WO2023061393A1 - 包含抗肿瘤药物的组合物及其制备方法和用途 - Google Patents

包含抗肿瘤药物的组合物及其制备方法和用途 Download PDF

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WO2023061393A1
WO2023061393A1 PCT/CN2022/124787 CN2022124787W WO2023061393A1 WO 2023061393 A1 WO2023061393 A1 WO 2023061393A1 CN 2022124787 W CN2022124787 W CN 2022124787W WO 2023061393 A1 WO2023061393 A1 WO 2023061393A1
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albumin
composition
content
cholesterol
lipid
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PCT/CN2022/124787
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English (en)
French (fr)
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柯天一
檀琳
温佰刚
劳芳
刘岩
王宁霞
居尚
崔冬雪
杜旭召
欧阳芳幸
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昆山新蕴达生物科技有限公司
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Priority to AU2022367142A priority Critical patent/AU2022367142A1/en
Priority to KR1020247015421A priority patent/KR20240099264A/ko
Priority to CN202280025871.1A priority patent/CN117098536A/zh
Priority to JP2024502452A priority patent/JP2024533941A/ja
Priority to MX2024003496A priority patent/MX2024003496A/es
Priority to EP22880329.2A priority patent/EP4417204A1/en
Priority to CA3231432A priority patent/CA3231432A1/en
Priority to CN202311513706.3A priority patent/CN117547535A/zh
Publication of WO2023061393A1 publication Critical patent/WO2023061393A1/zh
Priority to US18/436,704 priority patent/US20240252482A1/en

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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
    • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
    • A61K47/42—Proteins; Polypeptides; Degradation products thereof; Derivatives thereof, e.g. albumin, gelatin or zein
    • 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
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/4738—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/4745—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems condensed with ring systems having nitrogen as a ring hetero atom, e.g. phenantrolines
    • 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
    • A61K47/22—Heterocyclic compounds, e.g. ascorbic acid, tocopherol or pyrrolidones
    • 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
    • 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
    • A61K47/28—Steroids, e.g. cholesterol, bile acids or glycyrrhetinic acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/19—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles lyophilised, i.e. freeze-dried, solutions or dispersions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A61K9/5107—Excipients; Inactive ingredients
    • A61K9/5123—Organic compounds, e.g. fats, sugars
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A61K9/5107—Excipients; Inactive ingredients
    • A61K9/513—Organic macromolecular compounds; Dendrimers
    • A61K9/5169—Proteins, e.g. albumin, gelatin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A61K9/5192—Processes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • This application relates to the composition of 7-ethyl-10-hydroxycamptothecin (SN-38), which comprises SN-38, lipid, albumin and Span 20, and also relates to its preparation method and use.
  • SN-38 7-ethyl-10-hydroxycamptothecin
  • SN-38 is the active metabolite of the marketed drug irinotecan hydrochloride (CPT-11) in vivo, and its efficacy on some tumor cells is about 100-1000 times that of CPT-11 (Zhang J A, Xuan T, Parmar M, et al. Development and characterization of a novel liposome-based formulation of SN-38, [J]. International journal of pharmaceuticals, 2004, 270(1): 93-107).
  • SN-38 has inhibitory effects on a variety of tumor cells, such as colorectal cancer, small cell lung cancer, lymphoma, breast cancer, esophageal cancer, uterine cancer, ovarian cancer, etc.
  • the modification can be divided into water-soluble modification and fat-soluble modification.
  • the former can be found in, for example, WO1995022549A1, and the latter can be found in, for example, US20060229359A.
  • the method of fat-soluble modification generally involves preparing SN-38 into liposomes.
  • the preparation process generally includes modifying camptothecin molecules with hydrophobic molecules such as long-chain fatty acids or cholesterol, vitamin E, and lipophilic organic acids, and then mixing them with a certain proportion Excipients such as phospholipids and cholesterol are prepared into camptothecin-liposome products.
  • CN108567742A did not carry out SN-38 hydrophobic modification, but obtained camptothecin-liposome product by improving auxiliary materials. None of the above-mentioned reported products contain albumin, but lipid modification allows camptothecin to maintain a larger proportion of closed-loop active structures, so that it can be entrapped by albumin after entering the body, thereby prolonging the drug effect.
  • SN-38 compared with paclitaxel drugs, has more special physical and chemical properties, such as easier crystallization in water, positive charge on the surface, etc.
  • the stability of the preparation process and the expansion of the process have always been the pain points of its drug production.
  • the prepared nanoliposomes and nanosuspensions all have unsatisfactory drug loading of SN-38, complicated preparation process, or poor repeatability after process amplification. Disadvantages of formulation instability. How to obtain SN-38 nano-preparation with higher drug loading capacity, easy and stable process scale-up, and cost-effective SN-38 nano-preparation by optimizing the formulation composition and process is still an urgent problem to be solved.
  • PCT/CN2021/102332 The inventor's previous application (PCT/CN2021/102332) described a composition comprising SN-38, lipid and albumin with a high drug loading, and obtained a SN-38 preparation suitable for drug production.
  • adding Span 20 to the composition can reduce the number of high-pressure homogenization in the preparation process, effectively reduce the particle size of the preparation, improve filtration flux, reduce raw material loss and The cost is reduced, and the stability of the pharmaceutical preparation obtained at the same time is better, the particle size control after disintegration is better, and the drug effect is more stable.
  • the content of PCT/CN2021/102332 is incorporated herein by reference in its entirety.
  • compositions comprising SN-38, lipids and albumin.
  • the inventors have found that the addition of Span 20 to the composition when prepared on a larger scale (e.g., hundreds of milligrams or more of SN-38 starting material), including in scale-up processes, such as pilot-scale preparations, can Reduce the number of high-pressure homogenization in the preparation process, effectively reduce the particle size of nanoparticles in the composition, improve filtration flux, reduce raw material loss and reduce costs, and at the same time control the particle size of nanoparticles after disintegration; through further control The content of the albumin in the composition can control the size of the nanoparticles, making it closer to the size suitable for making medicines.
  • composition of the present invention also has the advantages of the composition of PCT/CN2021/102332, including: (1) increased drug loading and encapsulation efficiency of SN-38; (2) SN-38 containing a low level of ring-opening structure (3) does not contain albumin polymers, low immunogenicity, high safety; (4) the particle size of nanoparticles is small, and the particle size distribution is narrow; (5) excellent stability (including excellent dilution stability and storage stability); (6) SN-38 exists in amorphous form and/or nanocrystals, which has the advantages of fast dissolution rate and high bioavailability; and (7) excellent antitumor efficacy in vivo.
  • the present application provides a composition comprising SN-38, lipids, albumin and Span 20, characterized in that the composition comprises nanoparticles, wherein in the nanoparticles, the albumin Encapsulating at least part of said SN-38 and optionally at least part of said lipid, wherein said lipid is selected from the group consisting of cholesterol, cholesterol derivatives, cholesterol analogs and fatty acid esters and two or more thereof any combination of .
  • the present application provides a method of preparing a composition according to the first aspect.
  • the present application provides a method of preparing a composition comprising SN-38, a lipid, albumin and Span 20.
  • the present application provides a composition prepareable by the method according to the third aspect.
  • the present application also provides methods of preparing compositions with improved properties.
  • compositions comprising the compositions described above and uses thereof.
  • Fig. 1 is the HPLC typical pattern of content determination of SN-38 in the product prepared in Example 1.
  • Figure 2 is a typical HPLC profile for the determination of the content of cholesterol in the product prepared in Example 1.
  • Fig. 3 is the typical HPLC spectrum of the determination of the content of SN-38 of different structures in the goods prepared in Example 1
  • FIG. 4 is a typical SEC-HPLC profile for the determination of albumin aggregates in the product prepared in Example 1.
  • Figure 5 is the XRD pattern of the freeze-dried product prepared in Example 2, SN-38 and HSA.
  • FIG. 6 shows the results of the disintegration experiments performed in Example 19 under serial dilutions.
  • FIG. 7 shows the results of the disintegration experiments performed under serial dilutions in Example 20.
  • Fig. 8 shows the body weight change of the animals administered with the rHA-SN-38 preparation of Example 1 in Example 21.
  • Fig. 9 shows the results of in vivo experiments in which the preparation of Example 1 inhibits human triple-negative breast cancer MDA-MB-23.
  • Figure 10 shows the in vivo experimental results of the preparation of Example 2 inhibiting human colon cancer HT-29.
  • Fig. 11 shows the body weight changes of the test animals in Example 24.
  • FIG. 12 shows the changes in tumor volume of the test animals in Example 24.
  • Fig. 13 shows the body weight changes of the test animals in Example 25.
  • FIG. 14 shows the changes in tumor volume of the test animals in Example 25.
  • Fig. 15 shows the body weight changes of the test animals in Example 26.
  • FIG. 16 shows the changes in tumor volume of the test animals in Example 26.
  • Fig. 17 shows the body weight changes of the test animals in Example 27.
  • FIG. 18 shows the changes in tumor volume of the test animals in Example 27.
  • Fig. 19 shows the body weight changes of the test animals in Example 28.
  • FIG. 20 shows the changes in tumor volume of the test animals in Example 28.
  • Figure 21 shows the results of the disintegration test of the formulation containing Span 20 prepared in Example 30 under serial dilution.
  • Figure 22 shows the results of the disintegration test of the formulation prepared in Example 30 without Span 20 under gradient dilution.
  • Figure 23 shows the cryo-TEM observation results of the formulation containing Span 20 prepared in Example 30.
  • Figure 24 shows the cryo-TEM observation results of the formulation prepared in Example 30 without Span 20.
  • FIG. 25 shows the changes in tumor volume of the test animals in Example 34.
  • Fig. 26 shows the body weight changes of the test animals in Example 34.
  • Fig. 27 shows the tumor mass size of the test animals in Example 34.
  • Fig. 28 shows the body weight changes of the test animals in Example 38.
  • Figures 29-30 show the changes in tumor volume of the test animals in Example 38.
  • Fig. 31 shows the body weight changes of the test animals in Example 39.
  • Figures 32-33 show the changes in tumor volume of the test animals in Example 39.
  • nanoparticle means a particle having nanoscale dimensions in at least one dimension (e.g. one, two or three dimensions), e.g. a size of the order of about 1 nm, about 10 nm, about 100 nm or about 200-300 nm, preferably no more than 200nm-level dimensions.
  • nanoclaystal refers to a crystal with a size of 1 to 1000 nm, especially a crystal within 50 to 300 nm, which may be single crystal or polycrystalline.
  • vesicle refers to an ordered assembly of molecules with an outer layer structure that forms spontaneously when dispersed in an aqueous phase.
  • Span 20 also known as sorbitan monolaurate or Span 20, is a surfactant.
  • aqueous composition refers to a water-based composition, which is in liquid or semi-solid form, preferably in liquid form.
  • Liquid forms include, but are not limited to, solutions (eg, protein nanoparticle solutions), colloids, emulsions, and suspensions.
  • LD Drug Loading
  • EE Encapsulation Efficiency
  • organic acid includes saturated or unsaturated fatty acids having 1 to 24 carbon atoms, especially short chain fatty acids having 2 to 4 carbon atoms, medium chain fatty acids having 6 to 12 carbon atoms and fatty acids having 14-24 carbon long-chain fatty acids; and aromatic carboxylic acids.
  • An example that may be mentioned in particular is caprylic acid.
  • Long-chain fatty acids include, but are not limited to, palmitic acid (C16:0), stearic acid (C18:0), oleic acid (C18:1), linoleic acid (C18:2), alpha-linolenic acid (C18: 3), arachidonic acid (C20:4), eicosapentaenoic acid (C20:5), docosahexaenoic acid (C22:6). Examples that may in particular be mentioned are palmitic acid and stearic acid.
  • Numerical ranges recited herein are to be understood to encompass any and all subranges subsumed therein.
  • the range "1 to 10" should be understood to include not only the explicitly recited value of 1 to 10, but also any individual value within the range of 1 to 10 (e.g., 2, 3, 4, 5, 6, 7, 8, and 9) and subranges (eg, 1 to 2, 1.5 to 2.5, 1 to 3, 1.5 to 3.5, 2.5 to 4, 3 to 4.5, etc.). This principle also applies to ranges in which only one numerical value exists as a minimum or maximum value.
  • the present application provides a composition comprising SN-38, lipids, albumin and Span 20, characterized in that the composition comprises nanoparticles, wherein in the nanoparticles, the albumin encapsulating at least part of said SN-38 and optionally at least part of said lipid;
  • Lipid: SN-38 is about (0.1-10):1(w:w);
  • Albumin:SN-38 is about (1-100):1(w:w);
  • Span 20:SN-38 is about (3-60):100(w:w);
  • the lipid is selected from cholesterol, cholesterol derivatives, cholesterol analogs and fatty acid esters and any combination of two or more of them.
  • the lipid:SN-38 is about (0.5-6):1(w:w), such as about (0.5-5):1(w:w), about (0.5-3):1 (w:w), about (1-4):1(w:w), about (1.2-4):1(w:w), about (1.4-2):1(w:w), about ( 1.5-2.5):1(w:w) or about 1:1(w:w).
  • albumin:SN-38 is about (1-50):1 (w:w), for example about (3-25):1 (w:w), about (5-25):1 (w:w), about (5-20):1(w:w) or about (5-18):1(w:w), about (6-15):1(w:w), about ( 7-15):1(w:w), about (6-12):1(w:w), about (7-12):1(w:w), about (9-11):1(w :w) or about 10:1 (w:w).
  • Span 20:SN-38 is about (4-60):100 (w:w), such as about (5-60):100 (w:w), about (6-60):100 (w:w), about (7-55):100(w:w), about (8-50):100(w:w), about (10-45):100(w:w), about ( 12-40): 100(w:w), about (14-35): 100(w:w), about (15-30): 100(w:w), about (16-25): 100(w :w) or about (18-20):100(w:w).
  • albumin:lipid is about (1-100):1(w:w), such as about (2-20):1(w:w), about (3-15):1( w:w) or about (5-10):1(w:w), for example about 7:1(w:w) or about 10:1(w:w).
  • the content of SN-38 is about 1 w/w% to about 25 w/w%. In some embodiments, based on the total amount of SN-38, lipid and albumin in the composition, the content of the lipid is about 1 w/w% to about 35 w/w%. In some embodiments, the albumin is present in an amount of about 50 w/w% to about 98 w/w%, based on the total amount of SN-38, lipid and albumin in the composition.
  • the content of SN-38 is about 3w/w% to about 20w/w%. In some embodiments, based on the total amount of SN-38, lipid and albumin in the composition, the content of the lipid is about 2w/w% to about 30w/w%. In some embodiments, the albumin is present in an amount of about 55 w/w% to about 95 w/w%, based on the total amount of SN-38, lipid and albumin in the composition.
  • the content of SN-38 is about 3w/w% to about 15w/w%, for example about 4w /w%, about 5w/w%, about 6w/w%, about 6.5w/w%, about 7w/w%, about 7.5w/w%, about 8w/w%, about 8.5w/w%, about 9w/w%, about 10w/w%, about 11w/w%, about 12w/w%, about 13w/w%, or about 14w/w%.
  • the content of the lipid is about 3w/w% to about 30w/w%, such as about 4w/w%. w%, about 5w/w%, about 6w/w%, about 7w/w%, about 8w/w%, about 8.5w/w%, about 9w/w%, about 9.5w/w%, about 10w/ w%, about 10.5w/w%, about 11w/w%, about 11.5w/w%, about 12w/w%, about 12.5w/w%, about 13w/w%, about 13.5w/w%, about 14w/w%, about 15w/w%, about 16w/w%, about 17w/w%, about 18w/w%, about 19w/w%, about 20w/w%, about 21w/w%, about 24w/ w%, about 26w/w%, or about 28w/w%.
  • the content of albumin is about 60w/w% to about 94w/w%, such as about 64w/w%. w/w% to about 93w/w%, about 66w/w% to about 92w/w%, about 68w/w% to about 91w/w%, about 70w/w% to about 90w/w%, about 75w/w% to about 90w/w%, about 75w/w%, about 76w/w%, about 77w/w%, about 78w/w%, about 79w/w%, about 80w/w%, about 81w/w%, about 82w/w%, about 83w/w%, about 84w/w%, about 85w/w%, about 86w/w%, about 87w/w%, about 88w/w%, or about 89w/w%.
  • the inventors have found that as the amount of lipid increases, the composition tends to have a smaller average particle size, higher encapsulation efficiency and higher utilization of SN-38 ; Increase the amount of albumin may cause the reduction of drug loading, too high albumin content will also increase the particle size of nanoparticles, affecting the drug.
  • the content of the lipid is about 5 w/w% to about 24 w/w%. In one embodiment, based on the total amount of SN-38, lipid and albumin in the composition, the content of albumin is about 60w/w% to about 90w/w%.
  • an excessively high content of lipid components in the composition may have adverse effects on the individual.
  • the The lipid content is not more than about 18% w/w%, such as not more than about 16% w/w%, or not more than about 14 w/w%. However, this does not mean that compositions with higher lipid content are not desirable.
  • the content of Span 20 is about 0.03w/w% to about 12w/w%, such as about 0.06w/w% to about 10w/w%, 0.08w/w% to about 9w/w%, about 0.1w/w% to about 8w/w%, about 0.2w/w% to about 7w/w%, about 0.4w/w% to about 6w/w%, about 0.6w/w% to about 5w/w%, about 0.8w/w% to about 4w/w%, about 1w/w% to about 2w/w% .
  • the lipid:SN-38 is about (1-10):1(w:w), such as about (1-8):1(w:w), about (1-6):1 (w:w), about (1-5):1(w:w), about (1-4.5):1(w:w), about (1-4):1(w:w), about ( 1.2-3.8):1(w:w), about 1:1(w:w), about (1.4-3.6):1(w:w), about (1.6-3.4):1(w:w), About(1.8-3.2):1(w:w), About(2-3):1(w:w), About(2.2-2.8):1(w:w), About(2.4-2.6):1 (w:w), about 1:1(w:w), or about 2.5:1(w:w). In other embodiments, lipid:SN-38 is about (0.8-1.8):1 (w:w), about (0.9-1.7):1 (w:w), or about (1-1.4):1 .
  • albumin:SN-38 is about (5-40):1(w:w), for example about (5-less than 40):1(w:w), about (5-35): 1(w:w), about (5-30):1(w:w), about (8-25):1(w:w), about (10-22.5):1(w:w), about (12.5-20):1(w:w), about (15-17.5):1(w:w), about (16-18):1(w:w) or about 10:1(w:w) .
  • albumin:SN-38 is about (9-21):1 (w:w), about (9-20):1 (w:w), about (11-18):1 (w:w), or about (11.1-17.3):1(w:w).
  • Span 20:SN-38 is about (5-60):100 (w:w), such as about (6-60):100 (w:w), about (7-55):100 (w:w), about (8-50):100(w:w), about (10-45):100(w:w), about (12-40):100(w:w), about ( 14-35):100(w:w), about (15-30):100(w:w), about (16-25):100(w:w) or about (18-20):100(w :w).
  • Span 20:SN-38 is about (5-10):100 (w:w), about (5-9):100 (w:w), about (6-8.6):100 (w:w), about (6-8):100(w:w), or about (6.5-7):100(w:w).
  • albumin:lipid is about (1-40):1(w:w), for example about (1-less than 40):1(w:w), about (2-35):1 (w:w), about (3-15):1(w:w), about (5-10):1(w:w) or about (6-8):1(w:w), such as about 7:1(w:w) or about 10:1(w:w).
  • albumin:lipid is about (6-21):1(w:w), about (6.7-13):1(w:w), about (7-13):1(w :w), about (11-12.7):1(w:w).
  • the content of SN-38 is about 2w/w% to about 16w/w%. In some embodiments, based on the total amount of SN-38, lipid and albumin in the composition, the content of the lipid is about 2w/w% to about 35w/w%. In some embodiments, the albumin is present in an amount of about 75 w/w% to about 96 w/w%, based on the total amount of SN-38, lipid and albumin in the composition.
  • the content of SN-38 is about 2.5w/w% to about 15w/w%, such as about 4w/w% to about 10w/w%, about 4.5w/w% to about 9.5w/w%, about 5w/w% to about 9w/w%, or about 7.5w/w% to about 8w/w% .
  • the content of the lipid in the composition is about 2.5w/w% to about 30w/w% , for example about 4w/w% to about 12.5w/w%, about 4.5w/w% to about 12w/w%, about 7w/w% to about 10w/w%, or about 7.5w/w% to about 8w /w%.
  • the content of albumin in the composition is about 76w/w% to about 95w/w%, For example about 78w/w% to about 93w/w%, about 79w/w% to about 91.5w/w%, about 80w/w% to about 90w/w%, about 82w/w% to about 89w/w%, From about 84w/w% to about 88w/w%, or from about 84.5w/w% to about 87.5w/w%.
  • the content of SN-38 is about 3w/w% to about 14w/w%, for example about 3.5 w/w% to about 12w/w%, about 4w/w%, about 4.2w/w%, about 4.5w/w%, about 4.6w/w%, about 4.8w/w%, about 5w/w% , about 5.5w/w%, about 6w/w%, about 6.5w/w%, about 7w/w%, about 7.5w/w%, about 7.6w/w%, about 7.8w/w%, about 8w /w%, about 8.5w/w%, about 9w/w%, about 9.2w/w%, about 9.5w/w%, about 9.6w/w%, about 9.8w/w%, about 10w/w% , about 10.5 w/w%, about 11 w/w%, or about 11.5 w/w/w
  • the content of the lipid is about 4w/w% to about 25w/w%, such as about 4.5w /w% to about 20w/w%, about 4.3w/w%, about 4.5w/w%, about 4.7w/w%, about 5w/w%, about 5.5w/w%, about 6w/w%, About 6.5w/w%, about 7w/w%, about 7.5w/w%, about 7.6w/w%, about 7.8w/w%, about 8w/w%, about 8.5w/w%, about 9w/w %, about 9.5w/w%, about 10w/w%, about 10.5w/w%, about 11w/w%, about 11.5w/w%, about 12w/w%, about 12.1w/w%, about 12.3 w/w%, about 12.5w/w%, about 13w/w%, about 13.5w/
  • the content of albumin is about 78w/w% to about 92w/w%, such as about 79w/w%. w%, about 79.2w/w%, about 79.4w/w%, about 79.6w/w%, about 79.8w/w%, about 80w/w%, about 81w/w%, about 82w/w%, about 83w/w%, about 84w/w%, about 84.3w/w%, about 84.5w/w%, about 84.7w/w%, about 84.9w/w%, about 85w/w%, about 86w/w% , about 87w/w%, about 87.3w/w%, about 87.5w/w%, about 87.7w/w%, about 87.9w/w%, about 88w/w%, about 89w/w%, about 90w/ w% to about 92w/w%, such as about 79w/w%. w%, about
  • the content of Span 20 is about 0.14w/w% to about 5w/w%, such as about 0.2w/w% to about 2.5w/w%, about 0.22w/w% to about 2.0w/w%, about 0.24w/w% to about 2w/w%, about 0.26w/w% to about 1.5w /w%, about 0.28w/w% to about 1.0w/w%, about 0.3w/w% to about 0.9w/w%, about 0.32w/w% to about 0.8w/w%, about 0.34w/ w% to about 0.7w/w%, about 0.36w/w% to about 0.6w/w%, about 0.38w/w% to about 0.58w/w%, about 0.4w/w% to about 0.56w/w %, about 0.42w/w% to about 0.54w/w%, about 0.44w/w% to about 0.56w/w %, about 0.42w/w% to about 0.54w/w%, about 0.44w/w
  • the content of Span 20 is about 0.2w/w% to about 0.8w/w%, about 0.24w/w% to about 0.7w/w%, about 0.26w/w% to about 0.7w /w%, about 0.3w/w% to about 0.65w/w%, about 0.36w/w% to about 0.6w/w%, about 0.4w/w% to about 0.58w/w%, about 0.44w/ w % to about 0.56 w/w %, about 0.48 w/w % to about 0.54 w/w %, or about 0.5 w/w % to about 0.52 w/w %.
  • the SN-38 present in the nanoparticles is at least about 1 w/w % or at least about 2 w/w % of the total amount of SN-38, lipid and albumin in the composition , for example at least about 3w/w%, about 3w/w% to about 13w/w%, about 4w/w% to about 12w/w%, about 4w/w%, about 5w/w%, about 6w/w% , about 7w/w%, about 8w/w%, about 9w/w%, about 10w/w%, or about 11w/w%.
  • the SN-38 present in the nanoparticles is from about 80 w/w % to about 99 w/w % of the total amount of SN-38 in the composition, such as from about 88 w/w % to About 98w/w%, about 89w/w%, about 90w/w%, about 91w/w%, about 92w/w%, about 93w/w%, about 94w/w%, about 95w/w%, about 96w /w% or about 97w/w%.
  • the cholesterol derivative is selected from esters of cholesterol and organic acids, preferably selected from cholesterol palmitate and cholesterol caprylate and combinations thereof.
  • the cholesterol analog is selected from vitamin D2, vitamin D3, and combinations thereof.
  • the fatty acid ester is selected from fatty acid glycerides, preferably long-chain fatty acid glycerides, preferably glyceryl stearate, more preferably glyceryl monostearate.
  • the lipid is selected from any of cholesterol, cholesterol palmitate, cholesterol caprylate, vitamin D2, vitamin D3, glyceryl monostearate and two or more thereof combination.
  • the lipid is selected from cholesterol, cholesterol palmitate, vitamin D3, glyceryl monostearate, and any combination of two or more thereof;
  • the lipid is: cholesterol, cholesterol palmitate, vitamin D3, or glyceryl monostearate; a mixture of cholesterol and cholesterol palmitate; a mixture of cholesterol and vitamin D3; a mixture of cholesterol and monostearate; a mixture of glyceryl stearate; or a mixture of cholesterol palmitate and glyceryl monostearate.
  • the lipid is cholesterol.
  • cholesterol is used as the only lipid.
  • cholesterol:SN-38 is about (1-6):1(w:w), such as about (1.2-5):1(w:w), such as about (1.4-4) :1(w:w), about 3:1(w:w), about 2:1(w:w), or about 1:1(w:w).
  • cholesterol:SN-38 is about (0.8-1.8):1 (w:w), about (0.9-1.7):1 (w:w), about (1-1.4):1.
  • the albumin:SN-38 is about (3-25):1(w:w), such as about (4-20):1(w:w), about (5-15) :1(w:w), about (6-12):1(w:w), about (7-12):1(w:w), about (9-11):1(w:w) or John 10:1 (w:w).
  • albumin:SN-38 is about (9-21):1 (w:w), about (9-20):1 (w:w), about (11-18):1 (w:w), or about (11.1-17.3):1(w:w).
  • albumin:cholesterol is about (2-20):1 (w:w), such as about (3-15):1 (w:w), about (5-10):1 (w:w) or about 7:1(w:w). In other embodiments, albumin:cholesterol is about (6-21):1(w:w), about (6.7-13):1(w:w), about (7-13):1(w :w), about (11-12.7):1(w:w).
  • the content of SN-38 is about 3w/w% to about 15w/w%, such as about 4 w/w % to about 15 w/w %, about 6 w/w % to about 10 w/w %, or about 8 w/w % to about 12 w/w %. In other embodiments, the content of SN-38 is about 4w/w% to about 10w/w%, about 4.5w/w% to about 9.5w/w%, about 5w/w% to about 9w/ w%, or about 7.5w/w% to about 8w/w%.
  • the content of cholesterol is about 5w/w% to about 25w/w%, such as about 6w/w%. w% to about 22w/w%, or about 15w/w% to about 20w/w%. In other embodiments, the cholesterol content is about 4w/w% to about 12.5w/w%, about 4.5w/w% to about 12w/w%, about 7w/w% to about 10w/w% , or from about 7.5 w/w% to about 8 w/w%.
  • the amount of albumin is about 64w/w% to about 90w/w%, such as about 70w /w% to about 90w/w%.
  • the content of the albumin is about 78w/w% to about 93w/w%, about 79w/w% to about 91.5w/w%, about 80w/w% to about 90w/w% , about 82 w/w % to about 89 w/w %, about 84 w/w % to about 88 w/w %, or about 84.5 w/w % to about 87.5 w/w %.
  • the SN-38 present in the nanoparticles is at least about 3 w/w %, such as about 3 w/w, of the total amount of SN-38, cholesterol and albumin in the composition % to about 13w/w%, about 4w/w% to about 12w/w%, about 4w/w%, about 5w/w%, about 6w/w%, about 7w/w%, about 8w/w%, About 9 w/w%, about 10 w/w%, or about 11 w/w%.
  • the lipid is cholesterol palmitate. In some embodiments, the lipid is vitamin D3. In some embodiments, the lipid is glyceryl monostearate. In some embodiments, cholesterol palmitate, vitamin D3, or glyceryl monostearate are the sole lipids.
  • the lipid is a mixture of cholesterol and glyceryl monostearate, wherein cholesterol:glyceryl monostearate is for example about (0.2-5):1(w:w), about (0.5 -3):1(w:w), about (0.5-2):1(w:w), or about 1:1(w:w).
  • the lipid is a mixture of cholesterol palmitate and glyceryl monostearate; wherein cholesterol palmitate:glyceryl monostearate is, for example, about (0.2-5):1(w: w), about (0.5-3):1(w:w), about (0.5-2):1(w:w), or about 1:1(w:w).
  • the lipid is a mixture of cholesterol and cholesterol palmitate, wherein cholesterol:cholesterol palmitate is, for example, about (0.2-5):1 (w:w), about (0.5-3): 1(w:w), about (0.5-2):1(w:w), or about 1:1(w:w).
  • the lipid:SN-38 is, for example, about (1-6):1(w:w), about (1.2- 5):1(w:w), about (1.4-4.5):1(w:w), about 4.3:1(w:w), about 3:1(w:w), about 1:1(w :w) or about 2:1 (w:w).
  • the albumin:SN-38 is, for example, about (5-25):1 (w:w), about (10-20):1 (w:w), about (6-15) :1(w:w), about (7-15):1(w:w), about (9-12):1(w:w), about (9-11):1(w:w), About 9.5:1 (w:w) or about 10:1 (w:w).
  • albumin:the lipid is, for example, about (2-10):1 (w:w), about (3-7):1 (w:w) or about (4-6 ):1(w:w).
  • the amount of SN-38 is, for example, from about 5 w/w % to about 15 w/w based on the total amount of SN-38, the lipid and albumin in the composition %, about 6w/w% to about 12w/w%, about 7w/w% to about 10w/w%, about 8w/w% or about 9w/w%.
  • the lipid is present, for example, in an amount from about 5 w/w% to about 32 w/w%, based on the total amount of SN-38, the lipid and albumin in the composition , about 10w/w% to about 30w/w%, about 18w/w%, about 20w/w%, about 22w/w%, about 24w/w%, about 26w/w%, about 28w/w% or about 29w/w%.
  • the albumin is present in an amount of, for example, from about 60 w/w% to about 90 w/w%, based on the total amount of SN-38, the lipid and albumin in the composition , about 64w/w% to about 85w/w%, about 70w/w% to about 80w/w%, or about 75w/w%.
  • the SN-38 present in the nanoparticles represents at least about 3 w/w % of the total amount of SN-38, the lipid and albumin in the composition, such as about 3w/w% to about 10w/w%, about 4w/w% to about 9w/w%, about 4w/w%, about 5w/w%, about 6w/w%, about 7w/w% or about 8w/ w%.
  • the composition is in liquid, semi-solid or solid form.
  • the composition is in solid form, preferably in powder form. More preferably, the composition is freeze-dried powder.
  • SN-38 is preferably present in the composition in amorphous and/or nanocrystalline form, as determined by electron microscopy or X-ray diffraction (Cu-K ⁇ ) analysis.
  • the particle size range of the nanocrystals may be about 30-500 nm, preferably about 50-200 nm.
  • Said nanocrystalline form of SN-38 comprises 75% or more of the total SN-38, such as 80%, 85%, 90% or more.
  • Span 20 plays a role in improving the stability of the composition, adjusting the particle size of the nanoparticles and making them more dispersed.
  • the composition contains no additional stabilizers.
  • the composition preferably further comprises an additional stabilizer, such as a lyophilized stabilizer, in an amount such that when the composition is reconstituted to form an aqueous composition (including a solution) and emulsion), the content of the additional stabilizer is at least about 2w/v%, such as at least about 3w/v%, at least about 5w/v%, about 5w/v% to about 30w/v%, about 10w /v% to about 25w/v% or about 15w/v% to about 20w/v%.
  • an additional stabilizer such as a lyophilized stabilizer
  • the composition further comprises an additional stabilizer in an amount of about 60 w/w% to about 98 w/w% based on the total amount in the composition , for example about 65w/w% to about 97w/w%, about 68w/w% to about 96w/w%, about 69w/w% to about 95w/w%, about 70w/w% to about 94w/w%, About 71w/w% to about 93w/w%, about 72w/w% to about 92w/w%, about 73w/w%, about 74w/w%, about 75w/w%, about 76w/w%, about 77w /w%, about 78w/w%, about 79w/w%, about 80w/w%, about 81w/w%, about 82w/w%, about 83w/w%, about 84w/w%, about 85w/w %, about 86w/w%, about 87w/
  • the composition when the composition is reconstituted to form an aqueous composition (including solutions and emulsions) to have about 0.1 ⁇ g/mL to about 30.0 mg/mL (or SN-38 as described below for aqueous compositions)
  • the content of SN-38 is about 50 to 200 nm, such as about 90 to 150 nm, about 95 to 140 nm, about 100 to 130 nm, about 105 to 125 nm, or about 110 to 120 nm
  • the nanoparticles have an average particle size of about 50 to 200 nm. path.
  • compositions are aqueous compositions in liquid form, including solutions and emulsions.
  • the liquid form composition comprises SN-38 in the form of nanocrystals and/or vesicles.
  • the particle size range of the nanocrystals may be about 30-500 nm, preferably about 50-200 nm.
  • the composition is in the form of a solution; in some such embodiments, the composition is in the form of an emulsion.
  • the aqueous compositions of the present application have excellent dilution stability.
  • the composition when the composition is diluted (eg, diluted with 1 x PBS, pH about 7.4) such that the amount of SN-38 in the diluted composition reaches about 4 ⁇ g/mL or less, such as about 2 ⁇ g/mL mL or less, about 1 ⁇ g/mL or less, or about 0.4 ⁇ g/mL or less, such as about 0.1 ⁇ g/mL or 0.04 ⁇ g/mL, the nanoparticles do not disintegrate. Due to the excellent stability, the aqueous composition can be present as a concentrate or a dilution.
  • various components of the aqueous composition may have a wide content range.
  • SN-38 is present in an amount of about 0.1 ⁇ g/mL to about 30.0 mg/mL, about 0.2 ⁇ g/mL to about 27.0 mg/mL, about 0.5 ⁇ g/mL to about 24.0mg/mL, about 1.0 ⁇ g/mL to about 21.0mg/mL, about 5.0 ⁇ g/mL to about 18.0mg/mL, about 10.0 ⁇ g/mL to about 15.0mg/mL, about 20.0 ⁇ g/mL mL to about 12 mg/mL, about 25.0 ⁇ g/mL to about 9 mg/mL, about 50.0 ⁇ g/mL to about 6.0 mg/mL, or about 100.0 ⁇ g/mL to about 3.0 mg/mL.
  • the content of the lipid is about 0.05 ⁇ g/mL to about 100.0 mg/mL, about 0.1 ⁇ g/mL to about 90.0 mg/mL, about 0.25 ⁇ g /mL to about 80.0 mg/mL, about 0.5 ⁇ g/mL to about 70.0 mg/mL, about 2.5 ⁇ g/mL to about 60.0 mg/mL, about 5.0 ⁇ g/mL to about 50.0 mg/mL, about 10.0 ⁇ g/mL to about 40.0 mg/mL, about 12.5 ⁇ g/mL to about 30.0 mg/mL, about 25.0 ⁇ g/mL to about 20.0 mg/mL, or about 50.0 ⁇ g/mL to about 10.0 mg/mL.
  • the content of albumin is about 3.0 ⁇ g/mL to about 300.0 mg/mL, about 6.0 ⁇ g/mL to about 270.0 mg/mL, about 15.0 ⁇ g/mL to about 240.0 mg/mL, about 30.0 ⁇ g/mL to about 210.0 mg/mL, about 150.0 ⁇ g/mL to about 180.0 mg/mL, about 300.0 ⁇ g/mL to about 150.0 mg/mL, about 600.0 ⁇ g/mL to about 120.0 mg/mL, about 750.0 ⁇ g/mL to about 90.0 mg/mL, about 1500.0 ⁇ g/mL to about 60.0 mg/mL, or about 3.0 mg/mL to about 30.0 mg/mL.
  • the content of SN-38 can be about 100.0 ⁇ g/mL to about 3.0 mg/mL, such as about 200.0 ⁇ g/mL to about 2.5 mg/mL, about 300.0 ⁇ g/mL mL to about 2.0 mg/mL, about 400.0 ⁇ g/mL to about 1.5 mg/mL, about 500.0 ⁇ g/mL to about 1.0 mg/mL, or about 600 ⁇ g/mL to about 800 ⁇ g/mL; and/or
  • the content of the lipid can be about 50.0 ⁇ g/mL to about 10.0 mg/mL, such as about 100.0 ⁇ g/mL to about 8.0 mg/mL, about 200.0 ⁇ g/mL to about 6.0 mg/mL, about 300.0 ⁇ g/mL to about 4.0 mg/mL, about 400.0 ⁇ g/mL to about 3.0 mg/mL, about 500.0 ⁇ g/mL to about 2.5 mg/mL, about 600.0 ⁇ g/mL to about 2.0 mg/mL, about 700.0 ⁇ g/mL to about 1.5 mg/mL, about 800 ⁇ g/mL to about 1.0 mg/mL, or about 200 ⁇ g/mL to about 1.5 mg/mL; and/or
  • the content of albumin may be from about 3.0 mg/mL to about 30.0 mg/mL, such as from about 4.0 mg/mL to about 25.0 mg/mL, from about 5.0 mg/mL to about 20.0 mg/mL, from about 6.0 mg/mL to about 15.0 mg/mL, about 7.0 mg/mL to about 12.0 mg/mL, or about 8.0 mg/mL to about 10.0 mg/mL.
  • the nanoparticles in aqueous compositions (including solutions and emulsions), have an average particle size of no more than about 200 nm, such as no more than about 150 nm, preferably after storage for a certain period of time prior to application. still meet the above requirements.
  • the inventors of the present invention have found that the aqueous composition of the present application has such excellent properties.
  • the nanoparticles have an average particle size of about 50 to 200 nm, such as about 90 to 150 nm, or about 100 to 130 nm.
  • the nanoparticles have an average particle size of about 50 to 200 nm, such as about 90 to 150 nm, or about 100 to 130 nm, when stored at 4°C for 24 hours.
  • the nanoparticles have a particle size distribution index (PDI) of no greater than about 0.30, such as no greater than about 0.2, no greater than about 0.10, or no greater than about 0.01.
  • PDI particle size distribution index
  • the composition has a Zeta potential of about -35 mV to about -20 mV, eg, about -31 mV.
  • the composition contains no additional stabilizers.
  • the composition further comprises an additional stabilizer, wherein the additional stabilizer is present in an amount of at least about 2w/v%, preferably at least about 3w%, based on the total amount of the composition /v%, for example at least about 5w/v%, about 5w/v% to about 30w/v%, about 10w/v% to about 25w/v% or about 15w/v% to about 20w/v%.
  • the above-mentioned additional stabilizer can be selected from: albumins (such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder), monosaccharides, disaccharides, polysaccharides and any combination thereof; preferably selected from glucose and sucrose; preferably sucrose.
  • albumins such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • additional stabilizers is beneficial to maintain the average particle size of the nanoparticles.
  • the inventors have found that the presence of additional stabilizer reduces the increase in the average particle size of the nanoparticles in the aqueous composition when stored at 4°C for 24 h compared to the absence of additional stabilizer.
  • additional stabilizers especially sugar stabilizers, also provides additional advantages for the composition finally provided in the form of lyophilized powder, because they can be used during the freeze-drying of the aqueous composition.
  • the composition preferably contains no additional lyophilization excipients.
  • the composition may also contain additional lyophilized excipients, such as one or more of sucrose, mannitol, lactose, maltose, trehalose, and dextran.
  • the present application provides a composition comprising SN-38, a lipid, albumin and Span 20, wherein the lipid is cholesterol, characterized in that the composition comprises nano Particles, wherein in said nanoparticle said albumin encapsulates at least part of said SN-38 and optionally at least part of said lipid,
  • Cholesterol: SN-38 in some embodiments, about (1-3):1(w:w), such as about (1.2-2.5):1(w:w), about (1.4-2):1 (w:w), about (1.5-2):1(w:w), about (1.3-1.8):1(w:w), about (1.4-1.6):1(w:w), about ( 1.5-1.7):1(w:w), about (1.2-1.5):1(w:w), about 1:1(w:w) or about (1.4-1.5):1(w:w), Or, in other embodiments, about (0.8-1.8):1(w:w), about (0.9-1.7):1(w:w), or about (1-1.4):1;
  • Albumin:SN-38 in some embodiments, is about (5-15):1(w:w), such as about (5-12):1(w:w), about (6-12): 1(w:w) or about (7-12):1(w:w), about (9-11):1(w:w), about (10-12):1(w:w), or About 11:1(w:w), or, in other embodiments, about (9-21):1(w:w), about (9-20):1(w:w), about ( 11-18):1(w:w), or about (11.1-17.3):1(w:w);
  • Albumin:cholesterol in some embodiments, is about (3-10):1(w:w), about (4-8):1(w:w), or about (5-7):1(w :w), or, in other embodiments, about (6-21):1(w:w), about (6.7-13):1(w:w), about (7-13):1 (w:w), about (11-12.7):1(w:w); and
  • the content of SN-38 is about 6w/w% to about 14w/w%, such as about 6.5w /w% to about 13w/w%, about 7w/w% to about 12w/w%, about 7.5w/w% to about 12w/w%, about 8w/w% to about 11w/w%, about 8.5w /w% to about 10w/w%, or about 9w/w%.
  • the content of SN-38 is about 4w/w% to about 10w/w%, about 4.5w/w% to about 9.5w/w%, about 5w/w% to about 9w/ w%, or about 7.5w/w% to about 8w/w%.
  • the content of cholesterol is about 8w/w% to about 18w/w%, such as 8.5w/w% to about 17w/w%, about 9w/w% to about 16w/w%, about 9.5w/w% to about 16w/w%, about 10w/w% to about 16w/w%, about 10.5w/w% to about 16w/w%, about 11w/w% to about 15w/w%, about 11.5w/w% to about 15w/w%, about 12w/w% to about 15w/w%, about 12.5w/w% to about 14w/w%, or about 13w/w% to about 13.5w/w%.
  • the cholesterol content is about 4w/w% to about 12.5w/w%, about 4.5w/w% to about 12w/w%, about 7w/w% to about 10w/w% , or from about 7.5 w/w% to about 8 w/w%.
  • the content of albumin is about 66w/w% to about 90w/w%, such as about 68w/w % to about 89w/w, about 70w/w% to about 88w/w%, about 70w/w% to about 87w/w%, about 70w/w% to about 86w/w%, about 70w/w% to about 85w/w%, about 75w/w% to about 85w/w%, about 76w/w%, about 77w/w%, about 78w/w%, about 79w/w%, about 80w/w%, about 81w/ w%, about 82w/w%, about 83w/w%, or about 84w/w%.
  • the albumin content is about 78w/w% to about 93w/w%, about 79w/w% to about 91.5w/w%, about 80w/w% to about 90w/w% , about 82w/w% to about 89w/w%, about 84w/w% to about 88w/w%, or about 84.5w/w% to about 87.5w/w%.
