Co.,
  • the drugs in each group were injected into the tail vein, and the tumor length and diameter were measured before each injection.
  • the drug was administered every other day, 5 times for a total of 9 days.
  • Each dose of 100 ⁇ L, the drug concentrations of each group were SAC5A (1.40mM), PTX (1.17mM), PTX/SAC5A (1.17/1.40mM), PTX/SAC5A (0.59/0.70mM), Lipo-PTX (1.17mM) ), HSA-PTX (1.17 mM).
  • the mouse tumor volume and mouse body weight were measured from the first day of administration, and the results are shown in Fig. 7 and Fig. 8, respectively.
  • the survival rates of tumor-bearing mice in different treatment groups are shown in FIG. 9 .
  • the survival rate of mice in the PTX/SAC5A group was higher than that of other groups, proving that PTX/SAC5A can significantly improve the survival rate and have a prognostic effect of long-term treatment.
  • mice After administration, the survival of the mice was continuously monitored, and the tumor volume reached 2000 mm 3 by default, the mice died. After the mice died, the tumors were taken out for section staining analysis.
  • TUNLE staining procedure Frozen sections of mouse tumors were processed, and stained according to the experimental manual provided by Roach Company after moistening with PBS for 15 minutes.
  • Ki67 staining steps Take the frozen section out from -80°C, return to room temperature, treat with 0.1% Triton X-100 for 15 minutes, wash off Triton with PBS, block with 5% BSA (Sigma) for 1 hour, and add primary antibody (Ki67 primary antibody) , Rat source, Solebao), 4°C protected from light overnight. After 12 hours, the primary antibody was washed off with PBS, and then a fluorescently labeled secondary antibody (donkey anti-rabbit, alexa Fluro488, Soleibo) was added. After 1 hour, the slides were photographed and analyzed by confocal microscopy.
  • H&E staining steps 1) Dewax the paraffin sections to water: put the sections in xylene I for 30 minutes, xylene II for 30 minutes, anhydrous ethanol I for 10 minutes, anhydrous ethanol II for 10 minutes, 95% alcohol for 5 minutes, and 90% alcohol for 5 2) Hematoxylin staining of nuclei: Sections were stained with Harris hematoxylin for 5-10 minutes, washed with tap water, differentiated with 1% hydrochloric acid alcohol for several seconds, washed with tap water for 10 minutes, PBS Back to blue for 5 minutes, rinsed with running water; 3) Eosin stained cytoplasm: slices were stained in eosin staining solution for 1-3 minutes; 4) Dehydrated and mounted: the slices were sequentially placed in 95% alcohol I for 5 minutes to 95% alcohol II 5 minutes - absolute ethanol I 5 minutes - absolute ethanol II 5 minutes - xylene I 5 minutes - xylene II 5 minutes, dehydrated and transparent, take out the sections from
