Development of novel chimeric receptors for delivery of costimulation to tumor-reactive engineered T cells

Abstract

Introduction: Manipulation of the immune system to eliminate cancer, known as cancer immunotherapy, is an emerging field that has shown impressive clinical success and promise. Adoptive transfer of T cells engineered for tumor reactivity is an avenue of therapy for patients with previously untreatable disease. Our lab has developed a novel chimeric receptor, called a T cell antigen coupler (TAC), which redirects T cell cytotoxicity towards a tumor target. Although considerably effective, this receptor does not provide T cell costimulation necessary for optimal anti-tumor effectiveness. Methods: We explored two methods to deliver costimulation to TAC-engineered T cells. First, we designed a receptor to be utilized in conjunction with the TAC in a dual receptor system. This chimeric costimulatory receptor (CCR) was generated by fusion of the T cell TIGIT and CD28 receptors. In our second approach, we investigated direct incorporation of costimulatory domains into the TAC design. To do so, we substituted in regions from the CD28 or 4-1BB costimulatory receptors. Results: Three TIGIT/CD28 chimeras were successfully generated. Of these, two were well surface-expressed on primary human T cells. Despite testing of these receptors in several biological assays, we were unable to confirm functionality of these receptors in transmitting CD28 signals. We next generated the 4-1BB and CD28TAC variants. The 4-1BBTAC was poorly surface-expressed M.Sc Thesis – Arya Afsahi McMaster University – Medical Sciences iv and was difficult to introduce into T cells at high efficiency. The CD28TAC-variant was virtually absent from the T cell surface membrane. Further analysis indicated that the CD28TAC was retained in the endoplasmic reticulum (ER) and the 4-1BBTAC was produced at an extremely low amount. Conclusions: Our investigation into delivery of costimulation through a novel CCR or TAC receptor was inconclusive. We recommend several optimizations to both receptor design and experimental analysis to further elucidate the potential of these receptors.