  • the present application provides a composition comprising SN-38, a lipid, albumin and Span 20, wherein the lipid is cholesterol, characterized in that the composition comprises nano Particles, wherein in said nanoparticle said albumin encapsulates at least part of said SN-38 and optionally at least part of said lipid,
  • Cholesterol: SN-38 in some embodiments, about (1-5):1(w:w), such as about (1-4.5):1(w:w), about (1-4):1 (w:w), about (1.2-3.8):1(w:w), about (1.4-3.6):1(w:w), about (1.6-3.4):1(w:w), about ( 1.8-3.2):1(w:w), about (2-3):1(w:w), about (2.2-2.8):1(w:w), about (2.4-2.6):1(w :w), about 2.5:1(w:w), or about 1:1(w:w), or, in other embodiments, about (0.8-1.8):1(w:w), about ( 0.9-1.7):1(w:w) or about (1-1.4):1; and/or
  • Albumin:SN-38 in some embodiments, is about (5-25):1(w:w), such as about (5-20):1(w:w), about (6-19): 1(w:w), about (7-18):1(w:w), about (8-16):1(w:w), about (9-14):1(w:w) or about (10-12):1(w:w), or, in other embodiments, about (9-21):1(w:w), about (9-20):1(w:w) , about (11-18):1(w:w), or about (11.1-17.3):1(w:w); and/or
  • Albumin:cholesterol in some embodiments, is about (5-25):1(w:w), such as about (6-20):1(w:w), about (7-18):1( w:w), about (8-16):1(w:w), about (9-14):1(w:w), or about (10-12):1(w:w), or, in In other embodiments, about (6-21):1(w:w), about (6.7-13):1(w:w), about (7-13):1(w:w), about (11-12.7):1(w:w); and/or
  • Span 20:SN-38 in some embodiments, is about (5-15):100(w:w), such as about (6-12):100(w:w), about (7-10): 100(w:w) or about 7.5:100(w:w), or, in other embodiments, about (5-10):100(w:w), about (5-9):100( w:w), about (6-8.6):100(w:w), about (6-8):100(w:w), about (6.5-7):100(w:w).
  • the content of SN-38 is about 3w/w% to about 10w/w%, such as about 3.5w/w% to about 9.5w/w%, about 4w/w%, about 4.5w/w%, about 5w/w%, about 5.5w/w%, about 6w/w%, about 6.5w/w %, about 7w/w%, about 7.5w/w%, about 8w/w%, about 8.5w/w%, or about 9w/w%.
  • the content of SN-38 is about 4w/w% to about 10w/w%, about 4.5w/w% to about 9.5w/w%, about 5w/w% to about 9w/ w%, or about 7.5w/w% to about 8w/w%.
  • the cholesterol is present in an amount of about 4w/w% to about 18w/w%, for example about 4.5w /w% to about 17.5w/w%, about 5w/w%, about 5.5w/w%, about 6w/w%, about 6.5w/w%, about 7w/w%, about 7.5w/w%, About 8w/w%, about 8.5w/w%, about 9w/w%, about 9.5w/w%, about 10w/w%, about 10.5w/w%, about 11w/w%, about 11.5w/w% , about 12w/w%, about 12.5w/w%, about 13w/w%, about 13.5w/w%, about 14w/w%, about 14.5w/w%, about 15w/w%, about 15.5w/ w%, about 16w/w%, about 16.5w/w%, or about 17w/w/w
  • the cholesterol content is about 4w/w% to about 12.5w/w%, about 4.5w/w% to about 12w/w%, about 7w/w% to about 10w/w% , or from about 7.5 w/w% to about 8 w/w%.
  • the amount of albumin is about 78w/w% to about 92w/w%, such as about 79w /w%, about 80w/w%, about 81w/w%, about 82w/w%, about 83w/w%, about 84w/w%, about 85w/w%, about 86w/w%, about 87w/w %, about 88w/w%, about 89w/w%, about 90w/w%, or about 91w/w%.
  • the albumin content is about 78w/w% to about 93w/w%, about 79w/w% to about 91.5w/w%, about 80w/w% to about 90w/w% , about 82w/w% to about 89w/w%, about 84w/w% to about 88w/w%, or about 84.5w/w% to about 87.5w/w%.
  • the present invention provides a composition as described above, wherein:
  • Cholesterol: SN-38 is about (1-2.5):1(w:w), such as about 1.25:1(w:w), about 1:1(w:w); and/or
  • Albumin: SN-38 is about (8-25):1(w:w), such as about (10-20):1(w:w), about (12-18):1(w:w), about (12.5-17):1(w:w), about (15-16):1(w:w), about 10:1(w:w) or about 16.7:1(w:w); and/or or
  • Albumin:Cholesterol is about (5-10):1(w:w), such as about (6-9):1(w:w), about (7-8):1(w:w), about 10 :1(w:w) or about 6.7:1(w:w); and/or
  • Span 20:SN-38 is about (5-40):100(w:w), such as about (6-30):100(w:w), about (7-25):100(w:w), About (8-20):100(w:w), about (9-15):100(w:w) or about (10-12):100(w:w).
  • the content of SN-38 is about 3w/w% to about 9w/w%, such as about 3.5w /w% to about 8.5w/w%, about 4w/w%, about 4.5w/w%, about 5w/w%, about 5.5w/w%, about 6w/w%, about 6.5w/w%, About 7w/w%, about 7.5w/w%, about 8w/w%.
  • the content of cholesterol is about 8w/w% to about 18w/w%, such as about 8.5w/w % to about 17.5w/w%, about 9w/w%, about 9.5w/w%, about 10w/w%, about 10.5w/w%, about 11w/w%, about 11.5w/w%, about 12w /w%, about 12.5w/w%, about 13w/w%, about 13.5w/w%, about 14w/w%, about 14.5w/w%, about 15w/w%, about 15.5w/w%, About 16w/w%, about 16.5w/w%, or about 17w/w%.
  • the content of Span 20 is about 0.2w/w% to about 0.6w/w%, for example About 0.22w/w% to about 0.58w/w%, about 0.24w/w% to about 0.56w/w%, about 0.26w/w% to about 0.54w/w%, about 0.28w/w% to about 0.52w/w%, about 0.3w/w% to about 0.5w/w%, about 0.32w/w% to about 0.48w/w%, about 0.34w/w% to about 0.46w/w%, about 0.36 w/w % to about 0.44 w/w %, about 0.38 w/w % to about 0.42 w/w %, or about 0.4 w/w %.
  • Cholesterol: SN-38 is about (0.8-1.8):1(w:w), about (0.9-1.7):1(w:w), or about (1-1.4):1; and/or
  • Albumin: SN-38 is about (9-21):1(w:w), about (9-20):1(w:w), about (11-18):1(w:w), or about (11.1-17.3):1(w:w); and/or
  • Span 20:SN-38 is about (5-10):100(w:w), about (5-9):100(w:w), about (6-8.6):100(w:w), about (6-8):100(w:w), about (6.5-7):100(w:w); and/or
  • the content of the SN-38 is about 4w/w% to about 10w/w%, about 4.5w/w% to about 9.5w/w%, about 5w/w% to about 9w/w%, or about 7.5w /w% to about 8w/w%; and/or
  • the cholesterol content is about 4w/w% to about 12.5w/w%, about 4.5w/w% to about 12w/w%, about 7w/w% to about 10w/w%, or about 7.5w/w % to about 8w/w%; and/or
  • the content of the albumin is about 78w/w% to about 93w/w%, about 79w/w% to about 91.5w/w%, about 80w/w% to about 90w/w%, about 82w/w% to about 89w/w%, about 84w/w% to about 88w/w%, or about 84.5w/w% to about 87.5w/w%; and/or
  • the content of Span 20 is about 0.2w/w% to about 0.8w/w%, about 0.24w/w% to about 0.7w/w%, about 0.26w/w% to about 0.7w/w%, about 0.3w /w% to about 0.65w/w%, about 0.36w/w% to about 0.6w/w%, about 0.4w/w% to about 0.58w/w%, about 0.44w/w% to about 0.56w/ w %, about 0.48 w/w % to about 0.54 w/w %, or about 0.5 w/w % to about 0.52 w/w %.
  • compositions according to the second subset have advantages in preparation on a larger scale (e.g., hundreds of milligrams or more of SN-38 starting material), including in scale-up processes, e.g., pilot-scale preparations, e.g. Reduce the number of high-pressure homogenization in the process, effectively reduce the particle size of nanoparticles in the composition, improve the filtration flux, control the particle size after disintegration under physiological conditions, reduce raw material loss and reduce costs; and have suitable albumin content, so that the particle size of nanoparticles can be controlled to make it closer to the size suitable for drug making.
  • scale-up processes e.g., pilot-scale preparations, e.g. Reduce the number of high-pressure homogenization in the process, effectively reduce the particle size of nanoparticles in the composition, improve the filtration flux, control the particle size after disintegration under physiological conditions, reduce raw material loss and reduce costs; and have suitable albumin content, so that the particle size of nanoparticles can be controlled to make it closer to the size suitable
  • SN-38 present in the nanoparticles accounts for at least about 6w of the total amount of SN-38, cholesterol and albumin in the composition /w% to about 12w/w%, for example about 7w/w% to about 11w/w%, about 8w/w% to about 10w/w%, about 8.3% or about 9w/w%.
  • the SN-38 present in the nanoparticles is from about 95 w/w % to about 99 w/w %, such as from about 96 w/w % to about 99 w/w % of the total amount of SN-38 in the composition. About 99w/w%, about 97w/w% to about 99w/w%, about 98w/w% to about 99w/w%, or about 99w/w% or higher.
  • the composition is in liquid, semi-solid or solid form.
  • the composition is in the form of a solid, preferably a powder, more preferably a lyophilized powder.
  • SN-38 is preferably present in the composition in amorphous and/or nanocrystalline form, as determined by electron microscopy and X-ray diffraction (Cu-K ⁇ ) analysis.
  • the composition contains no additional stabilizers.
  • the composition further comprises an additional stabilizer in an amount such that when the composition is reconstituted to form an aqueous composition (including solutions and emulsions), the additional The content of stabilizer is at least about 2w/v%, such as at least about 3w/v%, at least about 5w/v%, about 5w/v% to about 30w/v%, about 10w/v% to about 25w/v% Or about 15w/v% to about 20w/v%.
  • the composition further comprises an additional stabilizer in an amount of about 70 w/w% to about 96 w/w%, based on the total amount of the composition, For example about 70w/w% to about 90w/w%, about 72w/w% to about 89w/w%, about 74w/w% to about 88w/w%, about 76w/w% to about 87w/w%, about 80w/w% to about 86w/w%, about 81w/w% to about 86w/w%, about 82w/w% to about 85w/w%, or about 83w/w% to about 84w/w%.
  • the composition further comprises an additional stabilizer, based on the total amount of the composition, the content of the additional stabilizer is about 80w/w% to about 96w/w% or about 84w/w% to about 95w/w%.
  • the additional stabilizer may be selected from mannitol, lactose, maltose, trehalose, dextran, glucose and sucrose and any combination thereof, preferably sucrose.
  • compositions are aqueous compositions in liquid form, including solutions and emulsions.
  • the liquid form composition comprises SN-38 in the form of nanocrystals and/or vesicles.
  • the composition is in the form of a solution; in some embodiments, the composition is in the form of an emulsion.
  • SN-38 is present in an amount from about 500.0 ⁇ g/mL to about 1.0 mg/mL, eg, from about 600 ⁇ g/mL to about 800 ⁇ g/mL, based on the total amount of the composition.
  • albumin is present in an amount of about 5.0 mg/mL to about 10.0 mg/mL, such as about 6.0 mg/mL to about 10 mg/mL, or about 7.0 mg/mL, based on the total amount of the composition. mg/mL to about 8.0 mg/mL.
  • the nanoparticles in the composition have an average particle size of about 90 to 160 nm, such as about 95 to 150 nm, about 100 to 140 nm, about 105 to 130 nm, about 110 to 125 nm, about 110 nm, about 115 nm , about 120 nm, about 125 nm, about 130 nm, 135 nm, about 140 nm, or about 145 nm.
  • the nanoparticles have a particle size distribution index (PDI) of no greater than about 0.30, such as no greater than about 0.2, no greater than about 0.10, or no greater than about 0.01.
  • PDI particle size distribution index
  • the composition has a Zeta potential of about -35 mV to about -20 mV, eg, about -31 mV.
  • the composition when the composition is diluted (e.g., diluted with 1x PBS at a pH of about 7.4) such that the amount of SN-38 in the diluted composition is about 4 ⁇ g/mL or less, such as about 2 ⁇ g/mL or less, about 1 ⁇ g/mL or less, about 0.4 ⁇ g/mL or less, about 0.1 ⁇ g/mL or less, about 0.04 ⁇ g/mL or less, about 0.02 ⁇ g/mL or less, or about At 0.01 ⁇ g/mL or lower, the nanoparticles do not disintegrate.
  • the nanoparticles do not disintegrate.
  • the composition contains no additional stabilizers.
  • the composition further comprises an additional stabilizer, wherein the additional stabilizer is present in an amount of at least about 2 w/v %, such as at least about 3 w/v, based on the total amount of the composition %, at least about 5w/v%, about 5w/v% to about 30w/v%, about 10w/v% to about 25w/v% or about 15w/v% to about 20w/v%.
  • the additional stabilizer may be selected from mannitol, lactose, maltose, trehalose, dextran, glucose and sucrose and any combination thereof, preferably sucrose.
  • Opened SN-38 is the inactive form of SN-38.
  • the inventors have surprisingly found that ring-opened SN-38 is only present in very low amounts in the compositions of the present application.
  • the ring-opened SN-38 accounts for about 2w/w% or less of the total amount of SN-38 in the composition, preferably about 1.8w/w% or less.
  • Albumin as a carrier may form multimers, including dimers, trimers, multimers, etc.
  • the presence of albumin polymers increases the risk of immunogenicity of drugs, especially parenterally administered drugs. Therefore, it is advantageous to contain as little albumin polymer as possible.
  • the inventors have surprisingly found that albumin multimers are absent or substantially absent in the compositions of the present application.
  • the monomeric form of albumin in the composition comprises at least about 95% w/w of the total amount of albumin, preferably at least about 96%, more preferably at least about 98%, more preferably at least about 99%, at least about 99.2% %, at least about 99.4%, or at least about 99.5%. It can be expected that the composition of the present application has the advantage of low immunogenicity and thus high safety.
  • the albumin that can be used in this application is selected from: human serum albumin (HSA), recombinant human albumin (rHA), bovine serum albumin and porcine serum albumin.
  • HSA human serum albumin
  • rHA recombinant human albumin
  • bovine serum albumin bovine serum albumin
  • porcine serum albumin the albumin comprises the amino acid sequence shown in SEQ ID NO:1.
  • the albumin is selected from human serum albumin (HSA) and recombinant human albumin (rHA).
  • the present application provides a method for preparing the above-mentioned composition according to the first aspect, characterized in that the method comprises the following steps:
  • the method comprises the steps of:
  • step (3) the product obtained in step (3) is sterilized.
  • the volume ratio of the second organic solvent to the DMSO or C 1-3 alcohol is about 1:20 (v/v) to about 20:1 (v/v), for example about 1:5 to about 5:1 (v/v), about 1:2 to about 4:1 (v/v), about 1:1 to about 4: 1 (v/v), about 1.5:1 (v/v) to about 3:1 (v/v), or about 2:1 (v/v) to about 7:3 (v/v).
  • the organic phase: the aqueous phase is about 1:2 (v/v) to about 1:50 (v/v), such as about 1:5 (v/v) to about 1:20(v/v), about 1:7(v/v) to about 1:15(v/v), 1:10(v/v) to about 1:12( v/v); for example about 1:5 (v/v) to about 1:12 (v/v), about 1:5 (v/v) to about 1:12 (v/v), about 1:6 (v/v), about 1:7(v/v), or about 1:10(v/v).
  • step (2) includes the following steps:
  • the C 1-3 alcohol includes methanol, ethanol and isopropanol and any combination thereof, for example, ethanol (EtOH).
  • the mixed organic solvent comprises about 7:3 (v/v) of the second organic solvent and ethanol. In some embodiments, the mixed organic solvent comprises about 1:1 (v/v) of the second organic solvent and DMSO.
  • the present invention provides the method described above, wherein said method comprises the steps of:
  • the organic phase is mixed with the aqueous phase at a ratio of about 1:10 (v/v) to about 1:15 (v/v), for example about 1:12 (v/v) to prepare an emulsion , the emulsion comprises the nanoparticles, wherein in the nanoparticles, the albumin encapsulates at least part of the SN-38 and optionally at least part of the lipid;
  • step (3) the product obtained in step (3) is sterilized.
  • the second organic solvent is CHCl 3 .
  • the second organic solvent is a mixture of CH2Cl2 and CHCl3 , wherein preferably, the volume ratio of CH2Cl2 and CHCl3 in the mixture is about 2:5-1: 1, preferably about 2:5.
  • using the mixture of CH 2 Cl 2 and CHCl 3 as the second organic solvent has the advantage of reducing the residual level of CHCl 3 in the final product, thereby reducing the limitation of solvent residue on clinical dosage.
  • the aqueous phase contains no additional stabilizers.
  • the aqueous phase already contains additional stabilizers; in other embodiments, the method further comprises adding additional stabilizers in step (2).
  • the amount of the additional stabilizer is such that in the product obtained in step (3) or (4), the content of the additional stabilizer is at least about 2w/v%, such as at least about 3w/v%, at least About 5w/v%, about 5w/v% to about 30w/v%, about 10w/v% to about 25w/v%, or about 15w/v% to about 20w/v%.
  • the additional stabilizer may be selected from albumins (such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder), monosaccharides, disaccharides, polysaccharides, mannitol and any Combination; preferably selected from: mannitol, lactose, maltose, trehalose, dextran, glucose and sucrose and any combination thereof; preferably sucrose.
  • albumins such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides such as
  • step (2) before the organic phase is mixed with the aqueous phase, the mixed organic solvent described in step (1) is added to the aqueous phase.
  • the volume of the mixed solvent added is equal to or smaller than the volume of the organic phase.
  • the volume ratio of the added mixed organic solvent to the organic phase is about 1:1 (v/v) to about 1:5 (v/v), such as about 1:2 (v/v) to about 1 :4(v/v) or about 1:3(v/v).
  • the concentration of SN-38 is about 5-17 mg/mL, such as about 5.25-12 mg/mL, about 7-12 mg/mL, such as about 10 mg/mL mL.
  • the concentration of SN-38 is 4-10 mg/mL, such as about 6-8 mg/mL.
  • the concentration of the lipid is about 3-50 mg/mL, such as about 5-45 mg/mL or about 7.5-30 mg/mL, about 10- 25mg/mL or about 15-20mg/mL.
  • the concentration of the lipid is about 10-20 mg/mL, such as about 15 mg/mL.
  • the concentration of the Span 20 is about 0.3-6 mg/mL, such as about 0.3-2 mg/mL or about 0.6-1 mg/mL.
  • the concentration of albumin in the aqueous phase is about 5-15 mg/mL, such as about 6-12 mg/mL, preferably about 6-10 mg/mL.
  • the concentration of albumin in the aqueous phase is about 8-30 mg/mL, such as about 12-20 mg/mL or about 16-18 mg/mL.
  • the method comprises step (4) sterilizing the product obtained in step (3).
  • sterilization is performed by filtering the product obtained in step (3) through a filter membrane of about 0.2 ⁇ m.
  • the method further comprises the steps of:
  • step (3) or (4) drying the product obtained in step (3) or (4), preferably spray drying or freeze drying, to provide a solid form, preferably a powder, more preferably a freeze-dried powder composition;
  • SN-38 is present in the composition in amorphous and/or nanocrystalline form, as determined by electron microscopy or X-ray diffraction (Cu-K ⁇ ) analysis.
  • step (5) also includes: before drying, adding an additional stabilizer to the product obtained in step (3) or (4), wherein the amount of the additional stabilizer is such that when step (5) When the obtained solid form is reconstituted to form an aqueous composition (including solutions and emulsions), the content of the additional stabilizer is at least about 2w/v%, such as at least about 3w/v%, at least about 5w/ v %, about 5 w/v % to about 30 w/v %, about 10 w/v % to about 25 w/v % or about 15 w/v % to about 20 w/v %.
  • the additional stabilizer may be selected from: albumins (such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder), monosaccharides, disaccharides, polysaccharides, mannitol and any combination thereof; preferably is selected from: mannitol, lactose, maltose, trehalose, dextran, glucose and sucrose and any combination thereof; preferably sucrose.
  • albumins such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides such as human serum albumin, recombinant human albumin, bovine serum albumin, skimmed milk powder
  • monosaccharides
  • the application provides a method for preparing a composition comprising SN-38, lipids, albumin and Span 20, wherein the method comprises the following steps:
  • Embodiments of the method are as described above for the method of the second aspect.
  • part of the albumin may encapsulate part of the SN-38 to form a nanoparticle, or may encapsulate a part of the lipid to form a nanoparticle. Therefore, in some embodiments, the composition according to the first aspect and the fourth aspect below may comprise nanoparticles formed by encapsulating a part of the SN-38 with part of the albumin; and/or composed of a part of the albumin A nanoparticle formed by encapsulating a portion of the lipid with a protein.
  • the present application provides a composition, which can be prepared by the method according to the third aspect as described above.
  • the present application also provides a method for preparing a composition with improved properties, wherein the composition comprises SN-38, lipids and albumin, and the albumin encapsulates at least part of the SN-38 and optionally at least part of said lipids to form nanoparticles,
  • the method is characterized in that Span 20 is added during the preparation of the composition.
  • the composition contains no additional stabilizers.
  • the improved properties include increased stability.
  • the increased stability comprises reduced production or levels of albumin multimers during manufacture, storage and/or use of the composition , and/or reduced nanoparticle size.
  • no or substantially no albumin multimers are present in the composition, or albumin multimers account for at most 5 w/w% of the total albumin, for example at most about 4%, at most about 2%, at most about 1.5%, up to about 1.2%, up to about 1.1%, up to about 1%, or up to about 0.8%.
  • composition is as described above in the first aspect.
  • the method comprises the steps of:
  • the present application provides a pharmaceutical composition, which comprises the composition described above and optionally a pharmaceutically acceptable carrier.
  • the present application provides a pharmaceutical composition, which comprises the dried composition described above and optionally a pharmaceutically acceptable carrier.
  • the drying is preferably freeze-drying or spray-drying, more preferably freeze-drying.
  • the pharmaceutical composition is in solid form, preferably lyophilized powder.
  • SN-38 is present in the composition in amorphous and/or nanocrystalline form.
  • the route of administration of the composition or pharmaceutical composition of the present application includes, but are not limited to, oral, nasal, topical, and parenteral.
  • the pharmaceutical composition is for parenteral administration, including but not limited to intravenous, intraarterial, subcutaneous, intradermal and intramuscular injection administration, more preferably intravenous injection (e.g., bolus injection or infusion ) administration.
  • a pharmaceutically acceptable carrier depends on the dosage form of the drug or pharmaceutical composition, above all on the route of administration of the dosage form (e.g. a dosage form for oral, nasal, intradermal, subcutaneous, topical, intramuscular or intravenous administration), Second depends on the formulation of the dosage form.
  • the pharmaceutically acceptable carrier can include water (such as water for injection), buffer, isotonic saline solution such as PBS (phosphate buffer saline), glucose, mannitol, dextrose, lactose, starch, stearic acid Magnesium acid, cellulose, magnesium carbonate, 0.3% glycerin, hyaluronic acid, ascorbic acid, lactic acid, ethanol, polyalkylene glycols such as polyethylene glycol (e.g. polyethylene glycol 4000) or polypropylene glycol, triglycerides, etc. .
  • water such as water for injection
  • buffer isotonic saline solution
  • glucose mannitol
  • dextrose mannitol
  • lactose lactose
  • starch starch
  • the present application provides the use of the above-mentioned composition or pharmaceutical composition in the preparation of a medicament for treating an individual's tumor sensitive to SN-38.
  • the present application provides the composition or pharmaceutical composition described above, which is used for treating tumors sensitive to SN-38 in an individual.
  • the present application provides a method for treating a tumor sensitive to SN-38 in an individual, comprising administering to the individual a therapeutically effective amount of the above-described composition or pharmaceutical composition.
  • the individual is a mammal, including but not limited to mice, rats, rabbits, guinea pigs, dogs, cats, sheep, cows, sheep, horses. In some embodiments, the individual is a human.
  • SN-38 sensitive tumor refers to a tumor that responds to the administration of SN-38, said response including reduction of tumor cells, reduction in tumor size, elimination of tumor metastasis, inhibition of tumor growth, etc. .
  • the tumor sensitive to SN-38 is selected from colorectal cancer, small cell lung cancer, lymphoma, breast cancer (preferably triple negative breast cancer), esophageal cancer, gastric cancer, liver cancer, kidney cancer, pancreatic cancer, uterine cancer and ovarian cancer.
  • the composition or pharmaceutical composition of the present application has favorable anti-tumor efficacy.
  • the composition of the present application has an antitumor activity superior to that of the commercially available irinotecan hydrochloride injection.
  • the present application provides a kit comprising the above-mentioned composition or pharmaceutical composition. If necessary, the kit may also include instructions for use, packaging, and a container for containing the composition or pharmaceutical composition.
  • the embodiments of the present invention have been described above with SN-38 as the active ingredient in the first to eleventh aspects, these are only one aspect of the concept of the present invention.
  • the concept of the present invention also covers the technical solutions in which other camptothecin drugs are used as active ingredients. It is expected that the embodiments described above will still apply when SN-38 is replaced by other camptothecins.
  • the present application also includes any and all of the embodiments described in any one of the first to tenth aspects above, except that SN-38 is replaced by other camptothecin drugs, unless there is a conflict in the context.
  • camptothecin drugs can be selected from: irinotecan (CPT-11), 10-hydroxycamptothecin (HCPT), topotecan (TPT), rubitecan (9-NC), 9- Aminocamptothecin (9-AC), Belotecan (Cas.No.: 256411-32-2), Dxd (Cas.No.: 1599440-33-1), DX-8951 (Exatecan) , CKD602 (belotecan), lertotecan (lurtotecan), Namitecan (Cas.No.:372105-27-6), ST1481 (gimatecan, Cas.No.:292618-32-7), BNP-1350 (Cas. No. 203923-89-1) and BN80915 (Diflomotecan).
  • CPT-11 10-hydroxycamptothecin
  • HCPT topotecan
  • rubitecan 9-NC
  • Embodiment 1 A composition comprising SN-38, lipids, albumin and Span 20, characterized in that said composition comprises nanoparticles, wherein in said nanoparticles said albumin encapsulates at least part of said SN-38 and optionally at least part of said lipid;
  • Lipid: SN-38 is about (0.1-10):1(w:w), about (0.5-6):1(w:w), about (0.5-5):1(w:w), about (0.5-3):1(w:w), about (1-4):1(w:w), about (1.2-4):1(w:w), about (1.4-2):1( w:w), about (1.5-2.5):1(w:w) or about 1:1;
  • Albumin: SN-38 is about (1-100):1(w:w), about (1-50):1(w:w), about (3-25):1(w:w), about (5-25): 1(w:w), about (5-20): 1(w:w), about (5-18): 1(w:w), about (6-15): 1( w:w), about (7-15):1(w:w), about (6-12):1(w:w), about (7-12):1(w:w), about (9 -11):1(w:w), about 10:1(w:w); and
  • Span 20:SN-38 is about (3-60):100(w:w), about (4-60):100(w:w), about (5-60):100(w:w), about (6-60):100(w:w), about (7-55):100(w:w), about (8-50):100(w:w), about (10-45):100( w:w), approx.(12-40):100(w:w), approx.(14-35):100(w:w), approx.(15-30):100(w:w), approx.(16 -25):100(w:w), or about (18-20):100(w:w); and
  • the lipid is selected from cholesterol, cholesterol derivatives, cholesterol analogs and fatty acid esters and any combination of two or more of them.
  • Embodiment 2 The composition of embodiment 1, characterized in that albumin:lipid is about (1-100):1(w:w), for example about (2-20):1(w:w), About (3-15):1(w:w), about (5-10):1(w:w), about 7:1(w:w), or about 10:1(w:w).
  • albumin:lipid is about (1-100):1(w:w), for example about (2-20):1(w:w), About (3-15):1(w:w), about (5-10):1(w:w), about 7:1(w:w), or about 10:1(w:w).
  • Embodiment 3 The composition of embodiment 1 or 2, characterized in that, based on the total amount of SN-38, lipid and albumin in the composition,
  • the content of the SN-38 is about 1w/w% to about 25w/w%; and/or
  • the content of the lipid is about 1w/w% to about 35w/w%; and/or
  • the albumin content is about 50w/w% to about 98w/w%
  • the content of the SN-38 is about 3w/w% to about 20w/w%; and/or
  • the content of the lipid is about 2w/w% to about 30w/w%; and/or
  • the albumin content is about 55w/w% to about 95w/w%;
  • the content of SN-38 is about 3w/w% to about 15w/w%, about 4w/w%, about 5w/w%, about 6w/w%, about 6.5w/w%, about 7w/w% , About 7.5w/w%, About 8w/w%, About 8.5w/w%, About 9w/w%, About 10w/w%, About 11w/w%, About 12w/w%, About 13w/w% or about 14w/w%; and/or
  • the lipid content is about 3w/w% to about 30w/w%, about 4w/w%, about 5w/w%, about 6w/w%, about 7w/w%, about 8w/w%, about 8.5w/w%, about 9w/w%, about 9.5w/w%, about 10w/w%, about 10.5w/w%, about 11w/w%, about 11.5w/w%, about 12w/w% , About 12.5w/w%, About 13w/w%, About 13.5w/w%, About 14w/w%, About 15w/w%, About 16w/w%, About 17w/w%, About 18w/w% , about 19w/w%, about 20w/w%, about 21w/w%, about 24w/w%, about 26w/w%, or about 28w/w%; and/or
  • the content of the albumin is about 60w/w% to about 94w/w%, about 64w/w% to about 93w/w%, about 66w/w% to about 92w/w%, about 68w/w% to about 91w/w%, about 70w/w% to about 90w/w%, about 75w/w% to about 90w/w%, about 76w/w%, about 77w/w%, about 78w/w%, about 79w/ w%, about 80w/w%, about 81w/w%, about 82w/w%, about 83w/w%, about 84w/w%, about 85w/w%, about 86w/w%, about 87w/w% , about 88w/w%, or about 89w/w%.
  • Embodiment 4 The composition of any one of embodiments 1 to 3, characterized in that, based on the total amount of SN-38, lipid, albumin and Span 20 in the composition, the content of Span 20 is About 0.03w/w% to about 12w/w%, about 0.06w/w% to about 10w/w%, about 0.08w/w% to about 9w/w%, about 0.1w/w% to about 8w/w% , about 0.2w/w% to about 7w/w%, about 0.4w/w% to about 6w/w%, about 0.6w/w% to about 5w/w%, about 0.8w/w% to about 4w/ w%, about 1 w/w% to about 2 w/w%.
  • Embodiment 5 The composition of any one of embodiments 1 to 4, characterized in that,
  • Lipids: SN-38 is about (1-10):1(w:w), about (1-8):1(w:w), about (1-6):1(w:w), about (1-5):1(w:w), about (1-4.5):1(w:w), about (1-4):1(w:w), about (1.2-3.8):1( w:w), about (1.4-3.6):1(w:w), about (1.6-3.4):1(w:w), about (1.8-3.2):1(w:w), about (2 -3):1(w:w), about (2.2-2.8):1(w:w), about (2.4-2.6):1(w:w), about 1:1(w:w), about 2.5:1(w:w), or about (0.8-1.8):1(w:w), about (0.9-1.7):1(w:w), about (1-1.4):1; and/or
  • Albumin: SN-38 is about (5-40):1(w:w), about (5-less than 40):1(w:w), about (5-35):1(w:w), About (5-30): 1 (w: w), about (8-25): 1 (w: w), about (10-22.5): 1 (w: w), about (12.5-20): 1 (w:w), about (15-17.5):1(w:w), about (16-18):1(w:w), about 10:1(w:w), or about (9-21 ):1(w:w), about (9-20):1(w:w), about (11-18):1(w:w), about (11.1-17.3):1(w:w) ;and / or
  • Span 20:SN-38 is about (5-60):100(w:w), about (6-60):100(w:w), about (7-55):100(w:w), about (8-50):100(w:w), about (10-45):100(w:w), about (12-40):100(w:w), about (14-35):100( w:w), about (15-30):100(w:w), about (16-25):100(w:w), about (18-20):100(w:w), or about ( 5-10): 100(w:w), about (5-9): 100(w:w), about (6-8.6): 100(w:w), about (6-8): 100(w :w), about (6.5-7):100(w:w); and/or
  • Lipid is about (1-40):1(w:w), about (1-less than 40):1(w:w), about (2-35):1(w:w), about (3-15):1(w:w), about (5-10):1(w:w), about (6-8):1(w:w) or about 7:1(w:w) , or about (6-21):1(w:w), about (6.7-13):1(w:w), about (7-13):1(w:w), about (11-12.7) :1(w:w).
  • Embodiment 6 The composition of any one of embodiments 1 to 5, characterized in that, based on the total amount of SN-38, lipid and albumin in the composition,
  • the content of SN-38 is about 2w/w% to about 16w/w%; and/or
  • the content of the lipid is about 2w/w% to about 35w/w%; and/or
  • the albumin content is about 75w/w% to about 96w/w%
  • the content of the SN-38 is about 2.5w/w% to about 15w/w%, about 4w/w% to about 10w/w%, about 4.5w/w% to about 9.5w/w%, about 5w/ w% to about 9w/w%, or about 7.5w/w% to about 8w/w%; and/or
  • the lipid content in the composition is about 2.5w/w% to about 30w/w%, about 4w/w% to about 12.5w/w%, about 4.5w/w% to about 12w/w%, about 7w/w% to about 10w/w%, or about 7.5w/w% to about 8w/w%; and/or
  • the content of albumin in the composition is about 76w/w% to about 95w/w%, about 78w/w% to about 93w/w%, about 79w/w% to about 91.5w/w%, about 80w /w% to about 90w/w%, about 82w/w% to about 89w/w%, about 84w/w% to about 88w/w%, or about 84.5w/w% to about 87.5w/w%;
  • the content of the SN-38 is about 3w/w% to about 14w/w%, about 3.5w/w% to about 12w/w%, about 4w/w%, about 4.2w/w%, about 4.5w/ w%, about 4.6w/w%, about 4.8w/w%, about 5w/w%, about 5.5w/w%, about 6w/w%, about 6.5w/w%, about 7w/w%, about 7.5w/w%, about 7.6w/w%, about 7.8w/w%, about 8w/w%, about 8.5w/w%, about 9w/w%, about 9.2w/w%, about 9.5w/ w%, about 9.6w/w%, about 9.8w/w%, about 10w/w%, about 10.5w/w%, about 11w/w%, or about 11.5w/w%; and/or
  • the lipid content is about 4w/w% to about 25w/w%, about 4.5w/w% to about 20w/w%, about 4.3w/w%, about 4.5w/w%, about 4.7w/ w%, about 5w/w%, about 5.5w/w%, about 6w/w%, about 6.5w/w%, about 6.7w/w%, about 6.9w/w%, about 7w/w%, about 7.5w/w%, about 7.6w/w%, about 7.8w/w%, about 8w/w%, about 8.5w/w%, about 9w/w%, about 9.5w/w%, about 10w/w %, about 10.5w/w%, about 11w/w%, about 11.5w/w%, about 12w/w%, about 12.1w/w%, about 12.3w/w%, about 12.5w/w%, about 13w/w%, about 13.5w/w%, about 14w/w%, about 14.5w
  • the content of the albumin is about 78w/w% to about 92w/w%, about 79w/w%, about 79.2w/w%, about 79.4w/w%, about 79.6w/w%, about 79.8w/ w%, about 80w/w%, about 81w/w%, about 82w/w%, about 83w/w%, about 84w/w%, about 84.3w/w%, about 84.5w/w%, about 84.7w /w%, about 84.9w/w%, about 85w/w%, about 86w/w%, about 87w/w%, about 87.3w/w%, about 87.5w/w%, about 87.7w/w%, About 87.9w/w%, about 88w/w%, about 89w/w%, about 90w/w%, about 91w/w%, about 91.3w/w%, or about 91.5w/w%.
  • Embodiment 7 The composition of any one of embodiments 1 to 6, characterized in that, based on the total amount of SN-38, lipid, albumin and Span 20 in the composition, the content of Span 20 is About 0.14w/w% to about 5w/w%, about 0.2w/w% to about 2.5w/w%, about 0.22w/w% to about 2.0w/w%, about 0.24w/w% to about 2w /w%, about 0.26w/w% to about 1.5w/w%, about 0.28w/w% to about 1.0w/w%, about 0.3w/w% to about 0.9w/w%, about 0.32w/ w% to about 0.8w/w%, about 0.34w/w% to about 0.7w/w%, about 0.36w/w% to about 0.6w/w%, about 0.38w/w% to about 0.58w/w %, about 0.4w/w% to about 0.56w/w%, about 0.42w/w% to about 0.54
  • Embodiment 8 The composition of any one of embodiments 1 to 7, characterized in that,
  • the SN-38 present in the nanoparticles accounts for at least about 1 w/w % or at least about 2 w/w %, such as at least about 3 w/w % of the total amount of SN-38, lipid and albumin in the composition. w%, about 3w/w% to about 13w/w%, about 4w/w% to about 12w/w%, about 4w/w%, about 5w/w%, about 6w/w%, about 7w/w% , about 8w/w%, about 9w/w%, about 10w/w%, or about 11w/w%; and/or
  • SN-38 present in the nanoparticles represents from about 80 w/w % to about 99 w/w % of the total amount of SN-38 in the composition, such as from about 88 w/w % to about 98 w/w %, about 89w/w%, about 90w/w%, about 91w/w%, about 92w/w%, about 93w/w%, about 94w/w%, about 95w/w%, about 96w/w%, or about 97w /w%.
  • Embodiment 9 The composition of any one of embodiments 1 to 8, characterized in that,
  • the cholesterol derivative is selected from esters of cholesterol and organic acids, including cholesterol palmitate and cholesterol caprylate and combinations thereof; and/or;
  • the cholesterol analog is selected from vitamin D2, vitamin D3 and combinations thereof; and/or
  • the fatty acid ester is selected from fatty acid glycerides, such as long-chain fatty acid glycerides, including glyceryl stearate, such as glyceryl monostearate.
  • Embodiment 10 The composition of any one of embodiments 1 to 9, wherein the lipid is cholesterol.
  • Embodiment 11 The composition of embodiment 10, characterized in that,
  • Cholesterol: SN-38 is about (1-6):1(w:w), about (1.2-5):1(w:w), about (1.4-4):1(w:w), about 3 :1(w:w), about 2:1(w:w), about 1:1(w:w), or about (0.8-1.8):1(w:w), about (0.9-1.7): 1(w:w), about (1-1.4):1; and/or
  • Albumin: SN-38 is about (3-25):1(w:w), about (4-20):1(w:w), about (5-15):1(w:w), about (6-12):1(w:w), about (7-12):1(w:w), about (9-11):1(w:w) or about 10:1(w:w) , or about (9-21):1(w:w), about (9-20):1(w:w), about (11-18):1(w:w), about (11.1-17.3) :1(w:w); and/or
  • the content of the SN-38 is about 3w/w% to about 15w/w%, about 4w/w% to about 15w/w%, about 6w/w% to about 10w/w%, or about 8w/w% to about About 12w/w%, or about 4w/w% to about 10w/w%, about 4.5w/w% to about 9.5w/w%, about 5w/w% to about 9w/w%, about 7.5w/w % to about 8w/w%; and/or
  • the cholesterol content is about 5w/w% to about 25w/w%, about 6w/w% to about 22w/w%, about 15w/w% to about 20w/w%, or about 4w/w% to about 12.5w/w%, about 4.5w/w% to about 12w/w%, about 7w/w% to about 10w/w%, about 7.5w/w% to about 8w/w%; and/or
  • the content of the albumin is about 64w/w% to about 90w/w%, about 70w/w% to about 90w/w%, or about 78w/w% to about 93w/w%, about 79w/w% to About 91.5w/w%, about 80w/w% to about 90w/w%, about 82w/w% to about 89w/w%, about 84w/w% to about 88w/w%, about 84.5w/w% to about 87.5w/w%; and/or
  • SN-38 present in the nanoparticles represents at least about 3 w/w %, such as about 3 w/w % to about 13 w/w %, of the total amount of SN-38, cholesterol and albumin in the composition, About 4w/w% to about 12w/w%, about 4w/w%, about 5w/w%, about 6w/w%, about 7w/w%, about 8w/w%, about 9w/w%, about 10w /w% or about 11w/w%.
  • Embodiment 12 The composition of embodiment 10, wherein cholesterol:SN-38 is about (1-3):1(w:w), about (1.2-2.5):1(w:w), about (1.4- 2):1(w:w), about (1.5-2):1(w:w), about (1.3-1.8):1(w:w), about (1.4-1.6):1(w:w) ), about (1.5-1.7):1(w:w), about (1.2-1.5):1(w:w), about 1:1(w:w), about (1.4-1.5):1(w :w), or about (0.8-1.8):1(w:w), about (0.9-1.7):1(w:w), about (1-1.4):1;
  • Albumin: SN-38 is about (5-15):1(w:w), about (5-12):1(w:w), about (6-12):1(w:w) or about (7-12):1(w:w), about (9-11):1(w:w), about (10-12):1(w:w), about 11:1(w:w) , or about (9-21):1(w:w), about (9-20):1(w:w), about (11-18):1(w:w), or about (11.1-17.3 ):1(w:w); and
  • Albumin Cholesterol is about (3-10):1(w:w), about (4-8):1(w:w), about (5-7):1(w:w), or about ( 6-21):1(w:w), about (6.7-13):1(w:w), about (7-13):1(w:w), about (11-12.7):1(w :w).
