Engineered cell preparations for treatment of hemophilia
AssigneeBe Biopharma, Inc.
InventorsTimothy James Mullen, Hanlan Liu, Anja Fides Hohmann, Sean Philip Leary Arlauckas, Swati Singh, Sean Patrick Keegan, Richard A. Morgan
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Abstract
The present disclosure provides methods and compositions for culturing, engineering, and administering engineered B lineage cell populations that produce Factor IX (FIX).
Claims48
A method of preparing an engineered B lineage cell population that express Factor IX (FIX), the method comprising the steps of:
- (a) isolating primary B cells so that a primary B cell population is obtained;
- (b) activating the primary B cell population;
- and
- (c) during or after the activating step, engineering the primary B cell population by integrating a transgene encoding FIX at a site selected from a CCR5 site, a IgH site, and a JCHAIN site,
- thereby generating an engineered B lineage cell population.
The method of any one of the above claims, further comprising a step of transferring the engineered B lineage cell population to cell culture media without human or bovine serum for about 24 hours.
The method of any one of the above claims, wherein the step of engineering further comprises introducing a donor construct comprising the transgene into the primary B cell population.
The method of any one of the above claims, wherein the step of engineering comprises contacting the primary B cell population with a targeted nuclease capable of introducing double-stranded breaks.
The method of any one of the above claims, wherein the step of engineering comprises:
- (a) electroporation of the primary B cell population, and/or
- (b) transduction of the primary B cell population with a donor construct.
A population of genetically modified B lineage cells comprising a transgene sequence encoding a FIX protein, wherein the transgene is expressed from an endogenous CCR5 locus.
A population of genetically modified B lineage cells comprising a transgene sequence encoding a FIX protein, wherein the transgene is expressed from an endogenous JCHAIN locus.
A population of genetically modified B lineage cells comprising a transgene sequence encoding a FIX protein, wherein the transgene is expressed from an endogenous IgH locus.
A pharmaceutical composition comprising one or more B lineage cells selected from the population of genetically modified B lineage cells of any of the above claims, and one or more pharmaceutically acceptable excipients.
A method of administering a pharmaceutical composition, wherein the pharmaceutical composition comprises:
- (a) one or more B lineage cells selected from the population of genetically modified B lineage cells of any one of the above claims;
- and
- (b) one or more pharmaceutically acceptable excipients;
- wherein the pharmaceutical composition is administered to a subject.
A method of monitoring engraftment of genetically modified B lincage cells within a subject, comprising one or more of:
- bioluminescence, ELISpot, flow cytometry, and enzyme-linked immunosorbent assay.
Dependencies are inferred from the claim text.