Nanoparticle-hydrogel composite for nucleic acid molecule delivery
Inventors
Schneider, Joel • Majumder, Poulami • Hoang, Chuong • Singh, Anand
Assignees
US Department of Health and Human Services
Publication Number
US-11904057-B2
Publication Date
2024-02-20
Expiration Date
2039-02-08
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Abstract
This disclosure provides novel peptide hydrogels containing encapsulated nanoparticles comprising nucleic acid molecules (such as miRNA) that can undergo multiple gel-to-solution (gel-sol) and solution-to-gel (sol-gel) phase transitions, and their use, such as for controlled delivery of nucleic acid molecules to a subject.
Core Innovation
The invention provides a novel peptide hydrogel containing encapsulated nanoparticles that comprise nucleic acid molecules, such as microRNA (miRNA) or their mimics and mimetics. These peptide hydrogels form fibrillar networks from amphiphilic cationic peptides folded into a β-hairpin conformation. The nanoparticles are formed by complexing nucleic acid molecules with a first amphiphilic cationic peptide that is unfolded and not in a β-hairpin conformation, then encapsulating these nanoparticles into the hydrogel matrix formed by a second amphiphilic cationic peptide in a β-hairpin conformation.
The key properties of the nanoparticle-hydrogel composite include shear-thinning upon application of shear stress and rheological recovery upon removal of shear stress. This allows the composite to be delivered locally to a target location within a subject, including routes such as syringe injection or spray delivery to coat anatomical surfaces. After delivery, nanoparticles are slowly released from the hydrogel matrix and taken up by cells, where the nucleic acid molecule is released to affect cellular function, for example, by gene silencing or regulating cellular pathways.
The background problem addressed is that while injectable hydrogels permit local delivery of therapeutics to reduce systemic off-target toxicity, previous hydrogel-mediated delivery vehicles face challenges for clinical viability, especially for nucleic acid-based therapeutics like RNA. Existing approaches could not effectively deliver functional miRNA to target cells in vivo with controlled release and stability within complex tissue environments such as serosal surfaces. The invention overcomes these challenges by designing a two-stage nanoparticle-hydrogel composite with defined peptide conformations, charge characteristics, and robust mechanical properties tailored for efficient nucleic acid delivery.
Claims Coverage
The patent includes 31 inventive features derived from multiple independent claims related to a peptide hydrogel with encapsulated nanoparticles for nucleic acid delivery, their compositions, structures, and methods of use in treating cancer.
Encapsulated nanoparticles comprising nucleic acid molecules complexed with unfolded cationic β-hairpin peptide
The hydrogel contains nanoparticles comprising a nucleic acid molecule complexed with a first amphiphilic cationic β-hairpin peptide that is unfolded and not in a β-hairpin conformation.
Peptide hydrogel formed from fibrillar network of β-hairpin folded amphiphilic cationic peptide
The hydrogel scaffold is formed from a second amphiphilic cationic β-hairpin peptide that is folded in a β-hairpin conformation, creating a fibrillar network.
Hydrogel shear-thinning and rheological recovery properties
The peptide hydrogel undergoes shear-thinning upon application of shear stress and recovers rheologically upon removal of shear stress, enabling flowable delivery and in situ gelation.
Nucleic acid molecule as antisense nucleic acid or microRNA
The nucleic acid molecule complexed in the nanoparticles is an antisense nucleic acid molecule or a microRNA or a mimic and/or mimetic thereof, including specific human mature miRNAs.
Electrostatic charge relationship between first and second amphiphilic cationic β-hairpin peptides
The net electrostatic charge of the first amphiphilic cationic β-hairpin peptide is equal to or more positive than that of the second amphiphilic cationic β-hairpin peptide at neutral pH, contributing to complex stability and release kinetics.
Specific amino acid sequences for peptides forming nanoparticles and hydrogel matrix
The first amphiphilic cationic β-hairpin peptide comprises or consists of the sequence VKVKVKVKVDPPTKVKVKVKV-NH2 (MAX1 peptide), and the second amphiphilic cationic β-hairpin peptide comprises or consists of the sequence VLTKVKTKVDPPTKVEVKVLV-NH2 (HLT2 peptide).