  • Embodiment 13 The composition of embodiment 12, characterized in that, based on the total amount of SN-38, cholesterol and albumin in the composition,
  • the content of the SN-38 is about 6w/w% to about 14w/w%, about 6.5w/w% to about 13w/w%, about 7w/w% to about 12w/w%, about 7.5w/w % to about 12w/w%, about 8w/w% to about 11w/w%, about 8.5w/w% to about 10w/w%, about 9w/w%, or about 4w/w% to about 10w/w %, about 4.5w/w% to about 9.5w/w%, about 5w/w% to about 9w/w%, or about 7.5w/w% to about 8w/w%; and/or
  • the cholesterol content is about 8w/w% to about 18w/w%, about 8.5w/w% to about 17w/w%, about 9w/w% to about 16w/w%, about 9.5w/w% to about About 16w/w%, about 10w/w% to about 16w/w%, about 10.5w/w% to about 16w/w%, about 11w/w% to about 15w/w%, about 11.5w/w% to about 15w/w%, about 12w/w% to about 15w/w%, about 12.5w/w% to about 14w/w%, about 13w/w% to about 13.5w/w%, or about 4w/w% to about 12.5w/w%, about 4.5w/w% to about 12w/w%, about 7w/w% to about 10w/w%, or about 7.5w/w% to about 8w/w%; and/or
  • the content of the albumin is about 66w/w% to about 90w/w%, about 68w/w% to about 89w/w, about 70w/w% to about 88w/w%, about 70w/w% to about 87w /w%, about 70w/w% to about 86w/w%, about 70w/w% to about 85w/w%, about 75w/w% to about 85w/w%, about 76w/w%, about 77w/w %, about 78w/w%, about 79w/w%, 80w/w%, 81w/w%, about 82w/w%, about 83w/w%, about 84w/w%, about 78w/w% to about 93w/ w%, about 79w/w% to about 91.5w/w%, about 80w/w% to about 90w/w%, about 82w/w% to about 89w/w%, about 84w/w% to about
  • Embodiment 14 The composition of embodiment 10, characterized in that,
  • Cholesterol: SN-38 is about (1-5):1(w:w), about (1-4.5):1(w:w), about (1-4):1(w:w), about ( 1.2-3.8):1(w:w), about (1.4-3.6):1(w:w), about (1.6-3.4):1(w:w), about (1.8-3.2):1(w :w), about (2-3):1(w:w), about (2.2-2.8):1(w:w), about (2.4-2.6):1(w:w), about 2.5:1 (w:w), about 1:1(w:w), or about (0.8-1.8):1(w:w), about (0.9-1.7):1(w:w), about (1-1.4 ):1; and/or
  • Albumin: SN-38 is about (5-25):1(w:w), about (5-20):1(w:w), about (6-19):1(w:w), about (7-18):1(w:w), about (8-16):1(w:w), about (9-14):1(w:w) or about (10-12):1( w:w), or about (9-21):1(w:w), about (9-20):1(w:w), about (11-18):1(w:w), or about (11.1-17.3):1(w:w); and/or
  • Span 20:SN-38 is about (5-40):100(w:w), about (6-30):100(w:w), about (7-25):100(w:w), about (8-20):100(w:w), about (9-15):100(w:w), about (10-12):100(w:w), or about (5-10):100 (w:w), about (5-9):100(w:w), about (6-8.6):100(w:w), about (6-8):100(w:w), about ( 6.5-7):100(w:w); and/or
  • the content of the SN-38 is about 3w/w% to about 10w/w%, about 3.5w/w% to about 9.5w/w%, about 4w/w%, about 4.5w/w%, about 5w/ w%, about 5.5w/w%, about 6w/w%, about 6.5w/w%, about 7w/w%, about 7.5w/w%, about 8w/w%, about 8.5w/w%, about 9w/w%, or about 4w/w% to about 10w/w%, about 4.5w/w% to about 9.5w/w%, about 5w/w% to about 9w/w%, about 7.5w/w% to about 8 w/w %; and/or
  • the cholesterol content is about 4w/w% to about 18w/w%, about 4.5w/w% to about 17.5w/w%, about 5w/w%, about 5.5w/w%, about 6w/w% , about 6.5w/w%, about 7w/w%, about 7.5w/w%, about 8w/w%, 8.5w/w%, about 9w/w%, about 9.5w/w%, about 10w/w %, about 10.5w/w%, about 11w/w%, about 11.5w/w%, about 12w/w%, about 12.5w/w%, about 13w/w%, about 13.5w/w%, about 14w /w%, about 14.5w/w%, about 15w/w%, about 15.5w/w%, about 16w/w%, about 16.5w/w%, about 17w/w%, or about 4w/w% to about 12.5w/w%, about 4.5w/w% to about 12w/w%,
  • the content of the albumin is about 78w/w% to about 92w/w%, about 79w/w%, about 80w/w%, about 81w/w%, about 82w/w%, about 83w/w%, about 84w/w%, about 85w/w%, about 86w/w%, about 87w/w%, about 88w/w%, about 89w/w%, about 90w/w%, about 91w/w%, or about 78w /w% to about 93w/w%, about 79w/w% to about 91.5w/w%, about 80w/w% to about 90w/w%, about 82w/w% to about 89w/w%, about 84w/ w% to about 88w/w%, or about 84.5w/w% to about 87.5w/w%.
  • Embodiment 15 The composition of any one of embodiments 12 to 14, characterized in that,
  • the SN-38 present in the nanoparticles accounts for at least about 6 to about 12 w/w % of the total amount of SN-38, cholesterol and albumin in the composition, for example about 7 to 7 w/w % about 11w/w%, about 8w/w% to about 10w/w%, about 8.3% or about 9w/w%; and/or
  • SN-38 present in the nanoparticles represents from about 95 w/w % to about 99 w/w % of the total amount of SN-38 in the composition, such as from about 96 w/w % to about 99 w/w %, About 97w/w% to about 99w/w%, about 98w/w% to about 99w/w%, about 99w/w% or higher.
  • Embodiment 16 The composition according to any one of embodiments 1 to 15, characterized in that said composition is in liquid, semi-solid or solid form.
  • Embodiment 17 The composition according to any one of embodiments 1 to 16, wherein the composition is in solid form, preferably in powder form, more preferably in freeze-dried powder;
  • SN-38 is present in the composition in amorphous and/or nanocrystalline form.
  • Embodiment 18 The composition according to any one of embodiments 1 to 17, characterized in that said composition does not comprise additional stabilizers; or
  • the composition further comprises an additional stabilizer, such as a lyophilized stabilizer, in an amount such that when the composition is reconstituted to form an aqueous composition (including solutions and emulsions), the
  • additional stabilizer is at least about 2w/v%, for example at least about 3w/v%, for example at least about 5w/v%, about 5w/v% to about 30w/v%, about 10w/v% to about 25w/v% or about 15w/v% to about 20w/v%.
  • Embodiment 19 The composition of embodiment 17 or 18, characterized in that the composition further comprises an additional stabilizer, based on the total amount of the composition,
  • the content of the additional stabilizer is about 60w/w% to about 98w/w%, for example about 65w/w% to about 97w/w%, about 68w/w% to about 96w/w%, about 69w/w % to about 95w/w%, about 70w/w% to about 94w/w%, about 71w/w% to about 93w/w%, about 72w/w% to about 92w/w%, about 73w/w%, About 74w/w%, about 75w/w%, about 76w/w%, about 77w/w%, about 78w/w%, about 79w/w%, about 80w/w%, about 81w/w%, about 82w /w%, about 83w/w%, about 84w/w%, about 85w/w%, about 86w/w%, about 87w/w%, about 88w/w%, about 89w/w%, about 90
  • the content of the additional stabilizer is about 70w/w% to about 96w/w%, about 70w/w% to about 90w/w%, about 72w/w% to about 89w/w%, about 74w/w% w% to about 88w/w%, about 76w/w% to about 87w/w%, or about 80w/w% to about 96w/w%, about 80w/w% to about 86w/w%, about 81w/w % to about 86w/w%, about 82w/w% to about 85w/w%, about 83w/w% to about 84w/w%, about 84w/w% to about 95w/w%.
  • composition of embodiment 18 or 19, wherein the additional stabilizer is selected from the group consisting of: albumins (e.g. human serum albumin, recombinant human albumin, bovine serum albumin, skim milk powder), monosaccharides, disaccharides , polysaccharide, mannitol and any combination thereof; preferably selected from: mannitol, lactose, maltose, trehalose, dextran, glucose and sucrose and any combination thereof; preferably sucrose.
  • albumins e.g. human serum albumin, recombinant human albumin, bovine serum albumin, skim milk powder
  • monosaccharides e.g. human serum albumin, recombinant human albumin, bovine serum albumin, skim milk powder
  • monosaccharides e.g. human serum albumin, recombinant human albumin, bovine serum albumin, skim milk powder
  • monosaccharides e.g. human serum albumin, recombinant
  • Embodiment 21 The composition of any one of embodiments 17 to 20, wherein when said composition is reconstituted to form an aqueous composition (including solutions and emulsions), it has from about 0.1 ⁇ g/mL to about 30.0 mg/mL , the nanoparticles have an average particle diameter of about 50 to 200 nm, such as about 90 to 150 nm, about 95 to 140 nm, about 100 to 130 nm, about 105 to 125 nm, or about 110 to 120 nm.
  • Embodiment 22 The composition of any one of embodiments 1 to 16, wherein said composition is an aqueous composition in liquid form, including solutions and emulsions;
  • said composition in liquid form comprises SN-38 in the form of nanocrystals and/or vesicles.
  • Embodiment 23 The composition of embodiment 22, characterized in that, based on the total amount of the composition,
  • SN-38 is present in an amount of about 0.1 ⁇ g/mL to about 30.0 mg/mL, about 0.2 ⁇ g/mL to about 27.0 mg/mL, about 0.5 ⁇ g/mL to about 24.0 mg/mL, about 1.0 ⁇ g/mL to about 21.0 mg/mL, about 5.0 ⁇ g/mL to about 18.0 mg/mL, about 10.0 ⁇ g/mL to about 15.0 mg/mL, about 20.0 ⁇ g/mL to about 12 mg/mL, about 25.0 ⁇ g/mL to about 9 mg/mL, about 50.0 ⁇ g/mL to about 6.0 mg/mL or about 100.0 ⁇ g/mL to about 3.0 mg/mL; and/or
  • the content of the lipid is about 0.05 ⁇ g/mL to about 100.0 mg/mL, about 0.1 ⁇ g/mL to about 90.0 mg/mL, about 0.25 ⁇ g/mL to about 80.0 mg/mL, about 0.5 ⁇ g/mL to about 70.0 mg/mL, about 2.5 ⁇ g/mL to about 60.0 mg/mL, about 5.0 ⁇ g/mL to about 50.0 mg/mL, about 10.0 ⁇ g/mL to about 40.0 mg/mL, about 12.5 ⁇ g/mL to about 30.0 mg /mL, about 25.0 ⁇ g/mL to about 20.0 mg/mL, or about 50.0 ⁇ g/mL to about 10.0 mg/mL; and/or
  • Embodiment 24 The composition according to embodiment 22 or 23, characterized in that, based on the total amount of the composition,
  • the content of SN-38 is about 100.0 ⁇ g/mL to about 3.0 mg/mL, such as about 200.0 ⁇ g/mL to about 2.5 mg/mL, about 300.0 ⁇ g/mL to about 2.0 mg/mL, about 400.0 ⁇ g/mL to about 1.5 mg/mL, about 500.0 ⁇ g/mL to about 1.0 mg/mL, or about 600 ⁇ g/mL to about 800 ⁇ g/mL; and/or
  • the lipid content is from about 50.0 ⁇ g/mL to about 10.0 mg/mL, such as from about 100.0 ⁇ g/mL to about 8.0 mg/mL, from about 200.0 ⁇ g/mL to about 6.0 mg/mL, from about 300.0 ⁇ g/mL to About 4.0 mg/mL, about 400.0 ⁇ g/mL to about 3.0 mg/mL, about 500.0 ⁇ g/mL to about 2.5 mg/mL, about 600.0 ⁇ g/mL to about 2.0 mg/mL, about 700.0 ⁇ g/mL to about 1.5 mg/mL, about 800 ⁇ g/mL to about 1.0 mg/mL, or about 200 ⁇ g/mL to about 1.5 mg/mL; and/or
  • the content of albumin is about 3.0 mg/mL to about 30.0 mg/mL, such as about 4.0 mg/mL to about 25.0 mg/mL, about 5.0 mg/mL to about 20.0 mg/mL, about 6.0 mg/mL to about 15.0 mg/mL, about 7.0 mg/mL to about 12.0 mg/mL, or about 8.0 mg/mL to about 10.0 mg/mL.
  • Embodiment 25 The composition of any one of embodiments 22 to 24, wherein the nanoparticles have a particle size of about 50 to 200 nm, such as about 90 to 150 nm, about 95 to 140 nm, about 100 to 130 nm, about 105 to 125 nm or an average particle size of about 110 to 120 nm.
  • Embodiment 26 The composition of any one of embodiments 22 to 25, characterized in that, when stored at 4° C. for 24 h, the nanoparticles have an the average particle size.
  • Embodiment 27 The composition of any one of Embodiments 22 to 26, wherein the nanoparticles have a particle size distribution of no more than about 0.30, such as no more than about 0.2, no more than about 0.10, or no more than about 0.01 Index (PDI).
  • PDI 0.01 Index
  • Embodiment 28 The composition of any one of Embodiments 22 to 27, characterized in that said composition has a Zeta potential of from about -35 mV to about -20 mV, for example about -31 mV.
  • Embodiment 29 The composition of any one of embodiments 22 to 28, characterized in that when diluted (for example with 1x PBS at pH about 7.4) said composition is such that the amount of SN-38 in the diluted composition Levels of about 4 ⁇ g/mL or less, such as about 2 ⁇ g/mL or less, about 1 ⁇ g/mL or less, about 0.4 ⁇ g/mL or less, about 0.1 ⁇ g/mL or less, about 0.04 ⁇ g/mL or less, about 0.02 ⁇ g/mL or less, or about 0.01 ⁇ g/mL or less, the nanoparticles do not disintegrate.
  • Embodiment 30 The composition of any one of embodiments 22 to 29, characterized in that the composition does not comprise additional stabilizers; or
  • the composition further comprises an additional stabilizer, wherein based on the total amount of the composition, the amount of the additional stabilizer is at least about 2w/v%, such as at least about 3w/v%, at least About 5w/v%, about 5w/v% to about 30w/v%, about 10w/v% to about 25w/v%, or about 15w/v% to about 20w/v%.
  • the additional stabilizer is at least about 2w/v%, such as at least about 3w/v%, at least About 5w/v%, about 5w/v% to about 30w/v%, about 10w/v% to about 25w/v%, or about 15w/v% to about 20w/v%.
  • Embodiment 31 The composition of embodiment 30, wherein the additional stabilizer is selected from the group consisting of albumins (e.g., human serum albumin, recombinant human albumin, bovine serum albumin, skim milk powder), monosaccharides, disaccharides , polysaccharide, mannitol and any combination thereof; preferably selected from: mannitol, lactose, maltose, trehalose, dextran, glucose and sucrose and any combination thereof; preferably sucrose.
  • albumins e.g., human serum albumin, recombinant human albumin, bovine serum albumin, skim milk powder
  • Embodiment 32 The composition of any one of embodiments 1 to 31, wherein the ring-opened SN-38 in the composition accounts for about 2 w/w% or less of the total amount of SN-38, for example about 1.8w/w% or less; and/or
  • Albumin polymers are absent or substantially absent in the composition; for example, albumin in monomeric form in the composition comprises at least about 95% w/w of the total amount of albumin, such as at least about 96%, at least About 98%, at least about 99%, at least about 99.2%, at least about 99.4%, or at least about 99.5%.
  • composition according to any one of embodiments 1 to 32 characterized in that the albumin is selected from the group consisting of human serum albumin (HSA), recombinant human albumin (rHA), bovine serum albumin and porcine serum albumin Protein;
  • the albumin comprises the amino acid sequence shown in SEQ ID NO:1;
  • the albumin is selected from human serum albumin (HSA) and recombinant human albumin (rHA).
  • HSA human serum albumin
  • rHA recombinant human albumin
  • Embodiment 34 A method of preparing the composition of any one of embodiments 1 to 33, characterized in that said method comprises the steps of:
  • Embodiment 35 The method of embodiment 34, characterized in that the method comprises the steps of:
  • the volume ratio of said DMSO or C 1-3 alcohol is about 1:20 (v/v) to about 20:1 (v/v), for example about 1:5 to about 5:1 (v/v), about 1 :2 to about 4:1 (v/v), about 1:1 to about 4:1 (v/v), about 1.5:1 (v/v) to about 3:1 (v/v) or about 2 :1 (v/v) to about 7:3 (v/v); and preparing an aqueous solution of albumin as the aqueous phase;
  • step (3) sterilizing the product obtained in step (3), preferably by filtering through a filter membrane of about 0.2 ⁇ m;
  • the second organic solvent is CHCl 3 ; or a mixture of CH 2 Cl 2 and CHCl 3 , wherein optionally, the volume ratio of CH 2 Cl 2 and CHCl 3 in the mixture is about 2:5 - 1:1, preferably about 2:5.
  • Embodiment 36 The method of embodiment 34 or 35, characterized in that, in step (2), the organic phase: the aqueous phase is about 1:2 (v/v) to about 1:50 (v/ v), for example about 1:5 (v/v) to about 1:20 (v/v), about 1:7 (v/v) to about 1:15 (v/v), about 1:10 (v /v) to about 1:12 (v/v); for example about 1:5 (v/v) to about 1:12 (v/v), about 1:5 (v/v) to about 1:12 ( v/v), about 1:6(v/v), about 1:7(v/v) or about 1:10(v/v).
  • the organic phase: the aqueous phase is about 1:2 (v/v) to about 1:50 (v/ v), for example about 1:5 (v/v) to about 1:20 (v/v), about 1:7 (v/v) to about 1:15 (v/v), about 1:10 (v /v) to
  • Embodiment 37 The method according to any one of embodiments 34 to 36, wherein step (2) comprises the following steps:
  • Embodiment 38 The method of any one of embodiments 34 to 37, characterized in that,
  • the aqueous phase does not contain additional stabilizers;
  • the aqueous phase already contains additional stabilizers
  • the method also includes adding additional stabilizers in step (2);
  • the amount of the additional stabilizer is such that in the product obtained in step (3) or (4), the content of the additional stabilizer is at least about 2w/v%, such as at least about 3w/v%, at least about 5w/v%, about 5w/v% to about 30w/v%, about 10w/v% to about 25w/v%, or about 15w/v% to about 20w/v%.
  • Embodiment 39 The method of embodiment 38, wherein the additional stabilizer is selected from the group consisting of albumins (e.g., human serum albumin, recombinant human albumin, bovine serum albumin, skim milk powder), monosaccharides, disaccharides, Polysaccharide, mannitol and any combination thereof; preferably selected from: mannitol, lactose, maltose, trehalose, dextran, glucose and sucrose and any combination thereof; preferably sucrose.
  • albumins e.g., human serum albumin, recombinant human albumin, bovine serum albumin, skim milk powder
  • monosaccharides e.g., disaccharides, Polysaccharide, mannitol and any combination thereof
  • mannitol preferably selected from: mannitol, lactose, maltose, trehalose, dextran, glucose and sucrose and any combination thereof; preferably sucrose.
  • Embodiment 40 The method of any one of embodiments 34 to 39, characterized in that, in step (2), the step ( 1) Mixed organic solvents described in.
  • Embodiment 41 The method of embodiment 40, characterized in that the volume of the added mixed organic solvent is equal to or less than the volume of the organic phase; for example, the volume ratio of the added mixed organic solvent to the organic phase is about 1:1 (v/v) to about 1:5 (v/v), for example about 1:2 (v/v) to about 1:4 (v/v) or about 1:3 (v/v).
  • Embodiment 42 The method according to any one of embodiments 34 to 41, characterized in that, in the organic phase described in step (1),
  • SN-38 at a concentration of about 5-17 mg/mL, such as about 5.25-12 mg/mL, about 7-12 mg/mL, or about 10 mg/mL; and/or
  • the concentration of the lipid is about 3-50 mg/mL, such as about 5-45 mg/mL or about 7.5-30 mg/mL, about 10-25 mg/mL or about 15-20 mg/mL; and/or
  • the concentration of albumin is about 5-15 mg/mL, such as about 6-12 mg/mL, or about 6-10 mg/mL.
  • Embodiment 43 The method according to any one of embodiments 34 to 41, characterized in that, in the organic phase described in step (1),
  • the concentration of SN-38 is about 4-10 mg/mL, such as about 6-8 mg/mL; and/or
  • the concentration of the lipid is about 10-20 mg/mL, such as about 15 mg/mL;
  • the concentration of the Span 20 is about 0.3-6mg/mL, such as about 0.3-2mg/mL or about 0.6-1mg/mL; and/or
  • the concentration of albumin is about 8-30 mg/mL, such as about 12-20 mg/mL or about 16-18 mg/mL.
  • Embodiment 44 The method of any one of embodiments 34 to 43, comprising the step (4) of sterilizing the product obtained in step (3), preferably by filtration through a membrane filter of about 0.2 ⁇ m.
  • Embodiment 45 The method of any one of embodiments 34 to 44, further comprising the steps of:
  • step (3) or (4) drying the product obtained in step (3) or (4), preferably spray drying or freeze drying, to provide a solid form, preferably a powder, more preferably a freeze-dried powder composition;
  • SN-38 is present in the composition in an amorphous form.
  • step (5) further comprises: before drying, adding additional stabilizer, wherein the amount of the additional stabilizer is such that when the solid form obtained in step (5) forms an aqueous composition (including solutions and emulsions) through reconstitution, the content of the additional stabilizer is at least about 2w/ v%, such as at least about 3w/v%, at least about 5w/v%, about 5w/v% to about 30w/v%, about 10w/v% to about 25w/v% or about 15w/v% to about 20w /v%.
  • Embodiment 47 A pharmaceutical composition comprising the composition of any one of embodiments 1 to 33 and optionally a pharmaceutically acceptable carrier.
  • Embodiment 48 A pharmaceutical composition comprising the dried composition of any one of embodiments 1 to 33 and optionally a pharmaceutically acceptable carrier.
  • Embodiment 49 The pharmaceutical composition of embodiment 48, wherein said drying is freeze drying or spray drying, preferably freeze drying.
  • Embodiment 50 The pharmaceutical composition according to embodiment 48 or 49, which is in solid form, preferably lyophilized powder; preferably for parenteral administration, more preferably intravenous injection administration.
  • Embodiment 51 Use of the composition of any one of embodiments 1 to 33 or the pharmaceutical composition of any one of embodiments 47 to 50 for the manufacture of a medicament for treating a tumor sensitive to SN-38 in an individual;
  • the tumor is selected from colorectal cancer, small cell lung cancer, lymphoma, breast cancer (preferably triple negative breast cancer), esophageal cancer, gastric cancer, liver cancer, renal cancer, pancreatic cancer, uterine cancer and ovarian cancer.
  • Embodiment 52 The composition of any one of embodiments 1 to 33 or the pharmaceutical composition of any one of embodiments 47 to 50 for use in the treatment of a tumor sensitive to SN-38 in an individual; preferably, the The tumor is selected from colorectal cancer, small cell lung cancer, lymphatic cancer, breast cancer (preferably triple negative breast cancer), esophageal cancer, gastric cancer, liver cancer, renal cancer, pancreatic cancer, uterine cancer and ovarian cancer.
  • Embodiment 53 A method of treating an SN-38 sensitive tumor in an individual comprising administering to said individual a therapeutically effective amount of the composition of any one of embodiments 1 to 33 or any one of embodiments 47 to 50
  • the tumor is selected from colorectal cancer, small cell lung cancer, lymphoma, breast cancer (preferably triple negative breast cancer), esophageal cancer, gastric cancer, liver cancer, kidney cancer, pancreatic cancer, uterine cancer and ovarian cancer.
  • Embodiment 54 A kit comprising the composition of any one of embodiments 1 to 33 or the pharmaceutical composition of any one of embodiments 47 to 50.
  • Embodiment 55 A method of preparing a composition with improved properties, wherein said composition comprises SN-38, a lipid, and albumin, and said albumin encapsulates at least part of said SN-38 and optionally at least part of the lipids to form nanoparticles,
  • the method is characterized in that Span 20 is added during the preparation of the composition;
  • composition does not contain additional stabilizers; and/or
  • the improved properties include: increased stability, for example when the composition is in liquid form, the improved stability includes: during the preparation, storage and/or use of the composition , reduced production or content of albumin multimers (for example albumin multimers are absent or substantially absent from the composition, or albumin multimers account for up to 5 w/w% of the total amount of albumin, for example up to about 4%, at most about 2%, at most about 1.5%, at most about 1.2%, at most about 1.1%, at most about 1%, or at most about 0.8%), and/or reduced nanoparticle size; and/or
  • albumin multimers for example albumin multimers are absent or substantially absent from the composition, or albumin multimers account for up to 5 w/w% of the total amount of albumin, for example up to about 4%, at most about 2%, at most about 1.5%, at most about 1.2%, at most about 1.1%, at most about 1%, or at most about 0.8%), and/or reduced nanoparticle size; and/or
  • composition is as defined in any one of embodiments 1-33.
  • Embodiment 56 The method of embodiment 55, characterized in that the method comprises the steps of:
  • Embodiment 57 The method of embodiment 56, comprising the steps of:
  • the volume ratio of said DMSO or C 1-3 alcohol is about 1:20 (v/v) to about 20:1 (v/v), for example about 1:5 to about 5:1 (v/v), about 1 :2 to about 4:1 (v/v), about 1:1 to about 4:1 (v/v), about 1.5:1 (v/v) to about 3:1 (v/v) or about 2 :1 (v/v) to about 7:3 (v/v); and preparing an aqueous solution of albumin as the aqueous phase;
  • step (3) sterilizing the product obtained in step (3), preferably by filtering through a filter membrane of about 0.2 ⁇ m;
  • the second organic solvent is CHCl 3 ; or a mixture of CH 2 Cl 2 and CHCl 3 , wherein optionally, the volume ratio of CH 2 Cl 2 and CHCl 3 in the mixture is about 2:5 - 1:1, preferably about 2:5.
  • the present invention can reduce the number of high-pressure homogenization in the preparation process, effectively reduce the particle size of the nanoparticles in the composition, improve the filtration flux, stabilize the particle size of the nanoparticles after disintegration, reduce the loss of raw materials and reduce the Cost, especially in scale-up processes such as pilot-scale preparations.
  • the present invention can control the particle size of nanoparticles by further controlling the content of albumin in the composition. Therefore, the particle size of the nanoparticles of the composition of the present invention is closer to the size suitable for drug making.
  • the composition of the present invention has low immunogenicity, high safety, and excellent storage stability.
  • the particle size and particle size distribution of the nanoparticles in the sample were measured using a Malvern Nano ZSE particle size potentiometer.
  • the wavelength of the laser beam of the instrument is 633nm, and the angle between the incident light and the scattered light is 173°.
  • the Zeta potential of the nanoparticles in the sample was measured using a Malvern Nano ZSE particle size potentiometer. Setting parameters: the sample material is protein, and the dispersant is water. The DTS1070 sample cell is selected, the measurement temperature is 25°C, and the automatic scanning detection is performed. The test samples were diluted with deionized water to 10 times the volume before detection. Each sample was measured three times in parallel, and the results were averaged.
  • the albumin content in the system was determined by BCA method.
  • the sample was diluted 10 times with BSA as the standard. Take 25 ⁇ L of the diluted sample, add 200 ⁇ L of detection solution, shake and mix on the shaker, seal the microwell plate, and incubate at 37°C for 120min. The absorbance was measured at 562nm with a microplate reader, and the concentration of albumin in the sample was calculated according to the standard curve.
  • the cholesterol content in the samples was determined by HPLC.
  • the sample dilution method is the same as the above-mentioned SN-38 content determination.
  • the chromatographic conditions are shown in Table 3.
  • the total SN-38 content in the system is determined by the method described above in "3. Determination of SN-38 content in the system".
  • the content of free SN-38 in the system was determined by HPLC after extraction by solid phase extraction.
  • the specific method of solid phase extraction is as follows:
  • the processing method of the sample is the same as the content determination of the above-mentioned SN-38, and the chromatographic conditions of the HPLC method are shown in Table 4.
  • the ring-opening ratio of SN-38 in the sample can be calculated.
  • the ring-opening ratios of SN-38 are all ⁇ 2.0%.
  • the aggregation of albumin in the samples was determined by SEC-HPLC. Take 5 ⁇ l of the prepared sample directly for detection, and the chromatographic conditions are shown in Table 6.
  • the stability of the sample is mainly achieved by storing the prepared samples at room temperature and 4°C, and observing whether there is any obvious precipitation or precipitation in the sample at regular intervals; at the same time, the samples are taken to detect the particle size and particle size distribution, and to study the nanoparticles in the sample. changes in particle size.
  • the disintegration test is to study the stability of the combination of albumin and SN-38 in the sample. Dilute the sample with 1 ⁇ PBS with pH ⁇ 7.4, measure the particle size and particle size distribution of the sample at different dilution times, and study how many times the nanoparticle will disintegrate and precipitate the SN-38 raw material. The higher the dilution factor, the better the stability of the nanoparticles.
  • the crystal form change of the active pharmaceutical ingredient (API) in the samples was evaluated by X-ray diffraction method (Bruker, D8 ADVANCE).
  • the crystal forms of albumin lyophilized powder, albumin-SN-38 lyophilized powder and SN-38 were detected respectively.
  • Sample preparation conditions each sample was adjusted to an API concentration of 2 mg/mL with water for injection.
  • the temperature is 4 degrees, the humidity is 100%, the blot time (adsorption time) is 9 seconds, and the blot force (adsorption strength) is 3.
  • the morphology of vesicles and crystals in the samples was observed with cryo-TEM (Talos L120C) at 120Kv.
  • SN-38 used in the following examples was from Sichuan Xieli Pharmaceutical Co., Ltd.; cholesterol was from Jiangsu Southeast Nanomaterials Co., Ltd.; rHA was from North China Pharmaceutical Co., Ltd.; HSA was from Guangzhou Shuanglin Biopharmaceutical Co., Ltd.; hydrochloric acid Irinotecan injection (CPT-11, 60mg/kg) was from Jiangsu Hengrui Medicine Co., Ltd.
  • rHA-SN-38 or HSA-SN-38 preparations and CPT-11 were based on the active ingredient; the vehicle was water for injection, used as a blank control.
  • Embodiment 1 Preparation of rHA-SN-38 product 1
  • step 2) Weigh 21mg of SN-38 and 30mg of cholesterol, add 3mL of the organic solvent in step 1) to completely dissolve them, and obtain a drug solution;
  • Table 7 The measurement result of each parameter of the sample prepared in embodiment 1
  • particle size increase ratio ((particle size after placement - particle size before placement) ⁇ 100%/particle size before placement)
  • Embodiment 2 Preparation of HSA-SN-38 product 1 freeze-dried agent and its complex solution
  • step 2) Weigh 21mg of SN-38 and 30mg of cholesterol, add 3mL of the organic solvent in step 1) to completely dissolve them, and obtain a drug solution;
  • step 6 Add sucrose to the product obtained in step 6), and stir to dissolve it completely to obtain a sucrose concentration of 30 mg/mL;
  • Table 8 shows the parameter measurement results of the complex solution of the freeze-dried product prepared in Example 2.
  • the determination method of free SN-38 content is: centrifuge the sample at high speed (centrifuge at 21000rpm for 1h), take the supernatant, and then add 9 times the volume of acetonitrile for extraction for detection .
  • the freeze-dried agent of the present application can be diluted into different concentrations for use as required.
  • Embodiment 3 Preparation of rHA-SN-38 product 2
  • step 2) Weigh 42mg of SN-38 and 60mg of cholesterol, add 3mL of the organic solvent in step 1) to completely dissolve it, and obtain a drug solution;
  • step 2) Weigh 42mg of SN-38 and 60mg of cholesterol, add 3mL of the organic solvent in step 1) to completely dissolve it, and obtain a drug solution;
  • Embodiment 5 Preparation of rHA-SN-38 product 4
  • step 2) Weigh 10mg of SN-38 and 60mg of cholesterol, add 3mL of the organic solvent in step 1) to completely dissolve it, and obtain a drug solution;
  • Table 11 shows the measurement results of Product 4.
  • Table 11 The measurement result of each parameter of the sample prepared in embodiment 5
  • Example 12 shows the measurement results of various parameters of the rHA-SN-38 preparation.
  • Embodiment 8 Research on different organic solvent systems
  • the obtained rHA-SN-38 product was prepared and tested according to the preparation process of Example 1 .
  • the particle size of the obtained product was also measured after overnight storage in a refrigerator at 4°C.
  • Table 14 shows the measurement results of various parameters of the prepared rHA-SN-38 preparation.
  • the recovery rate of SN-38 is calculated according to the following formula:
  • Embodiment 9 the investigation to the different ratio of organic solvent
  • step 1) of the preparation process of Example 1 was replaced by EtOH/CHCl 3 in different proportions shown in Table 15, and the amount of rHA in the system was adjusted to 300 mg, and the others were prepared according to the preparation process of Example 1,
  • the obtained rHA-SN-38 preparations were prepared and tested.
  • the measurement results of various parameters of the obtained rHA-SN-38 preparation are shown in Table 15.
  • the obtained rHA-SN-38 product was prepared and tested according to the preparation process of Example 1.
  • the measurement results of various parameters of the obtained rHA-SN-38 preparation are shown in Table 15.
  • Table 16 Parameter determination results of rHA-SN-38 preparations prepared using different concentrations of cholesterol
  • the obtained rHA-SN-38 product was prepared and tested according to the preparation process of Example 1 .
  • the measurement results of various parameters of the obtained rHA-SN-38 preparation are shown in Table 17.
  • Table 17 Parameter determination results of rHA-SN-38 preparations prepared using different concentrations of SN-38
  • Example 12 Investigation of different rHA concentrations in the aqueous phase
  • Table 18 Parameter determination results of rHA-SN-38 preparations prepared using different concentrations of rHA
  • Embodiment 13 the investigation to different organic solvent/aqueous phase volume ratios
  • step 2) Weigh 21mg of SN-38 and 30mg of cholesterol, add 3mL of the organic solvent in step 1) to completely dissolve them, and obtain a drug solution;
  • the present inventors also measured the effect of cyclodextrin (5%, 10% and 15%) as a stabilizer, and found that after the prepared product was placed at room temperature for 24 hours, the product was turbid or the particle size was larger than 250nm. Thus, cyclodextrins may not be suitable as stabilizers.
  • step 2) Weigh 21mg of SN-38 and 30mg of cholesterol, add 3mL of the organic solvent in step 1) to completely dissolve them, and obtain a drug solution;
  • Shear dispersion take 1 mL of the organic solvent in step 1), add it to the water phase in step 3), and shear and disperse for 5 minutes, then add the drug solution in step 2), and continue to shear and disperse for 5 minutes to obtain a coarse emulsion ;
  • Table 21 Determination results of various parameters of HSA-SN-38 products prepared by different preparation processes
  • step 2 Except that the cholesterol in step 2) of the preparation process of Example 1 is replaced by lipids in the following table 22 (cholesterol palmitate (Chol-PA, TCI Reagent Co., Ltd.); glyceryl monostearate (SA-Gly , damas-beta Reagent Co., Ltd.); Vitamin D3 (Aladdin Reagent Co., Ltd.))
  • the obtained rHA-SN-38 product was prepared and tested according to the preparation process of Example 1. The particle size of the obtained product was also measured after overnight storage in a refrigerator at 4°C. Table 22 shows the measurement results of various parameters of these rHA-SN-38 preparations.
  • the obtained rHA-SN-38 preparation was prepared and tested according to the preparation process of Example 1, except that the cholesterol in step 2) of the preparation process of Example 1 was replaced by the lipid combination in Table 23 below.
  • the particle size of the obtained product was also measured after overnight storage in a refrigerator at 4°C. Table 23 shows the measurement results of various parameters of these rHA-SN-38 preparations.
  • Table 23 Determination results of various parameters of rHA-SN-38 preparations prepared using lipid combinations
  • Example 18 Preparation of rHA-SN-38 preparations under conditions of increased lipid ratio
  • Example 2 other raw materials are the same as in Example 1 except that the amount of cholesterol is 60 mg.
  • the raw materials were divided into two groups, and the liquid preparation was prepared according to the method of Example 1, and the lyophilized powder preparation was prepared according to the method of Example 2 (without step 7).
  • the measurement results of each parameter of the obtained rHA-SN-38 liquid preparation and freeze-dried powder preparation are shown in the following table:
  • step 2) Weigh 3.36g of SN-38 and 4.8g of cholesterol, completely dissolve them in the organic solvent in step 1), and then incubate at 50°C for more than 30min to obtain a drug solution;
  • sucrose solution 100g/L, 2.16L
  • step 7) Evaporate the mixture obtained in step 6) with rotary evaporation at 60-70mbar, and then concentrate the sample with tangential flow ultrafiltration (Suzhou Sainz Instrument Co., Ltd.);
  • the relative recovery rate (%) of SN-38 represents the percentage of the concentration difference of SN-38 in the sample before and after passing through the membrane to the concentration of SN-38 in the sample before passing through the membrane;
  • HSA absolute recovery rate represents the percentage of the amount of HSA in the product after passing through the membrane to the amount of HSA charged.
  • step 2) Weigh 4.41g of SN-38 and 6.30g of cholesterol, dissolve them completely in 480mL of the organic solvent in step 1), incubate at 50°C for more than 30min, and mix with the remaining organic solvent to obtain a drug solution;
  • sucrose solution (36g/L, 6L) to the product obtained in step 5) and mix;
  • step 7) Evaporate the mixture obtained in step 6) with rotary evaporation at 60-70mbar, and then concentrate the sample with tangential flow ultrafiltration (Suzhou Sainz Instrument Co., Ltd.);
  • the prepared lyophilized preparation was reconstituted with water for injection to a concentration of SN-38 of 1 mg/mL, and then serially diluted to 100 ⁇ g/mL, 10 ⁇ g/mL, 1 ⁇ g/mL, 0.1 ⁇ g/mL and 0.01 ⁇ g/mL, and determined Particle size and particle size distribution at different concentrations.
  • Example 21 Toxicity study of the rHA-SN-38 preparation of Example 1
  • the rHA-SN-38 preparation prepared in Example 1 was subjected to pharmacodynamic experiments in the BALB/c nude mouse subcutaneous xenograft tumor model of human Hep 3B cells (ATCC HB-8064 cells), and the safety was evaluated at the same time.
  • the rHA-SN-38 product (30 mg/kg) prepared in Example 1 was administered to the qualified tumor-bearing BALB/c nude mice (5 in each group), administered once a week, for a total of consecutive Dosing 6 times.
  • the body weight of each animal was measured on the 13th, 16th, 20th, 23rd, 27th, 30th, 34th, 37th and 41st day after the administration, and the results are shown in FIG. 8 .
  • Example 1 (30 mg/kg) had no effect on the animal's body weight, had good safety, and had good efficacy in inhibiting tumors.
  • the rHA-SN-38 preparation prepared in Example 1 was used for pharmacodynamic experiments in the BALB/c nude mouse subcutaneous xenograft tumor model of human triple-negative breast cancer MDA-MB-231 (ATCC: CRM-HTB-26 TM ) , to study the application of the composition of the present invention in cancer treatment.
  • 21 qualified tumor-bearing BALB/c nude mice were randomly divided into 3 groups, 7 in each group, and were given sterile water for injection, commercially available irinotecan hydrochloride injection (60mg/kg), rHA-SN-38 Products (15mg/kg).
  • Tail vein injection administration administration twice a week, continuous administration for 3 weeks. The day of the first administration was defined as day 0, and the tumor volume of each animal was measured on that day. During the dosing period, the general clinical conditions of the animals were observed daily, and the body weight and tumor volume were measured twice a week.
  • the experimental results are shown in Table 26.
  • the rHA-SN-38 product of the present invention has a very significant antitumor effect in the BALB/c nude mouse subcutaneous transplanted tumor model of human triple-negative breast cancer MDA-MB-231, and is obviously better than the commercially available Irikang hydrochloride injection liquid.
  • the tumor inhibition rate of rHA-SN-38 preparation (15 mg/kg) was 95%, and that of commercially available irinotecan hydrochloride injection (60 mg/kg) was 71%.
  • Table 26 The curative effect of rHA-SN-38 preparations on human triple-negative breast cancer MDA-MB-231 subcutaneously transplanted tumors in nude mice
  • mice 42 qualified tumor-bearing BALB/c nude mice were randomly divided into 6 groups, 7 in each group, and were given sterile water for injection, commercially available irinotecan hydrochloride injection (60mg/kg, 80mg/kg), rHA (220mg/kg) and rHA-SN-38 preparations (1.67mg/kg, 5mg/kg, 15mg/kg).
  • Tail vein injection administration administration once a week, continuous administration for 3 weeks. The day of the first administration was defined as day 0, and the tumor volume of each animal was measured on that day. During the dosing period, the general clinical conditions of the animals were observed daily, and the body weight and tumor volume were measured twice a week.
  • the rHA-SN-38 product of the present invention has a small dosage, high tumor inhibition rate, and its therapeutic effect is significantly better than that of commercially available irinotecan hydrochloride injection, and has an excellent therapeutic effect on human triple-negative breast cancer .
  • Example 23 Study on anti-colon cancer activity (HT-29 model) of rHA-SN-38 preparations
  • the rHA-SN-38 freeze-dried powder product prepared in Example 2 was reconstituted, and the pharmacodynamics experiment was carried out in the BALB/c nude mouse subcutaneous xenograft tumor model of human colon cancer HT-29 (ATCC: HTB 3B TM ), To study the application of the composition of the present invention in cancer treatment.
  • 21 qualified tumor-bearing BALB/c nude mice were randomly divided into 3 groups, 7 in each group, and were given sterile water for injection, commercially available irinotecan hydrochloride injection (60mg/kg), rHA-SN-38 Products (30mg/kg).
  • Tail vein injection administration administration once a week, continuous administration for 3 weeks. The day of the first administration was defined as day 0, and the tumor volume of each animal was measured on that day. During the dosing period, the general clinical conditions of the animals were observed daily, and the body weight and tumor volume were measured twice a week.
  • the experimental results are shown in Table 27.
  • the rHA-SN-38 preparation (30mg/kg) of the present invention has a very significant tumor-inhibiting effect in the BALB/c nude mouse subcutaneous xenograft tumor model of human colon cancer HT-29, and is obviously better than the commercially available Irikan hydrochloride Injection (60mg/kg).
  • the tumor inhibition rate of rHA-SN-38 preparation (30mg/kg) was 72%, and that of commercially available irinotecan hydrochloride injection (60mg/kg) was 47%.
  • Table 27 The curative effect of rHA-SN-38 products on subcutaneous transplantation of human colon cancer HT-29 in nude mice
  • Fifteen qualified tumor-bearing BALB/c nude mice were randomly divided into 3 groups, 5 in each group, and were given sterile water for injection, commercially available irinotecan hydrochloride injection (60mg/kg, 80mg/kg)), rHA-SN-38 preparations (3.3 mg/kg, 10 mg/kg, 30 mg/kg).
  • Tail vein injection administration administration once a week, continuous administration for 4 weeks. The day of the first administration was defined as day 0, and the tumor volume of each animal was measured on that day. During the dosing period, the general clinical conditions of the animals were observed daily, and the body weight and tumor volume were measured twice a week.
  • the rHA-SN-38 product of the present invention has a small dosage, a high tumor inhibition rate, and its therapeutic effect is significantly better than that of the commercially available irinotecan hydrochloride injection, and has an excellent therapeutic effect on human colon cancer.
  • A01S human serum albumin (Guangzhou Shuanglin Biopharmaceutical Co., Ltd.), used as a negative control.
  • mice Forty-nine Balb/c female nude mice aged 6-8 weeks were selected, and the MDA-MB-231 tumor mass was inoculated into the right scapula of the nude mice. Sixteen days after the inoculation of the tumor mass, they were randomly divided into groups, and 7 animals in each group were injected with the test agent through the tail vein. One week after the last administration, tumors were removed from all mice and weighed.
  • Tumor volume was calculated by the following formula:
  • Tumor volume (mm 3 ) 1/2 ⁇ (a ⁇ b 2 ) (where a represents the long diameter and b represents the short diameter).
  • the relative tumor proliferation rate (T/C (%)) represents the percentage value of the relative tumor volume or tumor weight of the treatment group and the control group at a certain time point, and is calculated by the following formula:
  • T/C(%) T RTV /C RTV ⁇ 100%
  • T RTV the average RTV of the treatment group
  • C RTV the average RTV of the vehicle control group
  • V 0 is the tumor volume of the animal when grouping
  • V t is the tumor volume of the animal after treatment
  • T/C% T TW /C TW ⁇ 100%
  • T TW average tumor weight at the end of the experiment in the treatment group
  • C TW average tumor weight at the end of the experiment in the vehicle control group.
  • TGI (%) The relative tumor inhibition rate (TGI (%) was calculated by the following formula:
  • TGI(%) (1-T/C) ⁇ 100%
  • T and C are the relative tumor volume (RTV) or tumor weight (TW) of the treatment group and the control group at a specific time point, respectively.
  • Table 29 below shows the evaluation indexes of the tumor inhibitory efficacy of agents such as HSA-SN-38 and CPT-11 on the MDA-MB-231 xenograft tumor model.
  • High dose of HSA-SN-38 (15mg/kg) has obvious anti-tumor effect, better than CPT-11.
  • the results of tumor weight analysis were basically consistent with tumor volume.
  • Figure 11 shows the effect on the weight change of the tumor-bearing mice in each group.
