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293 Development of a MUC16-specific CAR-T cell exhibiting potent antitumor activity for the treatment of patients with recurrent ovarian cancer
  1. Nicole A Protheroe1,
  2. Kelly DellaGrotte1,
  3. Harini Kantamneni1,
  4. Sumati Sundaram1,
  5. Scott F Heller1,
  6. Christina Ferren1,
  7. Collin Walter1,
  8. Prashant R Nambiar1,
  9. Corinne Decker2,
  10. Alison Crawford2 and
  11. Tina Chang Albershardt1
  1. 12seventy bio, Cambridge, MA, USA
  2. 2Regeneron Pharmaceuticals, Tarrytown, NY, USA
  • Journal for ImmunoTherapy of Cancer (JITC) preprint. The copyright holder for this preprint are the authors/funders, who have granted JITC permission to display the preprint. All rights reserved. No reuse allowed without permission.


Background Despite significant improvements in surgical and medical treatments, ovarian cancer remains the most lethal gynecologic malignancy, with a 2-to 3-year recurrence rate, highlighting an unmet need for effective therapeutic options for patients.1 Mucin 16 (MUC16) is a type I integral membrane glycoprotein that is overexpressed in more than 80% of ovarian cancers.2 Proteolytic cleavage of full length MUC16 results in the shedding of CA125 into the bloodstream, leading to a truncated ectodomain at the cell surface.3 Here, we report the development of a chimeric antigen receptor (CAR) T cell product (BB4015) targeting the membrane-retained MUC16 ectodomain for the treatment of patients with recurrent ovarian cancer.

Methods in vitro assessment: Human ovarian tumor samples were evaluated for MUC16 expression via immunohistochemistry. BB4015’s phenotypic and functional assessments were evaluated using anti-MUC16 CAR T cells derived from LVV transduction of healthy donor PBMCs. BB4015 phenotype was defined via flow cytometry by CD4, CD8, CD62L, and CD45RA expression. BB4015 function was assessed for antigen-dependent cytokine release/cytotoxicity where BB4015 was co-incubated with antigen positive/negative tumor or primary cells. To determine the off-target binding potential of BB4015’s scFv, we employed a cellular microarray platform (Retrogenix/Charles River) encoding 5,868 full-length human plasma membrane and secreted proteins. in vivo assessment: BB4015 efficacy was evaluated for tumor growth suppression in female NSG mice inoculated with OVCAR3 tumor cells via intraperitoneal injection. Tumor burden and animal weight were measured 3 times a week from tumor-bearing mice infused with BB4015. Live blood draws were taken at various time points following infusion to determine BB4015 pharmacokinetics.

Results 96% of ovarian tumor biopsies evaluated stained positive for MUC16 expression with an antibody raised against the membrane-retained MUC16 ectodomain. BB4015 reactivity to recombinant MUC16 ectodomain elicited specific and dose-dependent cytokine release, with no detectable reactivity to soluble CA125 protein. BB4015’s scFv displayed no observable off-target binding. BB4015 exhibited phenotypic skewing toward a more central memory/naïve-like T cell subtype. This has been attributed to increased antitumor efficacy/greater antigen recall, compared to more effector memory subtypes.4 5 In mice bearing OVCAR3 ovarian tumors, BB4015 infusion inhibited tumor growth in a dose-dependent manner, with complete tumor regression upon the highest dose.

Conclusions BB4015 is a highly reactive anti-MUC16 CAR T cell that exhibits a clean on-target profile and potent anti-tumor activity in ovarian cancer tumor models. Thus, these findings support the clinical evaluation of BB4015 CAR T cells in the treatment of patients with ovarian cancer.


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  2. Aithal A. MUC16 as a novel target for cancer therapy. Expert Opin Ther Targets, 2018;22:675–686.

  3. Das S. Membrane proximal ectodomain cleavage of MUC16 occurs in the acidifyingGolgi/post-Golgi compartments. Sci Rep 2015;5:9759.

  4. Gattinoni L. Wnt signaling arrests effector T cell differentiation and generates CD8+ memory stem cells. Nat Med, 2009;15(7):808–13.

  5. Klebanoff CA. Central memory self/tumor-reactive CD8+ T cells confer superior antitumor immunity compared with effector memory T cells. Proc Natl Acad Sci U S A, 2005;102(27):9571–6.

Ethics Approval All mouse studies were conducted with the approval and oversight of the 2seventy bio-Institutional Animal Care and Use Committee.

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