  • the fluorescent signal marks the fragmented DNA.
  • the stronger the fluorescent signal the higher the degree of apoptosis.
  • the fluorescent signal stained on the nucleus of the PTX/SAC5A group is the strongest.
  • the degree of apoptosis was significantly higher than the other three groups.
  • Ki67 marks cell proliferation-related antigens.
  • the degree of cell proliferation in the PTX/SAC5A group was lower than that in the control group.
  • pyknosis, nuclear cytoplasmic separation, and nuclear efflux were observed in the PTX/SAC5A group by a 20 ⁇ microscope.
  • the degree of cell necrosis in the PTX/SAC5A group was significantly higher than that in the other three groups, which proved that the PTX/SAC5A group had a good tumor-killing effect.
  • the invention provides a sulfonated azocalixarene supramolecular compound as a hypoxia-responsive targeted drug adjuvant.
  • the compounds of the present invention can be formulated into corresponding pharmaceutical compositions together with appropriate active drug molecules, suitable for industrial application.

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Abstract

本发明涉及一种包括式(I)的偶氮杯芳烃超分子化合物作为乏氧响应的靶向药用辅料的药物组合物、以及偶氮杯芳烃超分子化合物在制药中的新用途。

Description

一种偶氮杯芳烃药用辅料及其用途 技术领域
本申请涉及制药领域，特别是涉及一种包括偶氮杯芳烃超分子化合物作为乏氧响应的靶向药用辅料的药物组合物、以及偶氮杯芳烃超分子化合物在制药中的新用途。
背景技术
乏氧是许多疾病的重要病理、生理特征之一，例如癌症(Nature Reviews Cancer,2011,11:393-410)、心肌梗塞(N.Engl.J.Med.,2017,377：1240-1249)、中风(Age and Aging,2002,31:10-12)、动脉粥样硬化(Curr.Opin.Lipidol.2009,20:409-14)、类风湿性关节炎(J.Biomed Sci.,2016,23:62)、炎症性肠病(J.Inflamm Res.2014,7:113-120)、慢性缺氧性肺病(Lancet Respir Med.2016,4:225)、慢性肾病(Int.J.Mol.Sci.2017,18:950)等。同时，乏氧也是这些疾病的有效靶点。常见的响应策略是将药物与乏氧响应基团——如硝基、醌、偶氮等共价相连。在乏氧环境下，响应基团发生还原后，引起药物的释放。然而，共价策略面临着分子设计复杂、合成分离耗时且成本高、以及共价连接带来活性变化和毒性等问题。
鉴于此，在制药领域中存在开发新的主客体非共价策略用于乏氧治疗的需求。主客体相互作用是组装过程中重要的驱动力之一，其多样的键合选择性，使得其在医药、生物化工、材料科学等领域存在潜在的应用前景。杯芳烃及其衍生物能够与有机分子或金属离子形成配合物，因此被认为是极具发展潜力的第三代超分子主体分子，在主客体化学领域得到广泛的研究。杯芳烃是一类由苯酚单元通过亚甲基在酚羟基的邻位连接而形成的环状低聚物。现有技术中已经开发出采用两亲性磺化杯芳烃(参见CN103550156A；CN104174026A)应用于药物递送的技术。
偶氮杯芳烃化合物是一类重要的功能性杯芳烃衍生物，已经广泛应用于金属离子络合(参见CN108404854A；Lilin Lu等人，Analytica Chimica Acta 535(2005):183-187；Tae Hyun Kim等人，Bull.Korean  Chem.Soc.2013,Vol.34,No.11:3377-3380)、有机小分子结合(J.Cameron Tyson等人，Journal of Inclusion Phenomena and Molecular Recognition in Chemistry 29(1997):109-118)、离子选择电极(Jianquan Lu等人，Journal of Electroanalytical Chemistry,528(2002),33-38)、染料(Shobhana K.Menon等人，J.Incl.Phenom.Macrocycl.Chem.(2010)67:73-79；Serkan
Figure PCTCN2022071935-appb-000001
等人，J.Incl.Phenom.Macrocycl.Chem.(2012)77:259-267)等领域。有论文称该类化合物也有被应用于包络药物活性分子而用于药物递送的前景。然而，据报道该类化合物仅仅被用于包络抗菌类药物或抗精神病类药物并起到药物缓释作用，没有在体内的特定靶点定向释放药物的功能。
本发明人在先的研究中开发了一种偶氮杯芳烃化合物CAC4A(5,11,17,23-四[(对羧基苯基)偶氮杂]-25,26,27,28-四-羟基杯[4]芳烃)，发现其能够在乏氧环境下释放出显影剂Rho123，因此可应用于乏氧环境成像(Geng,Wen-Chao等人，A Noncovalent Fluorescence Turn-on Strategy for Hypoxia Imaging,Angewandte Chemie,131,2019,pp.2399-2403)。然而，CAC4A化合物在溶解度方面存在限制，可能无法将有效量的活性物质递送到目标靶点。
