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WO2025237331A1 - Utilisation de l'annexine dans l'amélioration de l'efficacité de pénétration tumorale d'un nanotransporteur comportant une surface phospholipidique - Google Patents

Utilisation de l'annexine dans l'amélioration de l'efficacité de pénétration tumorale d'un nanotransporteur comportant une surface phospholipidique

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Publication number
WO2025237331A1
WO2025237331A1 PCT/CN2025/094837 CN2025094837W WO2025237331A1 WO 2025237331 A1 WO2025237331 A1 WO 2025237331A1 CN 2025094837 W CN2025094837 W CN 2025094837W WO 2025237331 A1 WO2025237331 A1 WO 2025237331A1
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WO
WIPO (PCT)
Prior art keywords
annexin
liposomes
phospholipid
cholesterol
nanocarriers
Prior art date
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Pending
Application number
PCT/CN2025/094837
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English (en)
Chinese (zh)
Inventor
孟幻
秦蒙蒙
赵宇亮
刘燕燕
冯振汉
王琪凯
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
National Center for Nanosccience and Technology China
Original Assignee
National Center for Nanosccience and Technology China
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Filing date
Publication date
Application filed by National Center for Nanosccience and Technology China filed Critical National Center for Nanosccience and Technology China
Publication of WO2025237331A1 publication Critical patent/WO2025237331A1/fr
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Synthetic bilayered vehicles, e.g. liposomes or liposomes with cholesterol as the only non-phosphatidyl surfactant
    • 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
    • A61K31/13—Amines
    • A61K31/135—Amines having aromatic rings, e.g. ketamine, nortriptyline
    • A61K31/136—Amines having aromatic rings, e.g. ketamine, nortriptyline having the amino group directly attached to the aromatic ring, e.g. benzeneamine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/337—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having four-membered rings, e.g. taxol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • 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
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
    • A61K31/7034—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin
    • A61K31/704—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin attached to a condensed carbocyclic ring system, e.g. sennosides, thiocolchicosides, escin, daunorubicin
    • 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
    • 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
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • This invention belongs to the field of biomedical technology and relates to the application of annexin in improving the tumor entry efficiency of nanocarriers with phospholipid surfaces.
  • Phospholipid nanocarriers represent an innovative therapeutic modality for tumors, with increasingly widespread clinical applications. These carriers include liposomes, lipid nanoparticles (LNPs), and phospholipid-coated nanocarriers (such as phospholipid-coated, lipid prodrug-coated, and biomembrane-coated organic or inorganic core designs). Using these nanocarriers can improve the bioavailability and safety of active pharmaceutical ingredients (APIs). However, the antitumor activity enhancement of these nanocarrier designs is relatively limited; in many tumor scenarios, nanomedicines exhibit antitumor activity comparable to free drug delivery systems.
  • liposomes lipid nanoparticles
  • phospholipid-coated nanocarriers such as phospholipid-coated, lipid prodrug-coated, and biomembrane-coated organic or inorganic core designs.
  • APIs active pharmaceutical ingredients
  • the antitumor activity enhancement of these nanocarrier designs is relatively limited; in many tumor scenarios, nanomedicines exhibit antitumor activity
  • the unsatisfactory antitumor efficacy of nanomedicine delivery systems stems from numerous reasons, including the lack of sufficient consideration of pathological biological barriers in current nanodesigns (such as the tumor vascular endothelial cell barrier, tumor stroma, and tumor peripheral cells).
  • pathological biological barriers such as the tumor vascular endothelial cell barrier, tumor stroma, and tumor peripheral cells.
  • the tumor accumulation capacity of intravenously injected nanomedicine delivery systems is often attributed to the intercellular spaces of tumor vascular endothelial cells, explained by the so-called EPR effect (enhanced permeability and retention effects) to explain the high intratumoral concentration of nanomedicine carriers.
  • EPR effect enhanced permeability and retention effects
  • the precise explanation of the "enhanced permeability and retention” (EPR) effect is the subject of significant debate and new insights; the EPR effect enables cancer nanocarriers to penetrate solid tumors.
  • Traditional explanations of the EPR effect primarily consider leakage from the tumor vascular system as the main factor controlling nano
  • EPR electrospray
  • the inventors call the ETR effect (enhanced transcytosis and retention effect), a non-classical EPR effect. That is, the tumor entry mechanism of nanocarriers is accomplished through multiple mechanisms, including EPR and ETR.
  • ETR is the primary tumor entry mechanism, surpassing the EPR effect.
  • transcytosis does not depend on the tumor vascular endothelial space. From a morphological perspective, the interaction between nanoparticles and blood vessels can be simply understood as "passing through” rather than "leaking out.” However, the nanobiological essence, regulatory mechanisms, and applications behind transcytosis have not yet been systematically studied.
  • this invention provides the application of annexin in improving the tumor entry efficiency of nanocarriers with phospholipid surfaces. It explores proteins that can improve the tumor entry efficiency of phospholipid nanocarriers, thereby improving the biodistribution of loaded drugs in tumors and enhancing drug bioavailability and efficacy.
  • the present invention adopts the following technical solution:
  • the present invention provides the application of annexin in improving the tumor entry efficiency of phospholipid nanocarriers.
  • annexin can bind to the surface of phospholipid nanocarriers and enhance the transcytosis of phospholipid nanocarriers, thereby improving their efficiency in reaching tumor sites (tumor enrichment capacity).
  • the annexin includes any one or a combination of at least two of annexin A2, annexin A1, annexin A10, annexin A11, annexin A13, annexin A3, annexin A4, annexin A5, annexin A6, annexin A7, annexin A8, annexin A8L1, and annexin A9.
  • the annexin is of human origin.
  • the phospholipid nanocarriers refer to nanocarriers with phospholipid surfaces commonly used in the art, including any one or a combination of at least two of liposomes, lipid nanoparticles, or phospholipid-coated nanoparticles (such as inorganic nanoparticles, organic nanoparticles, naturally derived nanoparticles, etc.).
  • the application includes modifying the annexin onto the surface of the phospholipid nanocarrier.
  • the liposomes include any one or a combination of at least two of the following: doxorubicin liposomes, irinotecan liposomes, paclitaxel liposomes, mitoxantrone liposomes, daunorubicin cytarabine liposomes, mivamotide liposomes, or amphotericin B liposomes.
