Compositions and methods for delivering inhibitory oligonucleotides
Inventors
Arya, Bira • Olkhanud, Purevdorj • Espinoza, Juan
Assignees
US Department of Health and Human Services
Publication Number
US-10765694-B2
Publication Date
2020-09-08
Expiration Date
2029-04-15
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Abstract
The present invention features compositions and methods that make use of complexes comprising one or more inhibitory nucleic acids and a targeting polypeptide, wherein the targeting polypeptide consists of a cell surface receptor ligand. The compositions can be used in methods of silencing gene expression in a cell, delivering agents to a target cell, and in treating or preventing a disease or disorder in a subject.
Core Innovation
The invention provides novel compositions and methods that utilize complexes comprising one or more inhibitory nucleic acids and a targeting polypeptide, wherein the targeting polypeptide consists of a cell surface receptor ligand. The compositions enable targeted and efficient delivery of inhibitory oligonucleotides to cells expressing the corresponding cell surface receptor. The invention features fusion molecules and complexes that include a nucleic acid binding moiety associated with a ligand such as a chemokine, cytokine, antibody, or growth factor, facilitating gene expression silencing, delivery of agents to specific cells, and treatment or prevention of diseases or disorders.
The compositions solve the need for improved treatments that overcome limitations associated with existing siRNA delivery technologies, such as difficulties in in vivo delivery, lack of specificity, and safety concerns associated with viral vector-based systems. The invention addresses the demand for high specificity of delivery of inhibitory nucleic acids that current systems fail to provide by employing cell surface receptor-specific ligands to achieve direct targeting and internalization of the inhibitory nucleic acids into target cells.
Claims Coverage
The patent includes 18 independent claims, focusing primarily on fusion molecules comprising targeting polypeptides selected from specific chemokines or their receptor binding fragments combined with nucleic acid binding domains, complexes including inhibitory nucleic acids with such fusion molecules, and specific amino acid sequences. The claims cover structural features, nucleic acid binding, inhibitory nucleic acid types, spacer peptides, and target specificity.
Fusion molecule comprising specific chemokine targeting polypeptides and nucleic acid binding domain
A fusion molecule comprising a targeting polypeptide selected from MCP-1/CCL-2, TARC/CCL17, RANTES/CCL5, or receptor binding fragments thereof, fused with a nucleic acid binding domain comprising amino acids 87 through 102 of SEQ ID NO: 11.
Inhibitory nucleic acid-fusion molecule complex formation
The fusion molecule optionally includes one or more inhibitory nucleic acids forming a complex capable of gene expression inhibition.
Targeting polypeptide defined by specific amino acid sequences
Targeting polypeptides specified as MCP-1 (amino acids 3-77 of SEQ ID NO: 11), TARC (amino acids 1-72 of SEQ ID NO: 9), or RANTES (amino acids 1-68 of SEQ ID NO: 10) or their receptor binding fragments.
Inclusion of spacer peptides between targeting polypeptide and nucleic acid binding domain
The fusion molecule may include spacer peptides selected from SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, or SEQ ID NO: 8 to separate the targeting polypeptide and nucleic acid binding domain.
Inhibitory nucleic acids comprising various RNA types and modifications
The inhibitory nucleic acids include antisense nucleic acid, siNA, siRNA, dsRNA, miRNA, shRNA, and combinations thereof, with possible nucleotide modifications.
Specific targeting of cytokine and chemokine gene expression
The inhibitory nucleic acids hybridize to or are complementary to nucleic acids encoding cytokines or chemokines, including IL-10.
Specific fusion molecule and complex compositions
Fusion molecules comprising the amino acid sequences of SEQ ID NO: 9, SEQ ID NO: 10, and SEQ ID NO: 11 are claimed, as well as complexes of inhibitory nucleic acids with the fusion molecule comprising SEQ ID NO: 11.
The claims collectively cover fusion molecules composed of specific chemokine-derived targeting polypeptides fused to defined nucleic acid binding domains, optionally forming complexes with various inhibitory nucleic acids, including modified forms targeting cytokine genes like IL-10, with spacer sequences facilitating their assembly, enabling targeted gene silencing and therapeutic delivery.
Stated Advantages
The invention provides improved specificity and efficiency in delivering inhibitory nucleic acids to target cells expressing the relevant cell surface receptors.
The compositions enable targeted gene silencing in diseased cells such as cancer cells, enhancing therapeutic potential while minimizing off-target effects.
The technology facilitates delivery of labeled oligonucleotides for diagnostic purposes including tracing cells and determining tumor metastasis.
The described fusion molecules and complexes offer a simplified and scalable production process with increased yield, including expression in yeast systems.
Documented Applications
Silencing or knocking down gene expression in cells by targeted delivery of inhibitory nucleic acids using complexes comprising targeting polypeptides that are cell surface receptor ligands.
Delivering inhibitory RNA molecules, including siRNA, into cells expressing specific chemokine receptors.
Treating or preventing diseases or disorders in subjects by decreasing gene expression in target cells, including treatment of cancers such as leukemia and breast cancer.
Delivering therapeutic agents or imaging agents coupled to inhibitory nucleic acids to target cells, for therapy or diagnostic imaging to determine prognosis or course of treatment.
Modulating gene expression in ex vivo tissue explants or cultured cells prior to transplantation to confer desired phenotypes for therapeutic applications.
Using complexes to deliver inhibitory nucleic acids to various cell types including immune cells, epithelial cells, malignant cells, and stem cells in mammalian subjects, including humans.
Use in diagnosing metastasis and monitoring disease progression, particularly in neoplastic and immunological diseases.
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