WO2026033348A1 - Polysaccharides et oligosaccharides encapsulés dans des nanoparticules lipidiques - Google Patents

Polysaccharides et oligosaccharides encapsulés dans des nanoparticules lipidiques

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Publication number
WO2026033348A1
WO2026033348A1 PCT/IB2025/057812 IB2025057812W WO2026033348A1 WO 2026033348 A1 WO2026033348 A1 WO 2026033348A1 IB 2025057812 W IB2025057812 W IB 2025057812W WO 2026033348 A1 WO2026033348 A1 WO 2026033348A1
Authority
WO
WIPO (PCT)
Prior art keywords
lipid
oligosaccharide
process according
solution
polysaccharide
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/IB2025/057812
Other languages
English (en)
Inventor
Sofia NIZZOLO
Sabrina Bertini
Marco Guerrini
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.)
Centro Alta Tecnologia Istituto Di Ricerche Chimiche E Biochimiche "g Ronzoni" Srl
Original Assignee
Centro Alta Tecnologia Istituto Di Ricerche Chimiche E Biochimiche "g Ronzoni" Srl
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Centro Alta Tecnologia Istituto Di Ricerche Chimiche E Biochimiche "g Ronzoni" Srl filed Critical Centro Alta Tecnologia Istituto Di Ricerche Chimiche E Biochimiche "g Ronzoni" Srl
Publication of WO2026033348A1 publication Critical patent/WO2026033348A1/fr
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • 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
    • A61K9/5161—Polysaccharides, e.g. alginate, chitosan, cellulose derivatives; Cyclodextrin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
    • A61K31/726—Glycosaminoglycans, i.e. mucopolysaccharides
    • A61K31/727—Heparin; Heparan
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
    • A61K31/726—Glycosaminoglycans, i.e. mucopolysaccharides
    • A61K31/728—Hyaluronic acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
    • A61K31/737—Sulfated polysaccharides, e.g. chondroitin sulfate, dermatan sulfate
    • 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