  • one animal in the CPT-11 high-dose (80mg/kg) treatment group died after the second administration, and the body weight of the remaining 6 animals did not significantly decrease; when the experiment ended, the body weight increased compared with that before administration 1.95%.
  • the body weight increased in different degrees compared with before administration, and the body weight increases of the high, middle and low dose groups were 3.70%, 7.63% and 4.85% respectively.
  • the vehicle and A01S two control groups had the most obvious body weight gain. At the end of the experiment, they increased by 10.84% and 9.04% respectively compared with before administration.
  • HSA-SN-38 has a significant tumor growth inhibitory effect in the MDA-MB-231 human breast cancer model at a dose of 15 mg/kg.
  • CPT-11 also had an obvious tumor growth inhibitory effect at a dose of 80mg/kg, but it was weaker than that of the high-dose HSA-SN-38 group; and one animal died after the second administration.
  • the antitumor activity of HSA-SN-38 was stronger than that of CPT-11, and it was well tolerated. No animals died during the experiment, and no other toxic reactions were observed.
  • HSA-SN-38 prepared in Example 20 (administration after vehicle reconstitution) in the human colon cancer cell line HCT116 (ATCC CCL-247) subcutaneous xenograft BALB/c nude mouse model, and Compared with the commercially available irinotecan hydrochloride for injection (CPT-11).
  • HCT116 tumors in good condition were cut into small tumors of 20-30 mm 3 and inoculated into the right scapula of nude mice. A total of 70 nude mice were inoculated. When the average tumor volume reached about 121 mm 15 days after tumor block inoculation, the mice with too small or too large tumor volume were excluded, and the remaining 49 mice were randomly grouped according to the tumor volume (7 groups, 7 mice in each group) and began to give Medicine (tail vein injection), administered once a week, a total of 4 administrations. Efficacy was evaluated based on relative tumor inhibition rate (TGI), and safety was evaluated based on animal body weight changes and death. One week after the last administration, tumors were taken from all mice, weighed and photographed.
  • TGI tumor inhibition rate
  • the tumor volume changes in each group are shown in Table 31 and Figure 14.
  • Table 32 shows the evaluation indexes of tumor inhibitory efficacy of HSA-SN-38, CPT-11 and other agents on the HCT116 xenograft tumor model.
  • the results of tumor weight analysis were basically consistent with tumor volume.
  • each The reduction in body weight of animals treated with HSA-SN-38 may be related to the cachexia characteristics of the HCT116 model.
  • HSA-SN-38 (30mg/kg and 10mg/kg doses) has a significant inhibitory effect on tumor growth in human colon cancer models, better than the positive control CPT-11, and is better tolerated. No animal died during the experimental administration, and no other toxic reactions were observed.
  • Example 26 In vivo pharmacodynamic study on SKOV-3 human ovarian cancer model
  • mice Eighty BALB/c nude mice were subcutaneously inoculated with SKOV-3 tumor mass. 14 days after inoculation, 56 tumor-bearing mice were selected and divided into 7 groups on average, 8 mice in each group. The drug was administered by tail vein injection, once a week, for a total of 4 times. Efficacy was evaluated based on relative tumor inhibition rate (TGI), and safety was evaluated based on animal body weight changes and death.
  • TGI tumor inhibition rate
  • the tumor volume changes in each group are shown in Table 34 and Figure 16.
  • Table 35 shows the evaluation indexes of the tumor inhibitory efficacy of HSA-SN-38 and CPT-11 on the SKOV-3 xenograft tumor model.
  • High dose of HSA-SN-38 (30mg/kg) has obvious antitumor effect, which is better than low dose of CPT-11 (60mg/kg).
  • the tumor volume of the HSA-SN-38 high-dose group was 445 mm 3
  • the RTV was 2.65
  • There was no significant difference between the middle and low doses of HSA-SN-38 and the vehicle group (p values were 0.977 and 1.000, respectively).
  • the results of tumor weight analysis were basically consistent with tumor volume.
  • HSA-SN-38 has a significant inhibitory effect on tumor growth in the SKOV-3 human ovarian cancer model, which is significantly stronger than CPT-11, and is well tolerated, and no animal died during the experiment , and no other toxic reactions were seen.
  • Example 27 In vivo pharmacodynamic study on SW620 human colon cancer model
  • mice Seventy-five BALB/c nude mice were subcutaneously inoculated with SW620 tumor mass. On the 13th day after inoculation (D13), 49 tumor-bearing mice were selected and divided into 7 groups on average, 7 mice in each group. The drug was administered by tail vein injection, once a week, for a total of 4 times. Efficacy was evaluated based on relative tumor inhibition rate (TGI), and safety was evaluated based on animal body weight changes and death.
  • TGI tumor inhibition rate
  • the tumor volume changes in each group are shown in Table 37 and Figure 18.
  • Table 38 shows the evaluation indexes of the antitumor efficacy of rHA-SN-38 and CPT-11 on the SW620 xenograft tumor model.
  • High and medium doses (30, 10 mg/kg) of rHA-SN-38 have obvious tumor inhibitory effects, both of which are better than CPT-11.
  • the tumor volumes of the rHA-SN-38 high-dose and middle-dose groups were 21 mm 3 and 101 mm 3 , and the RTVs were 0.17 and 0.81, respectively,
  • the T/C values were 1% and 5%, respectively, which were significantly different from those of the vehicle group (p values were 0.040 and 0.047).
  • rHA-SN-38 at doses of 30 and 10 mg/kg can significantly inhibit tumor growth in the SW620 human colon cancer model, which is superior to CPT-11 at doses of 80 and 60 mg/kg. Furthermore, the animals in the two rHA-SN-38-treated groups gained significantly more body weight, whereas the animals in the two CPT-11-treated groups decreased or only slightly increased. Overall, the antitumor activity of rHA-SN-38 was significantly stronger than that of CPT-11, and it was well tolerated. No animals died during the experiment, and no other toxic reactions were observed.
  • Example 28 In vivo pharmacodynamic study on Hep3B human liver cancer model
  • mice Seventy-five BALB/c nude mice were subcutaneously inoculated with Hep3B tumor mass. On the 13th day after inoculation (D13), 49 tumor-bearing mice were selected and divided into 7 groups on average, 7 mice in each group. The drug was administered by tail vein injection, once a week, for a total of 4 times. Efficacy was evaluated based on relative tumor inhibition rate (TGI), and safety was evaluated based on animal body weight changes and death.
  • TGI tumor inhibition rate
  • the test product rHA-SN-38 has a significant tumor growth inhibitory effect in the Hep3B human liver cancer model at 30mg/kg and 10mg/kg, which is similar to the tumor inhibitory level of CPT-11 at 80mg/kg.
  • CPT-11 has anti-tumor activity at a dose of 60 mg/kg, which is similar to the anti-tumor effect of HSA-SN-38 at a dose of 3.33 mg/mL.
  • rHA-SN-38 was well tolerated, and no animal died or other toxic reactions were observed during the experiment.
  • Example 29 Effect of adding different surfactants on SN-38 formulations
  • step 2) Weigh 300mg of SN-38, 300mg of cholesterol and the amount of surfactant shown in Table 40 below, add 30mL of the mixed solvent in step 1), heat to completely dissolve, and obtain a drug solution;
  • Shear dispersion Mix the drug solution in step 2) with the water phase in step 3), and shear and disperse for 10 minutes to obtain a coarse emulsion;
  • the particle size of the samples was tested after shear dispersion, after high-pressure homogenization, and before and after filtration.
  • Example 30 Effect of adding Span 20 on filtration flux of SN-38 formulation
  • the disintegration experiments of the above two samples were also carried out with reference to the method described in item 9 of the preamble of the examples.
  • the sample was diluted with 1 ⁇ PBS with pH ⁇ 7.4, the particle size of the sample was measured at different dilution multiples, and the number of dilution times of the nanoparticles would disintegrate and the SN-38 raw material would be precipitated.
  • FIGS. 21 and 22 The results of the disintegration experiments are shown in FIGS. 21 and 22 .
  • the results show that no matter whether Span 20 is added or not, when diluted to 0.1ug/mL, the two samples are still in a relatively stable state. When further diluted to 10ng/mL, the particle size of the two samples increases significantly, and the nanoparticles disintegrate . After adding Span 20, the change in particle size after disintegration is smaller and more stable.
  • Example 31 Effects of adding different amounts of Span 20 on SN-38 preparations
  • step 2) Weigh SN-38, cholesterol and Span 20 shown in Table 42, add 30mL of the organic solvent in step 1) to completely dissolve it, and obtain a drug solution;
  • Shear dispersion Mix the drug solution in step 2) with the water phase in step 3), and shear and disperse for 10 minutes to obtain a coarse emulsion;
  • Embodiment 32 the influence of adding Span 20 on the stability of SN-38 preparation.
  • Example 31 1) Fill formulations 11 and 12 from Example 31 into vials, and vacuum freeze-dry to obtain a freeze-dried formulation of HSA-SN-38 nanoparticles. After 14 days of storage, it was diluted with deionized water to make the concentration of SN-38 the same as that before freeze-drying, and it was stored for 5 or 10 days under high temperature, high humidity or strong light as the sample to be tested.
  • the percentage content in the table indicates the percentage content of multimer in the sample, the smaller the value, the less multimer.
  • Example 33 Effect of albumin content on SN-38 formulations.
  • Example 29 According to the formulation shown in Table 45, the method of Example 29 was repeated to prepare the SN-38 formulation, except that Span 20 was not added to formulation 13 and 6 and 12 g of HSA were included in formulations 15 and 16, respectively.
  • the particle size of each step in the preparation of the SN-38 preparation was measured, and the API filtration recovery rate was also measured to investigate the influence of the HSA content on the preparation. The results are shown in Table 46.
  • Example 34 In vivo pharmacodynamic study on SW620 human colon cancer model
  • QW x 4 means administration once a week, 4 times in total.
  • the P2 generation tumor tissue was used to evaluate the antitumor activity of the test product. Seven days after inoculation, when the average volume of the tumor reached about 159 mm 3 , the mice with too small or too large tumor volume were excluded, and the remaining 36 mice were randomly grouped according to the tumor volume and started to be administered.
  • T/C (%) refers to the percentage of TV or TW in the treatment group (T) relative to the blank control group (C), the smaller the value, the better the tumor suppressing effect.
  • Example 35 The effect of replacing the EtOH/CHCl 3 mixed solvent with the EtOH/CH 2 Cl 2 /CHCl 3 mixed solvent on the preparation
  • step 2) Weigh 300 mg of SN-38, 300 mg of cholesterol and 18 mg of Span 20, add 30 mL of the mixed solvent in step 1), and dissolve it completely under heating to obtain a drug solution;
  • Shear dispersion mix the drug solution in step 2) with the water phase in step 3), and shear and disperse for 10-15 minutes to obtain a coarse emulsion;
  • the residual amount of CHCl 3 was significantly reduced, and at the same time, the residual level of roughly quantitative CH 2 Cl 2 was also low ( ⁇ 2ug/mg API ). Based on the residual amount of CHCl 3 in the filtered liquid and the maximum single-day exposure to CHCl 3 (600ug), the maximum clinical dosage is >100mg/m 2 , which greatly reduces the dose limitation caused by residual CHCl 3 .
  • Shear dispersion and homogenization Connect the online shearer (Fluke FDHS3/60) and the high-pressure homogenizer (ATS, AH12-150) in series in a continuous production mode, and set the speed of the online shearer to 8000- 10000rpm. Pump the aqueous phase of step 3) and the organic phase of step 2) into an online shearing machine at a certain ratio, then enter the high-pressure homogenizer, and homogenize 5 times under a pressure of 1300-1500 bar;
  • Evaporation remove chloroform and dichloromethane in the system by evaporating in a falling film evaporator (the evaporation tube temperature is 40°C-45°C);
  • Example 37 Effect of adding Span 20 on the stability of HSA-SN-38 preparations at different times
  • Span 20 The effect of Span 20 on albumin stability (polymer, particle size, etc.) in solution was studied by measuring the content of human serum albumin multimer in SN-38 preparation.
  • SKOV-3 tumor mass was subcutaneously inoculated into 40 BALB/c nude mice. On the 13th day after inoculation (D13), 18 tumor-bearing mice were selected and divided into 3 groups on average, 6 mice in each group. The drug was administered by tail vein injection, once a week, for a total of 4 times. The specific drug regimen is shown in Table 53. Efficacy was evaluated based on relative tumor inhibition rate (TGI), and safety was evaluated based on animal body weight changes and death.
  • TGI tumor inhibition rate
  • QW x 4 means administration once a week, 4 times in total.
  • Table 54 shows the evaluation indexes of the tumor inhibitory efficacy of HSA-SN-38 preparation 12 and preparation 18 on the SKOV-3 xenograft tumor model.
  • T/C (%) refers to the percentage of TV or TW in the treatment group (T) relative to the blank control group (C), the smaller the value, the better the tumor suppressing effect.
  • Preparation 12 and Preparation 18 had a significant effect of inhibiting tumor growth in the SKOV-3 human ovarian cancer model at a dose of 30 mg/kg, and the tumors were basically eliminated in the end, and no animals died during the experiment, and no other toxic reactions were seen.
  • the results showed that the antitumor activity of the formulation containing Span20 (Formulation 18) was comparable to that of the formulation without Span 20 (Formulation 12).
  • HCT116 tumors in good condition were cut into small tumors of 20-30 mm 3 and inoculated into the right scapula of 40 nude mice.
  • the mice with too small or too large tumor volume were excluded, and the remaining 18 mice were randomly grouped according to the tumor volume (3 groups, 6 mice in each group) and began to give Medicine (tail vein injection), administered once a week, a total of 4 administrations.
  • Efficacy was evaluated based on relative tumor inhibition rate (TGI), and safety was evaluated based on animal body weight changes and death.
  • TGI tumor inhibition rate
  • One week after the last administration tumors were taken from all mice, weighed and photographed.
  • QW x 4 means administration once a week, 4 times in total.
  • Preparation 12 and Preparation 18 had a significant effect of inhibiting tumor growth in the HCT116 human colon cancer model at a dose of 10 mg/kg, and the tumor basically regressed, and no animals died during the experiment, and no other toxic reactions were observed.
  • compositions, methods and uses of the present application have been described herein with reference to certain preferred embodiments. However, since certain variations thereof will be apparent to those skilled in the art based on the disclosure set forth herein, the application should not be considered limited thereto.

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Abstract

一种包含7-乙基-10-羟基喜树碱的组合物及其制备方法和用途，所述组合物包含SN-38、脂质、白蛋白和Span 20。

Description

包含抗肿瘤药物的组合物及其制备方法和用途 技术领域
本申请涉及7-乙基-10-羟基喜树碱(SN-38)的组合物，其包含SN-38、脂质、白蛋白和Span 20，还涉及其制备方法和用途。
背景技术
SN-38是已上市药物盐酸伊立替康(CPT-11)在体内的活性代谢物，其对某些肿瘤细胞的效力大约是CPT-11的100-1000倍(Zhang J A,Xuan T,Parmar M,et al.Development and characterization of a novel liposome-based formulation of SN-38,[J].International journal of pharmaceutics,2004,270(1):93-107)。SN-38对多种肿瘤细胞均有抑制作用，如结直肠癌、小细胞肺癌、淋巴癌、乳腺癌、食管癌、子宫癌、卵巢癌等。然而，CPT-11在体内转化为SN-38的效率很低，仅为2-8％(Rowinsky E K,Grochow L B,Ettinger D S,et al.Phase I and pharmacological study of the novel topoisomerase I inhibitor CPT-11 administered as a ninety-minute infusion every 3 weeks,[J].Cancer research,1994,54(2):427-436)。
有研究表明SN-38闭合的内酯环结构是其发挥抗癌活性的有效成份，但该闭环结构的SN-38在大部分生物相容和药剂学上可接受的溶剂中都很难溶解，成药性差。上述因素大大限制了其发展及临床应用。目前为止，还没有一个SN-38作为活性成分的药物制剂获得上市批准。
因此，目前仍急切需要解决SN-38的可溶性和成药性的问题。为了解决SN-38的可溶性问题，有的研究致力于对SN-38进行结构修饰。所述修饰又可分为水溶性修饰和脂溶性修饰，前者可见于例如WO1995022549A1，后者可见于例如US20060229359A等。脂溶性修饰的方式一般涉及将SN-38制备成为脂质体，其制备过程一般包括用长链脂肪酸或者胆固醇、维生素E、亲脂性有机酸等疏水性分子修饰喜树碱分子，然后与一定比例的磷脂、胆固醇等辅料制备成喜树碱-脂质体产品。CN108567742A未进行SN-38疏水性修饰，而是通过改进辅料来获得喜树碱-脂质体产品。上述这些报道的产物中均不含白蛋白，而是通过脂质修饰使得喜树碱能够维持更多比例的闭环活性结构，从而在进入体内后能够被白蛋白包载，从而延长药效。
另外，SN-38与紫杉醇类药物相比，由于其更特殊的理化特性，如更易在水中结晶、表面携带正电荷等，制剂制备工艺的稳定和工艺的放大一直是其成药的痛点。
无论是水溶性还是脂溶性修饰/脂质体制备，制备获得的纳米脂质体、纳米混悬液等均存在SN-38的载药量不理想，制备工艺复杂，或工艺放大后重复性差，制剂不稳定的缺点。如何通过优化制剂组成和工艺获得具有更高载药量的SN-38、易于稳定工艺放大、成本经济的SN-38纳米制剂，仍然是亟待解决的问题。
本发明人的在先申请(PCT/CN2021/102332)记载了一种高载药量的包含SN-38、脂质和白蛋白的组合物，获得了适合成药的SN-38制剂。在上述工作的基础上，本发明人意外地发现，向所述组合物中添加Span 20能够减少制备过程中高压均质的次数，有效减小制剂粒径，提高过滤通量，减少原料损耗和降低成本，同时获得的药物制剂稳定性更好，崩解后粒径控制更好，药效更稳定。PCT/CN2021/102332的内容以其整体通过援引加入本文。
发明内容
概述
国际专利申请PCT/CN2021/102332公开了包含SN-38、脂质和白蛋白的组合物。本发明人发现，当以较更大规模(例如，百毫克级或更多的SN-38原料)的制备，包括在放大工艺，例如中试规模的制备中，向该组合物添加Span 20能够减少制备过程中高压均质的次数，有效减小组合物中纳米颗粒的粒径，提高过滤通量，减少原料损耗和降低成本，同时在崩解后能控制纳米粒子的粒径；通过进一步控制所述组合物中白蛋白的含量，能够控制纳米颗粒的粒径，使其更接近于适合成药的尺寸。本发明的组合物同样具有PCT/CN2021/102332的组合物的优点，包括：(1)提高的SN-38载药 量和包封率；(2)包含低水平的开环结构的SN-38；(3)不包含白蛋白多聚体，免疫原性低，安全性高；(4)纳米粒子的粒径小，粒径分布窄；(5)优异的稳定性(包括优异的稀释稳定性和储存稳定性)；(6)SN-38以无定形形式和/或纳米晶体存在，具有溶出速率快和生物利用度高的优点；以及(7)优异的体内抗肿瘤效力。
在第一方面，本申请提供组合物，其包含SN-38、脂质、白蛋白和Span 20，其特征在于，所述组合物包含纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质，其中所述脂质选自胆固醇、胆固醇衍生物、胆固醇类似物和脂肪酸酯以及它们中的两者或更多者的任意组合。
在第二方面，本申请提供制备根据第一方面的组合物的方法。
在第三方面，本申请提供制备包含SN-38、脂质、白蛋白和Span 20的组合物的方法。
在第四方面，本申请提供可由根据第三方面所述的方法制备的组合物。
在第五方面，本申请还提供制备具有改善的性质的组合物的方法。
本申请的其他方面还提供包含上文所述的组合物的药物组合物及其用途。
附图简要说明：
图1为实施例1制备的制品中SN-38的含量测定的HPLC典型图谱。
图2为实施例1制备的制品中胆固醇的含量测定的HPLC典型图谱。
图3为实施例1制备的制品中不同结构的SN-38的含量测定的HPLC典型图谱
图4为实施例1制备的制品中白蛋白聚集体测定的SEC-HPLC典型图谱。
图5为实施例2制备的冻干制品、SN-38以及HSA的XRD图谱。
图6显示了实施例19中在梯度稀释下进行的崩解实验的结果。
图7显示了实施例20中在梯度稀释下进行的崩解实验的结果。
图8显示了实施例21中接受实施例1的rHA-SN-38制品给药的动物体重变化。
图9显示实施例1的制品抑制人三阴性乳腺癌MDA-MB-23的体内实验的结果。
图10显示实施例2的制品抑制人结肠癌HT-29的体内实验结果。
图11显示了实施例24中试验动物的体重变化。
图12显示了实施例24中试验动物的肿瘤体积变化。
图13显示了实施例25中试验动物的体重变化。
图14显示了实施例25中试验动物的肿瘤体积变化。
图15显示了实施例26中试验动物的体重变化。
图16显示了实施例26中试验动物的肿瘤体积变化。
图17显示了实施例27中试验动物的体重变化。
图18显示了实施例27中试验动物的肿瘤体积变化。
图19显示了实施例28中试验动物的体重变化。
图20显示了实施例28中试验动物的肿瘤体积变化。
图21显示了实施例30中制备的包含Span 20的制剂在梯度稀释下进行的崩解实验的结果。
图22显示了实施例30中制备的不含Span 20的制剂在梯度稀释下进行的崩解实验的结果。
图23显示了实施例30中制备的包含Span 20的制剂的冷冻透射电镜观察结果。
图24显示了实施例30中制备的不含Span 20的制剂的冷冻透射电镜观察结果。
图25显示了实施例34中试验动物的肿瘤体积变化。
图26显示了实施例34中试验动物的体重变化。
图27显示了实施例34中试验动物的瘤块大小。
图28显示了实施例38中试验动物的体重变化。
图29-图30显示了实施例38中试验动物的肿瘤体积变化。
图31显示了实施例39中试验动物的体重变化。
图32-图33显示了实施例39中试验动物的肿瘤体积变化。
详述
定义
除非在下文中另有定义，本文中所用的所有技术术语和科学术语的含义意图与本领域技术人员通常所理解的相同。提及本文中使用的技术意图指在本领域中通常所理解的技术，包括那些对本领域技术人员显而易见的技术的变化或等效技术的替换。虽然相信以下术语对于本领域技术人员很好理解，但仍然阐述以下定义以更好地解释本发明。
术语“纳米粒子”表示在至少一个维度(例如一个、两个或三个维度)上具有纳米级尺寸的粒子，例如约1nm、约10nm、约100nm或约200-300nm级别的尺寸，优选不超过200nm级别的尺寸。
术语“纳米晶体”指尺寸为1至1000nm的晶体，尤其指50至300nm内的晶体，可以是单晶，也可以是多晶体。
术语“囊泡”是指分散于水相中时会自发形成的一类具有外层结构的分子有序组合体。
术语“Span 20”也称作山梨醇酐单月桂酸酯(sorbitan monolaurate)或司盘20，是一种表面活性剂。
术语“水性组合物”是指水基组合物，其为液体或半固体形式，优选液体形式。液体形式包括但不限于溶液(例如蛋白纳米粒子溶液)、胶体、乳液和悬浮液。
术语“载药量(LD)”和“包封率(EE)”可以按照实施例部分中提供的公式计算。
术语“有机酸”包括具有1至24个碳原子的饱和或不饱和脂肪酸，特别是具有2至4个碳原子的短链脂肪酸、具有6至12个碳原子的中链脂肪酸以及具有14-24个碳原子的长链脂肪酸；以及芳族羧酸。可以特别提及的一个例子是辛酸。“长链脂肪酸”包括但不限于棕榈酸(C16：0)、硬脂酸(C18：0)、油酸(C18：1)、亚油酸(C18：2)、α-亚麻酸(C18：3)、花生四烯酸(C20：4)、二十碳五烯酸(C20：5)、二十二碳六烯酸(C22：6)。可以特别提及的例子是棕榈酸和硬脂酸。
术语“包括”、“包含”、“具有”、“含有”或“涉及”及其在本文中的其它变体形式为包含性的(inclusive)或开放式的，且不排除其它未列举的元素或方法步骤，尽管其它未列举的元素或方法步骤不一定存在(即，这些术语也涵盖术语“基本上由……组成”和“由……组成”)。
术语“约”是指在所述数值的±10％范围内，优选±5％范围内，更优选±2％范围内。
本文中所述的数值范围应理解为涵盖其中包含的任何和所有子范围。例如，范围“1至10”应理解为不仅包括明确记载的1至10的值，而且还包括1至10范围内的任何单个值(例如2、3、4、5、6、7、8和9)和子范围(例如1至2、1.5至2.5、1至3、1.5至3.5、2.5至4、3至4.5等等)。该原则亦适用于仅用一个数值作为最小值或最大值的范围。
本文中提及的文献均以其整体援引加入本文中。
在第一方面，本申请提供组合物，其包含SN-38、脂质、白蛋白和Span 20，其特征在于，所述组合物包含纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
脂质:SN-38为约(0.1-10):1(w:w)；
白蛋白:SN-38为约(1-100):1(w:w)；以及
Span 20:SN-38为约(3-60):100(w:w)；并且
其中所述脂质选自胆固醇、胆固醇衍生物、胆固醇类似物和脂肪酸酯以及它们中的两者或更多者的任意组合。
在一些实施方案中，脂质:SN-38为约(0.5-6):1(w:w)，例如约(0.5-5):1(w:w)，约(0.5-3):1(w:w)，约(1-4):1(w:w)，约(1.2-4):1(w:w)、约(1.4-2):1(w:w)、约(1.5-2.5):1(w:w)或约1：1(w:w)。
在一些实施方案中，白蛋白:SN-38为约(1-50):1(w:w)，例如约(3-25):1(w:w)，约(5-25):1(w:w)，约(5-20):1(w:w)或约(5-18):1(w:w)，约(6-15):1(w:w)，约(7-15):1(w:w)、约(6-12):1(w:w)、约(7-12):1(w:w)、约(9-11):1(w:w)或约10:1(w:w)。
在一些实施方案中，Span 20:SN-38为约(4-60):100(w:w)，例如约(5-60):100(w:w)、约(6-60):100(w:w)、约(7-55):100(w:w)、约(8-50):100(w:w)、约(10-45):100(w:w)、约(12-40):100(w:w)、约(14-35):100(w:w)、约(15-30):100(w:w)、约(16-25):100(w:w)或约(18-20):100(w:w)。
在一些实施方案中，白蛋白:脂质为约(1-100):1(w:w)，例如约(2-20):1(w:w)，约(3-15):1(w:w)或约(5-10):1(w:w)，例如约7:1(w:w)或约10:1(w:w)。
在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述SN-38的含量为约1w/w％至约25w/w％。在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述脂质的含量为约1w/w％至约35w/w％。在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述白蛋白的含量为约50w/w％至约98w/w％。
在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述SN-38的含量为约3w/w％至约20w/w％。在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述脂质的含量为约2w/w％至约30w/w％。在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述白蛋白的含量为约55w/w％至约95w/w％。