  • the modification method specifically includes mixing the annexin with the phospholipid nanocarrier and incubating them.
  • the phospholipid nanocarrier comprises one, two, or more of the following lipids:
  • Phospholipids Lecithin (Egg phosphatidylcholine, EPC), soybean phosphatidylcholine (SPC), phosphatidylserine (PS), phosphatidylethanolamine (PE), dioleoyl-phosphatidylethanolamine (DOPE), phosphatidylglycerol (PG), such as DMPG, DSPG, and phosphatidylinositol.
  • PA Phosphatidic acid
  • DPPC dipalmitoylphosphatidylcholine
  • DSPC distearoylphosphatidylcholine
  • DOPC dioleoylphosphatidylcholine
  • DMPC dimyristoylphosphatidylcholine
  • HPC hydrogenated soy phosphatidylcholine
  • Cholesterol and its analogues Cholesterol, Cholesteryl hemisuccinate (CHEMS), Cholesteryl oleate, and Sitosterol;
  • PEGylated lipids DSPE-PEG2000, (1,2-Distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000]), DOPE-PEG2000, DMPE-PEG2000, DSPE-PEG-maleimide/biotin/amine, cholesterol-PEG derivatives, and PEGylated lipids with similar structures but different molecular weights;
  • Ionizable and cationic lipids Ionizable and cationic lipids: Ionizable lipids, DLin-MC3-DMA, SM-102, ALC-0315, DODMA (Dimethyldioctadecylammonium), Dlin-DMA;
  • DOTAP Dioleoyltrimethylammonium propane
  • DOTMA Dioleoyltrimethylammonium
  • DDAB Dimethyldioctadecylammonium bromide
  • CTAB Cityltrimethylammonium bromide
  • DMPG/DSPG phosphatidylglycerol, anionic
  • DOPS/POPS phosphatidylserine, anionic
  • cardiolipin sphingomyelin
  • DSPE-PEG-Maleimide DSPE-PEG-Folate
  • DSPE-PEG-RGD Glycolipids (e.g., GM1, GM3)
  • lipids include: monolaurin, lysolipids, squalene derivatives, and ceramides/sphingolipids.
  • the phospholipid nanoparticles comprise phospholipids composed of the following:
  • DSPC Distearate phosphatidylcholine
  • DOPA sodium dioleoyl phosphatidylcholine
  • DOTAP (2,3-dioleoyl-propyl)-trimethylammonium chloride
  • DSPC Distearate phosphatidylcholine
  • DOPA sodium dioleoyl phosphatidylcholine
  • Chol cholesterol
  • DSPE-PEG2000 Distearate phosphatidylcholine
  • EPC Egg yolk lecithin
  • cholesterol Chol
  • EPG Egg yolk phosphatidylglycerol
  • DMPC myristoyl phosphatidylcholine
  • DSPC Distearylphosphatidylcholine
  • DSPG distearylphosphatidylglycerol
  • cholesterol cholesterol
  • POPC Palmitoyl phosphatidylcholine
  • DOPS dioleoyl phosphatidylserine
  • xii Disorhodophospholipid (DEPC), palmitophosphatidylglycerol (DPPG), cholesterol (Chol), and tricaprylin;
  • HSPC Hydrogenated soybean phosphatidylcholine
  • cholesterol cholesterol
  • DSPG distearate phosphatidylglycerol
  • the present invention provides the use of annexins in the preparation of delivery vectors, wherein the annexins include any one or a combination of at least two of annexin A2, annexin A1, annexin A10, annexin A11, annexin A13, annexin A3, annexin A4, annexin A5, annexin A6, annexin A7, annexin A8, annexin A8L1, or annexin A9;
  • the application includes modifying the annexin onto the surface of a phospholipid nanocarrier.
  • the phospholipid nanocarrier includes any one or a combination of at least two of liposomes, lipid nanoparticles, or phospholipid-coated nanoparticles.
  • annexin which has a transcytotoxic enhancement effect, it can be further applied to prepare delivery vectors to obtain delivery vectors with enhanced tumor entry efficiency.
  • the present invention provides a delivery carrier comprising a phospholipid nanocarrier and a membrane annealing protein modified on the surface of the nanocarrier.
  • the annexin includes any one or a combination of at least two of annexin A2, annexin A1, annexin A10, annexin A11, annexin A13, annexin A3, annexin A4, annexin A5, annexin A6, annexin A7, annexin A8, annexin A8L1, or annexin A9.
  • the phospholipid nanocarrier includes any one or a combination of at least two of liposomes, lipid nanoparticles, or phospholipid-coated nanoparticles.
  • the present invention provides the use of annexin in improving the biodistribution of drugs at lesion sites or in the preparation of pharmaceutical compositions, the use of which includes modifying the annexin onto the surface of a phospholipid nanocarrier and loading drugs onto the phospholipid nanocarrier modified with annexin.
  • the annexin includes any one or a combination of at least two of the following: annexin A2, annexin A1, annexin A10, annexin A11, annexin A13, annexin A3, annexin A4, annexin A5, annexin A6, annexin A7, annexin A8, annexin A8L1, and annexin A9.
  • the phospholipid nanocarrier includes any one or a combination of at least two of liposomes, lipid nanoparticles, or phospholipid-coated nanoparticles.
  • the lesion site includes lesion sites that highly express ⁇ 5 integrin and/or ⁇ 1 integrin.
  • annexin can bind to the surface of phospholipid nanocarriers; secondly, annexin can also interact with integrins on the cell surface (such as ⁇ 5 integrin or ⁇ 1 integrin), promoting the transcytosis of phospholipid nanocarriers.
  • annexin can not only promote the accumulation of phospholipid nanoparticles at tumor sites, but also promote the accumulation of phospholipid nanoparticles in other lesion tissues with high expression of the ⁇ 5 and/or ⁇ 1 subunits.
  • tumor types include: breast cancer, pancreatic cancer, liver cancer, ovarian cancer, osteosarcoma, prostate cancer, glioma, melanoma, myxofibrosarcoma, skin cancer, lung cancer, and gastric cancer, etc.
