Manufacture of an Allogeneic CAR-T Stem Cell Memory Product Candidate for Multiple Myeloma, P-Bcma-ALLO1, Is Robust, Reproducible and Highly Scalable

Chimeric Antigen Receptor (CAR) T cell therapy has generated unprecedented efficacy in the treatment of multiple hematologic malignancies. For relapsed/refractory Multiple Myeloma (MM), autologous CAR-T products directed against the B cell maturation antigen (BCMA), such as Poseida's P-BCMA-101, have demonstrated significant efficacy. P-BCMA-101 is comprised of a high-percentage of stem cell memory T cells (TSCM), resulting in a product that is much safer and potentially more durable than other anti-BCMA autologous product candidates. However, as individualized products, all autologous CAR-T products are expensive to manufacture and dependent upon patient T-cells of variable quality. We are developing P-BCMA-ALLO1, an off-the-shelf allogeneic (allo) BCMA-specific CAR-T product candidate derived from healthy donor material, which provides numerous advantages over autologous products, increasing patient access by being immediately available and greatly reducing manufacturing cost and variability. P-BCMA-ALLO1 is produced using two key platform technologies: the nonviral piggyBac® (PB) DNA Modification System and the high-fidelity Cas-CLOVER™ (CC) Site-Specific Gene Editing System. The mRNA coding for hyperactive, or “Super PB” transposase (SPB), and CC enzymes are codelivered with the P-BCMA-ALLO1 PB-based DNA transgene via electroporation to healthy donor T cells to stably integrate the transgene, as well as to knockout (KO) several mediators of allo graft-versus-host and host-versus-graft responses to maximize patient safety and durability of response. The P-BCMA-ALLO1 transgene encodes three genes, a BCMA-specific single-domain variable heavy chain (VH)-CAR (VCAR) gene, a drug selection gene to generate a ~100% CAR+ product, as well as a caspase-based safety switch gene to reduce or eliminate the product in vivo, if desired. The CC System is used to KO the endogenous T Cell Receptor (TCR) and beta-2 microglobulin, thereby decreasing Major Histocompatibility Complex (MHC) class I expression. KO of these key targets is aimed to prevent graft-versus-host disease, as well as reduce host-versus-graft rejection of the product. The CC System can efficiently edit resting T cells, thereby maintaining a high-percentage of TSCM cells, and does not create unwanted off-target mutations, another important consideration when creating an allo product candidate. To maximize the number of doses produced from a single manufacturing run, we have developed a proprietary “booster molecule” that allows for significant expansion of TCR-KO CAR-TSCM cells to potentially produce hundreds of doses. To date, large-scale manufacturing of significant doses of potent allo CAR-T products has been challenging for the field. P-BCMA-ALLO1 manufacturing uses a potentially unlimited number of individual serial donors. We have currently produced P-BCMA-ALLO1 at both research and near-commercial scale from >35 donors with >97% manufacturing success. While a range of TCR-KO efficiencies was observed (~50-90%), the final product was always >99% homozygous TCR-KO after a purification step. Overall expansion of TCR-KO cells ranged from ~2-20 fold, and after removal of unedited TCR+ cells ~0.42-7.04x10e9 TCR-KO cells were recovered from 0.75x10e9 starting cells. However, working at clinical production scale (starting with ~3x10e9 cells), up to 250 doses of P-BCMA-ALLO1 could be manufactured per run, at a dose of 150x10e6 cells/patient. Importantly, with this level of donor and manufacturing robustness, no significant prior screening of donor material, other than to meet standard FDA requirements, would be needed. P-BCMA-ALLO1 made from multiple donors were comprised of an exceptionally high-percentage of the desirable TSCM cells (CD45RA+CD62L+CD45RO-) and had minimal to no expression of exhaustion markers, such as PD-1 or Lag3. Furthermore, P-BCMA-ALLO1 demonstrated potent efficacy in the RPMI-8226 xenograft model in NSG mice across multiple products generated from separate individual healthy donors. Altogether, these data demonstrate a robust, reproducible and highly scalable manufacturing process. Moreover, this manufacturing process can easily be expanded for use with additional CAR targets for treatment of other hematologic or solid tumor malignancies. Disclosures Cranert: Poseida Therapeutics: Employment, Equity Ownership. Richter: Poseida Therapeutics: Employment, Equity Ownership. Tong: Poseida Therapeutics: Employment, Equity Ownership. Weiss: Poseida Therapeutics, Inc.: Employment, Equity Ownership. Tan: Poseida Therapeutics: Employment, Equity Ownership. Ostertag: Poseida Therapeutics: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Coronella: Poseida Therapeutics, Inc: Employment, Equity Ownership. Shedlock: Poseida Therapeutics, Inc.: Employment, Equity Ownership.