本发明人经过研究，利用结构准确、分子量固定、批合成稳定、易于衍生化且具有独特空腔键合性质的磺化偶氮杯芳烃主体，作为乏氧响应的载药分子容器平台，可实现有效量的靶向药物在体内乏氧区域的释放。
发明内容
本发明的一个目的是提供一种乏氧响应的靶向载药分子容器平台，从而提高活性药物分子递送的有效性、靶向性和安全性。
更具体地，本发明提供一种药物组合物，包括至少一种药物活性物质和式(I)的偶氮杯芳烃化合物：
Figure PCTCN2022071935-appb-000002
其中，
n为4至8的整数，
M独立地选自H、Na和K。
在优选的实施方式中，M优选为Na。
另外优选地，n为4、5或6。
在优选的实施方式中，用于使用本发明的载药分子容器平台进行靶向给药的药物活性物质选自治疗以下一种或多种疾病的药物：癌症、心肌梗塞、中风、动脉粥样硬化、类风湿性关节炎、炎症性肠病、慢性缺氧性肺病和慢性肾病。
优选地，治疗癌症的药物选自：选自阿霉素、紫杉醇、喜树碱、羟基喜树碱、伊立替康、拓扑替康、贝洛替康、奥沙利铂、氯吡格雷、依利罗地、羟基氯喹、洛伐他汀、他达拉非、替拉替尼、乐伐替尼、索拉非尼、瑞戈非尼、吉非替尼和甲氧雌二醇。
治疗心肌梗塞的药物选自：阿司匹林、替格瑞洛、比索洛尔和氯吡格雷。
治疗中风的药物选自：阿司匹林、肝素、阿替普酶、尼莫地平、罗吡唑、替拉扎特和依利罗地。
治疗动脉粥样硬化的药物选自：洛伐他汀和依泽替米贝。
治疗类风湿性关节炎的药物选自：甲氨蝶呤、托法替尼、巴瑞替尼、依那西普、阿巴西普、来氟米特、羟基氯喹和柳氮磺胺吡啶。
治疗炎症性肠病的药物选自：美沙拉秦、硫唑嘌呤、巯嘌呤、甲氨蝶呤、英夫利昔单抗、阿达木单抗、赛妥珠单抗、那他珠单抗和沙利度。
治疗慢性缺氧性肺病的药物选自：他达拉非、利奥西呱和曲前列环素。
治疗慢性肾病的药物选自：罗沙司他、双嘧达莫、氢氯噻嗪和维达司他。
在本发明的优选的药物组合物中，药物活性物质与式(I)的偶氮杯芳烃化合物的摩尔比为1:(0.8-5.0)，优选为1:(0.9-3.0)，更优选为1:(1.0-1.5)，最优选为1:1。
本发明还涉及式(I)的偶氮杯芳烃化合物在制备乏氧响应的靶向药物辅料中的用途。偶氮杯芳烃化合物是一类乏氧响应的靶向药物辅料，可以改善药物分子的溶解性和稳定性，降低其对正常细胞的影响。乏氧是癌症、心肌梗塞、中风、动脉粥样硬化、类风湿性关节炎、炎症性肠病、慢性缺氧性肺病、慢性肾病等疾病的典型特征。为了在乏氧条件下生存，乏氧细胞往往通过糖酵解获得能量，成为抵抗缺氧的有效途径。然而，这种途径会导致乳酸水平和还原酶的升高，例如细胞色素b5还原酶、NADPH硝基还原酶和DT-硫辛酰胺脱氢酶等。当药物分子被包结在本发明的偶氮杯芳烃化合物主体空腔内时，在正常组织，药物分子不被释放；在乏氧环境中，遇到过度表达的还原酶(偶氮还原酶)，杯芳烃偶氮键断裂，药物分子脱离主体空腔，在乏氧区域可控地释放，实现在乏氧区域的靶向治疗，使偶氮杯芳烃化合物成为一类具有刺激响应的靶向药物辅料。
在又一方面，本发明提供式(I)的偶氮杯芳烃化合物，用于治疗以下一种或多种疾病的用途或方法：癌症、心肌梗塞、中风、动脉粥样硬化、类风湿性关节炎、炎症性肠病、慢性缺氧性肺病和慢性肾病，
Figure PCTCN2022071935-appb-000003
其中，
n为4至8的整数，
M独立地选自H、Na和K；
所述用途或方法包括将所述偶氮杯芳烃化合物作为药物辅料，与 用于治疗所述一种或多种疾病的药物活性物质混合，并施用给有需要的患者。
附图说明
图1显示PTX/SACnA主客体包合物乏氧响应的示意图。
图2显示加入SDT前后，SAC4A的紫外吸收曲线。
图3a显示将各浓度的SAC5A和药物分子PTX溶于PBS后得到的澄清溶液的照片；图3b显示SAC5A增溶PTX的相溶解度曲线。
图4a-f显示PBS和SAC5A给药后两组小鼠血液中ALT、AST、ALP、BUN、LDH和CRP变化的图。
图5显示PBS和SAC5A给药后两组小鼠中各器官的H&E染色照片。
图6a显示用PBS、PTX(5.00mg/kg)、SAC5A(15.61mg/kg)、SAC5A/PTX(PTX：5.00mg/kg，SAC5A：10.93mg/kg)处理的小鼠的淋巴细胞水平；图6b显示用PBS、PTX(5.00mg/kg)、SAC5A(15.61mg/kg)、SAC5A/PTX(PTX：5.00mg/kg，SAC5A：10.93mg/kg)处理的小鼠的白细胞水平。
图7显示PBS、SAC5A/PTX(PTX：5.00mg/kg，SAC5A：10.93mg/kg)、SAC5A/PTX(PTX：2.50mg/kg，SAC5A：5.46mg/kg)、PTX(5.00mg/kg)、Lipo-PTX(5.00mg/kg)、HSA-PTX(5.00mg/kg)、SAC5A(10.93mg/kg)处理的小鼠的肿瘤体积变化。
图8显示PBS、SAC5A/PTX(PTX：5.00mg/kg，SAC5A：10.93mg/kg)、SAC5A/PTX(PTX：2.50mg/kg，SAC5A：5.46mg/kg)、PTX(5.00mg/kg)、Lipo-PTX(5.00mg/kg)、HSA-PTX(5.00mg/kg)、SAC5A(10.93mg/kg)处理的荷瘤小鼠的体重变化。
图9示出PBS、SAC5A/PTX(PTX：5.00mg/kg，SAC5A：10.93mg/kg)、SAC5A/PTX(PTX：2.50mg/kg，SAC5A：5.46mg/kg)、PTX(5.00mg/kg)、Lipo-PTX(5.00mg/kg)、HSA-PTX(5.00mg/kg)、SAC5A(10.93mg/kg)治疗后荷瘤小鼠的存活率。
图10显示各组小鼠肿瘤的Ki67、TUNLE和H&E染色的共聚焦显微镜照片。
具体实施方式
术语
除非另外定义，所有本文使用的科技术语都具有与要求保护的主题所属领域的技术人员一般理解相同的含义。
除非另有说明，本发明采用本领域技术范围内的质谱、NMR、HPLC、蛋白质化学、生物化学、重组DNA技术和药理学等常规方法。除非提供具体的定义，否则与本文描述的分析化学、合成有机化学、以及医学和药物化学等化学上相关的命名和实验室操作和技术，是本领域技术人员已知的。一般而言，前述技术和步骤可以通过本领域众所周知的和在各种一般文献和更具体文献中描述的常规方法来实施，这些文献在本说明书中被引用和讨论。