  • non-tumor types include: psoriasis and diabetes, etc.
  • this invention conducts further experimental verification, analyzing the effect of annexin on the bioavailability and efficacy of drugs loaded on phospholipid nanocarriers, and finds that it can improve the bioavailability of drug delivery and optimize efficacy.
  • the present invention provides a nanodelivery system comprising a phospholipid nanocarrier, an annexin, and a drug, wherein the annexin is modified on the surface of the nanocarrier and the drug is loaded inside the phospholipid nanocarrier.
  • a nanodelivery system is further developed.
  • the drug is loaded onto a phospholipid nanocarrier, and then annexin is used to improve delivery efficiency, thereby improving drug bioavailability and efficacy.
  • drugs mentioned are those commonly used in the art for delivery via phospholipid nanocarriers, such as antitumor drugs doxorubicin, irinotecan, paclitaxel, mitoxantrone, daunorubicin, cytarabine, mivastatin, and antibacterial drugs amphotericin B, etc.
  • the annexin includes any one or a combination of at least two of the following: annexin A2, annexin A1, annexin A10, annexin A11, annexin A13, annexin A3, annexin A4, annexin A5, annexin A6, annexin A7, annexin A8, annexin A8L1, or annexin A9.
  • the phospholipid nanocarrier includes any one or a combination of at least two of liposomes, lipid nanoparticles, or phospholipid-coated nanoparticles.
  • annexin can spontaneously bind to the surface of phospholipid nanocarriers. Therefore, in practical applications, delivery carriers coated with annexin can be prepared in advance, or phospholipid nanocarriers and annexin can be mixed and incubated before use.
  • the present invention provides a combination pharmaceutical composition comprising a drug-loaded phospholipid nanocarrier and annexin, wherein the combination pharmaceutical composition is a single compound preparation or a combination of two separate preparations.
  • the annexin includes any one or a combination of at least two of the following: annexin A2, annexin A1, annexin A10, annexin A11, annexin A13, annexin A3, annexin A4, annexin A5, annexin A6, annexin A7, annexin A8, annexin A8L1, or annexin A9.
  • the phospholipid nanocarrier includes any one or a combination of at least two of liposomes, lipid nanoparticles, or phospholipid-coated nanoparticles.
  • the present invention provides the use of reagents for detecting ⁇ 5 integrin and/or ⁇ 1 integrin in evaluating the ability of annexin to enhance the tumor entry efficiency of phospholipid nanocarriers and/or improve drug biodistribution at lesion sites.
  • annexin enhancement effect is positively correlated with the expression levels of ⁇ 5 integrin and/or ⁇ 1 integrin.
  • integrin expression taking breast cancer as an example, log2 of FPKM ⁇ 4; FPKM (fragments per kilobase per million mapped reads) represents gene expression level
  • it can significantly enhance the tumor entry efficiency of phospholipid nanocarriers and the biodistribution and efficacy of drugs at lesion sites.
  • the enhancement effect is generally limited.
  • annexin coating to promote nanocarrier transport and enhance drug efficacy can be predicted, and beneficiaries of annexin coating technology can be screened, thereby achieving precision treatment.
  • the application includes: detecting the expression levels of ⁇ 5 integrin and/or ⁇ 1 integrin in patients, and evaluating the tumor entry efficiency of the delivery carrier described in the third aspect or evaluating the efficacy of the nanodelivery system described in the fifth aspect or the combination drug composition described in the sixth aspect based on the expression levels.
  • the present invention provides a reagent kit comprising:
  • the annexins include any one or a combination of at least two of the following: annexin A2, annexin A1, annexin A10, annexin A11, annexin A13, annexin A3, annexin A4, annexin A5, annexin A6, annexin A7, annexin A8, annexin A8L1, or annexin A9.
  • the phospholipid nanocarriers include any one or a combination of at least two of liposomes, lipid nanoparticles, or phospholipid-coated nanoparticles.
  • the drug-loaded phospholipid nanocarrier is mixed with annexin and incubated before use.
  • the present invention provides the use of annexin and phospholipid nanocarriers in the preparation of medicaments for treating diseases.
  • This aspect can also be described as the annexin and phospholipid nanocarriers of the present invention for treating diseases.
  • the lesion site of the disease is a lesion site with high expression of ⁇ 5 integrin and/or ⁇ 1 integrin;
  • the disease is selected from breast cancer, pancreatic cancer, liver cancer, ovarian cancer, osteosarcoma, prostate cancer, glioma, melanoma, myxofibrosarcoma, skin cancer, lung cancer, gastric cancer, psoriasis, diabetes, or combinations thereof.
  • the present invention provides a method for treating a disease, comprising administering the combination pharmaceutical composition of the present invention or the kit of the present invention to a subject in need.
  • the lesion site of the disease is a lesion site with high expression of ⁇ 5 integrin and/or ⁇ 1 integrin.
  • the disease is cancer
  • the drug is used to treat cancer
  • the cancer is selected from breast cancer, pancreatic cancer, liver cancer, ovarian cancer, osteosarcoma, prostate cancer, glioma, melanoma, myxofibrosarcoma, skin cancer, lung cancer, gastric cancer, or combinations thereof;
  • the drug includes doxorubicin, irinotecan, paclitaxel, mitoxantrone, or combinations thereof.
  • the disease is psoriasis, and the drug is used to treat psoriasis; in another embodiment, the disease is diabetes, and the drug is used to treat diabetes.
  • the method of treating a disease according to the present invention further includes the step of detecting the expression of ⁇ 5 integrin and/or ⁇ 1 integrin at lesion sites in a subject prior to administration.
  • the present invention has the following beneficial effects:
  • This invention creatively identifies proteins that can enhance the tumor entry efficiency of current phospholipid nanocarriers. It discovers that coating phospholipid nanocarriers with annexin promotes transcytosis, increasing tumor entry efficiency and thus improving the bioavailability of drugs loaded on the nanocarriers, thereby enhancing therapeutic efficacy. Furthermore, it was found that annexin's synergistic effect is based on a "phospholipid surface-annexin-integrin" structure and is positively correlated with the expression levels of ⁇ 5 or ⁇ 1 integrins. Therefore, annexin pre-coating can increase the biodistribution of phospholipid nanocarriers at lesion sites with high ⁇ 5 or ⁇ 1 integrin expression. Moreover, by pre-detecting and analyzing patients' ⁇ 5 or ⁇ 1 integrin expression levels, the ability of annexin coating technology to promote nanocarrier transport can be predicted, and beneficiaries of annexin coating technology can be screened.