Definitions

  • the present invention relates to lipid nanoparticles comprising a polysaccharide and/or an oligosaccharide.
  • lipid-based drug delivery systems have attracted significant attention from researchers. These systems offer various advantages over conventional dosage forms, such as greater stability and bioavailability.
  • liposome-based system consist of one or more lipid bilayers and a hydrophilic core. They can be used to deliver both hydrophobic substances, which localize within the lipid bilayer, and hydrophilic substances, which reside in the hydrophilic core.
  • the size of liposomes generally ranges from a minimum of 50 nm to a maximum of approximately 1000 nm, and they are also suitable for encapsulating relatively large molecules.
  • Glycosaminoglycans are natural products of considerable pharmaceutical interest. Among the most widely used are heparin, dermatan, heparan sulfate, chondroitins, and hyaluronic acid. The molecular weight of these natural products varies significantly, generally ranging from 5 to 40 kDa for heparin, dermatan, heparan sulfate, and chondroitins, while it can reach up to 4,000 kDa in the case of hyaluronic acid. The high molecular weight of these compounds often makes oral administration impractical, necessitating intravenous administration. However, it is well known that intravenous administration leads to rapid degradation of the molecule, with a consequent rapid reduction in biological activity.
  • LNPs lipid nanoparticles
  • Lipid nanoparticles are typically composed of four main lipids: an ionizable lipid in either a positively charged or neutral form, a saturated phospholipid (zwitterionic lipid) forming the bilayer, cholesterol, and a PEG-conjugated lipid.
  • an ionizable lipid in either a positively charged or neutral form a saturated phospholipid (zwitterionic lipid) forming the bilayer
  • cholesterol a PEG-conjugated lipid.
  • PatisiranTM the molar ratio among the listed components (DLin-MC3-DMA: 1,2-DSPC: Cholesterol: C-DMC-PEG(2000)) is 50: 10:38.5: 1.5.
  • WO 2022/115604 discloses novel PEGylated lipid compounds to be used in compositions comprising, in addition to the PEGylated lipid, cholesterol and phospholipids for the encapsulation of mRNA fragments.
  • the patent application specifies that the encapsulation may be used for RNA fragments or small molecules.
  • vaccines are typically administered intramuscularly, whereas certain polysaccharides —such as glycosaminoglycans, and in particular heparin — are administered intravenously. All pharmacokinetic data obtained for one route of administration cannot be directly translated to a different administration route.
  • WO2023084217A1 describes advanced payload delivery systems based on lipid nanoparticles functionalised with polymers (PF-LNPs), designed for intracellular delivery of nucleic acids, particularly self-amplifying RNA (saRNA).
  • PF-LNPs lipid nanoparticles functionalised with polymers
  • saRNA self-amplifying RNA
  • a distinguishing feature of the system described in the patent is the use of a preparation method different from standard microfluidic approaches, which adversely affects particle size distribution.
  • Experimental data e.g., Figure 3B, page 95
  • lipid nanoparticles comprising a polysaccharide or an oligosaccharide having a molecular weight ranging from 1,200 Da to 200,000 Da by mixing, using a microfluidic system, a solution comprising a neutral lipid compound, a neutral phospholipid, a sterol, and optionally an ionizable cationic lipid compound, with a solution comprising the oligosaccharide or the polysaccharide.
  • the polysaccharides and oligosaccharides incorporated in the lipid nanoparticles have a molecular weight ranging from 1,500 to 150,000 Da, more preferably from 2,000 to 120,000 Da.
  • the invention also relates to lipid nanoparticles comprising an oligosaccharide or a polysaccharide obtainable by the process defined above.
  • the neutral lipid compounds usable in the present invention are compounds containing at least one lipid chain bound to a polyglycol group.
  • the compounds are referred to as PEGylated lipids.
  • the bond between the lipid group and the glycol group may be formed by esterification between the carboxyl group of the lipid and one or two hydroxyl groups of the terminal glycol group, as in the case of DMG- PEG(2000), or may involve a functional group linking the polyglycolic part to the lipid part, as in DSPE-MPEG(2000).
  • DMG-PEG(2000) is used.
  • One of the purposes of the neutral lipid compounds is to prevent the adsorption of plasma proteins, thereby inhibiting uptake by the mononuclear phagocyte system, which is one of the main barriers to the efficacy of LBDD systems.
  • Lipidic sterols are generally used in LNP systems to fill membrane packing defects.
  • the most common is cholesterol, although similar molecules such as cholesterol sulfate may also be used, as well as cholesterol derivatives modified chemically to be pH-sensitive and/or capable of reacting with proteins and/or enzymes.
  • the phospholipid is preferably selected from 1,2-distearoylphosphatidylcholine (1,2-DSPC), phosphatidylcholine, and phosphatidylethanolamine.
  • 1,2-DSPC 1,2-distearoylphosphatidylcholine
  • phosphatidylcholine 1,2-distearoylphosphatidylcholine
  • phosphatidylethanolamine 1,2-distearoylphosphatidylcholine