本发明的偶氮杯芳烃化合物及其载药用途
本发明提供一种乏氧响应的靶向载药分子容器平台，其为式(I)的偶氮杯芳烃化合物：
Figure PCTCN2022071935-appb-000004
其中，
n为4至8的整数，
M独立地选自H、Na和K。
在优选的实施方式中，M优选为Na。
另外优选地，n为4、5或6。
特别优选地，本发明的作为乏氧响应的靶向载药分子容器平台的磺化偶氮杯芳烃化合物选自以下化合物：
Figure PCTCN2022071935-appb-000005
本发明的磺化偶氮杯芳烃化合物能够与药物活性物质通过氢键、静电、疏水等非共价相互作用形成主客体包合物，键合常数为10 3以上，更优选10 4以上，再更优选10 5以上，特别优选10 6以上，最优选10 7以上。如图1所示，在体内乏氧区域，本发明的磺化偶氮杯芳烃化合物的偶氮键在过表达的偶氮还原酶作用下被还原断裂，使得药物活性物质被原型释放出来，实现了药物的靶向性，减少了给药量，提高了安全性。
由于本发明的磺化偶氮杯芳烃化合物能够与各种小分子药物活性物质形成稳定的主客体非共价结合结构，并在体内乏氧区域将药物活性物质靶向释放出来，因此可以用作乏氧响应的靶向载药分子容器平台或药物辅料而在制药领域中应用。
本发明的药物组合物
本申请还提供药物组合物，其包括至少一种药物活性物质和式(I)的偶氮杯芳烃化合物。任选地，本发明的药物组合物还可以包括其它药学可接受的载体或赋形剂、以及其它治疗剂。
在优选的实施方式中，用于使用本发明的载药分子容器平台进行靶向给药的药物活性物质选自治疗以下一种或多种疾病的药物：癌症、心肌梗塞、中风、动脉粥样硬化、类风湿性关节炎、炎症性肠病、慢性缺氧性肺病和慢性肾病。
其中，优选的治疗癌症的药物选自：选自阿霉素、紫杉醇、喜树碱、羟基喜树碱、伊立替康、拓扑替康、贝洛替康、奥沙利铂、氯吡格雷、依利罗地、羟基氯喹、洛伐他汀、他达拉非、替拉替尼、乐伐替尼、索拉非尼、瑞戈非尼、吉非替尼和甲氧雌二醇。
优选的治疗心肌梗塞的药物选自：阿司匹林、替格瑞洛、比索洛尔和氯吡格雷。
优选的治疗中风的药物选自：阿司匹林、肝素、阿替普酶、尼莫地平、罗吡唑、替拉扎特和依利罗地。
优选的治疗动脉粥样硬化的药物选自：洛伐他汀和依泽替米贝。
优选的治疗类风湿性关节炎的药物选自：甲氨蝶呤、托法替尼、巴瑞替尼、依那西普、阿巴西普、来氟米特、羟基氯喹和柳氮磺胺吡啶。
优选的治疗炎症性肠病的药物选自：美沙拉秦、硫唑嘌呤、巯嘌呤、甲氨蝶呤、英夫利昔单抗、阿达木单抗、赛妥珠单抗、那他珠单抗和沙利度。
优选的治疗慢性缺氧性肺病的药物选自：他达拉非、利奥西呱和曲前列环素。
优选的治疗慢性肾病的药物选自：罗沙司他、双嘧达莫、氢氯噻嗪和维达司他。
在优选的实施方式中，至少一种药物活性物质与本发明式(I)的磺化偶氮杯芳烃化合物以1:(0.8-5.0)的摩尔比给予患者，优选以1:(0.9-3.0)的摩尔比，更优选以1:(1.0-1.5)的摩尔比，最优选约1:1的摩尔比给予患者。
在治疗过程中，可以根据情况单独或与一种或多种其它的治疗剂组合使用。可以通过常规的注射、口服、吸入、直肠和经皮施用中的至少一种将包含本发明的药物辅料和药物活性物质的药物施用给患者。其他可以适用的给药方式还包括，但不限于耳廓给药、关节内给药、肌肉内给药、鞘内给药、体外给药、颊部给药、支气管内给药、结膜给药、皮肤上给药、牙齿给药、子宫颈内给药、鼻窦内给药、气管内给药、肠内给药、硬膜外给药、腹内给药、胆内给药、囊内给药、口咽给药、间质给药、心脏内给药、软骨内给药、海绵体内给药、脑内给药、腔内给药、角膜内给药、颅内给药、颅骨给药、皮内给药、病灶内给药、鼓室内给药、牙龈内给药、卵巢内给药、眼内给药、椎间盘内给药、管内给药、十二指肠内给药、眼给药、硬膜内给药、表皮内给药、食管内给药、鼻饲给药、鼻腔给药、喉给药、心室内给药、胃内给药、肝内给药、管腔内给药、玻璃体内给药、淋巴内给药、乳房内给药、骨髓内给药、窦内给药、脑膜内给药、结内给药、肺内给 药、心包内给药、腹膜内给药、胸膜内给药、前列腺内给药、脊柱内给药、滑膜内给药、腱内给药、睾丸内给药、结膜下给药、脑室内给药、静脉内给药、眼球后给药、关节周给药、胸内给药、蛛网膜下给药、牙周给药、鼓膜给药、经气管给药、肿瘤内给药、阴道给药、输尿管给药、子宫内给药、口服给药、肠胃给药、肠胃外给药、舌下给药、经皮给药、跨粘膜给药、神经周给药、透皮给药、直肠给药、软组织给药、动脉内给药、皮下给药、局部给药、羊膜外给药、膀胱内输注等。
在本发明的实施方式中，在根据本发明对患者进行治疗时，给定药物的量取决于诸多因素，如具体的给药方案、疾病或病症类型及其严重性、需要治疗的受治疗者或宿主的独特性(例如体重)。但是，根据特定的周围情况，包括例如已采用的具体药物、给药途径、治疗的病症、以及治疗的受治疗者或宿主，施用剂量可由本领域已知的方法常规决定。通常，就成人治疗使用的剂量而言，施用剂量典型地在0.02-5000mg/天，例如约1-1500mg/天的范围。该所需剂量可以方便地被表现为一剂、或同时给药的(或在短时间内)或在适当的间隔的分剂量，例如每天二、三、四剂或更多分剂。本领域技术人员可以理解的是，尽管给出了上述剂量范围，但具体的有效量可根据患者的情况并结合医师诊断而适当调节。
除了能够在乏氧环境下靶向给药和提供低毒性之外，本发明的磺化偶氮杯芳烃化合物作为载药分子容器平台的优势还在于：其在溶液特别是水溶液中具有高溶解度，并且对于难溶药物具备额外的增溶效果，因此在用作药物递送平台时不会由于溶解度的限制而束缚活性药物的给药浓度，能够为给药浓度的选择提供更高的自由度。
为了增加活性药物在溶液中的溶解度，可以通过将活性药物与本发明的磺化偶氮杯芳烃化合物共研磨后溶解、或在溶液中共同超声、震荡、回流等方式而实现。
偶氮杯芳烃化合物的制备
使用本领域技术人员已知的标准合成技术或使用本领域已知的方法与本文描述的方法组合，可以合成式(I)的化合物。另外，本文给出 的溶剂、温度和其它反应条件可以根据本领域技术而改变。作为进一步的指导，也可以利用以下的合成方法。
所述反应可以按顺序使用，以提供本文描述的化合物；或它们可以用于合成片段，所述片段通过本文描述的方法和/或本领域已知的方法随后加入。