  • Figure 1 illustrates the mechanism of nanoparticle tumor entry.
  • Figure 2 illustrates an established in situ KPC model, a rigorous preclinical model used to study drug delivery.
  • the luciferase gene was introduced into KPC cells.
  • subsequent autopsy and bioluminescence imaging revealed primary tumor growth within 1–2 weeks.
  • Trichrome staining showed the stromal-rich primary tumor adjacent to normal pancreatic tissue.
  • Figure 3 shows the morphology of probes F1, F2, and F3, with a scale bar of 25 nm.
  • Figure 4 is a schematic diagram of the structure of the gold core nanocarrier.
  • ROI region of interest
  • Figure 6 shows the biodistribution of different types of phospholipid nanoparticles in mice with KPC orthotopic pancreatic cancer; F1 emitted the highest particle signal at the tumor site. Data are expressed as mean ⁇ standard deviation. Statistical significance was assessed by unpaired t-test *p ⁇ 0.05.
  • Figure 7 is a schematic diagram of serum incubation followed by mass spectrometry analysis of various particulate variants F1 to F3.
  • Figure 8 shows the results of protein crown analysis on the surface of the nanocarriers.
  • the heatmap shows the most abundant proteins detected on the nanocarrier surface. We present 46 proteins exhibiting high abundance.
  • the right panel is a Venn diagram of the identified proteins in F1–F3.
  • the unique protein species specific to the best-performing particle (F1) are highlighted by the ⁇ region (red).
  • the ⁇ region (cyan) includes protein types detected in F1, which also appear in F2 and F3.
  • Figure 9 shows the electrophoretic analysis results of proteins coated on the nanocarrier surface. Based on the findings of Example 1, ANX A2, ANX A3, ANX A5, ANX A7, and ANX A8, as well as pure VTN, albumin, and GAPDH, were tested. Electrophoretic analysis after incubating F1 particles with these protein types confirmed the effective attachment of these proteins to F1.
  • Figure 10 is a schematic diagram of using an EC transwell device to evaluate the potential for nanoparticle transport across a HUVEC monolayer.
  • Figure 12 illustrates the use of 70 kD dextran to confirm tight junctions in the transwell system.
  • FITC-labeled dextran with a molecular weight of 70 kD was introduced into the upper layer of the transport pore.
  • the fluorescence value of FITC leaking through the cell layer to the lower layer was assessed at 1 hour. This can serve as an indicator of cell monolayer integrity.
  • Figure 14 illustrates the A2 binding stability test.
  • A2-pre-coated F1 nanoparticles were incubated in serum-supplemented saline for 0, 3, 6, 12, 24, and 48 hours. Subsequently, after washing four times with PBS, electrophoretic analysis was performed to assess the remaining protein attached to the F1 surface.
  • Figure 16 shows representative ex vivo NIR fluorescence images of tumors and organs in tumor-bearing mice 24 hours after intravenous injection of NIR-labeled LC-MSNPs with different protein coatings (particle dose 50 mg/kg). Note that this study included multiple cancer models, and the ex vivo IVIS images represent snapshots taken at a single time point. Subtle differences exist in the distribution of normal organs across groups, which may require further investigation in the future.
  • Figure 17 shows the prepared F1 version of the gold nucleus labeled with A2 protein.
  • Figure 18 shows the morphological characteristics of the nanocarriers within the tumor.
  • Tumor samples were collected 1 hour and 6 hours after intravenous injection, followed by TEM studies of the tumor tissue.
  • EC endothelial cells
  • VVO vesicular organelles.
  • the inserted image confirms the presence of gold labeling within the particles.
  • Green arrows transcellular nanoparticles within EC vesicles.
  • Figure 19 shows TEM images confirming the presence of nanoparticles within cancer cells 6 hours after intravenous injection. Data are presented as mean ⁇ standard deviation. Statistical significance was assessed by an unpaired t-test *p ⁇ 0.05.
  • Figure 20 shows: a) TEM images of additional tumors in mice 1 hour after intravenous injection of LC-MSNPs embedded with gold A2. Notable phenomena observed within 1 hour include cell membrane protrusions, filaments, and internalization of nanoparticles into endothelial cells. These additional images demonstrate that the nanoparticles are undergoing endothelial metastasis. b) TEM images of an orthotopic EMT6 tumor in the control group without any treatment. No obvious endothelial metastasis features were observed in the control group tumors.
  • Figure 21 shows the structure of the Martini force field.
  • Figure 22 shows the results of the coarse-grained (CG) simulation analysis.
  • Figure 23 shows the results of the all-atom (AA) simulation analysis.
  • Figure 24 shows the contact area between annexin A2 and lipids.
  • Figure 25 is a sandwich model diagram of "phospholipid surface-annexin-integrin".
  • Figure 26 shows that while the interaction between A2 and ⁇ 5 ⁇ 1 integrin is controlled by hydrophobic interactions (a), the interaction between A2 and the nanosurface (lipid) is mediated by electrostatic interactions (c).
  • the inserted boxes show specific amino acids and their positions in the indicated proteins, b (for A2 and ⁇ 5 ⁇ 1 integrin) and d (for A2 and lipid).
  • Figure 27 shows the effect of knocking down ⁇ 5 ⁇ 1 integrin expression on nanoparticle transport; the nanoparticle translocation experiment was performed on HUVEC cells with knocked-out integrin ⁇ 5 subunits by transwell assay.
  • Figure 28 shows the transwell monolayer integrity check in an shRNA-mediated gene knockout assay.
  • FITC-Dextran fluorescence was observed through the intercellular spaces of the transwell endothelial cells.
  • shRNA did not alter the integrity of the monolayer.
  • Figure 30 shows the physicochemical characterization and protein binding stability analysis of LC-MSNP coated with mutant A2 (A2-M).
  • A2-M mutant A2
  • a Size, PDI, and zeta potential measurements of A2-M coated LC-MSNP.