  • An optional component in the preparation of the LNPs is an ionizable cationic lipid. These compounds are neutral at approximately pH 7 and acquire a positive charge at pH values below their pKa, facilitating the formation of reverse micelles that encapsulate the glycosaminoglycan in the LNP core. Their neutrality at pH 7 results in much lower cytotoxicity compared to permanently charged cationic lipids.
  • Preferred ionizable cationic lipids include: l,2-dioleoyl-3 -dimethylammonium propane (DODAP), l,2-dioleyl-3 -dimethylammonium propane (DODMA), 4-(dimethylamino)-butanoic acid-l-(9Z,12Z)-9,12-octadecadien-l-yl- 10, 13 -nonadecadi en-l-yl ester (DLin-MC3-DMA), 6-((2-hexyldecanoyl)oxy)-N-(6-((2- hexyldecanoyl)oxy)hexyl)-N-(4-hydroxybutyl)hexan-l-aminium (ALC-0315), and 306OH0 having formula
  • the preferred ionizable cationic lipids are DLin-MC3-DMA and 306OH0, with DLin- MC3-DMA being most preferred.
  • Microfluidic systems usable in the present invention are disclosed by Masatoshi Maeki et al., Microfluidic technologies and devices for lipid nanoparticle-based RNA delivery, Journal of Controlled Release 344 (2022) 80-96, incorporated herein by reference.
  • the nanoparticles of the present invention can efficiently encapsulate oligosaccharides and polysaccharides.
  • encapsulatable polysaccharides include glycosaminoglycans (hyaluronic acid, heparin, dermatan, dermatan sulfate, chondroitin 4 sulfate, chondroitin 6 sulfate, keratan sulfate, heparan sulfate, glycosplit heparin), inulin, pectin, alginate, dextrans, sulfated dextrans, chitosan, chitin, arabinoxylans, arabinogalactans, carrageenan, pentosan polysulfate, glucomannans, and galactomannans.
  • glycosaminoglycans hyaluronic acid, heparin, dermatan, dermatan sulfate, chondroitin 4 sulfate
  • the nanoparticles according to the invention are obtained through the following process. Three solutions are prepared: a first solution containing the lipid portion, i.e., a phospholipid, a neutral lipid, and a lipid sterol, and optionally an ionizable cationic lipid; a second solution containing a poly- or oligosaccharide; and a third solution containing a buffer.
  • the first two solutions are mixed together using a microfluidic system to obtain nanoparticles encapsulating the poly- or oligosaccharide.
  • the microfluidic mixing system is an IJM (Impingement Jet Mixer).
  • the nanoparticles obtained using these microfluidic systems are characterized by very high stability in their physical properties, and therefore exhibit high reproducibility, allowing for the production of a consistent product over time.
  • the volume-weighted average diameter of the nanoparticles according to the invention ranges from 40 nm to 200 nm, more preferably from 50 nm to 150 nm, and even more preferably from 60 nm to 120 nm.
  • the process according to the invention enables a high encapsulation efficiency of the poly- or oligosaccharide within the nanoparticles.
  • the encapsulation efficiency is greater than 50%, more preferably greater than 60%.
  • composition obtained by the process of the present invention may be used as such, i.e., containing non-encapsulated poly- or oligosaccharide, or, for certain applications, purification of the composition may be recommended to remove the non-encapsulated poly- or oligosaccharide. Additionally, both the purified and non-purified compositions may be used in combination with other poly- or oligosaccharides optionally modified by the introduction of functional groups that render them pH-sensitive and/or capable of interacting with proteins and/or enzymes.
  • Lipid nanoparticles were prepared using a Nano Scaler IJM device from Knauer as follows.
  • the polysaccharide solution was prepared by dissolving the polysaccharide in a 0.14 mM or 0.014 mM NaCl solution, or in a 0.01 mM or 0.1 mM citrate buffer, and maintaining the solution under stirring overnight.
  • the solution containing the lipid portion was obtained by dissolving the lipid compounds in absolute ethanol at the concentrations reported in Table 1.
  • PBS phosphate-buffered saline
  • pH 7.4 approximately 8 g of sodium chloride, 0.2 g of potassium chloride, approximately 1.44 g of disodium phosphate, and approximately 0.245 g of monopotassium phosphate were added to 1 L of deionized water.
  • the pH was adjusted to the desired final value using HC1 or NaOH.
  • a suitable amount of UFH was dissolved in 0.14 mM NaCl for 2 hours at room temperature to reach the final concentration indicated in the table.
  • PBS at pH 7.4 was used as the quench solution.
  • the lipid mixture was prepared by adding 16 pL of a stock solution of DLin-MC3-DMA (100 mg/mL), 16 pL of a stock solution of cholesterol (5 mg/mL), 16 pL of a stock solution of 1,2-DSPC (25 mg/mL), and 189 pL of a stock solution of 1,2-DMG- PEG (1 mg/mL). The mixture was brought to a final volume of 1 mL with ethanol.
  • LNPs were prepared using an IJM NanoScaler under condition 1.
  • the total flow rate (TFR) was set at 3 mL/min with the following settings: Pump 1 (UFH): 1.5 mL/min, Pump 2 (Lipids): 0.5 mL/min, Pump 3 (Quench): 1 mL/min.
  • Particle size and Zeta potential (Zp) were determined by dynamic light scattering (DLS) measurements on the formulations. A volume of 80 pL of solution was used for particle size analysis, while 800 pL were used for Zp determination.
  • Encapsulation efficiency was assessed by HP-SEC-TDA, after measuring the UFH concentration outside the LNPs (recovered concentration). The encapsulation efficiency (EE%) was then calculated using the following formula:
  • Table 2 reports the data from experiments conducted with UFH under various conditions and at different UFH concentrations, using lipid mixtures A to H as described in Table 1.