可以使用与下述类似的方法，通过使用适当的可选择的起始原料，合成化合物。用于合成本文描述的化合物的起始原料可以被合成或可以从商业来源获得。本文描述的化合物和其它相关具有不同取代基的化合物可以使用本领域技术人员已知的技术和原料合成。制备本文公开的化合物的一般方法可以来自本领域已知的反应，并且该反应可以通过由本领域技术人员所认为适当的试剂和条件修改，以引入本文提供的分子中的各种部分。
如果需要，反应产物可以使用常规技术分离和纯化，包括但不限于过滤、蒸馏、结晶、色谱等方法。这些产物可以使用常规方法表征，包括物理常数和图谱数据。
本发明的偶氮杯芳烃化合物的通用合成路线如下。
Figure PCTCN2022071935-appb-000006
SAC4A：将对氨基苯磺酸(5.19g，30.0mmol)和碳酸钠(1.59g，15.0mmol)溶于30mL水中，加热至溶解。稍冷却后加入亚硝酸钠(2.13g，31.5mmol)的24mL水溶液，冰浴冷却下滴加至10.5mL浓盐酸中。将所得的液体在冰浴条件下滴加至含有杯[4]芳烃(杯[n]芳烃CnA，n为4-8，的合成参照文献Org.Synth.1990,68,238；J.Am.Chem.Soc.1982,104,2652；J.Org.Chem.1998,63,489)(3.00g，7.2mmol)和三水合乙酸钠(12.24g，90.0mmol)的78mL甲醇与DMF的混合溶液(v:v＝5:8)中。冰浴2小时，逐渐升温至室温20摄氏度。加入稀盐酸(30mL HCl+450mL H 2O)调至pH＝2，加热至60摄氏度，持续30分钟。减压蒸去甲醇、水、DMF。残留固体利用水-甲醇体系在回流条件下重结晶，冷 却，抽滤，干燥得到产物。 1H NMR(400MHz,DMSO-d 6,δ):8.06(d,J＝8.06Hz,8H,ArH),7.87(m,16H,ArH),4.41(s,8H,Ar-CH 2-Ar)。
SAC5A：将对氨基苯磺酸(1.73g，10.0mmol)和碳酸钠(0.53g，5.0mmol)溶于10mL水中，加热至溶解。稍冷却后加入亚硝酸钠(0.71g，10.5mmol)的8mL水溶液，冰浴冷却下滴加至3.5mL浓盐酸中。将所得的液体在冰浴条件下滴加至含有杯[5]芳烃(1.01g，1.9mmol)和三水合乙酸钠(4.08g，30.0mmol)的26mL甲醇与DMF的混合溶液(v:v＝5:8)中。冰浴2小时，逐渐升温至室温20摄氏度。加入稀盐酸(1mL HCl+149mL H 2O)调pH至酸性，加热至60摄氏度，持续30分钟。减压蒸去甲醇、水、DMF。残留固体利用水-甲醇体系在回流条件下重结晶，冷却，抽滤，干燥得到产物。 1H NMR(400MHz,DMSO-d 6,δ):7.85-7.70(m,30H,ArH),4.41(s,10H,Ar-CH 2-Ar)。
SAC6A：将对氨基苯磺酸(1.73g，10.0mmol)和碳酸钠(0.53g，5.0mmol)溶于10·mL水中，加热至溶解。稍冷却后加入亚硝酸钠(0.71g，10.5mmol)的8mL水溶液，冰浴冷却下滴加至3.5mL浓盐酸中。将所得的液体在冰浴条件下滴加至含有杯[6]芳烃(1.02g，1.6mmol)和三水合乙酸钠(4.08g，30.0mmol)的26mL甲醇与DMF的混合溶液(v:v＝5:8)中。冰浴2小时，逐渐升温至室温20摄氏度。加入稀盐酸(1mL HCl+149mL H 2O)调pH至酸性，加热至60摄氏度，持续30分钟。减压蒸去甲醇、水、DMF。残留固体利用水-甲醇体系在回流条件下重结晶，冷却，抽滤，干燥得到产物。 1H NMR(400MHz,DMSO-d 6,δ):7.84-7.74(m,36H,ArH),4.04(s,12H,Ar-CH 2-Ar)。
SAC8A：将对氨基苯磺酸(0.87g，5.0mmol)和碳酸钠(0.27g，2.5mmol)溶于5mL水中，加热至溶解。稍冷却后加入亚硝酸钠(0.36g，5.3mmol)的4mL水溶液，冰浴冷却下滴加至10.5mL浓盐酸中。将所得的液体在冰浴条件下滴加至含有杯[8]芳烃(0.51g，0.6mmol)和三水合乙酸钠(2.04g，15.0mmol)的13mL甲醇与DMF的混合溶液(v:v＝5:8)中。冰浴2小时，逐渐升温至室温20摄氏度。加入稀盐酸(0.5·mL HCl+75·mL H 2O)调至pH酸性，加热至60摄氏度，持续30分钟。减压蒸去甲醇、水、DMF。残留固体利用水-甲醇体系在回流条件下重结晶，冷却，抽滤，干燥得到产物。1H NMR(400MHz,DMSO-d 6,δ):7.722(m, 48H,ArH),4.08(s,16H,Ar-CH 2-Ar)。
作为对比例，另合成CAC4A。
Figure PCTCN2022071935-appb-000007
将2.5mmol对氨基苯甲酸固体与500μL浓盐酸分别加入3.75mL水中，将所得溶液放置到冰盐浴中，冷却到2℃。将2.70g(40mmol)亚硝酸钠溶解于2.5mL水中，将溶液逐滴滴加到对氨基苯甲酸和盐酸的混合液中。反应放热，为保证重氮盐稳定，反应过程中控制温度于5℃以下，搅拌30分钟，可见溶液变为淡黄色，得到溶液1。将1mmol杯[4]芳烃与4.08g(30mmol)三水合乙酸钠溶于26mL甲醇与DMF的混合溶液(v:v＝5:8)中，置于冰水浴中冷却至5℃以下，得到溶液2。将溶液1缓慢逐滴滴加到溶液2中，滴加过程中控制温度不超过5℃。滴加完成后，在5℃下搅拌15分钟，转移至室温环境下，继续搅拌2.5小时，静置待悬浊液沉降完全后向其中加入37.5mL稀盐酸(0.25％)，搅拌并于60℃加热30分钟。过滤后用蒸馏水和MeOH洗涤数次，得到红色固体即为粗产物。将粗产物溶于25mL的热碳酸氢钠(0.5M)溶液中，过滤除去滤渣。冷却后向滤液中加入3mL浓盐酸，并于60℃下加热搅拌30分钟，过滤，滤饼蒸馏水洗涤数次后，放入真空干燥箱中干燥过夜，得到纯化产物。 1H NMR(400MHz,DMSO-d 6)δ8.02(d,8H,J＝8.6Hz,Ar-H),7.81(d,8H,J＝8.6Hz,ArH),7.80(s,8H,calix-H),4.37and 3.72(s,8H,Ar-CH 2-Ar)；MS(MALDI-TOF):calcd.for C 56H 39N 8O 12 -[M-H] -1015.269,found 1015.195。
测试实施例
实施例1：磺化偶氮杯芳烃化合物与药物分子键合常数的测定
测试方法：荧光滴定法。
测试工具：测试选用石英比色皿为样品池，测试光路10mm，仪器型号为Varian Cary Eclipse，配有型号为Cary Single-cuvette Peltier 的比色皿控温装置。