  • c Electrophoretic analysis showing the stability of A2 (left) and A2-M (middle) binding to LC-MSNP at specified time points. The gray values of the A2 and A2-M protein bands in the electrophoresis were quantified using ImageJ software.
  • Figure 31 shows the six amino acid residues on mutant A2, which were replaced with alanine.
  • the resulting A2 mutant (A2-M) was used to label pre-coated particles with NIR, and then the nanoparticle distribution was studied in an in situ EMT6 mouse model.
  • the A2 coating was used as a control.
  • Figure 32 shows the "zombie” model: In order to study the transport of nanoparticles in tumor sites, we established a “zombie” model using EMT6 orthotopic tumor mice according to the published protocol.
  • Figure 33 shows the effect of formalin fixation on tumor vessels on nanocarrier transport.
  • "zombie” mice blood vessels of EMT6 tumor-bearing mice were fixed, and blood containing A2-coated LC-MSNPs was circulated using a pump. Data are expressed as mean ⁇ standard deviation. Statistical significance was assessed by unpaired t-tests: *p ⁇ 0.05, **p ⁇ 0.01, ***p ⁇ 0.001. Using this method, we demonstrated that the abundance of nanoparticles at the "zombie" tumor site was reduced by approximately 70%.
  • Figure 34 illustrates the pharmacological inhibition of ⁇ 5 ⁇ 1 integrin, which hinders the entry of A2 pre-coated nanocarriers into tumors.
  • Figure 35 illustrates the enhanced drug delivery effects of A2 pre-coating on LC-MSNP and liposomes.
  • Body weight data are included in the efficacy data for groups g, b, c, e, and f. Note that animal studies evaluating IRIN-loaded LC MSNPs and liposomes were conducted concurrently, therefore both groups shared the same saline control. For clarity, we plotted the data according to vector type.
  • Figure 36 shows the assessment of tumor burden by IVIS biofluorescence signal in the orthotopic KPC-luc model receiving IRIN LC-MSNP (left) and IRIN liposomes (right).
  • Figure 38 shows representative H&E images of major organs obtained in the efficacy studies of the KPC and EMT6 models. Scale bar: 100 ⁇ m.
  • Figure 39 shows the results of A2-mediated increase in granule enrichment depending on the expression of ⁇ 5 ⁇ 1 integrin; the evidence of A2-mediated increase in granule pathway depends on the expression of ⁇ 5 ⁇ 1 integrin in EMT6 tumor-bearing mice.
  • Figure 40 shows the fluorescence imaging results of drug distribution in patient-derived breast cancer xenograft models with high and low integrin expression after intravenous injection of doxorubicin-loaded liposomes with and without A2 coating (CD31 indicates tumor vessels, DOX indicates doxorubicin).
  • Figure 41 shows the quantitative results of silicon accumulation in tumors after intravenous injection of silicon-based nanocarriers loaded with doxorubicin and without A2 coating in patient-derived breast cancer xenograft models with high and low integrin expression. Similar experiments were repeated using DOX LC-MSNP, followed by silicon analysis. Data are expressed as mean ⁇ standard deviation. Statistical significance was tested using unpaired t-tests: *p ⁇ 0.05, **p ⁇ 0.01, ***p ⁇ 0.001, ****p ⁇ 0.001.
  • Figure 42 illustrates the gating strategy of the flow cytometer in Figure 39.
  • Figure 43 illustrates our use of the BC PDX model to advance the study in order to validate the hypothesis that ⁇ 5 ⁇ 1-dependent efficacy improved the results of the EMT6 model.
  • the figure shows ⁇ 5 ⁇ 1 integrin expression data for both models, as well as H&E and Trichrome staining of PDX tumors.
  • b Gene expression levels of integrin ⁇ 5 and ⁇ 1 were further validated by qPCR (ACTB was used as an internal control).
  • Figure 44 shows the fluorescence intensity of DOX in randomly selected fields of view analyzed using ImageJ software.
  • the lower part of Figure 21 provides representative images and semi-quantitative results of A2-mediated liposome enrichment in PDX. Data are expressed as mean ⁇ standard deviation. Statistical significance was assessed using an unpaired t-test. *p ⁇ 0.05.
  • Figure 46 is a schematic diagram of the dosing regimen for animal experiments.
  • Figure 47 shows the binding capacity of A2 protein on LC-MSNP model particles with different PEG densities.
  • Various commercially available liposomes were included in this figure based on the PEG design.
  • the commercially available doxorubicin liposomes tested in Figure 6 The PEG density of batch number 692240505 was 5%.
  • b LC-MSNP model particles were cultured with A2 protein for 1 hour, followed by washing and immediate analysis using SDS-PAGE electrophoresis.
  • c Normalized gray values of the A2 protein band at the PEG density shown in b.
  • Figure 48 shows: a-b
  • We evaluated the stability of A2 pre-coating on low-density and high-density DSPE-PEG2000 LC-MSNPs. After A2 pre-coating and removal of unbound free protein, the particles were gently shaken on a shaker at specified time points, and protein surface adhesion was quantified. Data are expressed as mean ⁇ SD, n 3. d-e, The enhancement effect of A2 on LC-MSNPs containing 0% or 3% DSPE-PEG2000 was tested in the orthotopic EMT6 model. The results showed that the efficacy of the A2 coating was affected by the PEG density, i.e., 3% PEG instead of 0% PEG significantly improved efficacy.
  • Figure 49 illustrates the enhanced antitumor effect of commercial DOX liposomes in vivo with A2 coating.
  • A2 protein can effectively attach to commercial DOX liposomes.
  • D. Size and PDI values of A2-pre-coated commercial DOX liposomes measured in serum-supplemented saline at different time points (n 3).
  • Figure 50 illustrates the comparison of A2-mediated tumor pathway enhancement using size-controlled LC MSNPs.
  • a. Electrophoretic analysis showed that all LC MSNPs within this size range effectively bound to A2 (n 3).
  • Annexins or Annexin-like proteins, in this application belong to a family of calcium-dependent phospholipid-binding proteins.