在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述SN-38的含量为约3w/w％至约15w/w％，例如约4w/w％、约5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约8w/w％、约8.5w/w％、约9w/w％、约10w/w％、约11w/w％、约12w/w％、约13w/w％或约14w/w％。
在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述脂质的含量为约3w/w％至约30w/w％，例如约4w/w％、约5w/w％、约6w/w％、约7w/w％、约8w/w％、约8.5w/w％、约9w/w％、约9.5w/w％、约10w/w％、约10.5w/w％、约11w/w％、约11.5w/w％、约12w/w％、约12.5w/w％、约13w/w％、约13.5w/w％、约14w/w％、约15w/w％、约16w/w％、约17w/w％、约18w/w％、约19w/w％、约20w/w％、约21w/w％、约24w/w％、约26w/w％或约28w/w％。
在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述白蛋白的含量为约60w/w％至约94w/w％，例如约64w/w％至约93w/w％，约66w/w％至约92w/w％、约68w/w％至约91w/w％、约70w/w％至约90w/w％、约75w/w％至约90w/w％，约75w/w％、约76w/w％、约77w/w％、约78w/w％、约79w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约85w/w％、约86w/w％、约87w/w％、约88w/w％或约89w/w％。
不希望受任何理论的限制，本发明人发现，随着脂质用量的增加，所述组合物趋向于具有更小的平均粒径，更高的包封率和更高的SN-38利用率；增加白蛋白的用量可能会造成载药量的降低，过高的白蛋白含量还会增大纳米粒子粒径，影响成药。在一个实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述脂质的含量为约5w/w％至约24w/w％。在一个实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述白蛋白的含量为约60w/w％至约90w/w％。
另一方面，组合物中脂质成分含量过高可能会给个体带来不利的影响。从减少通过所述组合物的施用导致的脂质摄入的角度，预期优选的是，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述组合物中的脂质的含量不超过约18％w/w％，例如不超过约16％w/w％、或不超过约14w/w％。然而，这并不表示具有较高脂质含量的组合物不是合意的。
在一些实施方案中，以所述组合物中的SN-38、脂质、白蛋白和Span 20的总量计，Span 20的含量为约0.03w/w％至约12w/w％，例如约0.06w/w％至约10w/w％、0.08w/w％至约9w/w％、约0.1w/w％至约8w/w％、约0.2w/w％至约7w/w％、约0.4w/w％至约6w/w％、约0.6w/w％至约5w/w％、约0.8w/w％至约4w/w％、约1w/w％至约2w/w％。
在一些实施方案中，脂质:SN-38为约(1-10):1(w:w)，例如约(1-8):1(w:w)、约(1-6):1(w:w)、约(1-5):1(w:w)、约(1-4.5):1(w:w)、约(1-4):1(w:w)、约(1.2-3.8):1(w:w)、约1:1(w:w)、约(1.4-3.6):1(w:w)、约(1.6-3.4):1(w:w)、约(1.8-3.2):1(w:w)、约(2-3):1(w:w)、约(2.2-2.8):1(w:w)、约(2.4-2.6):1(w:w)、约1:1(w:w)或约2.5:1(w:w)。在另一些实施方案中，脂质:SN-38为约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)或约(1-1.4):1。
在一些实施方案中，白蛋白:SN-38为约(5-40):1(w:w)，例如约(5-小于40):1(w:w)、约(5-35):1(w:w)、约(5-30):1(w:w)、约(8-25):1(w:w)、约(10-22.5):1(w:w)、约(12.5-20):1(w:w)、约(15-17.5):1(w:w)、约(16-18):1(w:w)或约10:1(w:w)。在另一些实施方案中，白蛋白:SN-38为约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、或约(11.1-17.3):1(w:w)。
在一些实施方案中，Span 20:SN-38为约(5-60):100(w:w)，例如约(6-60):100(w:w)、约(7-55):100(w:w)、约(8-50):100(w:w)、约(10-45):100(w:w)、约(12-40):100(w:w)、约(14-35):100(w:w)、 约(15-30):100(w:w)、约(16-25):100(w:w)或约(18-20):100(w:w)。在另一些实施方案中，Span 20:SN-38为约(5-10):100(w:w)、约(5-9):100(w:w)、约(6-8.6):100(w:w)、约(6-8):100(w:w)、或约(6.5-7):100(w:w)。
在一些实施方案中，白蛋白:脂质为约(1-40):1(w:w)，例如约(1-小于40):1(w:w)、约(2-35):1(w:w)、约(3-15):1(w:w)、约(5-10):1(w:w)或约(6-8):1(w:w)，例如约7:1(w:w)或约10:1(w:w)。在一些实施方案中，白蛋白:脂质为约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)。
在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述SN-38的含量为约2w/w％至约16w/w％。在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述脂质的含量为约2w/w％至约35w/w％。在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述白蛋白的含量为约75w/w％至约96w/w％。
在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述SN-38的含量为约2.5w/w％至约15w/w％，例如约4w/w％至约10w/w％，约4.5w/w％至约9.5w/w％，约5w/w％至约9w/w％，或约7.5w/w％至约8w/w％。在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述组合物中的脂质的含量为约2.5w/w％至约30w/w％，例如约4w/w％至约12.5w/w％，约4.5w/w％至约12w/w％，约7w/w％至约10w/w％，或约7.5w/w％至约8w/w％。在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述组合物中的白蛋白的含量为约76w/w％至约95w/w％，例如约78w/w％至约93w/w％，约79w/w％至约91.5w/w％，约80w/w％至约90w/w％，约82w/w％至约89w/w％，约84w/w％至约88w/w％，或约84.5w/w％至约87.5w/w％。
在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述SN-38的含量为约3w/w％至约14w/w％，例如约3.5w/w％至约12w/w％、约4w/w％、约4.2w/w％、约4.5w/w％、约4.6w/w％、约4.8w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约7.6w/w％、约7.8w/w％、约8w/w％、约8.5w/w％、约9w/w％、约9.2w/w％、约9.5w/w％、约9.6w/w％、约9.8w/w％、约10w/w％、约10.5w/w％、约11w/w％或约11.5w/w％。
在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述脂质的含量为约4w/w％至约25w/w％，例如约4.5w/w％至约20w/w％、约4.3w/w％、约4.5w/w％、约4.7w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约7.6w/w％、约7.8w/w％、约8w/w％、8.5w/w％、约9w/w％、约9.5w/w％、约10w/w％、约10.5w/w％、约11w/w％、约11.5w/w％、约12w/w％、约12.1w/w％、约12.3w/w％、约12.5w/w％、约13w/w％、约13.5w/w％、约14w/w％、约14.5w/w％、约15w/w％、约15.5w/w％、约16w/w％、约16.5w/w％、约17w/w％、约17.5w/w％、约18w/w％、约18.5w/w％、约19w/w％或约19.5w/w％。
在一些实施方案中，以所述组合物中的SN-38、脂质和白蛋白的总量计，所述白蛋白的含量为约78w/w％至约92w/w％，例如约79w/w％、约79.2w/w％、约79.4w/w％、约79.6w/w％、约79.8w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约84.3w/w％、约84.5w/w％、约84.7w/w％、约84.9w/w％、约85w/w％、约86w/w％、约87w/w％、约87.3w/w％、约87.5w/w％、约87.7w/w％、约87.9w/w％、约88w/w％、约89w/w％、约90w/w％、约91w/w％、约91.3w/w％、或约91.5w/w％。
在一些实施方案中，以所述组合物中的SN-38、脂质、白蛋白和Span 20的总量计，Span 20的含量为约0.14w/w％至约5w/w％，例如约0.2w/w％至约2.5w/w％、约0.22w/w％至约2.0w/w％、约0.24w/w％至约2w/w％、约0.26w/w％至约1.5w/w％、约0.28w/w％至约1.0w/w％、约0.3w/w％至约0.9w/w％、约0.32w/w％至约0.8w/w％、约0.34w/w％至约0.7w/w％、约0.36w/w％至约0.6w/w％、约0.38w/w％至约0.58w/w％、约0.4w/w％至约0.56w/w％、约0.42w/w％至约0.54w/w％、约0.44w/w％至约0.52w/w％、约0.46w/w％、约0.48w/w％或约0.5w/w％。在另一些实施方案中，Span 20的含量为约0.2w/w％至约0.8w/w％、约0.24w/w％至约0.7w/w％、约0.26w/w％至约0.7w/w％、约0.3w/w％至约0.65w/w％、约0.36w/w％至约0.6w/w％、约0.4w/w％至约0.58w/w％、约0.44w/w％至约0.56w/w％、约0.48w/w％至约0.54w/w％、或约0.5w/w％至约0.52w/w％。
在一些实施方案中，存在于所述纳米粒子中的SN-38占所述组合物中的SN-38、脂质和白蛋白的总量的至少约1w/w％或至少约2w/w％，例如至少约3w/w％，约3w/w％至约13w/w％，约4w/w％至约12w/w％，约4w/w％、约5w/w％、约6w/w％、约7w/w％、约8w/w％、约9w/w％、约10w/w％或约11w/w％。
在一些实施方案中，存在于所述纳米粒子中的SN-38占所述组合物中的SN-38的总量的约80w/w％至约99w/w％，例如约88w/w％至约98w/w％，约89w/w％、约90w/w％、约91w/w％、约92w/w％、约93w/w％、约94w/w％、约95w/w％、约96w/w％或约97w/w％。
在一些实施方案中，所述胆固醇衍生物选自胆固醇与有机酸形成的酯，优选选自胆固醇棕榈酸酯和胆固醇辛酸酯及其组合。
在一些实施方案中，所述胆固醇类似物选自维生素D2、维生素D3及其组合。
在一些实施方案中，所述脂肪酸酯选自脂肪酸甘油酯，优选长链脂肪酸甘油酯，优选硬脂酸甘油酯，更优选单硬脂酸甘油酯。
在一些优选的实施方案中，所述脂质选自胆固醇、胆固醇棕榈酸酯、胆固醇辛酸酯、维生素D2、维生素D3、单硬脂酸甘油酯以及它们中的两者或更多者的任意组合。
在一些优选的实施方案中，所述脂质选自胆固醇、胆固醇棕榈酸酯、维生素D3、单硬脂酸甘油酯以及它们中的两者或更多者的任意组合；
在一些优选的实施方案中，所述脂质为：胆固醇、胆固醇棕榈酸酯、维生素D3或单硬脂酸甘油酯；胆固醇和胆固醇棕榈酸酯的混合物；胆固醇和维生素D3的混合物；胆固醇和单硬脂酸甘油酯的混合物；或胆固醇棕榈酸酯和单硬脂酸甘油酯的混合物。
在一些更优选的实施方案中，所述脂质为胆固醇。优选地，胆固醇作为唯一的脂质。在一些此类实施方案中，胆固醇:SN-38为约(1-6):1(w:w)，例如约(1.2-5):1(w:w)，例如约(1.4-4):1(w:w)、约3:1(w:w)、约2:1(w:w)或约1:1(w:w)。在另一些实施方案中，胆固醇:SN-38为约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)、约(1-1.4):1。在一些此类实施方案中，白蛋白:SN-38为约(3-25):1(w:w)，例如约(4-20):1(w:w)，约(5-15):1(w:w)，约(6-12):1(w:w)、约(7-12):1(w:w)、约(9-11):1(w:w)或约10:1(w:w)。在另一些实施方案中，白蛋白:SN-38为约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、或约(11.1-17.3):1(w:w)。在一些此类实施方案中，白蛋白:胆固醇为约(2-20):1(w:w)，例如约(3-15):1(w:w)、约(5-10):1(w:w)或约7:1(w:w)。在另一些实施方案中，白蛋白:胆固醇为约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)。
在一些此类实施方案中，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，所述SN-38的含量为约3w/w％至约15w/w％，例如约4w/w％至约15w/w％、约6w/w％至约10w/w％或约8w/w％至约12w/w％。在另一些实施方案中，所述SN-38的含量为约4w/w％至约10w/w％、约4.5w/w％至约9.5w/w％、约5w/w％至约9w/w％、或约7.5w/w％至约8w/w％。在一些此类实施方案中，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，所述胆固醇的含量为约5w/w％至约25w/w％，例如约6w/w％至约22w/w％、或约15w/w％至约20w/w％。在另一些实施方案中，所述胆固醇的含量为约4w/w％至约12.5w/w％、约4.5w/w％至约12w/w％、约7w/w％至约10w/w％、或约7.5w/w％至约8w/w％。在一些此类实施方案中，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，所述白蛋白的含量为约64w/w％至约90w/w％，例如约70w/w％至约90w/w％。在另一些实施方案中，所述白蛋白的含量为约78w/w％至约93w/w％、约79w/w％至约91.5w/w％、约80w/w％至约90w/w％、约82w/w％至约89w/w％、约84w/w％至约88w/w％、或约84.5w/w％至约87.5w/w％。在一些此类实施方案中，存在于所述纳米粒子中的SN-38占所述组合物中的SN-38、胆固醇和白蛋白的总量的至少约3w/w％，例如约3w/w％至约13w/w％，约4w/w％至约12w/w％，约4w/w％、约5w/w％、约6w/w％、约7w/w％、约8w/w％、约9w/w％、约10w/w％或约11w/w％。
在另一些实施方案中，所述脂质为胆固醇棕榈酸酯。在一些实施方案中，所述脂质为维生素D3。在一些实施方案中，所述脂质为单硬脂酸甘油酯。在一些实施方案中，胆固醇棕榈酸酯、维生素D3或单硬脂酸甘油酯作为唯一的脂质。
还可以涵盖的是，所述脂质为胆固醇和单硬脂酸甘油酯的混合物，其中胆固醇:单硬脂酸甘油酯为例如约(0.2-5):1(w:w)，约(0.5-3):1(w:w)，约(0.5-2):1(w:w)，或约1:1(w:w)。
在一些实施方案中，所述脂质为胆固醇棕榈酸酯和单硬脂酸甘油酯的混合物；其中胆固醇棕榈酸酯:单硬脂酸甘油酯为例如约(0.2-5):1(w:w)，约(0.5-3):1(w:w)，约(0.5-2):1(w:w)，或约1:1(w:w)。
在一些实施方案中，所述脂质为胆固醇和胆固醇棕榈酸酯的混合物，其中胆固醇:胆固醇棕榈酸酯为例如约(0.2-5):1(w:w)，约(0.5-3):1(w:w)，约(0.5-2):1(w:w)，或约1:1(w:w)。
对于上述包含除胆固醇之外的脂质或者胆固醇和其他脂质的组合的实施方案，所述脂质:SN-38为例如约(1-6):1(w:w)，约(1.2-5):1(w:w)，约(1.4-4.5):1(w:w)、约4.3:1(w:w)、约3:1(w:w)、约1:1(w:w)或约2:1(w:w)。在一些此类实施方案中，白蛋白:SN-38为例如约(5-25):1(w:w)，约(10-20):1(w:w)，约(6-15):1(w:w)，约(7-15):1(w:w)、约(9-12):1(w:w)、约(9-11):1(w:w)、约9.5:1(w:w)或约10:1(w:w)。在一些此类实施方案中，白蛋白:所述脂质为例如约(2-10):1(w:w)，约(3-7):1(w:w)或约(4-6):1(w:w)。在一些此类实施方案中，以所述组合物中的SN-38、所述脂质和白蛋白的总量计，所述SN-38的含量为例如约5w/w％至约15w/w％，约6w/w％至约12w/w％、约7w/w％至约10w/w％、约8w/w％或约9w/w％。在一些此类实施方案中，以所述组合物中的SN-38、所述脂质和白蛋白的总量计，所述脂质的含量为例如约5w/w％至约32w/w％，约10w/w％至约30w/w％、约18w/w％、约20w/w％、约22w/w％、约24w/w％、约26w/w％、约28w/w％或约29w/w％。在一些此类实施方案中，以所述组合物中的SN-38、所述脂质和白蛋白的总量计，所述白蛋白的含量为例如约60w/w％至约90w/w％，约64w/w％至约85w/w％、约70w/w％至约80w/w％、或约75w/w％。在一些此类实施方案中，存在于所述纳米粒子中的SN-38占所述组合物中的SN-38、所述脂质和白蛋白的总量的至少约3w/w％，例如约3w/w％至约10w/w％，约4w/w％至约9w/w％，约4w/w％、约5w/w％、约6w/w％、约7w/w％或约8w/w％。
在一些根据上文所述的实施方案中，所述组合物为液体、半固体或固体形式。
在一些实施方案中，所述组合物为固体形式，优选为粉末形式。更优选地，所述组合物为冻干粉。
在一些此类实施方案中，SN-38优选地以无定形和/或纳米晶体形式存在于所述组合物中，如通过电镜或X射线衍射法(Cu-Kα)分析所测定的。所述纳米晶体的粒径范围可以为约30-500nm，优选约50-200nm。所述纳米晶体形式的SN-38占总SN-38的75％或更高，例如80％、85％、90％或更高。
不希望受任何理论的限制，在本申请的组合物中，Span 20起到提高所述组合物的稳定性，调节纳米粒子的粒径以及使其更加分散的作用。
在一些实施方案中，所述组合物不包含额外的稳定剂。在另一些实施方案中，所述组合物优选地还包含额外的稳定剂，例如冻干稳定剂，所述额外的稳定剂的量使得当所述组合物通过复溶形成水性组合物(包括溶液和乳液)时，所述额外的稳定剂的含量为至少约2w/v％，例如至少约3w/v％，至少约5w/v％，约5w/v％至约30w/v％，约10w/v％至约25w/v％或约15w/v％至约20w/v％。
在一些此类实施方案中，所述组合物还包含额外的稳定剂，以所述组合物中的总量计，所述额外的稳定剂的含量为约60w/w％至约98w/w％，例如约65w/w％至约97w/w％，约68w/w％至约96w/w％，约69w/w％至约95w/w％，约70w/w％至约94w/w％、约71w/w％至约93w/w％、约72w/w％至约92w/w％、约73w/w％、约74w/w％、约75w/w％、约76w/w％、约77w/w％、约78w/w％、约79w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约85w/w％、约86w/w％、约87w/w％、约88w/w％、约89w/w％、约90w/w％或约91w/w％。
在一些实施方案中，当所述组合物通过复溶形成水性组合物(包括溶液和乳液)而具有约0.1μg/mL至约30.0mg/mL(或下文关于水性组合物所述的SN-38的含量值)的SN-38的含量时，所述纳米粒子具有约50至200nm，例如约90至150nm、约95至140nm、约100至130nm、约105至125nm或约110至120nm的平均粒径。
在另一些实施方案中，所述组合物为液体形式的水性组合物，包括溶液和乳液。
在一些实施方案中，所述液体形式的组合物包含以纳米晶体和/或囊泡的形式的SN-38。所述纳米晶体的粒径范围可以为约30-500nm，优选约50-200nm。
在一些此类实施方案中，所述组合物为溶液的形式；在一些此类实施方案中，所述组合物为乳液的形式。
本发明人发现，本申请的水性组合物具有优异的稀释稳定性。在一些实施方案中，当稀释(例如用pH约7.4的1 x PBS稀释)所述组合物致使SN-38在稀释后的组合物中的含量达到约4μg/mL或更低，例如约2μg/mL或更低，约1μg/mL或更低，或约0.4μg/mL或更低，例如约0.1μg/mL或0.04μg/mL时，所述纳米粒子不发生崩解。由于优异的稳定性，所述水性组合物可以作为浓缩液或稀释液存在。
因此，所述水性组合物的各种组分可以具有较宽的含量范围。在一些此类实施方案中，以所述组合物的总量计，SN-38的含量为约0.1μg/mL至约30.0mg/mL、约0.2μg/mL至约27.0mg/mL、约0.5μg/mL至约24.0mg/mL、约1.0μg/mL至约21.0mg/mL、约5.0μg/mL至约18.0mg/mL、约10.0μg/mL至约15.0mg/mL、约20.0μg/mL至约12mg/mL、约25.0μg/mL至约9mg/mL、约50.0μg/mL至约6.0mg/mL或约100.0μg/mL至约3.0mg/mL。
在一些实施方案中，以所述组合物的总量计，所述脂质的含量为约0.05μg/mL至约100.0mg/mL、约0.1μg/mL至约90.0mg/mL、约0.25μg/mL至约80.0mg/mL、约0.5μg/mL至约70.0mg/mL、约2.5μg/mL至约60.0mg/mL、约5.0μg/mL至约50.0mg/mL、约10.0μg/mL至约40.0mg/mL、约12.5μg/mL至约30.0mg/mL、约25.0μg/mL至约20.0mg/mL、或约50.0μg/mL至约10.0mg/mL。
在一些实施方案中，以所述组合物的总量计，白蛋白的含量为约3.0μg/mL至约300.0mg/mL、约6.0μg/mL至约270.0mg/mL、约15.0μg/mL至约240.0mg/mL、约30.0μg/mL至约210.0mg/mL、约150.0μg/mL至约180.0mg/mL、约300.0μg/mL至约150.0mg/mL、约600.0μg/mL至约120.0mg/mL、约750.0μg/mL至约90.0mg/mL、约1500.0μg/mL至约60.0mg/mL、或约3.0mg/mL至约30.0mg/mL。
一般而言，以所述组合物的总量计，SN-38的含量可以为约100.0μg/mL至约3.0mg/mL，例如约200.0μg/mL至约2.5mg/mL、约300.0μg/mL至约2.0mg/mL、约400.0μg/mL至约1.5mg/mL、约500.0μg/mL至约1.0mg/mL、或约600μg/mL至约800μg/mL；和/或
所述脂质的含量可以为约50.0μg/mL至约10.0mg/mL，例如约100.0μg/mL至约8.0mg/mL、约200.0μg/mL至约6.0mg/mL、约300.0μg/mL至约4.0mg/mL、约400.0μg/mL至约3.0mg/mL、约500.0μg/mL至约2.5mg/mL、约600.0μg/mL至约2.0mg/mL、约700.0μg/mL至约1.5mg/mL、约800μg/mL至约1.0mg/mL、或约200μg/mL至约1.5mg/mL；和/或
白蛋白的含量可以为约3.0mg/mL至约30.0mg/mL，例如约4.0mg/mL至约25.0mg/mL、约5.0mg/mL至约20.0mg/mL、约6.0mg/mL至约15.0mg/mL、约7.0mg/mL至约12.0mg/mL、或约8.0mg/mL至约10.0mg/mL。
理想的是，在水性组合物(包括溶液和乳液)中，所述纳米粒子有不超过约200nm、例如不超过约150nm的平均粒径，优选在施用之前在储存一定时间后，其平均粒径仍然满足上述要求。本发明人发现，本申请的水性组合物具有这样的优异性质。
在一些实施方案中，所述纳米粒子具有约50至200nm，例如约90至150nm，或约100至130nm的平均粒径。
在一些实施方案中，当在4℃下保存24h后，所述纳米粒子具有约50至200nm，例如约90至150nm，或约100至130nm的平均粒径。
在一些实施方案中，所述纳米粒子具有不超过约0.30、例如不超过约0.2、不超过约0.10或不超过约0.01的粒径分布指数(PDI)。
在一些实施方案中，所述组合物具有约-35mV至约-20mV，例如约-31mV的Zeta电位。
在一些此类实施方案中，所述组合物不包含额外的稳定剂。在一些此类实施方案中，所述组合物还包含额外的稳定剂，其中以所述组合物的总量计，所述额外的稳定剂的含量为至少约2w/v％，优选至少约3w/v％，例如至少约5w/v％，约5w/v％至约30w/v％，约10w/v％至约25w/v％或约15w/v％至约20w/v％。
上文所述的额外的稳定剂可以选自：白蛋白类(例如人血清白蛋白、重组人白蛋白、牛血清白蛋白、脱脂奶粉)、单糖、二糖、多糖及其任意组合；优选地选自葡萄糖和蔗糖；优选为蔗糖。
额外的稳定剂的使用有利于保持纳米粒子的平均粒径。本发明人发现，当在4℃下保存24h后，与不存在额外的稳定剂的情况相比，额外的稳定剂的存在减小了所述水性组合物中纳米粒子的平均 粒径的增加。同时，对于最终以冻干粉形式提供的组合物而言，额外的稳定剂、特别是糖类稳定剂的使用还提供额外的优势，因为它们在将所述水性组合物冷冻干燥的过程中可以同时起到冻干赋形剂的作用，从而能够避免其他冻干赋形剂、特别是在现有技术中用作冻干赋形剂的白蛋白(例如HSA)的使用，从而具有成本效益以及有利于降低发生药物过敏反应的风险。因此，在一些实施方案中，所述组合物优选不包含额外的冻干赋形剂。当然，必要时，所述组合物也可以包含额外的冻干赋形剂，例如蔗糖、甘露醇、乳糖、麦芽糖、海藻糖、右旋糖酐中的一种或多种。
在该第一方面的第一子集中，本申请提供组合物，其包含SN-38、脂质、白蛋白和Span 20，其中所述脂质为胆固醇，其特征在于，所述组合物包含纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质，
其中：
胆固醇:SN-38，在一些实施方案中，为约(1-3):1(w:w)，例如约(1.2-2.5):1(w:w)、约(1.4-2):1(w:w)、约(1.5-2):1(w:w)、约(1.3-1.8):1(w:w)、约(1.4-1.6):1(w:w)、约(1.5-1.7):1(w:w)、约(1.2-1.5):1(w:w)、约1:1(w:w)或约(1.4-1.5):1(w:w)，或者，在另一些实施方案中，为约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)或约(1-1.4):1；
白蛋白:SN-38，在一些实施方案中，为约(5-15):1(w:w)，例如约(5-12):1(w:w)、约(6-12):1(w:w)或约(7-12):1(w:w)，约(9-11):1(w:w)，约(10-12):1(w:w)，或约11:1(w:w)，或者，在另一些实施方案中，为约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、或约(11.1-17.3):1(w:w)；
白蛋白:胆固醇，在一些实施方案中，为约(3-10):1(w:w)、约(4-8):1(w:w)或约(5-7):1(w:w)，或者，在另一些实施方案中，为约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)；以及
Span 20:SN-38如上文所述。
在一些实施方案中，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，所述SN-38的含量为约6w/w％至约14w/w％，例如约6.5w/w％至约13w/w％，约7w/w％至约12w/w％，约7.5w/w％至约12w/w％、约8w/w％至约11w/w％、约8.5w/w％至约10w/w％，或约9w/w％。在另一些实施方案中，所述SN-38的含量为约4w/w％至约10w/w％，约4.5w/w％至约9.5w/w％，约5w/w％至约9w/w％，或约7.5w/w％至约8w/w％。在一些实施方案中，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，所述胆固醇的含量为约8w/w％至约18w/w％，例如8.5w/w％至约17w/w％、约9w/w％至约16w/w％、约9.5w/w％至约16w/w％、约10w/w％至约16w/w％、约10.5w/w％至约16w/w％，约11w/w％至约15w/w％，约11.5w/w％至约15w/w％，约12w/w％至约15w/w％、约12.5w/w％至约14w/w％、或约13w/w％至约13.5w/w％。在另一些实施方案中，所述胆固醇的含量为约4w/w％至约12.5w/w％，约4.5w/w％至约12w/w％，约7w/w％至约10w/w％，或约7.5w/w％至约8w/w％。在一些实施方案中，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，所述白蛋白的含量为约66w/w％至约90w/w％，例如约68w/w％至约89w/w、约70w/w％至约88w/w％、约70w/w％至约87w/w％、约70w/w％至约86w/w％，约70w/w％至约85w/w％，约75w/w％至约85w/w％，约76w/w％、约77w/w％、约78w/w％、约79w/w％、约80w/w％、约81w/w％、约82w/w％约83w/w％或约84w/w％。在另一些实施方案中，所述白蛋白的含量为约78w/w％至约93w/w％，约79w/w％至约91.5w/w％，约80w/w％至约90w/w％，约82w/w％至约89w/w％，约84w/w％至约88w/w％，或约84.5w/w％至约87.5w/w％。
在该第一方面的第二子集中，本申请提供组合物，其包含SN-38、脂质、白蛋白和Span 20，其中所述脂质为胆固醇，其特征在于，所述组合物包含纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质，
其中：
胆固醇:SN-38，在一些实施方案中，为约(1-5):1(w:w)，例如约(1-4.5):1(w:w)、约(1-4):1(w:w)、约(1.2-3.8):1(w:w)、约(1.4-3.6):1(w:w)、约(1.6-3.4):1(w:w)、约(1.8-3.2):1(w:w)、约(2-3):1(w:w)、约(2.2-2.8):1(w:w)、约(2.4-2.6):1(w:w)、约2.5:1(w:w)或约1:1(w:w)，或者，在另一些实施方案中，为约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)或约(1-1.4):1；和/或
白蛋白:SN-38，在一些实施方案中，为约(5-25):1(w:w)，例如约(5-20):1(w:w)、约(6-19):1(w:w)、 约(7-18):1(w:w)、约(8-16):1(w:w)、约(9-14):1(w:w)或约(10-12):1(w:w)，或者，在另一些实施方案中，为约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、或约(11.1-17.3):1(w:w)；和/或
白蛋白:胆固醇，在一些实施方案中，为约(5-25):1(w:w)，例如约(6-20):1(w:w)、约(7-18):1(w:w)、约(8-16):1(w:w)、约(9-14):1(w:w)或约(10-12):1(w:w)，或者，在另一些实施方案中，为约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)；和/或
Span 20:SN-38，在一些实施方案中，为约(5-15):100(w:w)，例如约(6-12):100(w:w)、约(7-10):100(w:w)或约7.5:100(w:w)，或者，在另一些实施方案中，为约(5-10):100(w:w)、约(5-9):100(w:w)、约(6-8.6):100(w:w)、约(6-8):100(w:w)、约(6.5-7):100(w:w)。
在一些此类实施方案中，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，所述SN-38的含量为约3w/w％至约10w/w％，例如约3.5w/w％至约9.5w/w％、约4w/w％、约4.5w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约8w/w％、约8.5w/w％或约9w/w％。在另一些实施方案中，所述SN-38的含量为约4w/w％至约10w/w％，约4.5w/w％至约9.5w/w％，约5w/w％至约9w/w％，或约7.5w/w％至约8w/w％。
在一些此类实施方案中，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，所述胆固醇的含量为约4w/w％至约18w/w％，例如约4.5w/w％至约17.5w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约8w/w％、8.5w/w％、约9w/w％、约9.5w/w％、约10w/w％、约10.5w/w％、约11w/w％、约11.5w/w％、约12w/w％、约12.5w/w％、约13w/w％、约13.5w/w％、约14w/w％、约14.5w/w％、约15w/w％、约15.5w/w％、约16w/w％、约16.5w/w％或约17w/w％。在另一些实施方案中，所述胆固醇的含量为约4w/w％至约12.5w/w％，约4.5w/w％至约12w/w％，约7w/w％至约10w/w％，或约7.5w/w％至约8w/w％。
在一些此类实施方案中，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，所述白蛋白的含量为约78w/w％至约92w/w％，例如约79w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约85w/w％、约86w/w％、约87w/w％、约88w/w％、约89w/w％、约90w/w％或约91w/w％。在另一些实施方案中，所述白蛋白的含量为约78w/w％至约93w/w％，约79w/w％至约91.5w/w％，约80w/w％至约90w/w％，约82w/w％至约89w/w％，约84w/w％至约88w/w％，或约84.5w/w％至约87.5w/w％。
在一些进一步的实施方案中，本发明提供上文所述的组合物，其中：
胆固醇:SN-38为约(1-2.5):1(w:w)，例如约1.25:1(w:w)、约1:1(w:w)；和/或
白蛋白:SN-38为约(8-25):1(w:w)，例如约(10-20):1(w:w)、约(12-18):1(w:w)、约(12.5-17):1(w:w)、约(15-16):1(w:w)、约10:1(w:w)或约16.7:1(w:w)；和/或
白蛋白:胆固醇为约(5-10):1(w:w)，例如约(6-9):1(w:w)、约(7-8):1(w:w)、约10:1(w:w)或约6.7:1(w:w)；和/或
Span 20:SN-38为约(5-40):100(w:w)，例如约(6-30):100(w:w)、约(7-25):100(w:w)、约(8-20):100(w:w)、约(9-15):100(w:w)或约(10-12):100(w:w)。
在一些实施方案中，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，所述SN-38的含量为约3w/w％至约9w/w％，例如约3.5w/w％至约8.5w/w％、约4w/w％、约4.5w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约8w/w％。
在一些实施方案中，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，所述胆固醇的含量为约8w/w％至约18w/w％，例如约8.5w/w％至约17.5w/w％、约9w/w％、约9.5w/w％、约10w/w％、约10.5w/w％、约11w/w％、约11.5w/w％、约12w/w％、约12.5w/w％、约13w/w％、约13.5w/w％、约14w/w％、约14.5w/w％、约15w/w％、约15.5w/w％、约16w/w％、约16.5w/w％或约17w/w％。
在一些实施方案中，以所述组合物中的SN-38、脂质、白蛋白和Span 20的总量计，Span 20的含量为约0.2w/w％至约0.6w/w％，例如约0.22w/w％至约0.58w/w％、约0.24w/w％至约0.56w/w％、约0.26w/w％至约0.54w/w％、约0.28w/w％至约0.52w/w％、约0.3w/w％至约0.5w/w％、约0.32w/w％至约0.48w/w％、约0.34w/w％至约0.46w/w％、约0.36w/w％至约0.44w/w％、约0.38w/w％至约0.42w/w％、或约0.4w/w％。
在另一些进一步的实施方案中，本发明提供上文所述的组合物，其中：
胆固醇:SN-38为约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)或约(1-1.4):1；和/或
白蛋白:SN-38为约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、或约(11.1-17.3):1(w:w)；和/或
白蛋白:胆固醇为约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)；和/或
Span 20:SN-38为约(5-10):100(w:w)、约(5-9):100(w:w)、约(6-8.6):100(w:w)、约(6-8):100(w:w)、约(6.5-7):100(w:w)；和/或
以所述组合物中的SN-38、胆固醇和白蛋白的总量计，
所述SN-38的含量为约4w/w％至约10w/w％，约4.5w/w％至约9.5w/w％，约5w/w％至约9w/w％，或约7.5w/w％至约8w/w％；和/或
所述胆固醇的含量为约4w/w％至约12.5w/w％，约4.5w/w％至约12w/w％，约7w/w％至约10w/w％，或约7.5w/w％至约8w/w％；和/或
所述白蛋白的含量为约78w/w％至约93w/w％，约79w/w％至约91.5w/w％，约80w/w％至约90w/w％，约82w/w％至约89w/w％，约84w/w％至约88w/w％，或约84.5w/w％至约87.5w/w％；和/或
Span 20的含量为约0.2w/w％至约0.8w/w％、约0.24w/w％至约0.7w/w％、约0.26w/w％至约0.7w/w％、约0.3w/w％至约0.65w/w％、约0.36w/w％至约0.6w/w％、约0.4w/w％至约0.58w/w％、约0.44w/w％至约0.56w/w％、约0.48w/w％至约0.54w/w％、或约0.5w/w％至约0.52w/w％。
根据所述第二子集的组合物在较大规模(例如，百毫克级或更多的SN-38原料)的制备，包括在放大工艺，例如中试规模的制备中具有优势，例如在制备过程中减少高压均质的次数，有效减小组合物中纳米颗粒的粒径，提高过滤通量，生理条件下崩解后粒径可控，减少原料损耗和降低成本；以及具有适合的白蛋白的含量，从而能够控制纳米颗粒的粒径，使其更接近于适合成药的尺寸。
在根据所述第一和第二子集的一些实施方案中，存在于所述纳米粒子中的SN-38占所述组合物中的SN-38、胆固醇和白蛋白的总量的至少约6w/w％至约12w/w％，例如约7w/w％至约11w/w％、约8w/w％至约10w/w％、约8.3％或约9w/w％。
在一些实施方案中，存在于所述纳米粒子中的SN-38占所述组合物中的SN-38的总量的约95w/w％至约99w/w％，例如约96w/w％至约99w/w％，约97w/w％至约99w/w％、约98w/w％至约99w/w％，或约99w/w％或更高。
在一些根据上文所述的实施方案中，所述组合物为液体、半固体或固体形式。
在一些实施方案中，所述组合物为固体形式，优选为粉末形式，更优选为冻干粉。
在一些此类实施方案中，SN-38优选地以无定形和/或纳米晶体形式存在于所述组合物中，如通过电镜和X射线衍射法(Cu-Kα)分析所测定的。
在一些实施方案中，所述组合物不包含额外的稳定剂。在一些实施方案中，所述组合物还包含额外的稳定剂，所述额外的稳定剂的量使得当所述组合物通过复溶形成水性组合物(包括溶液和乳液)时，所述额外的稳定剂的含量为至少约2w/v％，例如至少约3w/v％，至少约5w/v％，约5w/v％至约30w/v％，约10w/v％至约25w/v％或约15w/v％至约20w/v％。
在一些此类实施方案中，所述组合物还包含额外的稳定剂，以所述组合物的总量计，所述额外的稳定剂的含量为约70w/w％至约96w/w％，例如约70w/w％至约90w/w％，约72w/w％至约89w/w％、约74w/w％至约88w/w％，约76w/w％至约87w/w％，约80w/w％至约86w/w％，约81w/w％至约86w/w％，约82w/w％至约85w/w％、或约83w/w％至约84w/w％。在另一些实施方案中，所述组合物还包含额外的稳定剂，以所述组合物的总量计，所述额外的稳定剂的含量为约80w/w％至约96w/w％或约84w/w％至约95w/w％。
所述额外的稳定剂可以选自甘露醇、乳糖、麦芽糖、海藻糖、右旋糖苷、葡萄糖和蔗糖及其任意组合，优选为蔗糖。
在另一些实施方案中，所述组合物为液体形式的水性组合物，包括溶液和乳液。
在一些此类实施方案中，所述液体形式的组合物包含以纳米晶体和/或囊泡的形式存在的SN-38。
在一些此类实施方案中，所述组合物为溶液的形式；在一些实施方案中，所述组合物为乳液的形式。
在一些此类实施方案中，以所述组合物的总量计，SN-38的含量为约500.0μg/mL至约1.0mg/mL，例如约600μg/mL至约800μg/mL。
在一些此类实施方案中，以所述组合物的总量计，白蛋白的含量为约5.0mg/mL至约10.0mg/mL，例如约6.0mg/mL至约10mg/mL，或约7.0mg/mL至约8.0mg/mL。
在一些实施方案中，所述组合物中的纳米粒子具有约90至160nm的平均粒径，例如约95至150nm、约100至140nm、约105至130nm、约110至125nm、约110nm、约115nm、约120nm、约125nm、约130nm、135nm、约140nm或约145nm。
在一些实施方案中，所述纳米粒子具有不超过约0.30、例如不超过约0.2、不超过约0.10或不超过约0.01的粒径分布指数(PDI)。
在一些实施方案中，所述组合物具有约-35mV至约-20mV，例如约-31mV的Zeta电位。
在一些实施方案中，当稀释(例如用pH约7.4的1x PBS稀释)所述组合物致使SN-38在稀释后的组合物中的含量达到约4μg/mL或更低，例如约2μg/mL或更低，约1μg/mL或更低，约0.4μg/mL或更低，约0.1μg/mL或更低，约0.04μg/mL或更低，约0.02μg/mL或更低，或约0.01μg/mL或更低时，所述纳米粒子不发生崩解。
在一些此类实施方案中，所述组合物不包含额外的稳定剂。在一些实施方案中，所述组合物还包含额外的稳定剂，其中以所述组合物的总量计，所述额外的稳定剂的含量为至少约2w/v％，例如至少约3w/v％，至少约5w/v％，约5w/v％至约30w/v％，约10w/v％至约25w/v％或约15w/v％至约20w/v％。
所述额外的稳定剂可以选自甘露醇、乳糖、麦芽糖、海藻糖、右旋糖苷、葡萄糖和蔗糖及其任意组合，优选为蔗糖。
开环的SN-38是SN-38的无活性形式。本发明人令人惊讶地发现，在本申请的组合物中，开环的SN-38仅以非常低的量存在。在一些实施方案中，所述组合物中开环的SN-38占SN-38总量的约2w/w％或更低，优选约1.8w/w％或更低。
作为载体的白蛋白可能形成多聚体，包括二聚体、三聚体、多聚体等。白蛋白多聚体的存在增加了药物、特别是胃肠外给药的药物产生免疫原性的风险。因此，有利的是，含有尽可能地少的白蛋白多聚体。本发明人令人惊讶地发现，本申请的组合物中不存在或基本不存在白蛋白多聚体。优选地，所述组合物中单体形式的白蛋白占白蛋白总量的至少约95w/w％，优选至少约96％，更优选至少约98％，更优选至少约99％，至少约99.2％，至少约99.4％，或者至少约99.5％。可以预期的是，本申请的组合物具有免疫原性低的优势，因而安全性高。
可用于本申请的白蛋白选自：人血清白蛋白(HSA)、重组人白蛋白(rHA)、牛血清白蛋白和猪血清白蛋白。例如，所述白蛋白包含SEQ ID NO:1所示的氨基酸序列。优选地，所述白蛋白选自人血清白蛋白(HSA)和重组人白蛋白(rHA)。
在第二方面，本申请提供制备上文所述的根据第一方面的组合物的方法，其特征在于，所述方法包括以下步骤：
(1)将所述SN-38、所述脂质和Span 20溶解于有机溶剂中形成有机相；以及制备白蛋白的水溶液以作为水相；
(2)将所述有机相和所述水相混合以形成乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
(3)去除所述乳液中的有机溶剂以得到包含所述纳米粒子的产物。
在一些实施方案中，所述方法包括如下步骤：
(1)用包含选自DMSO和C 1-3醇中的第一有机溶剂和选自CHCl 3及CH 2Cl 2和CHCl 3的混合物的第二有机溶剂的混合有机溶剂溶解所述SN-38、所述脂质和Span 20以形成有机相；以及制备白蛋白的水溶液以作为水相；
(2)将所述有机相与所述水相混合以制备乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒 子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
(3)除去所述有机溶剂；
(4)任选地，将步骤(3)中得到的产物灭菌。
在一些优选的实施方案中，在步骤(1)所述的混合有机溶剂中，所述第二有机溶剂与所述DMSO或C 1-3醇的体积比为约1:20(v/v)至约20:1(v/v)，例如约1:5至约5:1(v/v)，约1:2至约4:1(v/v)，约1:1至约4:1(v/v)，约1.5:1(v/v)至约3:1(v/v)或约2:1(v/v)至约7:3(v/v)。
在一些优选的实施方案中，在步骤(2)中，所述有机相:所述水相为约1:2(v/v)至约1:50(v/v)，例如约1:5(v/v)至约1:20(v/v)，约1:7(v/v)至约1:15(v/v)，1:10(v/v)至约1:12(v/v)；例如约1:5(v/v)至约1:12(v/v)，约1:5(v/v)至约1:12(v/v)，约1:6(v/v)、约1:7(v/v)或约1:10(v/v)。
在一些优选的实施方案中，步骤(2)包括以下步骤：
(2-1)在剪切下，使所述有机相分散在所述水相中以得到粗乳液；和
(2-2)将所述粗乳液在高压下均质化以得到包含所述纳米粒子的细乳液。
所述C 1-3醇包括甲醇、乙醇和异丙醇及其任意组合，例如为乙醇(EtOH)。
在一些实施方案中，所述混合有机溶剂包含约7:3(v/v)的所述第二有机溶剂和乙醇。在一些实施方案中，所述混合有机溶剂包含约1:1(v/v)的所述第二有机溶剂和DMSO。
在一些进一步的实施方案中，本发明提供上文所述的方法，其中所述方法包括如下步骤：
(1)用1:1(v/v)的所述第二有机溶剂/DMSO混合有机溶剂或7:3(v/v)的所述第二有机溶剂/乙醇混合有机溶剂溶解所述SN-38、所述脂质和Span 20以形成有机相；以及制备白蛋白的水溶液以作为水相；
(2)将所述有机相与所述水相以约1:10(v/v)至约1:15(v/v)、例如约1:12(v/v)的比例混合以制备乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
(3)除去所述有机溶剂；
(4)任选地，将步骤(3)中得到的产物灭菌。