试剂：罗丹明B购置于上海麦克林生化科技有限公司。紫杉醇(PTX)购自美仑生物，其他活性药物均为市售产品。
偶氮杯芳烃和活性药物分子的荧光滴定实验均在室温(25℃)进行。以磺化偶氮杯芳烃化合物SAC5A为例。首先配制SAC5A、罗丹明B(RhB)的母液，将其分别溶于磷酸盐缓冲溶液(PBS，10mM，pH＝7.4)中，配置浓度均为100μM。测试时先将SAC5A-RhB(0.4/0.3μM)荧光传感对配置于荧光池内，PBS定容到体积2.5mL。将PTX溶于甲醇，配置浓度1mM。随后将紫杉醇分子溶液以预定体积加入荧光池内，记录荧光强度变化。紫杉醇加入体积小于总体积的0.8％，荧光滴定数据由主客体1:1竞争键合模型进行拟合，激发波长554nm，测定主客体包结的键合常数K a。结果示于下表1。
表1.磺化偶氮杯芳烃对PTX的键合常数
Figure PCTCN2022071935-appb-000008
采用同样的方法检测各类活性药物分子与SAC5A的键合常数。具体而言，SAC5A和活性药物分子的荧光滴定实验均在室温(25℃)进行。首先配制SAC5A、罗丹明B(RhB)的母液，将其分别溶于磷酸盐缓冲溶液(PBS，10mM，pH＝7.4)中，配置浓度均为100μM。测试时先将SAC5A-RhB(0.4/0.3μM)荧光传感对配置于荧光池内，PBS定容到体积2.5mL。将活性药物分子溶于磷酸盐缓冲溶液(PBS，10mM，pH＝7.4)，配置浓度100μM，并向其加入荧光传感对，使荧光传感对浓度与荧光池内一致，PBS定容至1mL。随后将活性药物分子溶液以预定体积加入荧光池内，记录荧光强度变化。荧光滴定数据由主客体1:1竞争键合模型进行拟合，测定主客体包结的键合常数K a。各种活性药物分子与SAC5A的键合常数测试结果如下表2所示。
表2.活性药物分子与SAC5A的键合常数
Figure PCTCN2022071935-appb-000009
Figure PCTCN2022071935-appb-000010
主客体的键合常数越大，说明SAC5A与药物的包结能力越强，形成的包合物越稳定，药物越不容易泄露。如实验结果所示，本发明的偶氮杯芳烃化合物与活性药物分子具有较强的结合强度，键合常数达到10 3以上，优选10 4以上，更优选10 5以上，再更优选10 6以上，特别优选10 7以上。因此，本发明的磺化偶氮杯芳烃化合物能够与活性药物分子形成稳定的主客体非共价结合。
实施例2：连二亚硫酸钠(SDT)还原实验
测试方法：紫外-可见分光光谱法。
测试工具：日本岛津UV-3600紫外-可见分光光度计，配有控温模块(型号：PTC-348WI)，测试样品选用岛津自带石英比色皿，光程10mm。荧光仪器型号为Varian Cary Eclipse，配有型号为Cary Single-cuvette Peltier的比色皿控温装置。测试选用石英比色皿为样品池，测试光路10mm。
首先配制SAC4A的母液，将其溶于磷酸盐缓冲溶液(PBS，10mM，pH＝7.4)中，配制浓度为100μM。取SAC4A母液稀释到10μM SAC4A，体积2.5mL，测试420nm处紫外吸收随时间变化，2分钟时加入1.0mM SDT，测试结果如图2所示。图2内部是加入SDT前后，SAC4A紫外吸收曲线。从图2中看出，加入SDT后，SAC4A中偶氮键对应的紫外吸收随时间逐渐下降，说明SAC4A能被SDT还原，具有乏氧响应性。
结果显示，磺化偶氮杯芳烃化合物中加入过量的偶氮还原酶的模 拟分子连二亚硫酸钠(SDT)后，偶氮吸收消失，表明SAC4A的四个偶氮基团都被完全还原。偶氮键在过表达的偶氮还原酶作用下被还原断裂，药物活性物质可以被原型释放出来，实现了药物的靶向性，减少了给药量，提高了安全性。
实施例3：偶氮杯芳烃化合物的溶解度测试
测试方法：紫外-可见分光光谱法。
测试工具：日本岛津UV-3600紫外-可见分光光度计，配有控温模块(型号：PTC-348WI)，测试样品选用岛津自带石英比色皿，光程10mm。荧光仪器型号为Varian Cary Eclipse，配有型号为Cary Single-cuvette Peltier的比色皿控温装置。测试选用石英比色皿为样品池，测试光路10mm。
以SAC5A为例，首先测试SAC5A的紫外标准曲线，配置一系列SAC5A的标准溶液，浓度分别为：1μM、2μM、4μM、6μM、8μM，得到标准曲线。配制过饱和SAC5A的溶液，将其溶于磷酸盐缓冲溶液(PBS，10mM，pH＝7.4)中，震荡过夜。将溶液以转速12000rpm离心5分钟，取上清液稀释2万倍于比色皿中，体积为2.5mL，测试紫外吸收强度，代入标准曲线计算可得SAC5A的溶解度。
通过同样的方法测试SAC4A、SAC6A、SAC8A和对比化合物CAC4A的溶解度，结果列于下表3。
表3. 25℃，PBS缓冲液(10mM，pH7.4)中各种杯芳烃的溶解度
Figure PCTCN2022071935-appb-000011
与对比杯芳烃化合物CAC4A相比，本发明的磺化偶氮杯芳烃化合物的溶解度显著提高。因此，本发明的磺化偶氮杯芳烃化合物在用作药物递送平台时不会由于溶解度的限制而束缚活性药物的给药浓度， 能够为给药浓度的选择提供更高的自由度。
以化疗药物喜树碱为例，其常用静脉给药浓度(每10克小鼠注射100微升)为1.44-2.87mM。偶氮杯芳烃化合物作为药物载体与化疗药物结合的摩尔浓度比至少为1:1，羧基偶氮杯芳烃CAC4A自身的溶解度限制了活性化合物的给药剂量，因此选择本发明的磺化偶氮杯芳烃化合物具有显著的优势。
实施例4：磺化偶氮杯芳烃化合物对活性药物分子的增溶实验
按相同摩尔浓度分别称取PTX固体和SAC5A固体于研钵研磨15分钟，加入定量PBS(10mM，pH7.4)溶解即得到澄清溶液。按相同方法制备一系列浓度样品，分别为：0.1mM、0.5mM、1.0mM、2.0mM、5.0mM、10.0mM，拍照，如图3a所示。样品放置2小时后取上清液使用液相色谱分离检测。液相条件为：Waters 2695 HPLC(Waters，Alliance 2695，USA)，在37℃下于
Figure PCTCN2022071935-appb-000012
色谱柱(4.6mm×250mm，5μm)上进行色谱分离，采用水(A)和甲醇(B)等度洗脱。流动相的流速设置为1.0mL min -1，进样量为10μL，检测波长为232nm。
以SAC5A浓度为横轴，以最终增溶的PTX浓度为纵轴得到相溶解度曲线图，如图3b所示。