  • the annexins include any one or a combination of at least two of the following: annexin A2, annexin A1, annexin A10, annexin A11, annexin A13, annexin A3, annexin A4, annexin A5, annexin A6, annexin A7, annexin A8, annexin A8L1, annexin A8L2, or annexin A9.
  • the annexins are preferably human-derived, and their detailed sequences are shown in the table below. However, in addition to humanized annexins, the annexins should also include monomers or combinations of similar proteins from different species.
  • the annexins should also include monomers or combinations of similar proteins across species (such as rodents, non-human primates, ungulates, birds, etc.).
  • IV LC-MSNPs intravenous (IV) LC-MSNPs can enter a KPC-derived orthotopic PDAC model, partly through a transcellular-mediated mechanism ( Figure 1 ).
  • LC-MSNPs we have demonstrated that intravenous (IV) LC-MSNPs can enter a KPC-derived orthotopic PDAC model, partly through a transcellular-mediated mechanism ( Figure 1 ).
  • our primary objective was to investigate how the design of particulate lipid coatings influences the abundance and mechanism of tumor biodistribution in PDAC.
  • LC-MSNPs as “model nanoparticles” because they are easy to visualize and offer flexibility in modulating lipid composition.
  • it allows for data generation on unsupported bilayers using LC-MSNPs, which can improve liposome performance under certain conditions.
  • This embodiment explores proteins that enhance the tumor entry efficiency of phospholipid nanocarriers.
  • MSNP mesoporous silicon
  • CTAC hexadecyltrimethylammonium chloride
  • MSNP MSNP at a ratio of 10% to APTES (120 mg of M001 plus 12 ⁇ L of APTES) into the sample vial, heat in an oil bath at 80°C, and stir overnight.
  • DMF dimethylformamide
  • the lipids required for encapsulating a neutral membrane are as follows (taking a total mass of 40 mg as an example):
  • the lipids required to form a positively charged membrane are as follows (taking a total mass of 40 mg as an example):
  • the lipids required to form a negatively charged membrane are as follows (taking a total mass of 40 mg as an example):
  • a KrasLSL-G12D/+Trp53LSL-R172H/+Pdx1-Cre (KPC) orthotopic pancreatic cancer mouse model was established. The specific procedure included: First, female B6/129 mice were anesthetized with isoflurane (for approximately 8 weeks). Anesthesia was maintained by intraperitoneal injection of ketamine 50 mg/kg and thiazide 10 mg/kg. The skin around the surgical site and within 1 cm of its margin was shaved and disinfected alternately with povidone-iodine and 75% ethanol. Then, the mice were placed on a heated pad for surgery. A sterile gauze was placed over the left abdominal incision, and a 0.7 cm incision was made to expose the pancreas.
  • DMEM/Matrigel (1:1 v/v) suspension containing 2 ⁇ 106 KPC-luc cells 50 ⁇ L of DMEM/Matrigel (1:1 v/v) suspension containing 2 ⁇ 106 KPC-luc cells was injected into the tail of the pancreas using a 27G needle syringe. Finally, the fascia layer was sutured with absorbable sutures, and the skin was sutured with non-absorbable sutures. The mouse was placed on a warm mat until fully awake, then transferred to a clean cage, and artificial tears ointment was used to protect the mouse's eyes during the operation.
  • mice bearing tumors for about 10 days were given intravenous injection of nanocarriers.
  • the tumor-bearing mice were randomly divided into 3 groups of 4 mice each. Each group was injected with the above-constructed nanocarriers via the tail vein, 0.5 mg/mouse. 24 hours later, the content of different nanocarriers (F1, F2 and F3) in the primary and metastatic tumors was analyzed by small animal in vivo imaging (IVIS) and near-infrared spectroscopy.
  • IVIS small animal in vivo imaging
  • LA-ICP-MS laser ablation inductively coupled plasma mass spectrometry
  • Nanocarriers that enter the bloodstream acquire not only their chemical identity but also biological identity, which is highly correlated with the nano-biological crowns (such as protein crowns) on the surface of the nanomaterials.
  • the basic experimental procedure for protein crown analysis is shown in Figure 7, including:
  • Nanocarriers F1, F2, and F3 (1 mg/mL, 50 ⁇ L) were incubated with tumor-bearing mouse plasma (200 ⁇ L) at 37°C for 3 h.
  • Mass spectrometry analysis was used to determine the search area for the "protein crown,” and the differences in protein crowns on the surfaces of different designed nanocarriers were compared. The results are shown in Figure 8, aiming to identify protein targets on the surface of nanocarriers with significant transcytosis activity. Unknown target proteins related to transcytosis were mined using proteomics databases and search engines, and the results are shown in Tables 2 and 3. Protein adsorption analysis was used to identify protein types unique to or shared by F1, F2, and F3. Library analysis further identified annexin as being associated with efficient tumor entry of F1, and its mechanism is related to transcytosis.
  • This embodiment further tests the effect of annexin on the tumor entrainment efficiency of phospholipid nanocarriers.
  • a series of annexin-coated F1 nanocarriers were prepared using different annexin isoforms (A2, A3, A5, A7, and A8).
  • the specific preparation process is as follows:
  • nanocarrier F1 (1 mg/mL, 50 ⁇ L) with strong transcytosis effect was incubated with annexin (0.5 mg/mL, 200 ⁇ L) at 37 °C for 3 h.
  • F1 nanocarriers coated with protein-free (w/o), plasma, vitronectin (VTN), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), or albumin (Alb) were prepared respectively.
  • nanocarriers pre-coated with different proteins were prepared at a concentration of 200 ⁇ g/mL in DMEM culture medium and added to the top of the chamber to simulate the vascular side. After 24 h, the lower layer solution was collected, and the Dy680 fluorescence value was detected using a multi-mode microplate reader to compare the number of different nanocarriers that penetrated the endothelial monolayer. As shown in Figure 11, it can be seen that coating nanocarriers with annexin A2, A3 or A5 can effectively improve the transcellular transport capacity of nanocarriers.
  • the F1 nanocarriers coated with each protein were intravenously injected into pancreatic cancer KPC mice at a dose of 50 mg/kg. 24 hours after injection, the mice's organs were dissected and placed in a small animal in vivo imaging system to observe the fluorescence signal. The biological distribution was compared by the intensity of the fluorescence signal.
