VCN enhancer compositions and methods of using the same
Inventors
Bonner, Melissa • NEGRE, Olivier
Assignees
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Publication Number
US-12234476-B2
Publication Date
2025-02-25
Expiration Date
Abstract
The invention provides improved gene therapy methods and compositions.
Core Innovation
The invention is directed to a method of increasing the vector copy number (VCN) in a population of CD34+ hematopoietic stem or progenitor cells by culturing the cells for at least two hours with a lentiviral vector, prostaglandin E2 (PGE2) or 16,16-dimethyl PGE2, and poloxamer 338. The increase in VCN achieved with poloxamer 338 present is further enhanced beyond the increased VCN achieved with the lentiviral vector and PGE2 or 16,16-dimethyl PGE2 in the absence of poloxamer 338.
The method is described in the context of improving gene therapy performance in hematopoietic stem/progenitor cells, with a focus on increasing transduction efficiency and vector copy number using retroviral/lentiviral vectors. The disclosure also states that vector performance needs improvement and that acceptable cell characteristics such as viability and engraftment and differentiation after transplant are desired.
The document further describes lentiviral vector compositions and architectures for use in the CD34+ hematopoietic stem or progenitor cell population, including a 5′ LTR, a Ψ packaging signal, a self-inactivating 3′ LTR, cPPT/FLAP, an RNA export element, and a promoter operably linked to a polynucleotide of interest. Therapeutic transgene strategies and expression control include erythroid cell specific expression for globin gene and antisickling protein strategies and microglial promoter-driven expression for ABCD1 in the context of adrenoleukodystrophy or adrenomyeloneuropathy.
Claims Coverage
The provided independent claims center on a method that increases vector copy number (VCN) in CD34+ hematopoietic stem or progenitor cells using a lentiviral vector together with a prostaglandin E2 component and poloxamer 338, where the VCN increase is further enhanced beyond conditions without poloxamer 338. Dependent claims refine the method by specifying CD34+ selection, named lentiviral vectors, lentiviral vector architecture, and concentration ranges.
Poloxamer 338 plus prostaglandin E2 with lentiviral vector to enhance VCN
A method of increasing vector copy number (VCN) in a population of CD34+ hematopoietic stem or progenitor cells by culturing for at least two hours with a lentiviral vector, prostaglandin E2 (PGE2) or 16,16-dimethyl PGE2, and poloxamer 338, wherein the increase of VCN achieved with poloxamer 338 present is further enhanced beyond the increased VCN achieved by culturing with the lentiviral vector and PGE2 or 16,16-dimethyl PGE2 in the absence of poloxamer 338.
CD34+ expression-based selection of hematopoietic stem or progenitor cells
The method selects hematopoietic stem or progenitor cells based on CD34+ expression prior to transduction.
Named lentiviral vector selections for use in the VCN-enhancing method
The method is performed using a lentiviral vector selected from specified named lentiviral vector types.
Lentiviral vector architecture with 5′ LTR, Ψ packaging signal, RNA export element, promoter, and SIN 3′ LTR
The method is carried out using a lentiviral vector that includes a 5′ LTR, Ψ packaging signal, RNA export element, lentiviral cPPT, a promoter operably linked to a polynucleotide of interest, and a SIN 3′ LTR.
Quantitative culture constraint for poloxamer 338 concentration
The method includes culturing with poloxamer 338 in the culture at a concentration of about 50 μg/mL to about 1000 μg/mL.
Quantitative culture constraint for PGE2 or 16,16-dimethyl PGE2 concentration
The method includes culturing with either PGE2 or 16,16-dimethyl PGE2 present in the culture in a concentration of about 10 μM to about 100 μM.
Overall claim coverage centers on the combination of a lentiviral vector with PGE2 or 16,16-dimethyl PGE2 and poloxamer 338 to further enhance VCN relative to conditions lacking poloxamer 338, with dependent refinements covering CD34+ selection, specific lentiviral vector types, vector architecture, and concentration ranges.
Stated Advantages
Further enhances the VCN increase achieved by culturing with a lentiviral vector and PGE2 or 16,16-dimethyl PGE2 beyond the increased VCN achieved in the absence of poloxamer 338.
Increases transduction efficiency and/or VCN without markedly reducing viability.
Maintains engraftment/differentiation after transplant.
Documented Applications
Hematopoietic gene therapy using lentiviral vectors in a population of CD34+ hematopoietic stem or progenitor cells, including erythroid cell specific expression strategies for globin/antisickling proteins and microglial promoter-driven expression for ABCD1 related to adrenoleukodystrophy/adrenomyeloneuropathy.
Disease-relevant gene therapy payloads described include TPP1, ADA, IL2Rγ, tripeptidyl peptidase 1, alpha-L-iduronidase, and iduronate 2-sulfatase.
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