结果显示，本发明的磺化偶氮杯芳烃化合物SAC5A对于难溶水的药物具有显著的增溶效果，因此对于难溶性药物分子可进行增溶，提升药物利用率及安全性，对于水溶性药物则通过强包结作用降低副作用，增强治疗效果。
实施例5：载体安全性评估
选取雌性Balb/c、6-8周小鼠(购买于北京斯贝福公司)，随机分为PBS和SAC5A两组，每组3只小鼠，每只小鼠重约20克。采取尾静脉注射给药，给药剂量按浓度1mM计，每次给药200μL，SAC5A的药物剂量是15.61mg/kg，隔天注射四次。安全性实验根据南京建成生物工程研究所提供的试剂盒测定。血生化测试结果参见图4a-4f。
各缩写字母代表含义如下：
Alanine aminotransferase(ALT)谷丙转氨酶
Aspartate transaminase(AST)谷草转氨酶
Alkaline phosphatase(ALP)碱性磷酸(酯)酶
Blood urea nitrogen(BUN)血尿素氮
Lactate dehydrogenase(LDH)乳酸脱氢酶
C-reactive protein(CRP)C反应蛋白
如图4a-4f所示，SAC5A各项指标与PBS组比较没有显著差异，证明其安全性较高。
安全性试验给药结束后，将小鼠解剖取出心脏、肝脏、脾脏、肺脏、肾脏。各器官组织置于4％多聚甲醛(sigma)中固定24小时后，送天津易生源生物技术有限公司进行石蜡切片及H&E染色实验。随后进行显微镜分析。
如图5所示，组织病理学检查显示出与血生化检测相似的结论，H&E病理切片显示SAC5A的心脏、肝脏、脾脏、肺脏和肾脏等主要器官与对照组相比保留了相似的形态，表明没有可检测到的副作用。
实施例6：递送体系安全性评估
选取雌性Balb/c、6-8周小鼠(购买于北京斯贝福公司)，随机分为PBS、PTX、SAC5A、PTX/SAC5A四组，每组3只小鼠，每只小鼠重约20克。采取尾静脉注射给药，给药剂量按PTX剂量5mg/kg计，每次给药100μL，各组药物浓度分别是SAC5A(2.00mM)，PTX/SAC5A(1.17/1.40mM)，PTX(1.17mM)，隔天注射四次。安全性实验根据南京建成生物工程研究所提供的试剂盒测定。血常规测试结果参见图6a和6b。数据代表三个独立实验(n＝3)的平均值±标准差(s.d.)，与PBS相比，显著性水平为*P<0.05，***P<0.001。
各缩写字母代表含义如下：
lymphocyte(Lymph)淋巴细胞
white blood cell(WBC)白细胞
PTX组Lymph、WBC指标高于PBS组，说明游离PTX会引发炎症反应，而SAC5A和PTX/SAC5A各项指标与PBS组比较没有显著性差异，证明其安全性较高。
实施例7：动物抑瘤实验
将1×10 6个4T1细胞皮下注射到6-8周的雌性Balb/C小鼠腋下，等到肿瘤体积大小长到100mm 3。肿瘤大小通过游标卡尺测量，并使用以下公式计算肿瘤体积：V＝W 2×L/2，其中W和L分别是肿瘤的最短和最长直径。将小鼠随机分为PBS、SAC5A(10.93mg/kg)、PTX(5.00mg/kg)、SAC5A/PTX(PTX：5.00mg/kg，SAC5A：10.93mg/kg)、SAC5A/PTX(PTX：2.50mg/kg，SAC5A：5.46mg/kg)、Lipo-PTX(力扑素(注射用紫杉醇脂质体)，购自南京绿叶制药有限公司，5.00mg/kg，按PTX算)、HSA-PTX(注射用紫杉醇(白蛋白结合型)，购自石药集团欧意药业有限公司，5.00mg/kg，按PTX算)七组，每组6只小鼠。尾静脉注射各组药物，每次注射前测量小鼠肿瘤长短直径。隔天给药，给药5次共9天。每次给药100μL，各组药物浓度分别是SAC5A(1.40mM)、PTX(1.17mM)、PTX/SAC5A(1.17/1.40mM)、PTX/SAC5A(0.59/0.70mM)、Lipo-PTX(1.17mM)、HSA-PTX(1.17mM)。从给药第一天开始测量小鼠肿瘤体积及小鼠体重，结果分别示于图7和图8。
如图7所示，与对照组相比，PTX/SAC5A组肿瘤生长速度降低，停止给药后有短期抑制作用，证明PTX/SAC5A具有良好的抑瘤效果。监测小鼠体重可知，PTX/SAC5A组无显著毒性。
不同治疗组的荷瘤小鼠的生存率示于图9。如图9所示，PTX/SAC5A组小鼠生存率高于其他组，证明PTX/SAC5A可以显著提升生存率，具有长期治疗的预后效果。
给药后持续监测小鼠生存情况，肿瘤体积达到2000mm 3默认小鼠死亡，小鼠死亡后取出肿瘤用于切片染色分析。
TUNLE染色步骤：将小鼠肿瘤进行冰冻切片处理，PBS湿润15分钟后遵循Roach公司提供的实验手册进行染色。
Ki67染色步骤：将冰冻切片从-80℃取出，恢复至室温，0.1％Triton X-100处理15分钟后，PBS洗掉Triton，5％BSA(Sigma)封闭1小时后加入一抗(Ki67一抗，大鼠来源，索莱宝)，4℃避光过夜。12小时后PBS洗掉一抗，然后加入荧光标记的二抗(驴抗兔，alexa Fluro488，索莱宝)。1小时后封片共聚焦显微镜拍照分析。
H&E染色步骤：1)石蜡切片脱蜡至水：依次将切片放入二甲苯Ⅰ30分钟-二甲苯Ⅱ30分钟-无水乙醇Ⅰ10分钟-无水乙醇Ⅱ10分钟-95％酒精5分钟-90％酒精5分钟-80％酒精5分钟-70％酒精5分钟-蒸馏水洗；2)苏木素染细胞核：切片入Harris苏木素染5-10分钟，自来水洗，1％的盐酸酒精分化数秒，自来水冲洗10分钟，PBS返蓝5分钟，流水冲洗；3)伊红染细胞质：切片入伊红染液中染色1-3分钟；4)脱水封片：将切片依次放入95％酒精I 5分钟-95％酒精II 5分钟-无水乙醇Ⅰ5分钟-无水乙醇Ⅱ5分钟-二甲苯Ⅰ5分钟-二甲苯Ⅱ5分钟中脱水透明，将切片从二甲苯拿出来稍晾干，中性树胶封片；5)显微镜镜检，图像采集分析。
如图10所示，在Tunle染色照片中，荧光信号标记断裂DNA，荧光信号越强，细胞凋亡程度越高，PTX/SAC5A组细胞核所染上的荧光信号最强，因此，PTX/SAC5A组细胞凋亡程度明显高于其它三组。
Ki67染色照片中，Ki67标记细胞增殖相关抗原。PTX/SAC5A组细胞增殖程度低于对照组。
如图10的H&E所示，由20×显微镜观察到PTX/SAC5A组细胞核固缩，核质分离，细胞核外流。PTX/SAC5A组细胞坏死程度明显高于其他三组，证明PTX/SAC5A组具有良好的杀伤肿瘤的效果。
以上染色照片均证明PTX/SAC5A具有良好的肿瘤杀伤效果。
工业应用性