  • TEM tissue transmission electron microscopy
  • Annexin in this invention can further improve the tumor entry efficiency of phospholipid nanocarriers.
  • This embodiment analyzes the mechanism by which annexin promotes the tumor entry of phospholipid nanocarriers, using annexin A2 as an example.
  • MD simulations were performed to analyze the interaction between A2 and lipid molecules.
  • the system structure was constructed using the Martini force field (Fig. 21), resulting in a well-equilibrium system.
  • MD simulations were conducted using a combination of coarse-grained (CG) and all-atom (AA) models.
  • CG coarse-grained
  • AA all-atom
  • CG simulation we observed that the A2 protein reached a stable position within 50 microseconds, and calculated the root mean square deviation (RMSD) of the protein's heavy atoms relative to its initial position, which was 22 ⁇ (Fig. 22).
  • RMSD root mean square deviation
  • the final CG structure became the initial structure for the all-atom (AA) simulation, and a 500-nanosecond MD analysis was performed.
  • the average RMSD of the C ⁇ atom of the A2 protein relative to its initial position fluctuated around 19 ⁇ (Fig. 23). Furthermore, we monitored the contact area between the A2 protein and lipids throughout the simulation trajectory; the average contact area remained stable at approximately 1000 square ⁇ (Fig. 24), indicating that the binding of A2 to lipids was very stable during the AA simulation. That is, the RMSD value indicates that the simulation system has reached a convergent state, and the interfacial contact area indicates that the contact between A2 and phospholipid molecules has reached a stable state, revealing a "phospholipid surface-annexin-integrin" sandwich model (Figure 25).
  • the specific experimental procedure includes:
  • EMT6 tumor-bearing mice were perfused cardiacally with PBS solution containing heparin sodium (1 mg/mL), and the outflowing blood was collected simultaneously.
  • the mice were then fixed by cardiac perfusion for 1 hour with a fixative solution (4% formaldehyde, 0.5% glutaraldehyde, PBS).
  • the fixative solution was washed away by perfusion with PBS for 20 minutes.
  • nanoparticles (1 mg/1.5 mL) were added to the collected mouse blood.
  • the nanoparticles were circulated in the fixed mice using a peristaltic pump at a physiological blood flow rate (5-7 mL/min) for 2 hours.
  • the nanoparticles were then removed from the blood vessels by perfusion with PBS.
  • the tumors were removed and imaged by IVIS.
  • the same concentration of nanoparticles was injected intravenously into live mice. After 2 hours, the blood was removed by cardiac perfusion with PBS, the tumors were removed, and IVIS imaging was performed to analyze the nanoparticle enrichment.
  • knocking down ⁇ 5 ⁇ 1 integrin expression or using formalin to fix tumor blood vessels can interfere with the role of A2 in promoting nanocarrier transport.
  • microscale thermophoresis (MST) analysis showed that the Kd values of A2 and A2-M were 7.93 nM and 30.88 nM, respectively ( Figure 30).
  • A2-M was used for pre-coating NIR-labeled LC-MSNPs for biodistribution studies in orthotopic EMT6 tumor-bearing mice, compared to particles coated with wild-type (WT) A2.
  • WT wild-type
  • the A2 coating was significantly impaired, resulting in a signal reduction of more than 50% at the tumor site ( Figure 31). Based on these findings, we attribute the reduced homing efficiency to decreased binding stability during circulation, which may impair the effective activation of ⁇ 5 ⁇ 1 integrin and subsequent downstream signaling events.
  • EMT6 orthotopic tumor-bearing mice received intravenous injections of DOX-loaded LC-MSNP nanocarriers with or without an A2 pre-coating (DOX 5 mg/kg; equivalent to an A2 dose of approximately 2 mg/kg).
  • DOX 5 mg/kg equivalent to an A2 dose of approximately 2 mg/kg.
  • Our results showed that the A2 pre-coating significantly enhanced the anti-BC activity of DOX LC-MSNPs, with a p-value of 0.0243 (Fig. 35b).
  • IRIN-LC-MSNP For IRIN-LC-MSNP, the A2 coating improved survival outcomes in the KPC in situ model, with a p-value of 0.0035 (IRIN-LC-MSNP vs A2 pre-coated IRIN-LC-MSNP) (Figure 35c). Note that tumor burden in the KPC model was assessed using IVIS, which may be affected by later ascites, as shown in Figure 36.
  • annexin pre-coating can increase the biodistribution of phospholipid nanocarriers (such as liposomes, lipid nanoparticles, phospholipid-coated nanoparticles, etc.) at lesion sites with high expression of ⁇ 5 or ⁇ 1 integrins.
  • phospholipid nanocarriers such as liposomes, lipid nanoparticles, phospholipid-coated nanoparticles, etc.
  • Annexin A2 pre-coating showed a significant increase in doxorubicin fluorescence in the BR00164 model (PDX with high ⁇ 5 ⁇ 1 integrin), but the increase was minimal in the BR00290 model (PDX with low ⁇ 5 ⁇ 1 integrin) (Figs. 40, 44).
  • phospholipid-coated silicon-based nanocarriers to conduct similar experiments in the above PDX models and obtained similar results (Figs. 41, 45; Table 6).
  • This embodiment further analyzes the effect of annexin on the therapeutic effect of drugs loaded onto phospholipid nanocarriers, using annexin A2 as an example.
  • the specific experimental procedure includes:
  • a mouse model of KPC orthotopic pancreatic cancer was constructed (referring to the aforementioned construction method).
  • EMT6 triple-negative breast cancer mouse model 3 ⁇ 106 EMT6 cells were resuspended in matrix gel and injected into the left quadruple mammary fat pad of 6-week-old Balb/c female mice.
  • Mesoporous silica coated with doxorubicin-loaded liposomes was prepared using ammonium sulfate ( NH4)2SO4 as the trapping agent.
  • Mesoporous silica coated with irinotecan and phospholipids was prepared, including a trapping agent of TEA 8 SOS.
  • Doxorubicin-loaded phospholipid-coated silica-based nanocarriers (1 mg/mL, 50 ⁇ L) and irinotecan-loaded phospholipid-coated silica-based nanocarriers (1 mg/mL, 50 ⁇ L) were incubated with A2 protein (0.5 mg/mL, 200 ⁇ L) at 37 °C for 3 h for coating.