Terminal peptide modifications
The N-terminus of the first and/or second amphiphilic cationic β-hairpin peptides may be acetylated and/or their C-terminus may be amidated or modified to have neutral or positive charge.
Inclusion of heterologous anti-cancer agents dispersed within the peptide hydrogel
The peptide hydrogel can further include one or more heterologous anti-cancer agents such as chemotherapeutic agents dispersed within the matrix for combinatorial therapy.
Hydrogel composition parameters including modulus, ionic strength and concentration
The peptide hydrogel has a storage modulus greater than 40 Pascal, typically contains about 0.25% to 4.0% w/v of the second amphiphilic cationic peptide, and is formed at physiological ionic strength (about 10 mM to 400 mM NaCl) and pH 7.0 to 9.0 (typically about pH 7.4, 150 mM NaCl).
Methods of nucleic acid molecule administration to subjects using the peptide hydrogel
Methods comprising administering the peptide hydrogel containing nanoparticles to a target location in a subject for delivery of nucleic acid molecules.
Methods of treating or inhibiting cancer by localized delivery of peptide hydrogel
Methods comprising administering an effective amount of the peptide hydrogel to a target location where the cancer is present or at risk, with nucleic acid molecules (such as miRNAs) that inhibit the cancer.
Targeting serosal surfaces and specific cancers with peptide hydrogel delivery
The peptide hydrogel can be delivered to serosal surfaces lined by mesothelial cells (e.g., pleural, pericardial, peritoneal cavities) for treatment of serosal neoplasms such as malignant pleural mesothelioma and other cancer types disclosed.
Spray delivery or injection to coat complex anatomical serosal surfaces
The peptide hydrogel can be administered by injection or spray delivery to coat all or a portion of the serosal surface where cancer is present or at risk.
The claims define a nanoparticle-hydrogel composite comprising nanoparticles formed from a nucleic acid molecule complexed with an unfolded amphiphilic cationic β-hairpin peptide, encapsulated within a peptide hydrogel formed by a β-hairpin folded amphiphilic cationic peptide, displaying shear-thinning and recovery behavior. The nucleic acid includes antisense oligonucleotides and miRNAs. The composition includes defined peptide sequences, electrostatic charge relationships, terminal modifications, and optionally heterologous anti-cancer agents. The claims cover compositions, methods of manufacture, administration, and therapeutic methods targeting cancers, especially serosal neoplasms including malignant pleural mesothelioma.
Stated Advantages
The nanoparticle-hydrogel composite allows locoregional, sustained, and controlled delivery of nucleic acid molecules such as miRNAs with shear-thinning injectable and sprayable properties to cover complex tissue surfaces.
The system minimizes systemic distribution thereby limiting off-target toxicity during therapeutic delivery.
Nanoparticles formed from unfolded peptides enable effective uptake, endosomal escape, and functional gene silencing of delivered nucleic acid molecules in cells.
The hydrogel exhibits biodegradability, allowing for eventual bioresorption without need for removal.
The positively charged hydrogel fibrillar network stabilizes nanoparticles, preventing premature release of free nucleic acids and preserving functional delivery.
Single administration of the composite produces sustained therapeutic effects with improved patient compliance.
MiRNA-based treatment can modulate multiple intracellular pathways coherently, potentially reducing tumor resistance compared to conventional single-target therapies.
Documented Applications
Controlled local delivery of nucleic acid molecules such as miRNA therapeutics to complex anatomical surfaces, including serosal surfaces in the body.
Treatment or inhibition of cancer, particularly serosal neoplasms such as malignant pleural mesothelioma.
Use as a delivery system for combination therapies including nucleic acid molecules and heterologous anti-cancer agents such as chemotherapeutics.
Application of the composite by syringe injection or spray delivery to coat target tissues such as pleural surfaces in the lung, peritoneal cavities, or other serosal body cavities.
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