在一些实施方案中，所述第二有机溶剂是CHCl 3。在另一些实施方案中，所述第二有机溶剂是CH 2Cl 2和CHCl 3的混合物，其中优选地，所述混合物中CH 2Cl 2和CHCl 3的体积比为约2:5-1:1，优选为约2:5。与单独的CHCl 3相比，使用CH 2Cl 2和CHCl 3的混合物作为所述第二有机溶剂具有降低最终产品中CHCl 3残留水平的优势，从而减少溶剂残留对临床用药剂量的限制。
在一些实施方案中，所述水相不包含额外的稳定剂。
在另一些实施方案中，所述水相已经包含额外的稳定剂；在另一些实施方案中，所述方法还包括在步骤(2)之中添加额外的稳定剂。例如，所述额外的稳定剂的量使得在步骤(3)或(4)得到的产物中，所述额外的稳定剂的含量为至少约2w/v％，例如至少约3w/v％，至少约5w/v％，约5w/v％至约30w/v％，约10w/v％至约25w/v％或约15w/v％至约20w/v％。优选地，所述额外的稳定剂可选自：白蛋白类(例如人血清白蛋白、重组人白蛋白、牛血清白蛋白、脱脂奶粉)、单糖、二糖、多糖、甘露醇及其任意组合；优选地选自：甘露醇、乳糖、麦芽糖、海藻糖、右旋糖苷、葡萄糖和蔗糖及其任意组合；优选地为蔗糖。
在一些实施方案中，在步骤(2)中，在将所述有机相与所述水相混合之前，向所述水相中添加步骤(1)中所述的混合有机溶剂。例如，所添加的混合溶剂的体积等于或小于所述有机相的体积。例如，所添加的混合有机溶剂与所述有机相的体积比为约1:1(v/v)至约1:5(v/v)，例如约1:2(v/v)至约1:4(v/v)或约1:3(v/v)。
在一些实施方案中，在步骤(1)所述的有机相中，SN-38的浓度为约5-17mg/mL，例如约5.25-12mg/mL，约7-12mg/mL，例如约10mg/mL。
在另一些实施方案中，在步骤(1)所述的有机相中，SN-38的浓度为4-10mg/mL，例如约6-8mg/mL。
在一些实施方案中，在步骤(1)所述的有机相中，所述脂质的浓度为约3-50mg/mL，例如约5-45mg/mL或约7.5-30mg/mL，约10-25mg/mL或约15-20mg/mL。
在另一些实施方案中，在步骤(1)所述的有机相中，所述脂质的浓度为约10-20mg/mL，例如约15mg/mL。
在一些实施方案中，在步骤(1)所述的有机相中，所述Span 20的浓度为约0.3-6mg/mL，例如约0.3-2mg/mL或约0.6-1mg/mL。
在一些实施方案中，在所述水相中，白蛋白的浓度为约5-15mg/mL，例如约6-12mg/mL，优选约6-10mg/mL。
在另一些实施方案中，在所述水相中，白蛋白的浓度为约8-30mg/mL，例如约12-20mg/mL或约16-18mg/mL。
在一些实施方案中，所述方法包括步骤(4)将步骤(3)中得到的产物灭菌。对用于灭菌的方法没有特别的限制。在优选的实施方案中，通过将步骤(3)中得到的产物通过约0.2μm的滤膜过滤来除菌。
在一些实施方案中，所述方法还包括以下步骤：
(5)将步骤(3)或(4)得到的产物干燥，优选喷雾干燥或冷冻干燥，以提供固体形式，优选为粉末，更优选为冻干粉的组合物；
优选地，SN-38以无定形和/或纳米晶体形式存在于所述组合物中，如通过电镜或X射线衍射法(Cu-Kα)分析所测定的。
在一些进一步的实施方案中，步骤(5)还包括：在干燥前，向步骤(3)或(4)得到的产物中添加额外的稳定剂，其中所述额外的稳定剂的量使得当步骤(5)所得的固体形式通过复溶形成水性组合物(包括溶液和乳液)时，所述额外的稳定剂的含量为至少约2w/v％，例如至少约3w/v％，至少约5w/v％，约5w/v％至约30w/v％，约10w/v％至约25w/v％或约15w/v％至约20w/v％。
所述额外的稳定剂可选自：白蛋白类(例如人血清白蛋白、重组人白蛋白、牛血清白蛋白、脱脂奶粉)、单糖、二糖、多糖、甘露醇及其任意组合；优选地选自：甘露醇、乳糖、麦芽糖、海藻糖、右旋糖苷、葡萄糖和蔗糖及其任意组合；优选为蔗糖。
在第三方面，本申请提供制备包含SN-38、脂质、白蛋白和Span 20的组合物的方法，其特征在于，所述方法包括以下步骤：
(1)将所述SN-38、所述脂质和Span 20溶解于有机溶剂中以形成有机相；以及制备白蛋白的水溶液以作为水相；
(2)将所述有机相和所述水相混合以形成乳液，所述乳液包含纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
(3)去除所述乳液中的有机溶剂以得到含有纳米粒子的产物。
所述方法的实施方案如上文关于第二方面的方法所述。
在根据第二和第三方面的方法中，部分白蛋白可以包封部分SN-38从而形成纳米粒子，或者可以包封部分脂质从而形成纳米粒子。因此，在一些实施方案中，根据第一方面和下文第四方面的组合物可包含由部分所述白蛋白包封部分所述SN-38而形成的纳米粒子；和/或由部分所述白蛋白包封部分所述脂质而形成的纳米粒子。
在第四方面，本申请提供组合物，其可以通过如上文述的根据第三方面的方法制备。
在第五方面，本申请还提供制备具有改善的性质的组合物的方法，其中所述组合物包含SN-38、脂质和白蛋白，并且所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质以形成纳米粒子，
所述方法的特征在于在制备所述组合物的过程中加入Span 20。
在一些实施方案中，所述组合物不包含额外的稳定剂。
在一些实施方案中，所述改善的性质包括提高的稳定性。
在一些实施方案中，当所述组合物为液体形式时，所述提高的稳定性包括：在所述组合物的制备、储存和/或使用中，减少的白蛋白多聚体的产生或含量，和/或降低的纳米粒子粒径。优选地，所述组合物中不存在或基本不存在白蛋白多聚体，或者白蛋白多聚体占白蛋白总量至多5w/w％，例如至多约4％、至多约2％、至多约1.5％、至多约1.2％、至多约1.1％、至多约1％、或至多约0.8％。
在一些实施方案中，所述组合物如在上文第一方面中所述。
在一些实施方案中，所述方法包括以下步骤：
(1)将所述SN-38、所述脂质和Span 20溶解于有机溶剂中形成有机相；以及制备白蛋白的水 溶液以作为水相；
(2)将所述有机相和所述水相混合以形成乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
(3)去除所述乳液中的有机溶剂以得到包含所述纳米粒子的产物。
进一步地，所述方法的实施方案如上文关于第二方面的方法所述。
在第六方面，本申请提供药物组合物，其包含上文所述的组合物以及任选存在的药学上可接受的载体。
在第七方面，本申请提供药物组合物，其包含经干燥的上文所述的组合物以及任选存在的药学上可接受的载体。所述干燥优选是冷冻干燥或喷雾干燥，更优选冷冻干燥。优选地，所述药物组合物为固体形式，优选为冻干粉。优选地，SN-38以无定形和/或纳米晶体形式存在于所述组合物中。
对本申请的组合物或药物组合物的给药途径没有特别的限制。可以考虑的给药途径包括但不限于口服、经鼻、局部和胃肠外给药。优选地，所述药物组合物用于胃肠外给药，包括但不限于静脉内、动脉内、皮下、皮内和肌内注射给药，更优选静脉内注射(例如，推注或输注)给药。
药学上可接受的载体的选择取决于药物或药物组合物的剂型，首先取决于剂型的施用途径(例如用于口服、经鼻、皮内、皮下、局部、肌内或静脉施用的剂型)，其次取决于剂型的配方。例如，所述药学上可接受的载体可以包括水(如注射用水)、缓冲液、等渗盐溶液如PBS(磷酸盐缓冲液)、葡萄糖、甘露醇、右旋葡萄糖、乳糖、淀粉、硬脂酸镁、纤维素、碳酸镁、0.3％甘油、透明质酸、抗坏血酸、乳酸、乙醇、聚亚烷基二醇如聚乙二醇(例如聚乙二醇4000)或聚丙二醇、甘油三酯等。
在第八方面，本申请提供上文所述的组合物或者药物组合物在制备用于治疗个体的对SN-38敏感的肿瘤的药物中的用途。
在第九方面，本申请提供上文所述的组合物或者药物组合物，其用于治疗个体的对SN-38敏感的肿瘤。
在第十方面，本申请提供治疗个体的对SN-38敏感的肿瘤的方法，其包括向所述个体施用治疗有效量的上文所述的组合物或者药物组合物。
在一些实施方案中，所述个体为哺乳动物，包括但不限于小鼠、大鼠、兔、豚鼠、狗、猫、绵羊、牛、羊、马。在一些实施方案中，所述个体为人。
“对SN-38敏感的肿瘤”指的是对SN-38的给药有应答的肿瘤，所述应答包括肿瘤细胞的减少、肿瘤尺寸的缩小、肿瘤转移的消除、肿瘤生长的抑制，等等。优选地，所述对SN-38敏感的肿瘤选自结直肠癌、小细胞肺癌、淋巴癌、乳腺癌(优选三阴性乳腺癌)、食管癌、胃癌、肝癌、肾癌、胰腺癌、子宫癌和卵巢癌。
实验研究表明抗肿瘤药物与白蛋白结合后，可以显著提高药物的抗肿瘤效果，例如纳米白蛋白结合型紫杉醇，具有肿瘤组织选择性和独特的转运机制，与溶液剂紫杉醇相比，肿瘤中紫杉醇药物的浓度增加33％(Desai N,Trieu V,Yao Z,et al.,Increased antitumor activity,intratumor paclitaxel concentrations,and endothelial celltransport of cremophor-free,albumin-bound paclitaxel,ABI-007,compared with cremophor-based paclitaxel,[J].Clinical cancer research,2006,12(4):1317-1324)。上述结果的产生是因为药物与白蛋白结合后，可以利用肿瘤组织gp60受体与白蛋白结合，达到肿瘤主动靶向作用，以及利用药物白蛋白结合物纳米粒的EPR效应，达到肿瘤被动靶向作用，从而大大提高了药物的肿瘤分布，提高疗效、减小毒副作用。可以预期的是，本申请的组合物或药物组合物具有有利的抗肿瘤功效。通过初步的实验，本发明人发现本申请的组合物具有优于市售盐酸伊立替康注射液的抗肿瘤活性。
在第十一方面，本申请提供药盒，其包含上文所述的组合物或者药物组合物。必要时，所述药盒还可包含使用说明书、包装、容纳所述组合物或者药物组合物的容器。
尽管在上文第一至十一方面中以SN-38作为活性成分描述了本发明的实施方案，但这些只是本发明概念的一个方面。本发明的概念还涵盖了其他喜树碱类药物作为活性成分的技术方案。预期上文所述的实施方案在SN-38被替换为其他喜树碱类药物时仍然适用。换言之，本申请还包括上文第一至十方面中任一方面所述的任意和全部实施方案，只是其中的SN-38被替换为其他喜树碱类药物，除非上下文中存在抵触。所述其他喜树碱类药物可选自：伊立替康(CPT-11)、10-羟基喜树碱(HCPT)、 拓扑替康(TPT)、鲁比替康(9-NC)、9-氨基喜树碱(9-AC)、贝洛替康(Cas.No.:256411-32-2)、Dxd(Cas.No.:1599440-33-1)、DX-8951(依沙替康)、CKD602(倍罗替康(belotecan))、勒托替康(lurtotecan)、Namitecan(Cas.No.:372105-27-6)、ST1481(gimatecan,Cas.No.:292618-32-7)、BNP-1350(Cas.No.203923-89-1)和BN80915(二氟替康(Diflomotecan))。本领域技术人员会理解，此类实施方案也会实现下文所述的有益效果。
如本申请的发明内容所述的本发明的实施方案包括：
实施方案1.组合物，其包含SN-38、脂质、白蛋白和Span 20，其特征在于，所述组合物包含纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
脂质:SN-38为约(0.1-10):1(w:w)，约(0.5-6):1(w:w)，约(0.5-5):1(w:w)，约(0.5-3):1(w:w)，约(1-4):1(w:w)，约(1.2-4):1(w:w)，约(1.4-2):1(w:w)，约(1.5-2.5):1(w:w)或约1:1；
白蛋白:SN-38为约(1-100):1(w:w)，约(1-50):1(w:w)，约(3-25):1(w:w)，约(5-25):1(w:w)，约(5-20):1(w:w)，约(5-18):1(w:w)，约(6-15):1(w:w)，约(7-15):1(w:w)，约(6-12):1(w:w)，约(7-12):1(w:w)、约(9-11):1(w:w)，约10:1(w:w)；以及
Span 20:SN-38为约(3-60):100(w:w)，约(4-60):100(w:w)，约(5-60):100(w:w)，约(6-60):100(w:w)，约(7-55):100(w:w)，约(8-50):100(w:w)，约(10-45):100(w:w)，约(12-40):100(w:w)，约(14-35):100(w:w)，约(15-30):100(w:w)，约(16-25):100(w:w)，或约(18-20):100(w:w)；并且
其中所述脂质选自胆固醇、胆固醇衍生物、胆固醇类似物和脂肪酸酯以及它们中的两者或更多者的任意组合。
实施方案2.实施方案1的组合物，其特征在于，白蛋白:脂质为约(1-100):1(w:w)，例如约(2-20):1(w:w)，约(3-15):1(w:w)，约(5-10):1(w:w)，约7:1(w:w)，或约10:1(w:w)。
实施方案3.实施方案1或2的组合物，其特征在于，以所述组合物中的SN-38、脂质和白蛋白的总量计，
所述SN-38的含量为约1w/w％至约25w/w％；和/或
所述脂质的含量为约1w/w％至约35w/w％；和/或
所述白蛋白的含量为约50w/w％至约98w/w％；
或者，
所述SN-38的含量为约3w/w％至约20w/w％；和/或
所述脂质的含量为约2w/w％至约30w/w％；和/或
所述白蛋白的含量为约55w/w％至约95w/w％；
或者，
所述SN-38的含量为约3w/w％至约15w/w％、约4w/w％、约5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约8w/w％、约8.5w/w％、约9w/w％、约10w/w％、约11w/w％、约12w/w％、约13w/w％或约14w/w％；和/或
所述脂质的含量为约3w/w％至约30w/w％、约4w/w％、约5w/w％、约6w/w％、约7w/w％、约8w/w％、约8.5w/w％、约9w/w％、约9.5w/w％、约10w/w％、约10.5w/w％、约11w/w％、约11.5w/w％、约12w/w％、约12.5w/w％、约13w/w％、约13.5w/w％、约14w/w％、约15w/w％、约16w/w％、约17w/w％、约18w/w％、约19w/w％、约20w/w％、约21w/w％、约24w/w％、约26w/w％或约28w/w％；和/或
所述白蛋白的含量为约60w/w％至约94w/w％、约64w/w％至约93w/w％、约66w/w％至约92w/w％、约68w/w％至约91w/w％、约70w/w％至约90w/w％、约75w/w％至约90w/w％、约76w/w％、约77w/w％、约78w/w％、约79w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约85w/w％、约86w/w％、约87w/w％、约88w/w％或约89w/w％。
实施方案4.实施方案1至3中任一项的组合物，其特征在于，以所述组合物中的SN-38、脂质、白蛋白和Span 20的总量计，Span 20的含量为约0.03w/w％至约12w/w％、约0.06w/w％至约10w/w％、0.08w/w％至约9w/w％、约0.1w/w％至约8w/w％、约0.2w/w％至约7w/w％、约0.4w/w％至约6w/w％、约0.6w/w％至约5w/w％、约0.8w/w％至约4w/w％、约1w/w％至约2w/w％。
实施方案5.实施方案1至4中任一项的组合物，其特征在于，
脂质:SN-38为约(1-10):1(w:w)、约(1-8):1(w:w)、约(1-6):1(w:w)、约(1-5):1(w:w)、约(1-4.5):1(w:w)、约(1-4):1(w:w)、约(1.2-3.8):1(w:w)、约(1.4-3.6):1(w:w)、约(1.6-3.4):1(w:w)、约(1.8-3.2):1(w:w)、约(2-3):1(w:w)、约(2.2-2.8):1(w:w)、约(2.4-2.6):1(w:w)、约1:1(w:w)、约2.5:1(w:w)、或约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)、约(1-1.4):1；和/或
白蛋白:SN-38为约(5-40):1(w:w)、约(5-小于40):1(w:w)、约(5-35):1(w:w)、约(5-30):1(w:w)、约(8-25):1(w:w)、约(10-22.5):1(w:w)、约(12.5-20):1(w:w)、约(15-17.5):1(w:w)、约(16-18):1(w:w)、约10:1(w:w)、或约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、约(11.1-17.3):1(w:w)；和/或
Span 20:SN-38为约(5-60):100(w:w)、约(6-60):100(w:w)、约(7-55):100(w:w)、约(8-50):100(w:w)、约(10-45):100(w:w)、约(12-40):100(w:w)、约(14-35):100(w:w)、约(15-30):100(w:w)、约(16-25):100(w:w)、约(18-20):100(w:w)、或约(5-10):100(w:w)、约(5-9):100(w:w)、约(6-8.6):100(w:w)、约(6-8):100(w:w)、约(6.5-7):100(w:w)；和/或
白蛋白:脂质为约(1-40):1(w:w)、约(1-小于40):1(w:w)、约(2-35):1(w:w)、约(3-15):1(w:w)、约(5-10):1(w:w)、约(6-8):1(w:w)或约7:1(w:w)、或约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)。
实施方案6.实施方案1至5中任一项的组合物，其特征在于，以所述组合物中的SN-38、脂质和白蛋白的总量计，
所述SN-38的含量为约2w/w％至约16w/w％；和/或
所述脂质的含量为约2w/w％至约35w/w％；和/或
所述白蛋白的含量为约75w/w％至约96w/w％；
或者，
所述SN-38的含量为约2.5w/w％至约15w/w％，约4w/w％至约10w/w％，约4.5w/w％至约9.5w/w％，约5w/w％至约9w/w％，或约7.5w/w％至约8w/w％；和/或
所述组合物中的脂质的含量为约2.5w/w％至约30w/w％，约4w/w％至约12.5w/w％，约4.5w/w％至约12w/w％，约7w/w％至约10w/w％，或约7.5w/w％至约8w/w％；和/或
所述组合物中的白蛋白的含量为约76w/w％至约95w/w％，约78w/w％至约93w/w％，约79w/w％至约91.5w/w％，约80w/w％至约90w/w％，约82w/w％至约89w/w％，约84w/w％至约88w/w％，或约84.5w/w％至约87.5w/w％；
或者，
所述SN-38的含量为约3w/w％至约14w/w％，约3.5w/w％至约12w/w％、约4w/w％、约4.2w/w％、约4.5w/w％、约4.6w/w％、约4.8w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约7.6w/w％、约7.8w/w％、约8w/w％、约8.5w/w％、约9w/w％、约9.2w/w％、约9.5w/w％、约9.6w/w％、约9.8w/w％、约10w/w％、约10.5w/w％、约11w/w％或约11.5w/w％；和/或
所述脂质的含量为约4w/w％至约25w/w％、约4.5w/w％至约20w/w％、约4.3w/w％、约4.5w/w％、约4.7w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约6.7w/w％、约6.9w/w％、约7w/w％、约7.5w/w％、约7.6w/w％、约7.8w/w％、约8w/w％、约8.5w/w％、约9w/w％、约9.5w/w％、约10w/w％、约10.5w/w％、约11w/w％、约11.5w/w％、约12w/w％、约12.1w/w％、约12.3w/w％、约12.5w/w％、约13w/w％、约13.5w/w％、约14w/w％、约14.5w/w％、约15w/w％、约15.5w/w％、约16w/w％、约16.5w/w％、约17w/w％、约17.5w/w％、约18w/w％、约18.5w/w％、约19w/w％或约19.5w/w％；和/或
所述白蛋白的含量为约78w/w％至约92w/w％、约79w/w％、约79.2w/w％、约79.4w/w％、约79.6w/w％、约79.8w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约84.3w/w％、约84.5w/w％、约84.7w/w％、约84.9w/w％、约85w/w％、约86w/w％、约87w/w％、约87.3w/w％、约87.5w/w％、约87.7w/w％、约87.9w/w％、约88w/w％、约89w/w％、约90w/w％、约91w/w％、约91.3w/w％、或约91.5w/w％。
实施方案7.实施方案1至6中任一项的组合物，其特征在于，以所述组合物中的SN-38、脂质、白蛋白和Span 20的总量计，Span 20的含量为约0.14w/w％至约5w/w％、约0.2w/w％至约2.5w/w％、约0.22w/w％至约2.0w/w％、约0.24w/w％至约2w/w％、约0.26w/w％至约1.5w/w％、约0.28w/w％至约1.0w/w％、约0.3w/w％至约0.9w/w％、约0.32w/w％至约0.8w/w％、约0.34w/w％至约0.7w/w％、约0.36w/w％至约0.6w/w％、约0.38w/w％至约0.58w/w％、约0.4w/w％至约0.56w/w％、约0.42w/w％至约0.54w/w％、约0.44w/w％至约0.52w/w％、约0.46w/w％、约0.48w/w％、约0.5w/w％、约0.2w/w％至约0.8w/w％、约0.24w/w％至约0.7w/w％、约0.26w/w％至约0.7w/w％、约0.3w/w％至约0.65w/w％、约0.36w/w％至约0.6w/w％、约0.4w/w％至约0.58w/w％、约0.44w/w％至约0.56w/w％、约0.48w/w％至约0.54w/w％、或约0.5w/w％至约0.52w/w％。
实施方案8.实施方案1至7中任一项的组合物，其特征在于，
存在于所述纳米粒子中的SN-38占所述组合物中的SN-38、脂质和白蛋白的总量的至少约1w/w％或至少约2w/w％，例如至少约3w/w％，约3w/w％至约13w/w％，约4w/w％至约12w/w％，约4w/w％、约5w/w％、约6w/w％、约7w/w％、约8w/w％、约9w/w％、约10w/w％或约11w/w％；和/或
存在于所述纳米粒子中的SN-38占所述组合物中的SN-38的总量的约80w/w％至约99w/w％，例如约88w/w％至约98w/w％，约89w/w％、约90w/w％、约91w/w％、约92w/w％、约93w/w％、约94w/w％、约95w/w％、约96w/w％或约97w/w％。
实施方案9.实施方案1至8中任一项的组合物，其特征在于，
所述胆固醇衍生物选自胆固醇与有机酸形成的酯，包括胆固醇棕榈酸酯和胆固醇辛酸酯及其组合；和/或；
所述胆固醇类似物选自维生素D2、维生素D3及其组合；和/或
所述脂肪酸酯选自脂肪酸甘油酯，例如长链脂肪酸甘油酯，包括硬脂酸甘油酯，例如单硬脂酸甘油酯。
实施方案10.实施方案1至9中任一项的组合物，其特征在于，所述脂质为胆固醇。
实施方案11.实施方案10的组合物，其特征在于，
胆固醇:SN-38为约(1-6):1(w:w)、约(1.2-5):1(w:w)，约(1.4-4):1(w:w)、约3:1(w:w)、约2:1(w:w)、约1:1(w:w)、或约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)、约(1-1.4):1；和/或
白蛋白:SN-38为约(3-25):1(w:w)、约(4-20):1(w:w)、约(5-15):1(w:w)、约(6-12):1(w:w)、约(7-12):1(w:w)、约(9-11):1(w:w)或约10:1(w:w)、或约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、约(11.1-17.3):1(w:w)；和/或
白蛋白:胆固醇为约(2-20):1(w:w)、约(3-15):1(w:w)、约(5-10):1(w:w)或约7:1(w:w)、或约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)；和/或
以所述组合物中的SN-38、胆固醇和白蛋白的总量计，
所述SN-38的含量为约3w/w％至约15w/w％、约4w/w％至约15w/w％、约6w/w％至约10w/w％或约8w/w％至约12w/w％、或约4w/w％至约10w/w％、约4.5w/w％至约9.5w/w％、约5w/w％至约9w/w％、约7.5w/w％至约8w/w％；和/或
所述胆固醇的含量为约5w/w％至约25w/w％、约6w/w％至约22w/w％、约15w/w％至约20w/w％、或约4w/w％至约12.5w/w％、约4.5w/w％至约12w/w％、约7w/w％至约10w/w％、约7.5w/w％至约8w/w％；和/或
所述白蛋白的含量为约64w/w％至约90w/w％、约70w/w％至约90w/w％、或约78w/w％至约93w/w％、约79w/w％至约91.5w/w％、约80w/w％至约90w/w％、约82w/w％至约89w/w％、约84w/w％至约88w/w％、约84.5w/w％至约87.5w/w％；和/或
存在于所述纳米粒子中的SN-38占所述组合物中的SN-38、胆固醇和白蛋白的总量的至少约3w/w％，例如约3w/w％至约13w/w％，约4w/w％至约12w/w％，约4w/w％、约5w/w％、约6w/w％、约7w/w％、约8w/w％、约9w/w％、约10w/w％或约11w/w％。
实施方案12.实施方案10的组合物，其中胆固醇:SN-38为约(1-3):1(w:w)、约(1.2-2.5):1(w:w)、约(1.4-2):1(w:w)、约(1.5-2):1(w:w)、约(1.3-1.8):1(w:w)、约(1.4-1.6):1(w:w)、约(1.5-1.7):1(w:w)、 约(1.2-1.5):1(w:w)、约1:1(w:w)、约(1.4-1.5):1(w:w)、或约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)、约(1-1.4):1；
白蛋白:SN-38为约(5-15):1(w:w)、约(5-12):1(w:w)、约(6-12):1(w:w)或约(7-12):1(w:w)、约(9-11):1(w:w)、约(10-12):1(w:w)、约11:1(w:w)、或约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、或约(11.1-17.3):1(w:w)；以及
白蛋白:胆固醇为约(3-10):1(w:w)、约(4-8):1(w:w)、约(5-7):1(w:w)、或约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)。
实施方案13.实施方案12的组合物，其特征在于，以所述组合物中的SN-38、胆固醇和白蛋白的总量计，
所述SN-38的含量为约6w/w％至约14w/w％，约6.5w/w％至约13w/w％，约7w/w％至约12w/w％，约7.5w/w％至约12w/w％、约8w/w％至约11w/w％、约8.5w/w％至约10w/w％，约9w/w％，或约4w/w％至约10w/w％，约4.5w/w％至约9.5w/w％，约5w/w％至约9w/w％，或约7.5w/w％至约8w/w％；和/或
所述胆固醇的含量为约8w/w％至约18w/w％，约8.5w/w％至约17w/w％、约9w/w％至约16w/w％、约9.5w/w％至约16w/w％、约10w/w％至约16w/w％、约10.5w/w％至约16w/w％，约11w/w％至约15w/w％，约11.5w/w％至约15w/w％，约12w/w％至约15w/w％、约12.5w/w％至约14w/w％、约13w/w％至约13.5w/w％，或约4w/w％至约12.5w/w％，约4.5w/w％至约12w/w％，约7w/w％至约10w/w％，或约7.5w/w％至约8w/w％；和/或
所述白蛋白的含量为约66w/w％至约90w/w％，约68w/w％至约89w/w、约70w/w％至约88w/w％、约70w/w％至约87w/w％、约70w/w％至约86w/w％，约70w/w％至约85w/w％，约75w/w％至约85w/w％，约76w/w％、约77w/w％、约78w/w％、约79w/w％、80w/w％、81w/w％、约82w/w％约83w/w％，约84w/w％，约78w/w％至约93w/w％，约79w/w％至约91.5w/w％，约80w/w％至约90w/w％，约82w/w％至约89w/w％，约84w/w％至约88w/w％，或约84.5w/w％至约87.5w/w％。
实施方案14.实施方案10的组合物，其特征在于，
胆固醇:SN-38为约(1-5):1(w:w)、约(1-4.5):1(w:w)、约(1-4):1(w:w)、约(1.2-3.8):1(w:w)、约(1.4-3.6):1(w:w)、约(1.6-3.4):1(w:w)、约(1.8-3.2):1(w:w)、约(2-3):1(w:w)、约(2.2-2.8):1(w:w)、约(2.4-2.6):1(w:w)、约2.5:1(w:w)、约1:1(w:w)、或约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)、约(1-1.4):1；和/或
白蛋白:SN-38为约(5-25):1(w:w)、约(5-20):1(w:w)、约(6-19):1(w:w)、约(7-18):1(w:w)、约(8-16):1(w:w)、约(9-14):1(w:w)或约(10-12):1(w:w)、或约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、或约(11.1-17.3):1(w:w)；和/或
白蛋白:胆固醇为约(5-25):1(w:w)、约(6-20):1(w:w)、约(7-18):1(w:w)、约(8-16):1(w:w)、约(9-14):1(w:w)、约(10-12):1(w:w)、或约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)；和/或
Span 20:SN-38为约(5-40):100(w:w)、约(6-30):100(w:w)、约(7-25):100(w:w)、约(8-20):100(w:w)、约(9-15):100(w:w)、约(10-12):100(w:w)、或约(5-10):100(w:w)、约(5-9):100(w:w)、约(6-8.6):100(w:w)、约(6-8):100(w:w)、约(6.5-7):100(w:w)；和/或
以所述组合物中的SN-38、胆固醇和白蛋白的总量计，
所述SN-38的含量为约3w/w％至约10w/w％、约3.5w/w％至约9.5w/w％、约4w/w％、约4.5w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约8w/w％、约8.5w/w％、约9w/w％、或约4w/w％至约10w/w％、约4.5w/w％至约9.5w/w％、约5w/w％至约9w/w％、约7.5w/w％至约8w/w％；和/或
所述胆固醇的含量为约4w/w％至约18w/w％、约4.5w/w％至约17.5w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约8w/w％、8.5w/w％、约9w/w％、约9.5w/w％、约10w/w％、约10.5w/w％、约11w/w％、约11.5w/w％、约12w/w％、约12.5w/w％、约13w/w％、约13.5w/w％、约14w/w％、约14.5w/w％、约15w/w％、约15.5w/w％、约16w/w％、 约16.5w/w％、约17w/w％、或约4w/w％至约12.5w/w％、约4.5w/w％至约12w/w％、约7w/w％至约10w/w％、约7.5w/w％至约8w/w％；和/或
所述白蛋白的含量为约78w/w％至约92w/w％、约79w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约85w/w％、约86w/w％、约87w/w％、约88w/w％、约89w/w％、约90w/w％、约91w/w％、或约78w/w％至约93w/w％、约79w/w％至约91.5w/w％、约80w/w％至约90w/w％、约82w/w％至约89w/w％、约84w/w％至约88w/w％、或约84.5w/w％至约87.5w/w％。
实施方案15.实施方案12至14中任一项的组合物，其特征在于，
存在于所述纳米粒子中的SN-38占所述组合物中的SN-38、胆固醇和白蛋白的总量的至少约6w/w％至约12w/w％，例如约7w/w％至约11w/w％、约8w/w％至约10w/w％、约8.3％或约9w/w％；和/或
存在于所述纳米粒子中的SN-38占所述组合物中的SN-38的总量的约95w/w％至约99w/w％，例如约96w/w％至约99w/w％，约97w/w％至约99w/w％、约98w/w％至约99w/w％，约99w/w％或更高。
实施方案16.实施方案1至15中任一项的组合物，其特征在于，所述组合物为液体、半固体或固体形式。
实施方案17.实施方案1至16中任一项的组合物，其特征在于，所述组合物为固体形式，优选为粉末形式，更优选为冻干粉；
优选地，SN-38以无定形和/或纳米晶体的形式存在于所述组合物中。
实施方案18.实施方案1至17中任一项的组合物，其特征在于，所述组合物不包含额外的稳定剂；或者
优选地，所述组合物还包含额外的稳定剂，例如冻干稳定剂，所述额外的稳定剂的量使得当所述组合物通过复溶形成水性组合物(包括溶液和乳液)时，所述额外的稳定剂的含量为至少约2w/v％，例如至少约3w/v％，例如至少约5w/v％，约5w/v％至约30w/v％，约10w/v％至约25w/v％或约15w/v％至约20w/v％。
实施方案19.实施方案17或18的组合物，其特征在于，所述组合物还包含额外的稳定剂，以所述组合物的总量计，
所述额外的稳定剂的含量为约60w/w％至约98w/w％，例如约65w/w％至约97w/w％，约68w/w％至约96w/w％，约69w/w％至约95w/w％，约70w/w％至约94w/w％、约71w/w％至约93w/w％、约72w/w％至约92w/w％、约73w/w％、约74w/w％、约75w/w％、约76w/w％、约77w/w％、约78w/w％、约79w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约85w/w％、约86w/w％、约87w/w％、约88w/w％、约89w/w％、约90w/w％或约91w/w％；
例如，所述额外的稳定剂的含量为约70w/w％至约96w/w％、约70w/w％至约90w/w％、约72w/w％至约89w/w％、约74w/w％至约88w/w％、约76w/w％至约87w/w％，或约80w/w％至约96w/w％、约80w/w％至约86w/w％、约81w/w％至约86w/w％、约82w/w％至约85w/w％、约83w/w％至约84w/w％、约84w/w％至约95w/w％。
20.实施方案18或19的组合物，其中所述额外的稳定剂选自：白蛋白类(例如人血清白蛋白、重组人白蛋白、牛血清白蛋白、脱脂奶粉)、单糖、二糖、多糖、甘露醇及其任意组合；优选地选自：甘露醇、乳糖、麦芽糖、海藻糖、右旋糖苷、葡萄糖和蔗糖及其任意组合；优选为蔗糖。
实施方案21.实施方案17至20中任一项的组合物，其中当所述组合物通过复溶形成水性组合物(包括溶液和乳液)而具有约0.1μg/mL至约30.0mg/mL时，所述纳米粒子具有约50至200nm，例如约90至150nm、约95至140nm、约100至130nm、约105至125nm或约110至120nm的平均粒径。
实施方案22.实施方案1至16中任一项的组合物，其特征在于，所述组合物为液体形式的水性组合物，包括溶液和乳液；
特别的，所述液体形式的组合物包含以纳米晶体和/或囊泡的形式存在的SN-38。
实施方案23.实施方案22的组合物，其特征在于，以所述组合物的总量计，
SN-38的含量为约0.1μg/mL至约30.0mg/mL、约0.2μg/mL至约27.0mg/mL、约0.5μg/mL至约24.0mg/mL、约1.0μg/mL至约21.0mg/mL、约5.0μg/mL至约18.0mg/mL、约10.0μg/mL至约15.0mg/mL、约20.0μg/mL至约12mg/mL、约25.0μg/mL至约9mg/mL、约50.0μg/mL至约6.0mg/mL或约100.0μg/mL至约3.0mg/mL；和/或
所述脂质的含量为约0.05μg/mL至约100.0mg/mL、约0.1μg/mL至约90.0mg/mL、约0.25μg/mL至约80.0mg/mL、约0.5μg/mL至约70.0mg/mL、约2.5μg/mL至约60.0mg/mL、约5.0μg/mL至约50.0mg/mL、约10.0μg/mL至约40.0mg/mL、约12.5μg/mL至约30.0mg/mL、约25.0μg/mL至约20.0mg/mL、或约50.0μg/mL至约10.0mg/mL；和/或
白蛋白的含量为约3.0μg/mL至约300.0mg/mL、约6.0μg/mL至约270.0mg/mL、约15.0μg/mL至约240.0mg/mL、约30.0μg/mL至约210.0mg/mL、约150.0μg/mL至约180.0mg/mL、约300.0μg/mL至约150.0mg/mL、约600.0μg/mL至约120.0mg/mL、约750.0μg/mL至约90.0mg/mL、约1500.0μg/mL至约60.0mg/mL、或约3.0mg/mL至约30.0mg/mL。
实施方案24.实施方案22或23的组合物，其特征在于，以所述组合物的总量计，
SN-38的含量为约100.0μg/mL至约3.0mg/mL，例如约200.0μg/mL至约2.5mg/mL、约300.0μg/mL至约2.0mg/mL、约400.0μg/mL至约1.5mg/mL、约500.0μg/mL至约1.0mg/mL、或约600μg/mL至约800μg/mL；和/或
所述脂质的含量为约50.0μg/mL至约10.0mg/mL，例如约100.0μg/mL至约8.0mg/mL、约200.0μg/mL至约6.0mg/mL、约300.0μg/mL至约4.0mg/mL、约400.0μg/mL至约3.0mg/mL、约500.0μg/mL至约2.5mg/mL、约600.0μg/mL至约2.0mg/mL、约700.0μg/mL至约1.5mg/mL、约800μg/mL至约1.0mg/mL、或约200μg/mL至约1.5mg/mL；和/或
白蛋白的含量为约3.0mg/mL至约30.0mg/mL，例如约4.0mg/mL至约25.0mg/mL、约5.0mg/mL至约20.0mg/mL、约6.0mg/mL至约15.0mg/mL、约7.0mg/mL至约12.0mg/mL、或约8.0mg/mL至约10.0mg/mL。
实施方案25.实施方案22至24中任一项的组合物，其特征在于，所述纳米粒子具有约50至200nm，例如约90至150nm、约95至140nm、约100至130nm、约105至125nm或约110至120nm的平均粒径。
实施方案26.实施方案22至25中任一项的组合物，其特征在于，当在4℃下保存24h后，所述纳米粒子具有约50至200nm、例如约90至150nm或约100至130nm的平均粒径。
实施方案27.实施方案22至26中任一项的组合物，其特征在于，所述纳米粒子具有不超过约0.30、例如不超过约0.2、不超过约0.10或不超过约0.01的粒径分布指数(PDI)。
实施方案28.实施方案22至27中任一项的组合物，其特征在于，所述组合物具有约-35mV至约-20mV，例如约-31mV的Zeta电位。
实施方案29.实施方案22至28中任一项的组合物，其特征在于，当稀释(例如用pH约7.4的1x PBS稀释)所述组合物致使SN-38在稀释后的组合物中的含量达到约4μg/mL或更低，例如约2μg/mL或更低，约1μg/mL或更低，约0.4μg/mL或更低，约0.1μg/mL或更低，约0.04μg/mL或更低，约0.02μg/mL或更低，或约0.01μg/mL或更低时，所述纳米粒子不发生崩解。
实施方案30.实施方案22至29中任一项的组合物，其特征在于，所述组合物不包含额外的稳定剂；或者
优选地，所述组合物还包含额外的稳定剂，其中以所述组合物的总量计，所述额外的稳定剂的含量为至少约2w/v％，例如至少约3w/v％，至少约5w/v％，约5w/v％至约30w/v％，约10w/v％至约25w/v％或约15w/v％至约20w/v％。
实施方案31.实施方案30的组合物，其中所述额外的稳定剂选自：白蛋白类(例如人血清白蛋白、重组人白蛋白、牛血清白蛋白、脱脂奶粉)、单糖、二糖、多糖、甘露醇及其任意组合；优选地选自：甘露醇、乳糖、麦芽糖、海藻糖、右旋糖苷、葡萄糖和蔗糖及其任意组合；优选为蔗糖。
实施方案32.实施方案1至31中任一项的组合物，其特征在于，所述组合物中开环的SN-38占SN-38总量的约2w/w％或更低，例如约1.8w/w％或更低；和/或
所述组合物中不存在或基本不存在白蛋白多聚体；例如，所述组合物中单体形式的白蛋白占白蛋白总量的至少约95w/w％，例如至少约96％，至少约98％，至少约99％，至少约99.2％，至少约99.4％，或者至少约99.5％。
33.实施方案1至32中任一项的组合物，其特征在于，所述白蛋白选自：人血清白蛋白(HSA)、重组人白蛋白(rHA)、牛血清白蛋白和猪血清白蛋白；例如，所述白蛋白包含SEQ ID NO:1所示的氨基酸序列；
优选地，所述白蛋白选自人血清白蛋白(HSA)和重组人白蛋白(rHA)。
实施方案34.制备实施方案1至33中任一项所述的组合物的方法，其特征在于，所述方法包括以下步骤：
(1)将所述SN-38、所述脂质和Span 20溶解于有机溶剂中形成有机相；以及制备白蛋白的水溶液以作为水相；
(2)将所述有机相和所述水相混合以形成乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
(3)去除所述乳液中的有机溶剂以得到包含所述纳米粒子的产物。
实施方案35.实施方案34所述的方法，其特征在于，所述方法包括以下步骤：
(1)用包含选自DMSO和C 1-3醇(包括甲醇、乙醇和异丙醇及其任意组合，优选乙醇(EtOH))中的第一有机溶剂和选自CHCl 3及CH 2Cl 2和CHCl 3的混合物的第二有机溶剂的混合有机溶剂溶解所述SN-38、所述脂质和Span 20以形成有机相，其中在所述混合有机溶剂中，所述第二有机溶剂与所述DMSO或C 1-3醇的体积比为约1:20(v/v)至约20:1(v/v)，例如约1:5至约5:1(v/v)，约1:2至约4:1(v/v)，约1:1至约4:1(v/v)，约1.5:1(v/v)至约3:1(v/v)或约2:1(v/v)至约7:3(v/v)；以及制备白蛋白的水溶液以作为水相；
(2)将所述有机相与所述水相混合以制备乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
(3)除去所述有机溶剂；
(4)任选地，将步骤(3)中得到的产物灭菌，优选通过约0.2μm的滤膜过滤来除菌；
任选地，其中所述第二有机溶剂是CHCl 3；或CH 2Cl 2和CHCl 3的混合物，其中任选地，所述混合物中CH 2Cl 2和CHCl 3的体积比为约2:5-1:1，优选为约2:5。
实施方案36.实施方案34或35的方法，其特征在于，在步骤(2)中，所述有机相:所述水相为约1:2(v/v)至约1:50(v/v)，例如约1:5(v/v)至约1:20(v/v)，约1:7(v/v)至约1:15(v/v)，约1:10(v/v)至约1:12(v/v)；例如约1:5(v/v)至约1:12(v/v)，约1:5(v/v)至约1:12(v/v)，约1:6(v/v)、约1:7(v/v)或约1:10(v/v)。
实施方案37.实施方案34至36中任一项的方法，其特征在于，步骤(2)包括以下步骤：
(2-1)在剪切下，使所述有机相分散在所述水相中以得到粗乳液；和
(2-2)将所述粗乳液在高压下均质化以得到包含所述纳米粒子的细乳液。
实施方案38.实施方案34至37中任一项的方法，其特征在于，
所述水相不包含额外的稳定剂；或者
所述水相已经包含额外的稳定剂；或者
所述方法还包括在步骤(2)之中添加额外的稳定剂；并且
其中所述额外的稳定剂的量使得在步骤(3)或(4)得到的产物中，所述额外的稳定剂的含量为至少约2w/v％，例如至少约3w/v％，至少约5w/v％，约5w/v％至约30w/v％，约10w/v％至约25w/v％或约15w/v％至约20w/v％。
实施方案39.实施方案38的方法，其中所述额外的稳定剂选自：白蛋白类(例如人血清白蛋白、重组人白蛋白、牛血清白蛋白、脱脂奶粉)、单糖、二糖、多糖、甘露醇及其任意组合；优选地选自：甘露醇、乳糖、麦芽糖、海藻糖、右旋糖苷、葡萄糖和蔗糖及其任意组合；优选为蔗糖。
实施方案40.实施方案34至39中任一项的方法，其特征在于，在步骤(2)中，在将所述有机相与所述水相混合之前，向所述水相中添加步骤(1)中所述的混合有机溶剂。
实施方案41.实施方案40的方法，其特征在于，所添加的混合有机溶剂的体积等于或小于所述 有机相的体积；例如，所添加的混合有机溶剂与所述有机相的体积比为约1:1(v/v)至约1:5(v/v)，例如约1:2(v/v)至约1:4(v/v)或约1:3(v/v)。
实施方案42.实施方案34至41中任一项的方法，其特征在于，在步骤(1)所述的有机相中，
SN-38的浓度为约5-17mg/mL，例如约5.25-12mg/mL，约7-12mg/mL，或约10mg/mL；和/或
所述脂质的浓度为约3-50mg/mL，例如约5-45mg/mL或约7.5-30mg/mL，约10-25mg/mL或约15-20mg/mL；和/或
在所述水相中，白蛋白的浓度为约5-15mg/mL，例如约6-12mg/mL，或约6-10mg/mL。
实施方案43.实施方案34至41中任一项的方法，其特征在于，在步骤(1)所述的有机相中，
SN-38的浓度为约4-10mg/mL，例如约6-8mg/mL；和/或
所述脂质的浓度为约10-20mg/mL，例如约15mg/mL；和/或
所述Span 20的浓度为约0.3-6mg/mL，例如约0.3-2mg/mL或约0.6-1mg/mL；和/或
在所述水相中，白蛋白的浓度为约8-30mg/mL，例如约12-20mg/mL或约16-18mg/mL。
实施方案44.实施方案34至43中任一项的方法，所述方法包括步骤(4)将步骤(3)中得到的产物灭菌，优选通过约0.2μm的滤膜过滤来除菌。
实施方案45.实施方案34至44中任一项的方法，其特征在于，所述方法还包括以下步骤：
(5)将步骤(3)或(4)得到的产物干燥，优选喷雾干燥或冷冻干燥，以提供固体形式，优选为粉末，更优选为冻干粉的组合物；
优选地，SN-38以无定形形式存在于所述组合物中。
实施方案46.实施方案45所述的方法，其特征在于，步骤(5)还包括：在干燥前，向步骤(3)或(4)得到的产物中添加如实施方案39所述的额外的稳定剂，其中所述额外的稳定剂的量使得当步骤(5)所得的固体形式通过复溶形成水性组合物(包括溶液和乳液)时，所述额外的稳定剂的含量为至少约2w/v％，例如至少约3w/v％，至少约5w/v％，约5w/v％至约30w/v％，约10w/v％至约25w/v％或约15w/v％至约20w/v％。
实施方案47.药物组合物，其包含实施方案1至33中任一项的组合物以及任选存在的药学上可接受的载体。
实施方案48.药物组合物，其包含经干燥的实施方案1至33中任一项的组合物以及任选存在的药学上可接受的载体。
实施方案49.实施方案48的药物组合物，其中所述干燥是冷冻干燥或喷雾干燥，优选冷冻干燥。
实施方案50.实施方案48或49的药物组合物，其为固体形式，优选为冻干粉；优选用于胃肠外给药，更优选静脉内注射给药。
实施方案51.实施方案1至33中任一项的组合物或者实施方案47至50中任一项的药物组合物在制备用于治疗个体的对SN-38敏感的肿瘤的药物中的用途；优选地，所述肿瘤选自结直肠癌、小细胞肺癌、淋巴癌、乳腺癌(优选三阴性乳腺癌)、食管癌、胃癌、肝癌、肾癌、胰腺癌、子宫癌和卵巢癌。
实施方案52.实施方案1至33中任一项的组合物或者实施方案47至50中任一项的药物组合物，其用于治疗个体的对SN-38敏感的肿瘤；优选地，所述肿瘤选自结直肠癌、小细胞肺癌、淋巴癌、乳腺癌(优选三阴性乳腺癌)、食管癌、胃癌、肝癌、肾癌、胰腺癌、子宫癌和卵巢癌。
实施方案53.治疗个体的对SN-38敏感的肿瘤的方法，其包括向所述个体施用治疗有效量的实施方案1至33中任一项的组合物或者实施方案47至50中任一项的药物组合物；优选地，所述肿瘤选自结直肠癌、小细胞肺癌、淋巴癌、乳腺癌(优选三阴性乳腺癌)、食管癌、胃癌、肝癌、肾癌、胰腺癌、子宫癌和卵巢癌。
实施方案54.药盒，其包含实施方案1至33中任一项的组合物或者实施方案47至50中任一项的药物组合物。
实施方案55.制备具有改善的性质的组合物的方法，其中所述组合物包含SN-38、脂质和白蛋白，并且所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质以形成纳米粒子，