  • mice bearing tumors were injected with the drug one week later, twice a week, with drug treatment every two or three days.
  • the groups were: irinotecan-loaded phospholipid-coated silicon-based nanocarriers with/without A2 coating (6 doses in total) and saline; and doxorubicin-loaded phospholipid-coated silicon-based nanocarriers with/without A2 coating (5 doses in total) and saline.
  • A2 enhanced the antitumor activity of doxorubicin-loaded phospholipid-coated silicon-based nanocarriers and irinotecan-loaded phospholipid-coated silicon-based nanocarriers compared with uncoated A2-loaded doxorubicin-loaded phospholipid-coated silicon-based nanocarriers and irinotecan-loaded phospholipid-coated silicon-based nanocarriers.
  • doxorubicin and irinotecan were directly loaded onto liposomes and coated with Annexin A2 to test the in vivo therapeutic effect.
  • Doxorubicin-loaded liposomes (1 mg/mL, 50 ⁇ L) and irinotecan-loaded liposomes (1 mg/mL, 50 ⁇ L) were incubated with A2 protein (0.5 mg/mL, 200 ⁇ L) at 37 °C for 3 h for coating.
  • this invention utilizes annexin-coated drug-loaded nanocarriers to effectively improve drug bioavailability and efficacy.
  • PEGylated doxorubicin liposomes (brand name: [Brand Name]) were tested in an in vivo BC model. ).
  • Fig. 49a bedside preparation protocol
  • the purchased DOX liposomes and their A2 pre-coated formulations were thoroughly characterized prior to use (Fig. 49b).
  • the A2-coated commercial DOX liposomes dispersed well in serum-supplemented saline and exhibited colloidal stability for at least 0–48 hours (Fig. 49d).
  • the amount of free DOX released from both uncoated and A2-coated liposomes was less than 0.05% (Fig. 49e).
  • ATR effects may be independent of vascular leakage and could play a dominant role in nanomedicine delivery to matrix-rich solid tumors.
  • A2 modification as a convenient means to activate tumor transcytosis and enhance the efficacy of nanocarriers in solid tumors.
  • Annexin A2 a calcium-dependent phospholipid-binding protein, plays a crucial role in various cellular processes, including membrane transport, endocytosis, and exocytosis.
  • this invention creatively identifies proteins that can enhance the tumor entry efficiency of current phospholipid nanocarriers. It discovers that coating phospholipid nanocarriers with annexin promotes transcytosis, increasing tumor entry efficiency and thus improving the bioavailability of drugs loaded on the nanocarriers, thereby enhancing therapeutic efficacy. Furthermore, it was found that annexin's synergistic effect is based on a "phospholipid surface-annexin-integrin" structure and is positively correlated with the expression levels of ⁇ 5 or ⁇ 1 integrins. Therefore, it is known that annexin pre-coating can also increase the biodistribution of phospholipid nanocarriers at lesion sites with high ⁇ 5 or ⁇ 1 integrin expression. Moreover, by pre-detecting and analyzing the expression levels of ⁇ 5 or ⁇ 1 integrins in patients, the ability of annexin coating technology to promote nanocarrier transport can be predicted, and beneficiaries of annexin coating technology can be screened.

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Abstract

L'invention concerne l'utilisation de l'annexine dans l'amélioration de l'efficacité de pénétration tumorale d'un nanotransporteur comportant une surface phospholipidique. La présente invention explore de manière innovante une protéine capable de favoriser l'efficacité de pénétration tumorale d'un nanotransporteur à base phospholipidique, et indique que le revêtement du nanotransporteur à base phospholipidique avec de l'annexine peut améliorer l'efficacité de pénétration tumorale, ce qui permet d'améliorer la biodisponibilité d'un médicament chargé avec le nanotransporteur et d'améliorer l'effet thérapeutique. De plus, il a également été découvert que la capacité d'amélioration de l'efficacité de l'annexine est corrélée positivement avec le niveau d'expression de l'intégrine α5 ou β1. Au moyen de la pré-détection et de l'analyse du niveau d'expression de l'intégrine α5 ou β1 chez un patient, la capacité de la technologie de revêtement d'annexine pour favoriser le transport de nanotransporteurs peut être prédite, et la population bénéficiaire appropriée pour l'utilisation de la technologie de revêtement d'annexine peut être criblée.
PCT/CN2025/094837 2024-05-17 2025-05-14 Utilisation de l'annexine dans l'amélioration de l'efficacité de pénétration tumorale d'un nanotransporteur comportant une surface phospholipidique Pending WO2025237331A1 (fr)

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WO2006003488A2 (fr) * 2004-07-07 2006-01-12 I.P. Randwijck B.V. Annexines, derives de ces dernieres, variantes d'annexines-cys et utilisations therapeutiques et diagnostiques de ces dernieres
CN113438945A (zh) * 2019-02-08 2021-09-24 德国癌症研究公共权益基金会 膜联蛋白涂覆的颗粒
CN117897399A (zh) * 2021-06-14 2024-04-16 Inserm(法国国家健康医学研究院) 突变的膜联蛋白a5多肽及其在治疗目的中的用途
CN118512400A (zh) * 2024-05-17 2024-08-20 国家纳米科学中心 膜联蛋白在提高具有磷脂表面的纳米载体入瘤效率中的应用

Patent Citations (4)

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Publication number Priority date Publication date Assignee Title
WO2006003488A2 (fr) * 2004-07-07 2006-01-12 I.P. Randwijck B.V. Annexines, derives de ces dernieres, variantes d'annexines-cys et utilisations therapeutiques et diagnostiques de ces dernieres
CN113438945A (zh) * 2019-02-08 2021-09-24 德国癌症研究公共权益基金会 膜联蛋白涂覆的颗粒
CN117897399A (zh) * 2021-06-14 2024-04-16 Inserm(法国国家健康医学研究院) 突变的膜联蛋白a5多肽及其在治疗目的中的用途
CN118512400A (zh) * 2024-05-17 2024-08-20 国家纳米科学中心 膜联蛋白在提高具有磷脂表面的纳米载体入瘤效率中的应用

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