本发明提供一种磺化偶氮杯芳烃超分子化合物作为乏氧响应的靶向药物辅料。本发明的化合物可以与适当的活性药物分子一起制成相应的药物组合物，适于工业应用。
尽管本文对本发明作了详细说明，但本发明不限于此，本技术领域的技术人员可以根据本发明的原理进行修改，因此，凡按照本发明的原理进行的各种修改都应当理解为落入本发明的保护范围。

Claims (15)

  1. 一种药物组合物，包括至少一种药物活性物质和式(I)的偶氮杯芳烃化合物：
    Figure PCTCN2022071935-appb-100001
    其中，
    n为4至8的整数，
    M独立地选自H、Na和K，
    所述至少一种药物活性物质选自治疗以下一种或多种疾病的药物：癌症、心肌梗塞、中风、动脉粥样硬化、类风湿性关节炎、炎症性肠病、慢性缺氧性肺病和慢性肾病。
  2. 如权利要求1所述的药物组合物，其中所述至少一种药物活性物质为治疗癌症的药物。
  3. 如权利要求1或2所述的药物组合物，其中M为Na。
  4. 如权利要求1-3中任一项所述的药物组合物，其中所述式(I)的偶氮杯芳烃化合物选自：
    Figure PCTCN2022071935-appb-100002
  5. 如权利要求1-4中任一项所述的药物组合物，其中所述药物活性物质选自以下一种或多种：选自阿霉素、紫杉醇、喜树碱、羟基喜 树碱、伊立替康、拓扑替康、贝洛替康、奥沙利铂、氯吡格雷、依利罗地、羟基氯喹、洛伐他汀、他达拉非、替拉替尼、乐伐替尼、索拉非尼、瑞戈非尼、吉非替尼和甲氧雌二醇的治疗癌症的药物；选自阿司匹林、替格瑞洛、比索洛尔和氯吡格雷的治疗心肌梗塞的药物；选自阿司匹林、肝素、阿替普酶、尼莫地平、罗吡唑、替拉扎特和依利罗地的治疗中风的药物；选自洛伐他汀和依泽替米贝的治疗动脉粥样硬化的药物；选自甲氨蝶呤、托法替尼、巴瑞替尼、依那西普、阿巴西普、来氟米特、羟基氯喹和柳氮磺胺吡啶的治疗类风湿性关节炎的药物；选自美沙拉秦、硫唑嘌呤、巯嘌呤、甲氨蝶呤、英夫利昔单抗、阿达木单抗、赛妥珠单抗、那他珠单抗和沙利度的治疗炎症性肠病的药物；选自他达拉非、利奥西呱和曲前列环素的治疗慢性缺氧性肺病的药物；选自罗沙司他、双嘧达莫、氢氯噻嗪和维达司他的治疗慢性肾病的药物。
  6. 如权利要求1-5中任一项所述的药物组合物，其中所述药物活性物质与式(I)的偶氮杯芳烃化合物的摩尔比为1:(0.8-5.0)，优选为1:(0.9-3.0)，更优选为1:(1.0-1.5)。
  7. 式(I)的偶氮杯芳烃化合物，用于制备乏氧响应的靶向药物辅料的用途，
    Figure PCTCN2022071935-appb-100003
    其中，
    n为4至8的整数，
    M独立地选自H、Na和K。
  8. 如权利要求7所述的式(I)的偶氮杯芳烃化合物的用途，其中M 为Na。
  9. 如权利要求7或8所述的式(I)的偶氮杯芳烃化合物的用途，其中所述式(I)的偶氮杯芳烃化合物选自：
    Figure PCTCN2022071935-appb-100004
    Figure PCTCN2022071935-appb-100005
  10. 如权利要求7-9中任一项所述的式(I)的偶氮杯芳烃化合物的用途，其中所述乏氧响应的靶向药物是选自治疗以下一种或多种疾病的药物：癌症、心肌梗塞、中风、动脉粥样硬化、类风湿性关节炎、炎症性肠病、慢性缺氧性肺病和慢性肾病。
  11. 如权利要求10所述的式(I)的偶氮杯芳烃化合物的用途，其中所述乏氧响应的靶向药物选自以下一种或多种：选自阿霉素、紫杉醇、喜树碱、羟基喜树碱、伊立替康、拓扑替康、贝洛替康、奥沙利铂、氯吡格雷、依利罗地、羟基氯喹、洛伐他汀、他达拉非、替拉替尼、乐伐替尼、索拉非尼、瑞戈非尼、吉非替尼和甲氧雌二醇的治疗癌症的药物；选自阿司匹林、替格瑞洛、比索洛尔和氯吡格雷的治疗心肌梗塞的药物；选自阿司匹林、肝素、阿替普酶、尼莫地平、罗吡唑、替拉扎特和依利罗地的治疗中风的药物；选自洛伐他汀和依泽替米贝的治疗动脉粥样硬化的药物；选自甲氨蝶呤、托法替尼、巴瑞替尼、依那西普、阿巴西普、来氟米特、羟基氯喹和柳氮磺胺吡啶的治疗类风湿性关节炎的药物；选自美沙拉秦、硫唑嘌呤、巯嘌呤、甲氨蝶呤、 英夫利昔单抗、阿达木单抗、赛妥珠单抗、那他珠单抗和沙利度的治疗炎症性肠病的药物；选自他达拉非、利奥西呱和曲前列环素的治疗慢性缺氧性肺病的药物；选自罗沙司他、双嘧达莫、氢氯噻嗪和维达司他的治疗慢性肾病的药物。
  12. 式(I)的偶氮杯芳烃化合物，用于治疗以下一种或多种疾病的用途：癌症、心肌梗塞、中风、动脉粥样硬化、类风湿性关节炎、炎症性肠病、慢性缺氧性肺病和慢性肾病，
    Figure PCTCN2022071935-appb-100006
    其中，
    n为4至8的整数，
    M独立地选自H、Na和K；
    所述用途包括将所述偶氮杯芳烃化合物作为药物辅料，与用于治疗所述一种或多种疾病的药物活性物质混合，并施用给有需要的患者。
  13. 如权利要求12所述的式(I)的偶氮杯芳烃化合物的用途，其中M为Na。
  14. 如权利要求12或13所述的式(I)的偶氮杯芳烃化合物的用途，其中所述式(I)的偶氮杯芳烃化合物选自：
    Figure PCTCN2022071935-appb-100007
    Figure PCTCN2022071935-appb-100008
  15. 如权利要求12-14中任一项所述的式(I)的偶氮杯芳烃化合物的用途，其中所述药物活性物质选自以下一种或多种：选自阿霉素、紫杉醇、喜树碱、羟基喜树碱、伊立替康、拓扑替康、贝洛替康、奥沙利铂、氯吡格雷、依利罗地、羟基氯喹、洛伐他汀、他达拉非、替拉替尼、乐伐替尼、索拉非尼、瑞戈非尼、吉非替尼和甲氧雌二醇的治疗癌症的药物；选自阿司匹林、替格瑞洛、比索洛尔和氯吡格雷的治疗心肌梗塞的药物；选自阿司匹林、肝素、阿替普酶、尼莫地平、罗吡唑、替拉扎特和依利罗地的治疗中风的药物；选自洛伐他汀和依泽替米贝的治疗动脉粥样硬化的药物；选自甲氨蝶呤、托法替尼、巴瑞替尼、依那西普、阿巴西普、来氟米特、羟基氯喹和柳氮磺胺吡啶的治疗类风湿性关节炎的药物；选自美沙拉秦、硫唑嘌呤、巯嘌呤、甲氨蝶呤、英夫利昔单抗、阿达木单抗、赛妥珠单抗、那他珠单抗和沙利度的治疗炎症性肠病的药物；选自他达拉非、利奥西呱和曲前列环素的治疗慢性缺氧性肺病的药物；选自罗沙司他、双嘧达莫、氢氯噻嗪和维达司他的治疗慢性肾病的药物。
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