Assessment of Bioreactor-Derived Human Platelet Quality, Storage Profile, Safety, and Function

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 DESCRIPTION (provided by applicant): Platelet BioGenesis is developing a microfluidic bioreactor to produce human platelets at clinical scale. Platelets are the 'band-aids' of the bloodstream, responsible for clot formation and blood vessel repair. Low platelet count is a significant consequence of cancer treatment, transplant, and surgery, for which platelets are a critical first-line therapy to prevent mortality due to uncontrolled bleeding. Platelet units comprising 3x1011 platelets per 200-400 mL are at present derived exclusively from human volunteer donors, and must be stored at or above 22oC to avoid irreversible temperature-related activation/aggregation. Risk of bacterial growth during room temperature storage limits shelf life to 5 days, 2 of which are consumed by bacterial screening, and 1 by transport. As a result blood centers typically do not have more than a 1.5-day platelet inventory avail- able for transfusion, which is rapidly depleted in emergencies [1, 2]. To address this major unmet need we have developed a platelet bioreactor that reproduces key features of adult bone marrow (physiological microenvironment) to trigger new platelet production at increasing scale. To date we have shown that: (1) It is feasible to generate functional megakaryocytes and platelets from human induced pluripotent stem cells (iP-SCs, a replenishable source of progenitor cells which can be stored frozen for years)[3] and (2) our bioreactor will trigger platelet formation and improve the rate and extent of platelet production from human iPSC-derived MKs above established static culture approaches[4, 5]. This Direct-to-Phase II SBIR proposal outlines 3 specific aims to determine quality, safety, and function of bioreactor-derived platelets (bdPLTs) in humanized mouse models of thrombocytopenia, and to scale platelet production to yield a human transfusion unit: Aim 1. Assess bdPLT quality, storage profile, and safety in vitro. We will evaluate platelet structure, biomarker expression, metabolic activity, and function under FDA-approved storage conditions, and determine their safety by enumerating residual leukocyte counts, and assessing viral, septic, and teratoma risk. Aim 2. Assess the hemostatic and thrombogenic potential of bdPLT in humanized mice as well as platelet clearance and circulation time. We will evaluate the in vivo behavior of bdPLTs in a novel murine model that permits human but not mouse platelet accrual at sites of arterial injury (non-GLP pilot study). Hu- man donor platelets will serve as a physiological control. Aim 3. Scale production 3,000-fold to yield a human transfusion unit (3x1011 bdPLTs per 300 mL). We will optimize our bioreactor design for continuous media perfusion and parallelization, maximize megakaryocyte zonal distribution and trapping, and equalize shear stress exposure throughout the device.