CAR T-cells that target acute B-lineage leukemia irrespective of CD19 expression
Kristen Fousek, Junji Watanabe, Sujith K. Joseph, Ann George, Xingyue An, Tiara T. Byrd, Jessica S. Morris, Annie Luong, Melisa Martinez-Paniagua, Khaled Sanber, Shoba A. Navai, Ahmed Z. Gad, Vita S. Salsman, Pretty Mathew, Hye Na Kim, Dimitrios L. Wagner, Lorenzo Brunetti, Albert Jang, Matthew L. Baker, Navin Varadarajan, Meenakshi Hegde, Yong‐Mi Kim, Nora Heisterkamp, Hisham Abdel‐Azim, Nabil Ahmed
Abstract
Chimeric antigen receptor (CAR) T-cells targeting CD19 demonstrate remarkable efficacy in treating B-lineage acute lymphoblastic leukemia (BL-ALL), yet up to 39% of treated patients relapse with CD19(-) disease. We report that CD19(-) escape is associated with downregulation, but preservation, of targetable expression of CD20 and CD22. Accordingly, we reasoned that broadening the spectrum of CD19CAR T-cells to include both CD20 and CD22 would enable them to target CD19(-) escape BL-ALL while preserving their upfront efficacy. We created a CD19/20/22-targeting CAR T-cell by coexpressing individual CAR molecules on a single T-cell using one tricistronic transgene. CD19/20/22CAR T-cells killed CD19(-) blasts from patients who relapsed after CD19CAR T-cell therapy and CRISPR/Cas9 CD19 knockout primary BL-ALL both in vitro and in an animal model, while CD19CAR T-cells were ineffective. At the subcellular level, CD19/20/22CAR T-cells formed dense immune synapses with target cells that mediated effective cytolytic complex formation, were efficient serial killers in single-cell tracking studies, and were as efficacious as CD19CAR T-cells against primary CD19(+) disease. In conclusion, independent of CD19 expression, CD19/20/22CAR T-cells could be used as salvage or front-line CAR therapy for patients with recalcitrant disease.