所述方法的特征在于在制备所述组合物的过程中加入Span 20；
其中任选地，所述组合物不包含额外的稳定剂；和/或
其中任选地，所述改善的性质包括：提高的稳定性，例如当所述组合物为液体形式时，所述提高的稳定性包括：在所述组合物的制备、储存和/或使用中，减少的白蛋白多聚体的产生或含量(例如所述组合物中不存在或基本不存在白蛋白多聚体，或者白蛋白多聚体占白蛋白总量至多5w/w％，例如至多约4％、至多约2％、至多约1.5％、至多约1.2％、至多约1.1％、至多约1％、或至多约0.8％)，和/或降低的纳米粒子粒径；和/或
其中任选地，所述组合物如实施方案1至33中任一项所定义。
实施方案56.实施方案55的方法，其特征在于，所述方法包括以下步骤：
(1)将所述SN-38、所述脂质和Span 20溶解于有机溶剂中形成有机相；以及制备白蛋白的水溶液以作为水相；
(2)将所述有机相和所述水相混合以形成乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
(3)去除所述乳液中的有机溶剂以得到包含所述纳米粒子的产物。
实施方案57.实施方案56所述的方法，其特征在于，所述方法包括以下步骤：
(1)用包含选自DMSO和C 1-3醇(包括甲醇、乙醇和异丙醇及其任意组合，优选乙醇(EtOH))中的第一有机溶剂和选自CHCl 3及CH 2Cl 2和CHCl 3的混合物的第二有机溶剂的混合有机溶剂溶解所述SN-38、所述脂质和Span 20以形成有机相，其中在所述混合有机溶剂中，所述第二有机溶剂与所述DMSO或C 1-3醇的体积比为约1:20(v/v)至约20:1(v/v)，例如约1:5至约5:1(v/v)，约1:2至约4:1(v/v)，约1:1至约4:1(v/v)，约1.5:1(v/v)至约3:1(v/v)或约2:1(v/v)至约7:3(v/v)；以及制备白蛋白的水溶液以作为水相；
(2)将所述有机相与所述水相混合以制备乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
(3)除去所述有机溶剂；
(4)任选地，将步骤(3)中得到的产物灭菌，优选通过约0.2μm的滤膜过滤来除菌；
任选地，其中所述第二有机溶剂是CHCl 3；或CH 2Cl 2和CHCl 3的混合物，其中任选地，所述混合物中CH 2Cl 2和CHCl 3的体积比为约2:5-1:1，优选为约2:5。
有益效果
本发明人发现，本发明能够减少制备过程中高压均质的次数，有效减小组合物中纳米颗粒的粒径，提高过滤通量，稳定崩解后纳米粒子的粒径，减少原料损耗和降低成本，尤其是在放大工艺，例如中试规模的制备中。另外，通过进一步控制组合物中白蛋白的含量，本发明能够控制纳米颗粒的粒径。因此，本发明组合物的纳米颗粒的粒径更接近于适合成药的尺寸。另外，在储存过程中，由于Span 20的存在，白蛋白多聚体的形成被阻止，因此本发明的组合物的免疫原性低，安全性高，同时具有优异的储存稳定性。
实施例
在以下的实施例中将进一步举例说明本发明。这些实施例仅用于说明本发明，但不以任何方式限制本发明。
在实施例中使用的各缩写具有以下含义。
缩写 中文名称 缩写 中文名称
rHA 重组人血清白蛋白 HSA 人血清白蛋白
HPLC 高效液相色谱法 EtOH 乙醇
SEC-HPLC 分子排阻色谱法 CHCl 3 氯仿
PDI 聚合物分散性指数 Chol 胆固醇
Chol-PA 胆固醇棕榈酸酯 SA-Gly 单硬脂酸甘油酯
测定实施例中制备的制品的各种参数的方法
1.粒径及粒径分布测定
使用Malvern Nano ZSE粒度电位仪测定样品中纳米粒子的粒径及粒径分布。仪器激光束波长为633nm，入射光与散射光夹角为173°。设定参数：样品材料为蛋白，分散剂为水，测定温度为25℃，自动扫描检测。每个样品平行测定三次，结果取平均值。
2.Zeta电位的测定
使用Malvern Nano ZSE粒度电位仪测定样品中纳米粒子的Zeta电位。设定参数：样品材料为蛋白，分散剂为水。选用DTS1070样品池，测定温度为25℃，自动扫描检测。测试样品均用去离子水稀释10倍体积后进行检测。每个样品平行测定三次，结果取平均值。
3.体系中SN-38的含量测定
取10mg制备得到的样品，用去离子水稀释5倍后，再用异丙醇稀释10倍，超声萃取15min，然后于10000rpm/min下离心12min。取上清液，用HPLC测定体系中的SN-38，将峰面积带入到标准曲线中，计算出体系中SN-38的含量。
HPLC色谱条件如表1所示。
表1：HPLC测定SN-38含量的色谱条件
色谱柱型号 Agilent Poroshell 120 EC-C18 2.7μm 3.0×150mm
检测器波长 265nm、381nm
柱温 50℃
样品盘温度 室温
流动相 A：25mM NaH 2PO4,pH 3.1；B：乙腈(ACN)
流速 0.6mL/min
洗脱方式 梯度洗脱
进样体积 5μL
运行时间 25min
梯度洗脱条件如表2所示。
表2：在HPLC测定SN-38含量中使用的梯度洗脱条件
时间(min) A％ B％
0 80 20
5 80 20
15 15 85
20 15 85
22 80 20
25 80 20
获得的SN-38含量测定的典型图谱如图1所示(以实施例1为例)。
4.体系中白蛋白的含量测定
使用BCA法测定体系中白蛋白含量。以BSA作为标椎品，将样品稀释10倍。取25μL稀释后的样品，加200μL检测液，在振荡器上震荡混匀，再将微孔板密封，在37℃孵育120min。用酶标仪于562nm处测定吸光度，根据标准曲线计算样品中白蛋白的浓度。
5.体系中胆固醇的含量测定
用HPLC测定样品中的胆固醇含量。样品的稀释方法同上述SN-38含量测定。色谱条件如表3所示。
表3：HPLC测定胆固醇含量的色谱条件
色谱柱型号 Agilent Eclipse XDB-C18 5μm 4.6×250mM
色谱柱编号 ZJ-RP-001
检测器波长 205nm
柱温 30℃
样品盘温度 15℃
流速 1mL/min
流动相 甲醇
洗脱方式 等度洗脱
进样体积 5μL
运行时间 20min
获得的胆固醇含量测定的典型图谱如图2所示(以实施例1为例)。
6.载药量(LD)及包封率(EE)的计算
Figure PCTCN2022124787-appb-000001
Figure PCTCN2022124787-appb-000002
其中，体系中总SN-38含量由上述“3.体系中SN-38的含量测定”中描述的方法测定。
体系中游离的SN-38的含量由固相萃取法萃取后进行HPLC测定。固相萃取的具体方法如下：
1)SPE小柱(Select Core TM HLB，1mL)活化：先用3mL甲醇，然后用3mL超纯水活化。
2)将200μL待分离样品上样，重力作用过柱。
3)用2mL水洗脱，重力洗脱即得。
4)用2mL甲醇洗脱，重力洗脱。收集甲醇相，得到游离SN-38的检测溶液。
7.SN-38的两种结构的检测
两种不同活性的SN-38的结构如下示意图所示
Figure PCTCN2022124787-appb-000003
样品的处理方法同上述SN-38的含量测定，HPLC方法色谱条件如表4。
表4：用于不同结构的SN-38的含量测定的HPLC色谱条件
色谱柱型号 Agilent Poroshell 120 EC-C18 4μm 3.0×150mM
色谱柱编号 QCCA-RP-007
检测器波长 265nm，Ref 460nm,40nm
柱温 40℃
样品盘温度 室温
流速 1mL/min
流动相 A:25mM KH 2PO 4；B:ACN
洗脱方式 梯度洗脱
进样体积 5μL
运行时间 20min
梯度洗脱条件如表5所示。
表5：在HPLC测定不同结构的SN-38的含量中使用的梯度洗脱条件
时间(min) A％ B％
0 95 5
15 40 60
18 40 60
18.1 95 5
20 95 5
获得的不同结构的SN-38的含量测定的典型图谱如图3所示(以实施例1为例)。
根据谱图中开环结构与内酯环结构的峰面积比可以计算出样品中SN-38的开环比例。在根据本发明的实施例所制备的样品中，SN-38的开环比例均<2.0％。
8.样品中白蛋白聚集体的测定
用SEC-HPLC测定样品中白蛋白的聚集情况。直接取制备获得的样品5μl进行检测，色谱条件如表6所示。
表6：测定样品中白蛋白聚集体的SEC-HPLC色谱条件
色谱柱型号 TSKgel G4000SWxl 7.8×300mm，8μm
色谱柱编号 QCCA-RP-007
检测器波长 280nm、260nm
柱温 30℃
样品盘温度 10℃
流速 0.5mL/min
流动相 0.05M Tris-HCl，pH 7.0
洗脱方式 等度洗脱
进样体积 5μL
运行时间 30min
所获得的白蛋白聚集体测定的典型图谱如图4所示(以实施例1为例)。结果表明，样品中无白蛋白多聚体存在，仅有少量二聚体，表明此制品不会产生由于白蛋白多聚体而产生的免疫原性。
9.稳定性及崩解实验
样品的稳定性主要通过将制备的样品分别置于室温以及4℃储存，每隔一段时间观察样品中是否有明显的析出或者沉淀；同时取样品检测粒径及粒径分布，研究样品中纳米粒子的粒径变化情况。
崩解实验则是研究样品中白蛋白与SN-38结合的稳定性。用pH～7.4的1×PBS稀释样品，测定不同稀释倍数下样品的粒径及粒径分布，研究纳米粒子在稀释多少倍的情况下会发生崩解，析出SN-38原料。稀释倍数越高，则表明纳米粒子的稳定性越好。
10.XRD检测方法
用X射线衍射法(Bruker，D8ADVANCE)评价样品中活性药物成分(API)的晶型变化。分别检测白蛋白冻干粉、白蛋白-SN-38冻干粉以及SN-38的晶型。用Cu-Kα射线，以2°/min的速度扫描2°-40°的2θ范围。
11.电镜检测方法
制样条件：将各个样品用注射用水调整到API浓度为2mg/mL。温度为4度，湿度为100％，blot time(吸附时间)为9秒，blot force(吸附力度)为3。用冷冻透射电镜(Talos L120C)在120Kv下观察样品中囊泡和晶体的形态。
根据具体情况，选择上述测定方法，对以下各实施例所制备的样品进行测定。
实验材料：
除非另有说明，以下实施例中使用的SN-38来自四川协力制药股份有限公司；胆固醇来自江苏东南纳米材料有限公司；rHA来自华北制药股份有限公司；HSA来自广州双林生物制药有限公司；盐酸伊立替康注射液(CPT-11，60mg/kg)来自江苏恒瑞医药股份有限公司。
除非另有说明，在下文所述的各个动物实验研究中，rHA-SN-38或HSA-SN-38制品以及CPT-11的剂量是以活性成分计；溶媒指注射用水，用作空白对照。
实施例1：rHA-SN-38制品1的制备
1.制备工艺
1)配制体积比为2/3的EtOH/CHCl 3有机溶剂；
2)称取21mg SN-38以及30mg胆固醇，加入3mL步骤1)中的有机溶剂使之完全溶解，得到药物溶液；
3)用去离子水配制总体积约为21mL的rHA水溶液，作为水相，使得水相中rHA的总含量为200mg；
4)剪切分散：将步骤2)的药物溶液与步骤3)的水相混合后，剪切分散10-15min，得到粗乳液；
5)将粗乳液转移到高压均质机中，在1300-1500bar的压力下均质2-7min；
6)在40-45℃旋蒸4-8min；
7)用0.2μm的PES针头滤膜(Sartorius药业)过滤。在过滤前后，分别测定制品样品的粒径以及包封率、载药量等参数。
2.测定结果
实施例1制备的样品的各参数的测定结果如表7所示。
表7：实施例1制备的样品的各参数的测定结果
Figure PCTCN2022124787-appb-000004
注：粒径增加比例＝((放置后粒径-放置前粒径)×100％/放置前粒径)
实施例2：HSA-SN-38制品1冻干剂及其复溶液的制备
1.制备工艺
1)配制体积比为2/3的EtOH/CHCl 3有机溶剂；
2)称取21mg SN-38以及30mg胆固醇，加入3mL步骤1)中的有机溶剂使之完全溶解，得到药物溶液；
3)用去离子水配制总体积约为32mL的HSA水溶液，作为水相，使得水相中HSA的总含量为200mg；
4)剪切分散：将步骤2)的药物溶液与步骤3)的水相混合后，剪切分散10-15min，得到粗乳液；
5)将粗乳液转移到高压均质机中，在1300-1500bar的压力下均质2-7min；
6)在40-45℃旋蒸4-8min；
7)向步骤6)中得到产物中添加蔗糖，搅拌使之完全溶解以得到30mg/mL的蔗糖浓度；
8)用0.2μm的PES针头滤膜过滤，然后灌装至西林瓶中，经真空冷冻干燥得到HSA-SN-38纳米粒子冻干制剂。
对冻干后的制品进行XRD分析。
取两支冻干样品，其中一支用去离子水稀释使得SN-38的浓度与冻干前浓度相同(复溶液1)，另一支用去离子水稀释使得SN-38的浓度是冻干前浓度的6倍(复溶液2)。分别测定两种复溶液的 粒径和包封率等参数。
2、测定结果
(1)复溶液的参数测定结果
实施例2制备的冻干制品的复溶液的参数测定结果如表8所示。
表8：实施例2制备的冻干制品的复溶液的参数测定结果
参数 复溶液1 复溶液2
pH 6.91 6.90
粒径(d,nm) 158.3±3.4 178.2±3.2
PDI 0.210±0.018 0.241±0.018
Zeta电位(mV) -31.4±0.6 -29.3±0.9
开环比例(％) 1.86 1.92
包封率(％) 98.64 98.97
载药量(％) 5.08 5.08
备注：在此处的包封率测定中，游离SN-38含量的测定方法为：将样品高速离心(以21000rpm离心1h)，取上清液，然后添加9倍体积的乙腈进行萃取用于检测。
结果表明，复溶后，当活性成分浓度增加到高达6倍时，粒径仅稍有增加且仍然小于200nm。样品其他性质未受到大的影响。因此，本申请的冻干剂可根据需要稀释成不同的浓度进行使用。
(2)XRD分析结果
取冻干制品以及SN-38晶体和HSA进行XRD检测，结果如图5所示。
结果显示，对比HSA-SN-38和SN-38的测定结果，冻干粉中的SN-38为无定形状态。据报道，与晶体形式相比，无定形形式的药物活性成分更易溶解，生物利用度更高(例如，Wang,D.,Liang,N.,Kawashima,Y.et al.Biotin-modified bovine serum albumin nanoparticles as a potential drug delivery system for paclitaxel.J Mater Sci 54,8613–8626(2019))。相较于晶体状态的SN-38，本申请的制剂具有SN-38溶出速率快和生物利用度高的优点。
实施例3：rHA-SN-38制品2的制备
1.制备工艺
1)配制体积比为2/3的EtOH/CHCl 3有机溶剂；
2)称取42mg SN-38以及60mg胆固醇，加入3mL步骤1)中的有机溶剂使之完全溶解，得到药物溶液；
3)用去离子水配制总体积约为21mL的rHA水溶液，作为水相，使得水相中rHA的总含量为500mg；
4)剪切分散：将步骤2)的药物溶液与步骤3)的水相混合后，剪切分散10-15min，得到粗乳液；
5)将粗乳液转移到高压均质机中，在1300-1500bar的压力下均质2-7min；
6)在40-45℃相旋蒸4-8min；
7)用0.2μm的PES针头滤膜过滤，在过滤前后，分别测定制品样品的粒径以及包封率、载药量等参数。
2.测定结果
制品2的测定结果如表9所示。
表9：实施例3制备的样品的各参数的测定结果
参数 过膜前样品 过膜后样品
粒径/PDI(nm/---) 177.6nm/0.24 124.4nm/0.214
zeta电位 -26.2 -25.2
载药量(％) 4.61 3.21
包封率(％) 92.21 91.01
实施例4：rHA-SN-38制品3的制备
1.制备工艺
1)配制体积比为2/3的EtOH/CHCl 3有机溶剂；
2)称取42mg SN-38以及60mg胆固醇，加入3mL步骤1)中的有机溶剂使之完全溶解，得到药物溶液；
3)用去离子水配制总体积约为21mL的rHA水溶液，作为水相，使得水相中rHA的总含量为200mg；
4)剪切分散：将步骤2)的药物溶液与步骤3)的水相混合后，剪切分散10-15min，得到粗乳液；
5)将粗乳液转移到高压均质机中，在1300-1500bar的压力下均质2-7min；
6)在40-45℃旋蒸4-8min；
7)用0.2μm的PES针头滤膜过滤。在过滤前后，分别测定制品样品的粒径以及包封率、载药量等参数。
2.测定结果
制品3的测定结果如表10所示。
表10：实施例4制备的样品的各参数的测定结果
参数 过膜前样品 过膜后样品
粒径/PDI(nm/---) 150.7nm/0.264 117.1nm/0.204
载药量(％) 10.6 9.3
包封率(％) 80.3 79.10
实施例5：rHA-SN-38制品4的制备
1.制备工艺
1)配制体积比为2/3的EtOH/CHCl 3有机溶剂；
2)称取10mg SN-38以及60mg胆固醇，加入3mL步骤1)中的有机溶剂使之完全溶解，得到药物溶液；
3)用去离子水配制总体积约为21mL的rHA水溶液，作为水相，使得水相中rHA的总含量为150mg；
4)剪切分散：将步骤2)的药物溶液与步骤3)的水相混合后，剪切分散10-15min，得到粗乳液；
5)将粗乳液转移到高压均质机中，在1300-1500bar的压力下均质2-7min；
6)在40-45℃旋蒸4-8min；
7)用0.2μm的PES针头滤膜过滤。在过滤前后，分别测定制品样品的粒径以及包封率、载药量等参数。
2.测定结果
制品4的测定结果如表11所示。
表11：实施例5制备的样品的各参数的测定结果
参数 过膜前样品 过膜后样品
粒径/PDI(nm/---) 180.7nm/0.226 146.3nm/0.207
zeta电位 -26.1 -35.6
载药量(％) 4.1 3.13
包封率(％) 92.1 90.4
实施例6：在不存在脂质的条件下制备rHA-SN-38制品
除了在实施例1的制备工艺的步骤2)中不添加任何脂质(例如胆固醇)之外，按照实施例1的制备工艺，制备和测试了实施例6的rHA-SN-38制品。该rHA-SN-38制品的各参数的测定结果如表12所示。
表12：实施例6制备的rHA-SN-38制品的各参数的测定结果
参数 过膜前样品 过膜后样品
粒径/PDI(nm/---) 248.5nm/0.205 160.3nm/0.157
24h后粒径/PDI(nm/---) —— 271.1nm/0.228
载药量(％) —— 2.15
包封率(％) —— 82.70
结果表明，在不添加脂质的条件下制备的样品稳定性较差，在4℃冰箱中保存24h后，样品粒径增加69.12％。相比之下，实施例1制备的样品(添加了脂质)在4℃冰箱中保存24h后，样品粒径仅增加10.91％。这表明，胆固醇的添加大幅度增加样品的稳定性。另外，在不添加脂质的条件下的载药量和包封率低于添加胆固醇的条件下的结果。
实施例7：在添加其它脂质的条件下制备rHA-SN-38制品
除了将实施例1的制备工艺之步骤2)中的胆固醇替换为胆酸或软脂酸之外，按照实施例1的制备工艺，制备和测试了所获得的两种rHA-SN-38制品。这些rHA-SN-38制品的各参数的测定结果如表13所示。
表13：实施例7制备的rHA-SN-38制品的各参数的测定结果
Figure PCTCN2022124787-appb-000005
结果显示，使用胆酸或者软脂酸作为脂质，在相同的制备工艺下制备得到的样品的粒径较大，添加软脂酸的样品的粒径比添加胆酸的样品的粒径更大，且粒径分布更宽。过膜处理后，样品中的纳米粒子的含量极少，粒径分布很大，粒径和PDI无法给出客观结果。
实施例8：对不同有机溶剂体系的研究
除了将实施例1的制备工艺之步骤1)中的有机溶剂替换为表14所示的有机溶剂体系之外，按照实施例1的制备工艺，制备和测试了所获得的rHA-SN-38制品。还测定了所得制品于4℃冰箱中保存过夜后的粒径。所制得的rHA-SN-38制品的各参数的测定结果如表14所示。
表14：使用不同有机溶剂体系制备的rHA-SN-38制品的各参数的测定结果
Figure PCTCN2022124787-appb-000006
注：SN-38回收率根据下式计算：
Figure PCTCN2022124787-appb-000007
结果显示，以氯仿/乙醇体系作为有机溶剂制备得到的样品的粒径最小，载药量最高。
实施例9：对有机溶剂的不同比例的考察
将实施例1的制备工艺之步骤1)中的有机溶剂替换为表15所示的不同比例的EtOH/CHCl 3，同时将体系中rHA的用量调整为300mg，其它按照实施例1的制备工艺，制备和测试了所获得的 rHA-SN-38制品。所获得的rHA-SN-38制品的各参数的测定结果如表15所示。
表15：使用不同比例的EtOH/CHCl 3制备的rHA-SN-38制品的各参数的测定结果
Figure PCTCN2022124787-appb-000008
可见，不同比例的EtOH/CHCl 3有机溶液均可获得粒径、载药量和包封率符合要求的rHA-SN-38制品。
实施例10：对不同胆固醇浓度的考察
除了将实施例1的制备工艺之步骤2)中的胆固醇用量调整至表16所示的量之外，按照实施例1的制备工艺，制备和测试了所获得的rHA-SN-38制品。所获得的rHA-SN-38制品的各参数的测定结果如表15所示。
表16：使用不同浓度的胆固醇制备的rHA-SN-38制品的参数测定结果
Figure PCTCN2022124787-appb-000009
结果表明，增加配方中胆固醇的用量，过膜后样品的粒径更小，样品中SN-38的包封率越高，载药量也较高。
实施例11：对不同SN-38浓度的考察
除了将实施例1的制备工艺之步骤2)中的SN-38用量调整至表17所示的量之外，按照实施例1的制备工艺，制备和测试了所获得的rHA-SN-38制品。所获得的rHA-SN-38制品的各参数的测定结果如表17所示。
表17：使用不同浓度的SN-38制备的rHA-SN-38制品的参数测定结果
Figure PCTCN2022124787-appb-000010
结果表明，配方中SN-38的浓度增加对粒径影响不大，而载药量增大。
实施例12：对水相中不同rHA浓度的考察
除了将实施例1的制备工艺之步骤3)中的rHA用量或水相体积调整至表18所示的数值之外，按照实施例1的制备工艺，制备和测试了所获得的rHA-SN-38制品。所获得的rHA-SN-38制品的各参数的测定结果如表18所示。
表18：使用不同浓度的rHA制备的rHA-SN-38制品的参数测定结果
Figure PCTCN2022124787-appb-000011
结果表明，白蛋白用量的增加使得载药量略有降低，而对包封率影响不大。
实施例13：对不同的有机溶剂/水相体积比的考察
除了将实施例1的制备工艺之步骤2)中的有机溶剂体积或步骤3)中的水相体积调整至表19所示的数值之外，按照实施例1的制备工艺，制备和测试了所获得的rHA-SN-38制品。所获得的rHA-SN-38制品的各参数的测定结果如表19所示。
表19：以不同的有机溶剂/水相体积比制备的rHA-SN-38制品的各参数的测定结果
Figure PCTCN2022124787-appb-000012
结果表明，有机溶剂:水相的体积比越小，制品的载药量越高，且SN-38的包封率提高。
实施例14：包含额外的稳定剂的HSA-SN-38制品的制备
1.制备工艺
1)配制体积比为2/3的EtOH/CHCl 3有机溶剂；
2)称取21mg SN-38以及30mg胆固醇，加入3mL步骤1)中的有机溶剂使之完全溶解，得到药物溶液；
3)用去离子水配制HSA溶液，并加入作为稳定剂的蔗糖或葡萄糖，作为水相(约32mL)，其中水相中HSA的总含量为200mg，并且蔗糖或葡萄糖在终制品中的浓度如表20所示；
4)剪切分散：将步骤2)的药物溶液与步骤3)的水相混合后，剪切分散10-15min，得到粗乳液；
5)将粗乳液转移到高压均质机中，在1300-1500bar的压力下均质2-7min；
6)在40-45℃旋蒸4-8min；
7)用0.2μm的PES针头滤膜过滤。在过滤前后，分别测定制品样品的粒径以及包封率、载药量等参数；
8)将所得制品置于4℃冰箱中保存过夜后再次测定其粒径。
2.测定结果
所制备的HSA-SN-38制品的各参数的测定结果如表20所示。
表20：包含额外的稳定剂的HSA-SN-38制品的各参数的测定结果
Figure PCTCN2022124787-appb-000013
注：*由于葡萄糖为还原性糖，会对BCA检测蛋白浓度的结果造成影响，因此，添加葡萄糖的制品的载药量测定结果可能偏低。
结果表明，相较于葡萄糖，在水相中添加蔗糖制备得到制品的粒径相对更小，稳定性更好，但载药量低于添加葡萄糖制备得到的制品。对于同一稳定剂，随着其浓度增加，各参数进一步改进。
本发明人还测定了环糊精(5％、10％和15％)作为稳定剂的作用，发现所制备的制品在室温下放置24h后，制品已浑浊析出或者粒径大于250nm。因而，环糊精可能不适合作为稳定剂。
实施例15：以不同制备工艺制备HSA-SN-38制品
1.制备工艺
1)配制体积比为2/3的EtOH/CHCl 3有机溶剂；
2)称取21mg SN-38以及30mg胆固醇，加入3mL步骤1)中的有机溶剂使之完全溶解，得到药物溶液；
3)用去离子水配制HSA溶液，并加入蔗糖，作为水相(约32mL)，其中水相中HSA的总含量为200mg，并且蔗糖在终制品中的浓度为10％(w/v)；
4)剪切分散：取1mL步骤1)中的有机溶剂，加入到步骤3)中的水相中，剪切分散5min，然后加入步骤2)的药物溶液，继续剪切分散5min，得到粗乳液；
5)将粗乳液转移到高压均质机中，在1300-1500bar的压力下均质2-7min；
6)在40-45℃旋蒸4-8min；
7)用0.2μm的PES针头滤膜过滤。在过滤前后，分别测定制品样品的粒径以及包封率、载药量等参数；
8)将所得制品置于4℃冰箱中保存过夜后再次测定其粒径。
2.测定结果
所制备的HSA-SN-38制品的各参数的测定结果如表21所示。
表21：以不同制备工艺制备的HSA-SN-38制品的各参数的测定结果
Figure PCTCN2022124787-appb-000014
结果表明，通过先用少量有机溶剂处理白蛋白溶液，然后添加药物溶液制备的HSA-SN-38制品的各参数良好，尤其是载药量较高。
实施例16：使用其它脂质制备rHA-SN-38制品
除了将实施例1的制备工艺之步骤2)中的胆固醇替换为下表22中的脂质(胆固醇棕榈酸酯(Chol-PA，TCI试剂有限公司)；单硬脂酸甘油酯(SA-Gly，damas-beta试剂有限公司)；维生素D3(阿拉丁试剂有限公司))之外，按照实施例1的制备工艺制备和测试了所获得的rHA-SN-38制品。还测定了所得制品于4℃冰箱中保存过夜后的粒径。这些rHA-SN-38制品的各参数的测定结果如表22所示。
表22：使用其它脂质制备的rHA-SN-38制品的各参数的测定结果
Figure PCTCN2022124787-appb-000015
结果表明，Chol-PA和SA-Gly均能获得粒径均一、载药量高的制品。Chol-PA的用量越高，制品的粒径越小，SN-38绝对回收率以及载药量越高，但粒径分布变宽。添加SA-Gly制备得到的制品的粒径较大，SN-38的绝对回收率和载药量较高，但制品的稳定性较差。
实施例17：使用脂质组合制备rHA-SN-38制品
除了将实施例1的制备工艺之步骤2)中的胆固醇替换为下表23中的脂质组合之外，按照实施例1的制备工艺制备和测试了所获得的rHA-SN-38制品。还测定了所得制品于4℃冰箱中保存过夜后的粒径。这些rHA-SN-38制品的各参数的测定结果如表23所示。
表23：使用脂质组合制备的rHA-SN-38制品的各参数的测定结果
Figure PCTCN2022124787-appb-000016
结果表明：配方中添加Chol-PA能降低制品粒径，且添加量越大，制品粒径越小，稳定性越好；配方中添加SA-Gly制备得到的制品粒径较大，但粒径分布降低；同时添加Chol-PA及SA-Gly制备得到的制品粒径分布较低且储存稳定性较好。
实施例18：在提高的脂质比例的条件下制备rHA-SN-38制品
在本实施例中，除了胆固醇的量为60mg之外，其它原料与实施例1相同。为了获得液体和冻干粉两种制剂，将原料分成两组，分别按照实施例1的方法制备液体制剂，按照实施例2的方法(不进行步骤7))制备冻干粉制剂。获得的rHA-SN-38液体制剂和冻干粉制剂的各参数的测定结果如下表所示：
Figure PCTCN2022124787-appb-000017
实施例19：大规模制备HSA-SN-38制品
在前述小试制备的基础上研究中试放大制备工艺，在大规模条件下制备本实施例的HSA-SN-38制品。
1.制备工艺
1)配制乙醇(152mL)和CHCl 3(228mL)的混合有机溶剂；
2)称取3.36g SN-38以及4.8g胆固醇，使之完全溶解在步骤1)中的有机溶剂中，然后在50℃孵育30min以上，得到药物溶液；
3)用去离子水配制HSA(32g)的水溶液(3360mL)，作为水相；
4)剪切分散：将步骤2)的药物溶液与步骤3)的水相混合后，剪切分散10-15min，得到粗乳液；
5)将粗乳液转移到高压均质机中，在900-1200bar的压力下均质10个循环；
6)向步骤5)中得到产物中添加蔗糖溶液(100g/L，2.16L)并混合；
7)用旋转蒸发在60-70mbar下蒸发步骤6)中得到的混合物，再用切向流超滤(苏州塞恩斯仪器有限公司)浓缩样品；
8)用囊式滤器过滤(Sartorius，SARTOBRAN P)过滤。在过滤前后分别取样品，用于测定粒径以及包合效果。将滤液灌装至西林瓶中，经真空冷冻干燥得到HSA-SN-38纳米粒子冻干制剂。
2.测定结果
(1)粒径结果
步骤8)中制备的滤液样品的粒径为124.6nm(PDI＝0.187)。
(2)包合效果
表24.实施例19步骤8)中制备的滤液样品的包合效果
Figure PCTCN2022124787-appb-000018
注：SN-38相对回收率(％)表示过膜前后样品中SN-38的浓度差占过膜前样品中SN-38浓度的百分比；
HSA绝对回收率(％)表示过膜后制品中HSA的量占HSA投料量的百分比。
3.崩解实验
将制备的冻干制剂用pH～7.4的1×PBS复溶至1mg/mL的SN-38浓度，然后继续梯度稀释至100μg/mL、10μg/mL、1μg/mL、0.1μg/mL和0.01μg/mL，并测定不同浓度下的粒径以及粒径分布。
获得的结果如图6所示。结果显示，当SN-38的浓度小于1μg/mL时，HSA-SN-38纳米粒子才开始发生崩解；当SN-38的浓度达到0.01ug/mL时才缓慢发生崩解。这表明实施例19制备的HSA-SN-38冻干制品具有优异的稳定性，大大优于目前市售的其它白蛋白纳米制剂。
实施例20：大规模制备HSA-SN-38制品
在前述小试制备的基础上研究中试放大制备工艺，在大规模条件下制备本实施例的HSA-SN-38制品。
1.制备工艺
1)配制乙醇(222mL)和CHCl 3(333mL)的混合有机溶剂；
2)称取4.41g SN-38以及6.30g胆固醇，使之完全溶解在480mL步骤1)的有机溶剂中，在50℃孵育30min以上之后，与剩余的有机溶剂混合，得到药物溶液；
3)用去离子水配制HSA(32g)的水溶液(3360mL)，作为水相；
4)剪切分散：将步骤2)的药物溶液与步骤3)的水相混合后，剪切分散10-15min，得到粗乳液；
5)将粗乳液转移到高压均质机中，在900-1200bar的压力下均质10个循环；
6)向步骤5)中得到产物中添加蔗糖溶液(36g/L，6L)并混合；
7)用旋转蒸发在60-70mbar下蒸发步骤6)中得到的混合物，再用切向流超滤(苏州塞恩斯仪器有限公司)浓缩样品；
8)用囊式滤器过滤(Sartorius，SARTOBRAN P)过滤。在过滤前后分别取样品，用于测定粒径以及包合效果。将滤液灌装至西林瓶中，经真空冷冻干燥得到HSA-SN-38纳米粒子冻干制剂。
2.测定结果
(1)粒径结果
步骤8)中制备的滤液样品粒径为146.9nm(PDI＝0.208)。
(2)包合效果
表25.实施例20步骤8)中制备的滤液样品的包合效果
Figure PCTCN2022124787-appb-000019
3.崩解实验
将制备的冻干制剂用注射用水复溶至1mg/mL的SN-38浓度，然后继续梯度稀释至100μg/mL、10μg/mL、1μg/mL、0.1μg/mL和0.01μg/mL，并测定不同浓度下的粒径以及粒径分布。
获得的结果如图7所示。结果显示，与实施例19的制品类似，本实施例的HSA-SN-38制品在SN-38的浓度小于约1μg/mL时开始发生崩解，粒径分布明显变宽；在SN-38的浓度达到0.1ug/mL时快速崩解。本实施例的HSA-SN-38制品仍具有良好的稳定性，明显优于市售的其它白蛋白纳米制剂。
实施例21：实施例1的rHA-SN-38制品的毒性研究
将实施例1制备的rHA-SN-38制品在人Hep 3B细胞(ATCC HB-8064细胞)的BALB/c裸鼠皮下移植瘤模型中进行药效学实验，同时评价安全性。
经尾静脉注射，向筛选合格的荷瘤BALB/c裸鼠(每组5只)给药实施例1制备的rHA-SN-38制品(30mg/kg)，每周给药1次，一共连续给药6次。分别在给药后第13、16、20、23、27、30、34、37和41天测定各动物的体重，结果如图8所示。
结果显示，实施例1的制品(30mg/kg)对动物的体重没有影响，安全性好，同时抑制肿瘤的效力好。
实施例22：rHA-SN-38制品的抗乳腺肿瘤活性研究
利用实施例1制备的rHA-SN-38制品在人三阴性乳腺癌MDA-MB-231(ATCC:CRM-HTB-26 TM)的BALB/c裸鼠皮下移植瘤肿瘤模型中进行药效学实验，以研究本发明的组合物在癌症治疗上的应用。
该研究分两种实验方案进行。
方案一：
筛选合格的荷瘤BALB/c裸鼠21只，随机分为3组，每组7只，分别给予无菌注射用水、市售盐酸伊立替康注射液(60mg/kg)、rHA-SN-38制品(15mg/kg)。尾静脉注射给药，每周给药2次，连续给药3周。第一次给药当天为第0天，当天测定各动物的肿瘤体积。给药期间每天观察动物的一般临床，每周进行2次体重和肿瘤体积的测量。
实验结果见表26。本发明的rHA-SN-38制品在人三阴性乳腺癌MDA-MB-231的BALB/c裸鼠皮下移植瘤模型中具有极显著的抑瘤效果，且明显优于市售盐酸伊立康注射液。给药3次后，rHA-SN-38制品(15mg/kg)的抑瘤率为95％，市售盐酸伊立替康注射液(60mg/kg)的抑瘤率为71％。
表26：rHA-SN-38制品对人三阴性乳腺癌MDA-MB-231裸鼠皮下移植瘤的疗效
Figure PCTCN2022124787-appb-000020
方案二：
筛选合格的荷瘤BALB/c裸鼠42只，随机分为6组，每组7只，分别给予无菌注射用水、市售盐酸伊立替康注射液(60mg/kg、80mg/kg)、rHA(220mg/kg)和rHA-SN-38制品(1.67mg/kg、5mg/kg、15mg/kg)。尾静脉注射给药，每周给药1次，连续给药3周。第一次给药当天为第0天，当天测定各动物的肿瘤体积。给药期间每天观察动物的一般临床，每周进行2次体重和肿瘤体积的测量。
实验结果见图9。如图9所示，所采用的不同剂量的rHA-SN-38制品显示出优异的抑瘤优势，且呈现剂量依赖性。
由以上实验结果可见，本发明的rHA-SN-38制品给药剂量小，抑瘤率高，治疗效果明显优于市售盐酸伊立替康注射液，对人三阴性乳腺癌具有优异的治疗效果。
实施例23：rHA-SN-38制品的抗结肠癌活性(HT-29模型)研究
将实施例2制备的rHA-SN-38冻干粉制品复溶，在人结肠癌HT-29(ATCC:HTB 3B TM)的BALB/c裸鼠皮下移植瘤肿瘤模型中进行药效学实验，以研究本发明的组合物在癌症治疗中的应用。
该研究分两种实验方案进行。
方案一：
筛选合格的荷瘤BALB/c裸鼠21只，随机分为3组，每组7只，分别给予无菌注射用水、市售盐酸伊立替康注射液(60mg/kg)、rHA-SN-38制品(30mg/kg)。尾静脉注射给药，每周给药1次，连续给药3周。第一次给药当天为第0天，当天测定各动物的肿瘤体积。给药期间每天观察动物的一般临床，每周进行2次体重和肿瘤体积的测量。
实验结果见表27。本发明的rHA-SN-38制品(30mg/kg)在人结肠癌HT-29的BALB/c裸鼠皮下移植瘤模型中具有极显著的抑瘤效果，且明显优于市售盐酸伊立康注射液(60mg/kg)。给药4次后，rHA-SN-38制品(30mg/kg)的抑瘤率为72％，市售盐酸伊立替康注射液(60mg/kg)的抑瘤率为47％。
表27：rHA-SN-38制品对人结肠癌HT-29裸鼠皮下移植瘤的疗效
Figure PCTCN2022124787-appb-000021
方案二：
筛选合格的荷瘤BALB/c裸鼠15只，随机分为3组，每组5只，分别给予无菌注射用水、市售盐酸伊立替康注射液(60mg/kg、80mg/kg))、rHA-SN-38制品(3.3mg/kg、10mg/kg、30mg/kg)。尾静脉注射给药，每周给药1次，连续给药4周。第一次给药当天为第0天，当天测定各动物的肿瘤体积。给药期间每天观察动物的一般临床，每周进行2次体重和肿瘤体积的测量。
如图10所示，不同剂量的rHA-SN-38制品也显示了优异的抑瘤优势，且呈现剂量依赖性。
由以上实验结果可见，本发明的rHA-SN-38制品的给药剂量小，抑瘤率高，治疗效果明显优于市售盐酸伊立替康注射液，对人结肠癌具有优异的治疗效果。
实施例24：对MDA-MB-231肿瘤的体内药效学研究
目的：评价不同剂量的实施例19(溶媒复溶后给药)的HSA-SN-38制品在人三阴性乳腺癌细胞系MDA-MB-231(ATCC:CRM-HTB-26 TM)皮下异种移植BALB/c裸鼠模型中的抗肿瘤活性，并与市售注射用盐酸伊立替康(CPT-11)进行比较。
1.实验设计
表28.测试药剂在MDA-MB-231人乳腺癌模型中的抗肿瘤作用实验设计
Figure PCTCN2022124787-appb-000022
注：QW：每周给药1次；
A01S：人血清白蛋白(广州双林生物制药有限公司)，用作阴性对照。
2.实验方法
选取49只6-8周龄的Balb/c雌性裸鼠，将MDA-MB-231瘤块接种到裸鼠的右侧肩胛处。瘤块接种16天后随机分组，每组7只动物，尾静脉注射给药测试药剂。最后一次给药1周后，从所有小鼠取肿瘤，并称重。
3.实验观察和数据收集
肿瘤细胞接种后，除了观察肿瘤生长情况，还检测治疗对动物行为的影响：实验动物的活动性，摄食和饮水，体重变化(体重每周测量2次)，眼睛、被毛及其它异常情况。实验过程中观察到的临床症状均记录在原始数据中。通过以下公式计算肿瘤体积：
肿瘤体积(mm 3)＝1/2×(a×b 2)(其中a表示长径，b表示短径)。
当单只动物的体重下降超过15％时(BWL>15％)，停止向该单只动物给药，并在体重下降恢复到10％以下，恢复给药。当单只动物体重下降>20％时，按照动物福利对其实施安乐死。
4.疗效评价标准
相对肿瘤增殖率(T/C(％))表示在某一时间点，治疗组和对照组的相对肿瘤体积或瘤重的百分比值，通过如下公式计算：
T/C(％)＝T RTV/C RTV×100％
其中T RTV：治疗组平均RTV；C RTV：溶媒对照组平均RTV；
其中：
RTV＝V t/V 0，
其中V 0为分组时该动物的瘤体积，V t为治疗后该动物的瘤体积；
或者
T/C％＝T TW/C TW×100％
T TW：治疗组实验终结时平均瘤重；C TW：溶媒对照组实验终结时平均瘤重。
相对肿瘤抑制率(TGI(％))通过如下公式计算：
TGI(％)＝(1-T/C)×100％，
其中T和C分别为治疗组和对照组在某一特定时间点的相对肿瘤体积(RTV)或瘤重(TW)。
5.统计分析
本实验用one-way ANOVA进行各组间肿瘤均值的比较。方差齐性分析得出F值有显著性差异，在ANOVA分析之后用Dunnet’s T3(方差不齐)法再进行多重比较。用SPSS 17.0进行所有数据分析。p<0.05认为有显著性差异。
6.实验结果
6.1体重变化：如图11所示。
6.2肿瘤体积变化：如图12所示。
6.3抗肿瘤效力评价指标
下表29显示HSA-SN-38及CPT-11等药剂对MDA-MB-231异种移植瘤模型的抑瘤效力评价指标。
表29.MDA-MB-231人乳腺癌模型中各治疗组效力分析
Figure PCTCN2022124787-appb-000023
注：a.代表给药前动物数/实验结束时动物数
7.总结
HSA-SN-38高剂量(15mg/kg)具有明显的抑瘤效果，优于CPT-11。在实验结束时(肿瘤接种后33天)，在给药3次后，HSA-SN-38高剂量组的肿瘤体积为128mm 3，RTV为0.89，T/C值为5％，p＝0.015。HSA-SN-38中剂量(5mg/kg)对肿瘤生长也有一定的抑制作用，但与溶媒组无显著性差异(p＝0.130)。HSA-SN-38低剂量(1.67mg/kg)治疗组未观察到明显的抗肿瘤活性。瘤重的分析结果与肿瘤体积基本一致。
各组荷瘤鼠的体重变化影响如图11所示。实验过程中，CPT-11高剂量(80mg/kg)治疗组在第二次给药后，1只动物死亡，剩余6只动物体重未见明显降低；结束实验时，体重相较给药前增加1.95％。CPT-11低剂量(60mg/kg)治疗组无动物死亡，末次给药后，体重相较给药前增加2.41％。HSA-SN-38的3个剂量组均无动物死亡，也未见其它任何异常的毒性反应。末次给药后体重相较给药前不同程度增加，高、中、低3个剂量组体重增加分别为3.70％、7.63％及4.85％。溶媒及A01S两个对照组的体重增加最明显，实验结束时，相较给药前分别增加了10.84％和9.04％。
综上所述，HSA-SN-38在15mg/kg剂量下在MDA-MB-231人乳腺癌模型中具有显著抑制肿瘤生长的作用。CPT-11在80mg/kg剂量下同样具有明显的抑制肿瘤生长的作用，但较HSA-SN-38高剂量组弱；而且在第二次给药后有1只动物死亡。整体而言，HSA-SN-38的抗肿瘤活性强于CPT-11，且耐受性较好，在实验过程中无动物死亡，也未见其它毒性反应。
实施例25：对HCT116肿瘤的体内药效学研究
目的：验证实施例20制备的HSA-SN-38(溶媒复溶后给药)在人结肠癌细胞系HCT116(ATCC CCL-247)皮下异种移植BALB/c裸鼠模型中的抗肿瘤活性，并与市售注射用盐酸伊立替康(CPT-11)进行比较。
1.实验设计
表30.测试药剂在人结肠癌细胞系HCT116中的抗肿瘤作用实验设计
Figure PCTCN2022124787-appb-000024
2.实验方法：
将状态较好的HCT116瘤块切成20-30mm 3的小瘤块，接种到裸鼠的右侧肩胛处，一共接种70只裸鼠。瘤块接种15天后肿瘤平均体积达到约121mm 3时，剔除瘤体积过小或过大的小鼠，将剩余的49只小鼠根据瘤体积随机分组(7组，每组7只)并开始给药(尾静脉注射)，每周给药1次，共给药4次。根据相对肿瘤抑制率(TGI)进行疗效评价，根据动物体重变化和死亡情况进行安全性评价。最后一次给药1周后，从所有小鼠取肿瘤，并称重、拍照。
3.统计分析
本实验用one-way ANOVA进行各组间肿瘤均值的比较。方差齐性分析得出F值有显著性差异，在ANOVA分析之后用Dunnet’s T3(方差不齐)法再进行多重比较。用SPSS 17.0进行所有数据分析。p<0.05认为有显著性差异。
4.实验结果
4.1体重变化：如图13所示。
4.2肿瘤体积变化
各组的肿瘤体积变化如表31和图14所示。
表31.各组动物在不同时间点的肿瘤体积
Figure PCTCN2022124787-appb-000025
4.3抗肿瘤效力评价指标
表32显示HSA-SN-38及CPT-11等药剂对HCT116异种移植瘤模型的抑瘤效力评价指标。
表32.HCT116人结肠癌模型中各治疗组效力分析
Figure PCTCN2022124787-appb-000026
注：a.代表给药前动物数/实验结束时动物数
5.总结与讨论
本实验中观察到，HSA-SN-38高剂量(30mg/kg)具有明显的抑瘤效果，优于CPT-11。在实验结束时(肿瘤接种后43天(D43))，在给药4次后，HSA-SN-38高剂量组的肿瘤体积为113mm 3，RTV为0.92，T/C值为7％，与溶媒组比较有显著性差异(p＝0.005)。HSA-SN-38中剂量(10mg/kg)的抗肿瘤活性与CPT-11低剂量组(60mg/kg)相当，肿瘤体积为433mm 3，RTV为3.75，T/C值为30％，与溶媒对照组比较，有显著性差异(p＝0.018)。瘤重的分析结果与肿瘤体积基本一致。
实验过程中，各治疗组均无动物死亡。但包括溶媒组在内的各组动物的体重都不同程度下降。实验结束时的动物体重相较于给药前的体重的降低在溶媒组为1.83％；在A01S组为5.41％，在CPT-11高、低剂量组(80和60mg/kg)分别为16.00％和14.41％；在HSA-SN-38的3个剂量组(30、10和3.3mg/kg)分别为11.84％、13.85％和12.57％。考虑到本实验中溶媒组及A01S组动物体重也不同程度降低，且在其它模型的药效实验中未观察到HSA-SN-38治疗组动物体重的明显降低，因此认为，在本实验中各HSA-SN-38治疗组动物体重的降低可能与HCT116模型的恶病质特征有关。
综上所述，HSA-SN-38(30mg/kg和10mg/kg剂量)在人结肠癌模型中具有显著抑制肿瘤生长的作用，优于阳性对照CPT-11，且耐受性较好，在实验给药过程中无动物死亡，也未见其它毒性反应。
实施例26：对SKOV-3人卵巢癌模型的体内药效学研究
目的：验证不同剂量的实施例19(溶媒复溶后给药)的HSA-SN-38在人卵巢癌细胞系SKOV-3(ATCC HTB 77)皮下异种移植NU/NU裸鼠模型中的抗肿瘤活性，并与市售注射用盐酸伊立替康(CPT-11)进行比较。
1.实验设计
表33.测试药剂在SKOV-3人卵巢癌模型中的抗肿瘤作用实验设计
Figure PCTCN2022124787-appb-000027
Figure PCTCN2022124787-appb-000028
2.实验方法
皮下接种SKOV-3瘤块至80只BALB/c裸小鼠。接种14天后，筛选出56只荷瘤小鼠，平均分为7组，每组8只。通过尾静脉注射给药，每周给药1次，共给药4次。根据相对肿瘤抑制率(TGI)进行疗效评价，根据动物体重变化和死亡情况进行安全性评价。
3.统计分析
本实验用one-way ANOVA进行各组间肿瘤均值的比较。方差齐性分析得出F值有显著性差异，在ANOVA分析之后用Dunnet’s T3(方差不齐)法再进行多重比较。用SPSS 17.0进行所有数据分析。p<0.05认为有显著性差异。
4.实验结果
4.1体重变化：如图15所示。
4.2肿瘤体积变化
各组的肿瘤体积变化如表34和图16所示。
表34.各组动物在不同时间点的肿瘤体积
Figure PCTCN2022124787-appb-000029
4.3抗肿瘤效力评价指标
表35显示HSA-SN-38及CPT-11对SKOV-3异种移植瘤模型的抑瘤效力评价指标。
表35.SKOV-3人卵巢癌模型中各治疗组效力分析
Figure PCTCN2022124787-appb-000030
Figure PCTCN2022124787-appb-000031
HSA-SN-38高剂量(30mg/kg)具有明显的抑瘤效果，优于CPT-11低剂量(60mg/kg)。在实验结束时(肿瘤接种后42天(D42))，在给药4次后，HSA-SN-38高剂量组的肿瘤体积为445mm 3，RTV为2.65，T/C值为18％，与溶媒组比较有显著性差异(p＝0.023)。CPT-11低剂量(60mg/kg)组与溶媒组无明显差异(p＝0.691)。HSA-SN-38中、低剂量与溶媒组比较无显著性差异(p值分别为0.977和1.000)。瘤重的分析结果与肿瘤体积基本一致。
实验过程中，CPT-11高剂量(80mg/kg)治疗组有6只动物死亡，其它组无动物死亡。试验结束时，动物体重较给药前均不同程度增加；HSA-SN-38高、中和低三个剂量组动物体重分别增加5.60％、5.65％及7.03％；CPT-11低剂量组动物体重相较给药前增加6.97％，高剂量组存活的两只动物体重增加11.94％。溶媒及A01S两个对照组的动物体重增加最明显，实验结束时，相较给药前分别增加了8.86％和11.99％。
结论：
HSA-SN-38在30mg/kg剂量下，在SKOV-3人卵巢癌模型中具有显著抑制肿瘤生长的作用，明显强于CPT-11，且耐受性较好，在实验过程中无动物死亡，也未见其它毒性反应。
实施例27：对SW620人结肠癌模型的体内药效学研究
目的：验证不同剂量的实施例4制备的rHA-SN-38在人结肠癌细胞系SW620(ATCC:CCL-227)皮下异种移植BALB/c裸鼠模型中的抗肿瘤活性，并与市售注射用盐酸伊立替康(CPT-11)进行比较。
1.实验设计
表36.测试药剂在SW620人结肠癌模型中的抗肿瘤作用实验设计
Figure PCTCN2022124787-appb-000032
2.实验方法
皮下接种SW620瘤块至75只BALB/c裸小鼠。接种后第13天(D13)，筛选出49只荷瘤小鼠，平均分为7组，每组7只。通过尾静脉注射给药，每周给药1次，共给药4次。根据相对肿瘤抑制率(TGI)进行疗效评价，根据动物体重变化和死亡情况进行安全性评价。
3.统计分析
本实验用one-way ANOVA进行各组间肿瘤均值的比较。方差齐性分析得出F值有显著性差异，在ANOVA分析之后用Dunnet’s T3(方差不齐)法再进行多重比较。用SPSS 17.0进行所有数据分析。p<0.05认为有显著性差异。
4.实验结果
4.1体重变化：如图17所示。
4.2肿瘤体积变化