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Public Health (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Epidemiology (AREA)
  • Veterinary Medicine (AREA)
  • General Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Molecular Biology (AREA)
  • Dermatology (AREA)
  • Physics & Mathematics (AREA)
  • Biomedical Technology (AREA)
  • Nanotechnology (AREA)
  • Optics & Photonics (AREA)
  • Medicinal Preparation (AREA)

Abstract

L'invention concerne un procédé de préparation d'une composition comprenant un oligosaccharide ou un polysaccharide encapsulé dans des nanoparticules lipidiques, le procédé comprenant les étapes suivantes consistant à : préparer une première solution d'une partie lipidique comprenant un phospholipide, un lipide neutre et un stérol lipidique ; préparer une deuxième solution comprenant un poly- ou oligosaccharide ; préparer une troisième solution contenant un tampon ; mélanger les première et deuxième solutions à l'aide d'un système microfluidique pour obtenir des nanoparticules lipidiques encapsulant l'oligosaccharide ou le polysaccharide ; combiner la troisième solution avec les première et deuxième solutions. L'invention concerne également une composition comprenant des nanoparticules lipidiques pouvant être obtenues par le procédé défini ci-dessus, ladite composition comprenant une partie lipidique comprenant un phospholipide, un composé lipidique neutre dans lequel une chaîne lipidique est liée à un groupe glycol, ainsi qu'un stérol lipidique ; un tampon ; et un oligosaccharide ou un polysaccharide.
PCT/IB2025/057812 2024-08-07 2025-07-31 Polysaccharides et oligosaccharides encapsulés dans des nanoparticules lipidiques Pending WO2026033348A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
IT202400018625 2024-08-07
IT102024000018625 2024-08-07

Publications (1)

Publication Number Publication Date
WO2026033348A1 true WO2026033348A1 (fr) 2026-02-12

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Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2149580A1 (fr) 2008-07-15 2010-02-03 Istituto Scientifico di Chimica E Biochimica "G Ronzoni Mimétiques d'oligosaccharides sulfatés
WO2020061457A1 (fr) * 2018-09-20 2020-03-26 Modernatx, Inc. Préparation de nanoparticules lipidiques et leurs méthodes d'administration
WO2022115604A2 (fr) 2020-11-24 2022-06-02 The Brigham And Women's Hospital, Inc. Véhicules d'administration à longue durée d'action et longue durée de circulation
WO2023282652A2 (fr) * 2021-07-07 2023-01-12 주식회사 제넥신 Dérivé d'acide hyaluronique-lipide, nanoparticule lipidique le comprenant et utilisation associée
WO2023084217A1 (fr) 2021-11-10 2023-05-19 Imperial College Innovations Limited Système de distribution de charge utile
WO2024044178A1 (fr) * 2022-08-24 2024-02-29 Popvax Private Limited Composition ou formulation de nanoparticules lipidiques (npl) pour agents thérapeutiques à base d'acides nucléiques

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2149580A1 (fr) 2008-07-15 2010-02-03 Istituto Scientifico di Chimica E Biochimica "G Ronzoni Mimétiques d'oligosaccharides sulfatés
WO2020061457A1 (fr) * 2018-09-20 2020-03-26 Modernatx, Inc. Préparation de nanoparticules lipidiques et leurs méthodes d'administration
WO2022115604A2 (fr) 2020-11-24 2022-06-02 The Brigham And Women's Hospital, Inc. Véhicules d'administration à longue durée d'action et longue durée de circulation
WO2023282652A2 (fr) * 2021-07-07 2023-01-12 주식회사 제넥신 Dérivé d'acide hyaluronique-lipide, nanoparticule lipidique le comprenant et utilisation associée
WO2023084217A1 (fr) 2021-11-10 2023-05-19 Imperial College Innovations Limited Système de distribution de charge utile
WO2024044178A1 (fr) * 2022-08-24 2024-02-29 Popvax Private Limited Composition ou formulation de nanoparticules lipidiques (npl) pour agents thérapeutiques à base d'acides nucléiques

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
MASATOSHI MAEKI ET AL.: "Microfluidic technologies and devices for lipid nanoparticle-based RNA delivery", JOURNAL OF CONTROLLED RELEASE, vol. 344, 2022, pages 80 - 96, XP055903700, DOI: 10.1016/j.jconrel.2022.02.017
T.D. KIM ET AL.: "Studies on liposome encapsulated heparin", THROMBOSIS RESEARCH, vol. 43, 1986, pages 603 - 612, XP022879967, DOI: 10.1016/0049-3848(86)90097-6

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