各组的肿瘤体积变化如表37和图18所示。
表37.各组动物在不同时间点的肿瘤体积
Figure PCTCN2022124787-appb-000033
4.3抗肿瘤效力评价指标
表38显示rHA-SN-38及CPT-11对SW620异种移植瘤模型的抑瘤效力评价指标。
表38.SW620人结肠癌模型中各治疗组效力分析
Figure PCTCN2022124787-appb-000034
注：a.代表给药前动物数/实验结束时动物数
rHA-SN-38高、中剂量(30、10mg/kg)具有明显的抑瘤效果，均优于CPT-11。在实验结束时(肿瘤接种后41天(D41))，在给药4次后，rHA-SN-38高、中剂量组的肿瘤体积分别为21mm 3和101mm 3，RTV分别为0.17和0.81，T/C值分别为1％和5％，与溶媒组比较有显著性差异(p值分别为0.040和0.047)。CPT-11高、低剂量组(80和60mg/kg)的效力相近，无明显量效关系，而且与溶媒组比较均无明显差异(p值分别为0.202和0.133)。rHA-SN-38低剂量(3.3mg/kg)组的肿瘤体积在D41时与溶媒组比较无显著性差异(p＝0.231)。瘤重的分析结果与肿瘤体积基本一致。
各组均无动物死亡，也未见其它异常反应。rHA-SN-38高、中剂量组动物在实验结束时体重相较给药前分别增加2.03％及4.01％。其它各组动物在实验结束时体重相较给药前均不同程度降低：在CPT-11高、低剂量组分别为5.81％和0.99％；在rHA-SN-38低剂量组为0.33％；以及在溶媒及A01S两个对照组分别为5.83％和4.39％。
综上所述，rHA-SN-38在30及10mg/kg剂量下在SW620人结肠癌模型中具有显著抑制肿瘤生长的作用，优于80及60mg/kg剂量的CPT-11。而且，这两个rHA-SN-38治疗组动物体重增加更明 显，而两个CPT-11治疗组动物则降低或仅轻微增加。整体而言，rHA-SN-38的抗肿瘤活性明显强于CPT-11，且耐受性较好，实验过程中无动物死亡，也未见其它毒性反应。
实施例28：对Hep3B人肝癌模型的体内药效学研究
目的：验证实施例5制备的rHA-SN-38在人肝癌细胞系Hep3B(ATCC-8064)皮下异种移植BALB/c裸鼠模型中的抗肿瘤活性，并与市售注射用盐酸伊立替康(CPT-11)进行比较。
1.实验设计
表39.测试药剂在Hep3B人肝癌模型中的抗肿瘤作用实验设计
Figure PCTCN2022124787-appb-000035
2.实验方法
皮下接种Hep3B瘤块至75只BALB/c裸小鼠。接种后第13天(D13)，筛选出49只荷瘤小鼠，平均分为7组，每组7只。通过尾静脉注射给药，每周给药1次，共给药4次。根据相对肿瘤抑制率(TGI)进行疗效评价，根据动物体重变化和死亡情况进行安全性评价。
3.统计分析
本实验用one-way ANOVA进行各组间肿瘤均值的比较。方差齐性分析得出F值有显著性差异，在ANOVA分析之后用Dunnet’s T3(方差不齐)法再进行多重比较。用SPSS 17.0进行所有数据分析。p<0.05认为有显著性差异。
4.实验结果
4.1体重变化：如图19所示。
4.2肿瘤体积变化：如图20所示。
结论
受试品rHA-SN-38在30mg/kg和10mg/kg剂量下在Hep3B人肝癌模型中具有显著抑制肿瘤生长的作用，与CPT-11在80mg/kg的剂量下的抑瘤水平类似。CPT-11在60mg/kg的剂量下，有抗肿瘤活性，与HSA-SN-38在3.33mg/mL剂量下的抑瘤效果类似。rHA-SN-38的耐受性较好，实验过程中无动物死亡，也未见其它毒性反应。
实施例29：添加不同表面活性剂对SN-38制剂的影响
制备工艺：
1)配制体积比为3/7的EtOH/CHCl 3混合溶剂；
2)称取300mg SN-38、300mg胆固醇以及下表40中所示量的表面活性剂，加入30mL步骤1)中的混合溶剂，加热使完全溶解，得到药物溶液；
3)用去离子水配制总体积约为370mL的HSA水溶液作为水相，使得水相中HSA的总含量为3g；
4)剪切分散：将步骤2)中的药物溶液与步骤3)的水相混合，剪切分散10min，得到粗乳液；
5)将粗乳液转移到高压均质机中，在1300-1500bar的压力下均质3-8遍；
6)将均质后的产物转移到烧瓶中；
7)在40-45℃下旋蒸4-6min以除去体系中的氯仿；
8)切向流超滤系统(TFF)(Millipore)换液除去体系中的EtOH；
9)用0.2μm的PES针头滤膜过滤；
10)将所得制品置于4℃冰箱中保存。
在制备过程中，在剪切分散后、高压均质后以及在过滤前后分别检测样品的粒径。
表40：具有不同表面活性剂的组合物的参数
Figure PCTCN2022124787-appb-000036
结果表明，在有机相中添加不同量的表面活性剂对制备工艺中的均质次数、粒径均有影响，其中Span 20在降低均质次数、减少纳米颗粒粒径方面均优于添加其它表面活性剂或未添加表面活性剂。
实施例30：添加Span 20对SN-38制剂过滤通量的影响
为进一步研究Span 20对制备工艺的影响，比较了在添加或不添加Span 20的条件下，在制备SN-38制剂的各个步骤的粒径、过滤通量。制备过程和Span 20的添加量同实施例29。结果示于表41。
表41：添加或不添加Span 20对粒径、过滤通量的影响
Figure PCTCN2022124787-appb-000037
结果表明，与不添加Span 20制备的制剂对比，在添加Span 20的条件下用同样的制备工艺得到的产物的粒径更小，通过PES针头滤器过滤的通量增加。
此外，还参照实施例的前序部分第9项所述的方法，进行了上述两个样品的崩解实验。具体而言，用pH～7.4的1×PBS稀释样品，测定不同稀释倍数下样品的粒径，研究纳米粒子在稀释多少倍的情况下会发生崩解，析出SN-38原料。稀释倍数越高，则表明纳米粒子的稳定性越好。
崩解实验的结果如图21和22所示。结果表明，无论是否加入Span 20，稀释到0.1ug/mL时，两个样品都仍然处于较为稳定的状态，当继续稀释至10ng/mL时，二者粒径明显增大，纳米粒子发生崩解。加入Span 20后，崩解后粒径变化幅度更小，更稳定。
另外，再参照实施例前序部分第11项所述的方法，用冷冻透射电镜观察上述两个样品的形态，电镜观察结果如图23和24所示。结果发现，两个样品中，API均存在纳米晶体和囊泡两种不同形态，且包含Span 20的样品中囊泡更多，纳米晶体更少。由于囊泡形式的SN-38进入体内后是快速起效的API，因此提示加入Span 20在更好地保持纳米颗粒粒径的同时，能帮助制剂在体内尽快发挥药效。
实施例31：添加不同量的Span 20对SN-38制剂的影响
制备工艺：
1)按下表42所示配制有机溶剂体系；
2)称取表42所示的SN-38、胆固醇以及Span 20，加入30mL步骤1)中的有机溶剂使之完全溶解，得到药物溶液；
3)用去离子水配制总体积约为370mL的HSA水溶液作为水相，使得水相中HSA的总含量为3g；
4)剪切分散：将步骤2)中的药物溶液与步骤3)的水相混合，剪切分散10min，得到粗乳液；
5)将粗乳液转移到高压均质机中，在1300-1500bar的压力下均质3-5遍；
6)将均质后的产物转移到烧瓶中；
7)在40-45℃下旋蒸4-6min以除去体系中的氯仿；
8)TFF换液除去体系中的DMSO或EtOH；
9)用0.2μm的PES针头滤膜过滤，并在过滤前后分别检测样品的粒径、API的过滤回收率和载药量等参数；
10)样品置于4℃冰箱中保存。
结果示于表42。
表42：添加不同量的Span 20对SN-38制剂的影响
Figure PCTCN2022124787-appb-000038
Figure PCTCN2022124787-appb-000039
结果表明，对于不同含量比例的SN-38和胆固醇，在制备过程中添加Span 20均能减少纳米颗粒的粒径，增加过滤效率和通量，提高过滤回收率。单因素对比表明，Span20能提高载药量和API回收率。
实施例32：添加Span 20对SN-38制剂稳定性的影响。
通过测定SN-38制剂中人血清白蛋白多聚体的含量来观察Span 20对溶液中白蛋白多聚体产生的影响。
实验方法：
1)将来自实施例31的制剂11和12灌装至西林瓶中，经真空冷冻干燥得到HSA-SN-38纳米粒子冻干制剂。存放14天后，后用去离子水稀释使得SN-38的浓度与冻干前浓度相同，在高温、高湿或强光条件下存放5天或10天后作为待测样品。
2)使用SEC-HPLC测定不同制备方法产生的SN-38制剂中人血白蛋白多聚体含量，直接取制备获得的样品5μl进行检测，色谱条件如表43所示。
表43：测定样品中白蛋白聚集体的SEC-HPLC色谱条件
Figure PCTCN2022124787-appb-000040
结果示于下表44：
表44：添加Span 20对白蛋白稳定性的影响
Figure PCTCN2022124787-appb-000041
注：表格中的百分含量表示样品中的多聚体的百分含量，数值越小，多聚体越少。
结果表明，与未添加Span 20的制剂12相比，添加了Span 20的制剂11的蛋白多聚体含量未发生明显的变化，表明Span 20能够抑制白蛋白聚集。因此，Span 20能够有效地防止溶液中白蛋白的聚集，延长药物的货架期，并且不会产生由于白蛋白多聚体而导致的免疫原性。
实施例33：白蛋白的含量对SN-38制剂的影响。
按照表45所示的配方，重复实施例29的方法制备SN-38制剂，除了制剂13中不添加Span 20以及制剂15和16中分别包含6和12g HSA。测定在制备SN-38制剂的各个步骤的粒径，还测定了API过滤回收率，以考察HSA含量对于制剂的影响。结果示于表46。
表45：制剂的组成
制剂 HSA(g) SN-38(mg) 胆固醇(mg) Span 20(mg)
13 3 300 300 0
14 3 300 300 18
15 6 300 300 18
16 12 300 300 18
表46：白蛋白的含量对SN-38制剂的影响
Figure PCTCN2022124787-appb-000042
结果表明，配方中增加白蛋白用量会增加样品的粒径，明显降低过滤通量，降低API过滤回收率。HSA:API的用量比控制在20:1以内比较合适；高于这个比例，尤其高达40:1以上时，不仅纳米颗粒粒径过大、不稳定，容易聚集产生免疫原性，而且导致过滤困难，API回收率低，无法大批量生产。
实施例34：对SW620人结肠癌模型的体内药效学研究
目的：验证实施例31中制备的具有不同量的Span 20和HSA的SN-38制剂在人结肠癌细胞系SW620(ATCC:CCL-227)皮下异种移植BALB/c裸鼠模型中的抗肿瘤活性及安全性，并与市售注射用盐酸伊立替康(CPT-11)进行比较。
表47：实验设计
Figure PCTCN2022124787-appb-000043
注：QW x 4表示每周给药一次，共计4次。
细胞培养
在37℃、含5％CO 2空气的培养箱中，在添加10％热灭活胎牛血清和琼脂的1640培养基中进行SW620细胞体外单层培养。一周两次用0.25％胰酶进行消化处理以传代。当细胞呈指数生长期时，收取细胞，计数和接种。
肿瘤细胞接种及瘤块传代
将5.0×10 6SW620肿瘤细胞悬浮于0.1mL PBS中，接种于5只裸鼠右侧肩胛处(P1代)。待肿瘤长至500-800mm 3时，将荷瘤小鼠用CO 2麻醉处死，取出瘤块，去除周围坏死的组织，将状态较好的瘤块切成20-30mm 3的小瘤块，接种到新的一批裸鼠(P2代)的右侧肩胛处，一共接种40只鼠。
瘤块接种及分组给药
在本试验中使用P2代肿瘤组织进行受试品的抗肿瘤活性评价。接种7天后肿瘤平均体积达到约159mm 3时，剔除瘤体积过小或过大的小鼠，将剩余的36只小鼠根据瘤体积随机分组并开始给药。
结果示于图25-27和表48。
表48
Figure PCTCN2022124787-appb-000044
注：T/C(％)是指治疗组(T)的TV或TW相对于空白对照组(C)的百分比，其值越小，说明抑瘤效果越好。
结果表明，具有不同量的Span 20的制剂与没有Span 20的制剂均具有优于CPT-11的抗肿瘤活性。
实施例35：将EtOH/CHCl 3混合溶剂替换成EtOH/CH 2Cl 2/CHCl 3混合溶剂对制剂的影响
1.制备工艺：
1)按照下表49所示的体积比配制EtOH/CH 2Cl 2/CHCl 3混合溶剂；
2)称取SN-38 300mg，胆固醇300mg以及Span 20 18mg，加入30mL步骤1)中的混合溶剂，在加热下使其完全溶解，得到药物溶液；
3)用去离子水配制总体积约为370mL的HSA水溶液作为水相，使得水相中HSA的总含量为3g；
剪切分散：将步骤2)的药物溶液与步骤3)的水相混合，剪切分散10-15min，得到粗乳液；
4)将粗乳液转移到高压均质机中，在1300-1500bar的压力下均质5遍，将均质后的样品转移到烧瓶中；
5)在40℃-45℃下旋蒸4-6min；
6)TFF换液除去体系中剩余的EtOH；
7)用0.2μm的PES针头滤膜过滤，并在过滤前后分别检测样品的粒径、API的过滤回收率和载药量等参数；
8)样品置于4℃冰箱中保存。
表49：不同有机溶剂体系对包载结果的影响
Figure PCTCN2022124787-appb-000045
2.结果：
将有机溶剂体系调整成EtOH/CH 2Cl 2/CHCl 3＝6/7/7后，CHCl 3残留量明显降低，同时，粗略定量CH 2Cl 2的残留水平也较低(～2ug/mg API)。按照过滤后液体中的CHCl 3残留量以及CHCl 3的单日最大曝露量(600ug)计算，临床最大使用剂量>100mg/m 2，这大大降低了CHCl 3残留导致的剂量限制。但是，由于SN-38在CH 2Cl 2中溶解度更低，有机相分散到水相中时析出晶体的速度更快，因此使用EtOH/CH 2Cl 2/CHCl 3混合溶剂制备得到的粗产品的粒径更大，API的过滤回收率相对较低，载药量相对较低。
当调整CH 2Cl 2的用量至EtOH/CH 2Cl 2/CHCl 3＝6/4/10后，粗产品的粒径降低，API的过滤回收率等参数与使用EtOH/CHCl 3混合溶剂制备得到的产品一致，但CHCl 3残留水平仍明显降低。
实施例36：大规模制备添加SP20的HSA-SN-38制剂
制备工艺：
1)按下表50所示配制混合有机溶剂300mL；
2)称取SN-38 3g，胆固醇3g以及Span 20 0.18g，加入30mL步骤1中的混合有机溶剂，加热使其完全溶解，得到药物溶液；
3)取150mL 20％的HSA溶液，加入3550mL去离子水稀释，配置成水相；
4)剪切分散和均质：用连续式生产方式将在线式剪切机(Fluke FDHS3/60)与高压均质机(ATS，AH12-150)串联，设置在线式剪切机转速为8000-10000rpm。将步骤3)的水相与步骤2)的有机相以一定比例泵入在线式剪切机，随后药液进入高压均质机，在1300-1500bar的压力下均质5遍；
5)蒸发：通过在降膜蒸发器(蒸发列管温度为40℃-45℃)中蒸发，除去体系中的氯仿以及二氯甲烷；
6)TFF换液除去体系中剩余的EtOH；
7)以80mg/ml的浓度添加蔗糖；
8)用0.2μm的囊式滤器过滤，并在过滤前后分别检测样品粒径、API的过滤回收率和载药量等参数；
9)将样品分装，每支西林瓶装15mL，然后冻干。检测经冻干的样品中的溶剂残留。
表50：10倍放大规模的制备工艺对包载结果的影响
Figure PCTCN2022124787-appb-000046
结果显示，使用EtOH/CH 2Cl 2/CHCl 3＝6/4/10作为混合有机溶剂制备得到的产品的氯仿残留水 平明显降低，且放大生产制备得到的产品的性质与小试批次一致。
实施例37：添加Span 20对HSA-SN-38制剂在不同时间的稳定性的影响
通过测定SN-38制剂中人血清白蛋白多聚体的含量来研究Span 20对溶液中白蛋白稳定性(多聚体、粒径等)产生的影响。
实验方法：
1)将来自实施例31的制剂12和实施例36的制剂18灌装至西林瓶中，经真空冷冻干燥得到HSA-SN-38制剂冻干制剂。稳定性试验开启前，所涉及到的样品存放在冷藏冰箱(2～8oC)中，分别存放0天、3个月和6个月后，后用去离子水稀释使得SN-38的浓度与冻干前浓度相同，而后检测水分、pH值、渗透压、粒径及粒径分布、多聚体和胆固醇含量，结果如表50(制剂18)和表51(制剂12)。
2)使用SEC-HPLC测定不同方法制备的SN-38制剂中人血清白蛋白多聚体含量。直接取制备获得的样品5μl进行检测，色谱条件如表43所示。
表51：制剂18的稳定性参数结果
Figure PCTCN2022124787-appb-000047
表52：制剂12的稳定性参数结果
Figure PCTCN2022124787-appb-000048
上述结果表明，随着时间推移，添加了Span 20的HSA-SN-38制剂的粒径变化、白蛋白多聚体变化更小，更趋于稳定。
实施例38：在SKOV-3人卵巢癌模型中的体内药效学研究
1、实验目的：验证实施例31中制备的制剂12和实施例36制备的制剂18在人卵巢癌细胞系SKOV-3(ATCC HTB 77)皮下异种移植BALB/c裸鼠模型中的抗肿瘤活性及安全性。
2.实验方法
皮下接种SKOV-3瘤块至40只BALB/c裸小鼠。接种后第13天(D13)，筛选出18只荷瘤小鼠，平均分为3组，每组6只。通过尾静脉注射给药，每周给药1次，共给药4次，具体给药方案见表53。根据相对肿瘤抑制率(TGI)进行疗效评价，根据动物体重变化和死亡情况进行安全性评价。
表53：测试制剂在SKOV-3人卵巢癌模型中的抗肿瘤作用
Figure PCTCN2022124787-appb-000049
注：QW x 4表示每周给药一次，共计4次。
3.统计分析
本实验用one-way ANOVA进行各组间肿瘤均值的比较。方差齐性分析得出F值有显著性差异，在ANOVA分析之后用Dunnet’s T3(方差不齐)法再进行多重比较。用SPSS 17.0进行所有数据分析。p<0.05认为有显著性差异。
4.实验结果
4.1体重变化：如图28所示。
4.2肿瘤体积变化：如图29-图30所示。
4.3表54显示HSA-SN-38制剂12和制剂18对SKOV-3异种移植瘤模型的抑瘤效力评价指标。
表54：测试制剂在SKOV-3人卵巢癌模型中的效力
Figure PCTCN2022124787-appb-000050
注：T/C(％)是指治疗组(T)的TV或TW相对于空白对照组(C)的百分比，其值越小，说明抑瘤效果越好。
结论：
制剂12和制剂18在30mg/kg剂量下在SKOV-3人卵巢癌模型中具有显著抑制肿瘤生长的作用，肿瘤最后都基本消除，且实验过程中无动物死亡，也未见其它毒性反应。结果表明，含有Span20的制剂(制剂18)与没有Span 20的制剂(制剂12)的抗肿瘤活性相当。
实施例39：在HCT116人结肠癌模型中的体内药效学研究
1.实验目的：验证实施例31中制备的制剂12和实施例36制备的制剂18在人结肠癌细胞系HCT116(ATCC CCL-247)皮下异种移植BALB/c裸鼠模型中的抗肿瘤活性及安全性。
2.实验方法
将状态较好的HCT116瘤块切成20-30mm 3的小瘤块，接种到40只裸鼠的右侧肩胛处。瘤块接种15天后肿瘤平均体积达到约121mm 3时，剔除瘤体积过小或过大的小鼠，将剩余的18只小鼠根据瘤体积随机分组(3组，每组6只)并开始给药(尾静脉注射)，每周给药1次，共给药4次。根据相对肿瘤抑制率(TGI)进行疗效评价，根据动物体重变化和死亡情况进行安全性评价。最后一次给药1周后，从所有小鼠取肿瘤，并称重、拍照。
表55：测试制剂在HCT116人结肠癌模型中的抗肿瘤作用
Figure PCTCN2022124787-appb-000051
注：QW x 4表示每周给药一次，共计4次。
3.统计分析
本实验用one-way ANOVA进行各组间肿瘤均值的比较。方差齐性分析得出F值有显著性差异，在ANOVA分析之后用Dunnet’s T3(方差不齐)法再进行多重比较。用SPSS 17.0进行所有数据分析。p<0.05认为有显著性差异。
4.实验结果
4.1体重变化：如图31所示。
4.2肿瘤体积变化：如图32-图33所示。
结论：
制剂12和制剂18在10mg/kg剂量下在HCT116人结肠癌模型中具有显著抑制肿瘤生长的作用，肿瘤基本消退，且实验过程中无动物死亡，也未见其它毒性反应。
等同以及援引加入
本文已经参考某些优选的实施方案描述了本申请的组合物、方法和用途。但是，由于基于本文所述的公开内容，其特定的变化对于本领域技术人员而言是显而易见的，所以本申请不应视为仅限于此。
除非另有定义，否则本文中所用的所有技术和科学术语的含义与本公开所属领域的普通技术人员通常所理解的含义相同。在说明书和权利要求书中，单数形式也包括复数，除非上下文另外明确指出。
此外，在某种程度上，本申请的方法不依赖于本文所阐述步骤的特定顺序，权利要求中所记载步骤的特定顺序不应被解释为对该权利要求的限制。
本文引用的所有专利、专利申请、参考文献和出版物都以其整体通过援引加入本文中。

Claims (20)

  1. 组合物，其包含SN-38、脂质、白蛋白和Span 20，其特征在于，所述组合物包含纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
    脂质:SN-38为约(0.1-10):1(w:w)，约(0.5-6):1(w:w)，约(0.5-5):1(w:w)，约(0.5-3):1(w:w)，约(1-4):1(w:w)，约(1.2-4):1(w:w)，约(1.4-2):1(w:w)，约(1.5-2.5):1(w:w)或约1:1；
    白蛋白:SN-38为约(1-100):1(w:w)，约(1-50):1(w:w)，约(3-25):1(w:w)，约(5-25):1(w:w)，约(5-20):1(w:w)，约(5-18):1(w:w)，约(6-15):1(w:w)，约(7-15):1(w:w)，约(6-12):1(w:w)，约(7-12):1(w:w)、约(9-11):1(w:w)，约10:1(w:w)；以及
    Span 20:SN-38为约(3-60):100(w:w)，约(4-60):100(w:w)，约(5-60):100(w:w)，约(6-60):100(w:w)，约(7-55):100(w:w)，约(8-50):100(w:w)，约(10-45):100(w:w)，约(12-40):100(w:w)，约(14-35):100(w:w)，约(15-30):100(w:w)，约(16-25):100(w:w)，或约(18-20):100(w:w)；并且
    其中所述脂质选自胆固醇、胆固醇衍生物、胆固醇类似物和脂肪酸酯以及它们中的两者或更多者的任意组合。
  2. 权利要求1的组合物，其特征在于，白蛋白:脂质为约(1-100):1(w:w)，例如约(2-20):1(w:w)，约(3-15):1(w:w)，约(5-10):1(w:w)，约7:1(w:w)，或约10:1(w:w)。
  3. 权利要求1或2的组合物，其特征在于，以所述组合物中的SN-38、脂质和白蛋白的总量计，
    所述SN-38的含量为约1w/w％至约25w/w％；和/或
    所述脂质的含量为约1w/w％至约35w/w％；和/或
    所述白蛋白的含量为约50w/w％至约98w/w％；
    或者，
    所述SN-38的含量为约3w/w％至约20w/w％；和/或
    所述脂质的含量为约2w/w％至约30w/w％；和/或
    所述白蛋白的含量为约55w/w％至约95w/w％；
    或者，
    所述SN-38的含量为约3w/w％至约15w/w％、约4w/w％、约5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约8w/w％、约8.5w/w％、约9w/w％、约10w/w％、约11w/w％、约12w/w％、约13w/w％或约14w/w％；和/或
    所述脂质的含量为约3w/w％至约30w/w％、约4w/w％、约5w/w％、约6w/w％、约7w/w％、约8w/w％、约8.5w/w％、约9w/w％、约9.5w/w％、约10w/w％、约10.5w/w％、约11w/w％、约11.5w/w％、约12w/w％、约12.5w/w％、约13w/w％、约13.5w/w％、约14w/w％、约15w/w％、约16w/w％、约17w/w％、约18w/w％、约19w/w％、约20w/w％、约21w/w％、约24w/w％、约26w/w％或约28w/w％；和/或
    所述白蛋白的含量为约60w/w％至约94w/w％、约64w/w％至约93w/w％、约66w/w％至约92w/w％、约68w/w％至约91w/w％、约70w/w％至约90w/w％、约75w/w％至约90w/w％、约76w/w％、约77w/w％、约78w/w％、约79w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约85w/w％、约86w/w％、约87w/w％、约88w/w％或约89w/w％；和/或
    以所述组合物中的SN-38、脂质、白蛋白和Span 20的总量计，Span 20的含量为约0.03w/w％至约12w/w％、约0.06w/w％至约10w/w％、0.08w/w％至约9w/w％、约0.1w/w％至约8w/w％、约0.2w/w％至约7w/w％、约0.4w/w％至约6w/w％、约0.6w/w％至约5w/w％、约0.8w/w％至约4w/w％、约1w/w％至约2w/w％。
  4. 权利要求1至3中任一项的组合物，其特征在于，
    脂质:SN-38为约(1-10):1(w:w)、约(1-8):1(w:w)、约(1-6):1(w:w)、约(1-5):1(w:w)、约(1-4.5):1(w:w)、约(1-4):1(w:w)、约(1.2-3.8):1(w:w)、约(1.4-3.6):1(w:w)、约(1.6-3.4):1(w:w)、约(1.8- 3.2):1(w:w)、约(2-3):1(w:w)、约(2.2-2.8):1(w:w)、约(2.4-2.6):1(w:w)、约1:1(w:w)、约2.5:1(w:w)、或约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)、约(1-1.4):1；和/或
    白蛋白:SN-38为约(5-40):1(w:w)、约(5-小于40):1(w:w)、约(5-35):1(w:w)、约(5-30):1(w:w)、约(8-25):1(w:w)、约(10-22.5):1(w:w)、约(12.5-20):1(w:w)、约(15-17.5):1(w:w)、约(16-18):1(w:w)、约10:1(w:w)、或约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、约(11.1-17.3):1(w:w)；和/或
    Span 20:SN-38为约(5-60):100(w:w)、约(6-60):100(w:w)、约(7-55):100(w:w)、约(8-50):100(w:w)、约(10-45):100(w:w)、约(12-40):100(w:w)、约(14-35):100(w:w)、约(15-30):100(w:w)、约(16-25):100(w:w)、约(18-20):100(w:w)、或约(5-10):100(w:w)、约(5-9):100(w:w)、约(6-8.6):100(w:w)、约(6-8):100(w:w)、约(6.5-7):100(w:w)；和/或
    白蛋白:脂质为约(1-40):1(w:w)、约(1-小于40):1(w:w)、约(2-35):1(w:w)、约(3-15):1(w:w)、约(5-10):1(w:w)、约(6-8):1(w:w)或约7:1(w:w)、或约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)。
  5. 权利要求1至4中任一项的组合物，其特征在于，以所述组合物中的SN-38、脂质和白蛋白的总量计，
    所述SN-38的含量为约2w/w％至约16w/w％；和/或
    所述脂质的含量为约2w/w％至约35w/w％；和/或
    所述白蛋白的含量为约75w/w％至约96w/w％；
    或者，
    所述SN-38的含量为约2.5w/w％至约15w/w％，约4w/w％至约10w/w％，约4.5w/w％至约9.5w/w％，约5w/w％至约9w/w％，或约7.5w/w％至约8w/w％；和/或
    所述组合物中的脂质的含量为约2.5w/w％至约30w/w％，约4w/w％至约12.5w/w％，约4.5w/w％至约12w/w％，约7w/w％至约10w/w％，或约7.5w/w％至约8w/w％；和/或
    所述组合物中的白蛋白的含量为约76w/w％至约95w/w％，约78w/w％至约93w/w％，约79w/w％至约91.5w/w％，约80w/w％至约90w/w％，约82w/w％至约89w/w％，约84w/w％至约88w/w％，或约84.5w/w％至约87.5w/w％；
    或者，
    所述SN-38的含量为约3w/w％至约14w/w％，约3.5w/w％至约12w/w％、约4w/w％、约4.2w/w％、约4.5w/w％、约4.6w/w％、约4.8w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约7.6w/w％、约7.8w/w％、约8w/w％、约8.5w/w％、约9w/w％、约9.2w/w％、约9.5w/w％、约9.6w/w％、约9.8w/w％、约10w/w％、约10.5w/w％、约11w/w％或约11.5w/w％；和/或
    所述脂质的含量为约4w/w％至约25w/w％、约4.5w/w％至约20w/w％、约4.3w/w％、约4.5w/w％、约4.7w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约6.7w/w％、约6.9w/w％、约7w/w％、约7.5w/w％、约7.6w/w％、约7.8w/w％、约8w/w％、约8.5w/w％、约9w/w％、约9.5w/w％、约10w/w％、约10.5w/w％、约11w/w％、约11.5w/w％、约12w/w％、约12.1w/w％、约12.3w/w％、约12.5w/w％、约13w/w％、约13.5w/w％、约14w/w％、约14.5w/w％、约15w/w％、约15.5w/w％、约16w/w％、约16.5w/w％、约17w/w％、约17.5w/w％、约18w/w％、约18.5w/w％、约19w/w％或约19.5w/w％；和/或
    所述白蛋白的含量为约78w/w％至约92w/w％、约79w/w％、约79.2w/w％、约79.4w/w％、约79.6w/w％、约79.8w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约84.3w/w％、约84.5w/w％、约84.7w/w％、约84.9w/w％、约85w/w％、约86w/w％、约87w/w％、约87.3w/w％、约87.5w/w％、约87.7w/w％、约87.9w/w％、约88w/w％、约89w/w％、约90w/w％、约91w/w％、约91.3w/w％、或约91.5w/w％；和/或
    以所述组合物中的SN-38、脂质、白蛋白和Span 20的总量计，Span 20的含量为约0.14w/w％至约5w/w％、约0.2w/w％至约2.5w/w％、约0.22w/w％至约2.0w/w％、约0.24w/w％至约2w/w％、 约0.26w/w％至约1.5w/w％、约0.28w/w％至约1.0w/w％、约0.3w/w％至约0.9w/w％、约0.32w/w％至约0.8w/w％、约0.34w/w％至约0.7w/w％、约0.36w/w％至约0.6w/w％、约0.38w/w％至约0.58w/w％、约0.4w/w％至约0.56w/w％、约0.42w/w％至约0.54w/w％、约0.44w/w％至约0.52w/w％、约0.46w/w％、约0.48w/w％、约0.5w/w％、约0.2w/w％至约0.8w/w％、约0.24w/w％至约0.7w/w％、约0.26w/w％至约0.7w/w％、约0.3w/w％至约0.65w/w％、约0.36w/w％至约0.6w/w％、约0.4w/w％至约0.58w/w％、约0.44w/w％至约0.56w/w％、约0.48w/w％至约0.54w/w％、或约0.5w/w％至约0.52w/w％。
  6. 权利要求1至5中任一项的组合物，其特征在于，
    所述胆固醇衍生物选自胆固醇与有机酸形成的酯，包括胆固醇棕榈酸酯和胆固醇辛酸酯及其组合；和/或；
    所述胆固醇类似物选自维生素D2、维生素D3及其组合；和/或
    所述脂肪酸酯选自脂肪酸甘油酯，例如长链脂肪酸甘油酯，包括硬脂酸甘油酯，例如单硬脂酸甘油酯。
  7. 权利要求1至6中任一项的组合物，其特征在于，所述脂质为胆固醇。
  8. 权利要求7的组合物，其特征在于，
    胆固醇:SN-38为约(1-6):1(w:w)、约(1.2-5):1(w:w)，约(1.4-4):1(w:w)、约3:1(w:w)、约2:1(w:w)、约1:1(w:w)、或约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)、约(1-1.4):1；和/或
    白蛋白:SN-38为约(3-25):1(w:w)、约(4-20):1(w:w)、约(5-15):1(w:w)、约(6-12):1(w:w)、约(7-12):1(w:w)、约(9-11):1(w:w)或约10:1(w:w)、或约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、约(11.1-17.3):1(w:w)；和/或
    白蛋白:胆固醇为约(2-20):1(w:w)、约(3-15):1(w:w)、约(5-10):1(w:w)或约7:1(w:w)、或约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)；和/或
    以所述组合物中的SN-38、胆固醇和白蛋白的总量计，
    所述SN-38的含量为约3w/w％至约15w/w％、约4w/w％至约15w/w％、约6w/w％至约10w/w％或约8w/w％至约12w/w％、或约4w/w％至约10w/w％、约4.5w/w％至约9.5w/w％、约5w/w％至约9w/w％、约7.5w/w％至约8w/w％；和/或
    所述胆固醇的含量为约5w/w％至约25w/w％、约6w/w％至约22w/w％、约15w/w％至约20w/w％、或约4w/w％至约12.5w/w％、约4.5w/w％至约12w/w％、约7w/w％至约10w/w％、约7.5w/w％至约8w/w％；和/或
    所述白蛋白的含量为约64w/w％至约90w/w％、约70w/w％至约90w/w％、或约78w/w％至约93w/w％、约79w/w％至约91.5w/w％、约80w/w％至约90w/w％、约82w/w％至约89w/w％、约84w/w％至约88w/w％、约84.5w/w％至约87.5w/w％；和/或
    存在于所述纳米粒子中的SN-38占所述组合物中的SN-38、胆固醇和白蛋白的总量的至少约3w/w％，例如约3w/w％至约13w/w％，约4w/w％至约12w/w％，约4w/w％、约5w/w％、约6w/w％、约7w/w％、约8w/w％、约9w/w％、约10w/w％或约11w/w％。
  9. 权利要求7的组合物，其中胆固醇:SN-38为约(1-3):1(w:w)、约(1.2-2.5):1(w:w)、约(1.4-2):1(w:w)、约(1.5-2):1(w:w)、约(1.3-1.8):1(w:w)、约(1.4-1.6):1(w:w)、约(1.5-1.7):1(w:w)、约(1.2-1.5):1(w:w)、约1:1(w:w)、约(1.4-1.5):1(w:w)、或约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)、约(1-1.4):1；
    白蛋白:SN-38为约(5-15):1(w:w)、约(5-12):1(w:w)、约(6-12):1(w:w)或约(7-12):1(w:w)、约(9-11):1(w:w)、约(10-12):1(w:w)、约11:1(w:w)、或约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、或约(11.1-17.3):1(w:w)；以及
    白蛋白:胆固醇为约(3-10):1(w:w)、约(4-8):1(w:w)、约(5-7):1(w:w)、或约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)。
  10. 权利要求7的组合物，其特征在于，
    胆固醇:SN-38为约(1-5):1(w:w)、约(1-4.5):1(w:w)、约(1-4):1(w:w)、约(1.2-3.8):1(w:w)、约(1.4-3.6):1(w:w)、约(1.6-3.4):1(w:w)、约(1.8-3.2):1(w:w)、约(2-3):1(w:w)、约(2.2-2.8):1(w:w)、约(2.4-2.6):1(w:w)、约2.5:1(w:w)、约1:1(w:w)、或约(0.8-1.8):1(w:w)、约(0.9-1.7):1(w:w)、约(1-1.4):1；和/或
    白蛋白:SN-38为约(5-25):1(w:w)、约(5-20):1(w:w)、约(6-19):1(w:w)、约(7-18):1(w:w)、约(8-16):1(w:w)、约(9-14):1(w:w)或约(10-12):1(w:w)、或约(9-21):1(w:w)、约(9-20):1(w:w)、约(11-18):1(w:w)、或约(11.1-17.3):1(w:w)；和/或
    白蛋白:胆固醇为约(5-25):1(w:w)、约(6-20):1(w:w)、约(7-18):1(w:w)、约(8-16):1(w:w)、约(9-14):1(w:w)、约(10-12):1(w:w)、或约(6-21):1(w:w)、约(6.7-13):1(w:w)、约(7-13):1(w:w)、约(11-12.7):1(w:w)；和/或
    Span 20:SN-38为约(5-40):100(w:w)、约(6-30):100(w:w)、约(7-25):100(w:w)、约(8-20):100(w:w)、约(9-15):100(w:w)、约(10-12):100(w:w)、或约(5-10):100(w:w)、约(5-9):100(w:w)、约(6-8.6):100(w:w)、约(6-8):100(w:w)、约(6.5-7):100(w:w)；和/或
    以所述组合物中的SN-38、胆固醇和白蛋白的总量计，
    所述SN-38的含量为约3w/w％至约10w/w％、约3.5w/w％至约9.5w/w％、约4w/w％、约4.5w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约8w/w％、约8.5w/w％、约9w/w％、或约4w/w％至约10w/w％、约4.5w/w％至约9.5w/w％、约5w/w％至约9w/w％、约7.5w/w％至约8w/w％；和/或
    所述胆固醇的含量为约4w/w％至约18w/w％、约4.5w/w％至约17.5w/w％、约5w/w％、约5.5w/w％、约6w/w％、约6.5w/w％、约7w/w％、约7.5w/w％、约8w/w％、8.5w/w％、约9w/w％、约9.5w/w％、约10w/w％、约10.5w/w％、约11w/w％、约11.5w/w％、约12w/w％、约12.5w/w％、约13w/w％、约13.5w/w％、约14w/w％、约14.5w/w％、约15w/w％、约15.5w/w％、约16w/w％、约16.5w/w％、约17w/w％、或约4w/w％至约12.5w/w％、约4.5w/w％至约12w/w％、约7w/w％至约10w/w％、约7.5w/w％至约8w/w％；和/或
    所述白蛋白的含量为约78w/w％至约92w/w％、约79w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约85w/w％、约86w/w％、约87w/w％、约88w/w％、约89w/w％、约90w/w％、约91w/w％、或约78w/w％至约93w/w％、约79w/w％至约91.5w/w％、约80w/w％至约90w/w％、约82w/w％至约89w/w％、约84w/w％至约88w/w％、或约84.5w/w％至约87.5w/w％。
  11. 权利要求9至10中任一项的组合物，其特征在于，
    存在于所述纳米粒子中的SN-38占所述组合物中的SN-38、胆固醇和白蛋白的总量的至少约6w/w％至约12w/w％，例如约7w/w％至约11w/w％、约8w/w％至约10w/w％、约8.3％或约9w/w％；和/或
    存在于所述纳米粒子中的SN-38占所述组合物中的SN-38的总量的约95w/w％至约99w/w％，例如约96w/w％至约99w/w％，约97w/w％至约99w/w％、约98w/w％至约99w/w％，约99w/w％或更高。
  12. 权利要求1至11中任一项的组合物，其特征在于，所述组合物为液体、半固体或固体形式；
    任选地，所述固体形式为粉末形式，优选为冻干粉；更优选地，SN-38以无定形和/或纳米晶体的形式存在于固体形式的所述组合物中。
  13. 权利要求1至12中任一项的组合物，其特征在于，所述组合物不包含额外的稳定剂；或者
    优选地，所述组合物还包含额外的稳定剂，例如冻干稳定剂，所述额外的稳定剂的量使得当所述组合物通过复溶形成水性组合物(包括溶液和乳液)时，所述额外的稳定剂的含量为至少约2w/v％，例如至少约3w/v％，例如至少约5w/v％，约5w/v％至约30w/v％，约10w/v％至约25w/v％或约15 w/v％至约20w/v％；和/或
    以所述组合物的总量计，所述额外的稳定剂的含量为约60w/w％至约98w/w％，例如约65w/w％至约97w/w％，约68w/w％至约96w/w％，约69w/w％至约95w/w％，约70w/w％至约94w/w％、约71w/w％至约93w/w％、约72w/w％至约92w/w％、约73w/w％、约74w/w％、约75w/w％、约76w/w％、约77w/w％、约78w/w％、约79w/w％、约80w/w％、约81w/w％、约82w/w％、约83w/w％、约84w/w％、约85w/w％、约86w/w％、约87w/w％、约88w/w％、约89w/w％、约90w/w％或约91w/w％；
    例如，所述额外的稳定剂的含量为约70w/w％至约96w/w％、约70w/w％至约90w/w％、约72w/w％至约89w/w％、约74w/w％至约88w/w％、约76w/w％至约87w/w％，或约80w/w％至约96w/w％、约80w/w％至约86w/w％、约81w/w％至约86w/w％、约82w/w％至约85w/w％、约83w/w％至约84w/w％、约84w/w％至约95w/w％；和/或
    所述额外的稳定剂选自：白蛋白类(例如人血清白蛋白、重组人白蛋白、牛血清白蛋白、脱脂奶粉)、单糖、二糖、多糖、甘露醇及其任意组合；优选地选自：甘露醇、乳糖、麦芽糖、海藻糖、右旋糖苷、葡萄糖和蔗糖及其任意组合；优选为蔗糖。
  14. 权利要求1至13中任一项的组合物，其特征在于，所述组合物中开环的SN-38占SN-38总量的约2w/w％或更低，例如约1.8w/w％或更低；和/或
    所述组合物中不存在或基本不存在白蛋白多聚体；例如，所述组合物中单体形式的白蛋白占白蛋白总量的至少约95w/w％，例如至少约96％，至少约98％，至少约99％，至少约99.2％，至少约99.4％，或者至少约99.5％。
  15. 权利要求1至14中任一项的组合物，其特征在于，所述白蛋白选自：人血清白蛋白(HSA)、重组人白蛋白(rHA)、牛血清白蛋白和猪血清白蛋白；例如，所述白蛋白包含SEQ ID NO:1所示的氨基酸序列；
    优选地，所述白蛋白选自人血清白蛋白(HSA)和重组人白蛋白(rHA)。
  16. 制备权利要求1至15中任一项所述的组合物的方法，其特征在于，所述方法包括以下步骤：
    (1)将所述SN-38、所述脂质和Span 20溶解于有机溶剂中形成有机相；以及制备白蛋白的水溶液以作为水相；
    (2)将所述有机相和所述水相混合以形成乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
    (3)去除所述乳液中的有机溶剂以得到包含所述纳米粒子的产物；
    优选地，所述方法包括以下步骤：
    (1)用包含选自DMSO和C 1-3醇(包括甲醇、乙醇和异丙醇及其任意组合，优选乙醇(EtOH))中的第一有机溶剂和选自CHCl 3及CH 2Cl 2和CHCl 3的混合物的第二有机溶剂的混合有机溶剂溶解所述SN-38、所述脂质和Span 20以形成有机相，其中在所述混合有机溶剂中，所述第二有机溶剂与所述DMSO或C 1-3醇的体积比为约1:20(v/v)至约20:1(v/v)，例如约1:5至约5:1(v/v)，约1:2至约4:1(v/v)，约1:1至约4:1(v/v)，约1.5:1(v/v)至约3:1(v/v)或约2:1(v/v)至约7:3(v/v)；以及制备白蛋白的水溶液以作为水相；
    (2)将所述有机相与所述水相混合以制备乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
    (3)除去所述有机溶剂；
    (4)任选地，将步骤(3)中得到的产物灭菌，优选通过约0.2μm的滤膜过滤来除菌；
    任选地，其中所述第二有机溶剂是CHCl 3；或CH 2Cl 2和CHCl 3的混合物，其中任选地，所述混合物中CH 2Cl 2和CHCl 3的体积比为约2:5-1:1，优选为约2:5。
  17. 权利要求16的方法，其特征在于，在步骤(2)中，所述有机相:所述水相为约1:2(v/v)至约1:50(v/v)，例如约1:5(v/v)至约1:20(v/v)，约1:7(v/v)至约1:15(v/v)，约1:10(v/v)至约1:12(v/v)； 例如约1:5(v/v)至约1:12(v/v)，约1:5(v/v)至约1:12(v/v)，约1:6(v/v)、约1:7(v/v)或约1:10(v/v)。
  18. 药物组合物，其包含权利要求1至15中任一项的组合物以及任选存在的药学上可接受的载体。
  19. 权利要求1至15中任一项的组合物或者权利要求17的药物组合物在制备用于治疗个体的对SN-38敏感的肿瘤的药物中的用途；优选地，所述肿瘤选自结直肠癌、小细胞肺癌、淋巴癌、乳腺癌(优选三阴性乳腺癌)、食管癌、胃癌、肝癌、肾癌、胰腺癌、子宫癌和卵巢癌。
  20. 制备具有改善的性质的组合物的方法，其中所述组合物包含SN-38、脂质和白蛋白，并且所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质以形成纳米粒子，
    所述方法的特征在于在制备所述组合物的过程中加入Span 20；
    其中任选地，所述组合物不包含额外的稳定剂；和/或
    其中任选地，所述改善的性质包括提高的稳定性，例如当所述组合物为液体形式时，所述提高的稳定性包括：在所述组合物的制备、储存和/或使用中，减少的白蛋白多聚体的产生或含量(例如所述组合物中不存在或基本不存在白蛋白多聚体，或者白蛋白多聚体占白蛋白总量至多5w/w％，例如至多约4％、至多约2％、至多约1.5％、至多约1.2％、至多约1.1％、至多约1％、或至多约0.8％)，和/或降低的纳米粒子粒径；
    和/或
    其中任选地，所述组合物如权利要求1至15中任一项所定义；
    优选地，所述方法包括以下步骤：
    (1)将所述SN-38、所述脂质和Span 20溶解于有机溶剂中形成有机相；以及制备白蛋白的水溶液以作为水相；
    (2)将所述有机相和所述水相混合以形成乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
    (3)去除所述乳液中的有机溶剂以得到包含所述纳米粒子的产物；
    更优选地，所述方法包括以下步骤：
    (1)用包含选自DMSO和C 1-3醇(包括甲醇、乙醇和异丙醇及其任意组合，优选乙醇(EtOH))中的第一有机溶剂和选自CHCl 3及CH 2Cl 2和CHCl 3的混合物的第二有机溶剂的混合有机溶剂溶解所述SN-38、所述脂质和Span 20以形成有机相，其中在所述混合有机溶剂中，所述第二有机溶剂与所述DMSO或C 1-3醇的体积比为约1:20(v/v)至约20:1(v/v)，例如约1:5至约5:1(v/v)，约1:2至约4:1(v/v)，约1:1至约4:1(v/v)，约1.5:1(v/v)至约3:1(v/v)或约2:1(v/v)至约7:3(v/v)；以及制备白蛋白的水溶液以作为水相；
    (2)将所述有机相与所述水相混合以制备乳液，所述乳液包含所述纳米粒子，其中在所述纳米粒子中，所述白蛋白包封至少部分所述SN-38和任选的至少部分所述脂质；
    (3)除去所述有机溶剂；
    (4)任选地，将步骤(3)中得到的产物灭菌，优选通过约0.2μm的滤膜过滤来除菌；
    任选地，其中所述第二有机溶剂是CHCl 3；或CH 2Cl 2和CHCl 3的混合物，其中任选地，所述混合物中CH 2Cl 2和CHCl 3的体积比为约2:5-1:1，优选为约2:5。
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