Harnessing B Cells after Stem Cell Transplant to Produce Novel CAR-Ts

Duke University, Durham NC

Investigators

Abstract

Abstract: Hematological malignancies like leukemias, lymphomas and multiple myeloma are all too often incurable without highly toxic immunotherapy like allogeneic hematopoietic stem cell transplantation (HCT). The advent of less toxic and more targeted, engineered chimeric antigen receptor (CAR) cytotoxic cells has been a major advance. Unfortunately, use of CAR-engineered cells is available only to a small subset of cancer patients whose tumor cells express known antigens. This limitation is due to the inability to produce the tumor-targeting, antigen- binding part of the chimeric receptor. This critical cell surface antigen-targeting part of a CAR, the single chain fragment variable region (or scFv) is not derived from a T cell. The scFv is derived from antibody-producing, B Cell Receptor (BCR)-Activated B cells because antibodies, unlike T Cell Receptors, bind to three-dimensional structures on tumor cell surfaces in an unrestricted manner. Our objective is to leverage what we have learned about the mechanisms of B cell tolerance in patients who develop immune toxicity and have decreased cancer relapsed rates to ask whether anti-tumor B cells can be cloned from patients after HCT to engineer less toxic, more targeted immune cell that eradicate hematological malignancies. Important findings regarding B cells by our group and others over the last 15 years have revealed that host and tumor-reactive B cells survive after HCT when immune tolerance mechanisms are broken in patient with chronic graft versus host disease (cGVHD). Altered immune homeostasis/balance after HCT, donor B cells develop under constant activation of the B Cell Receptor (BCR) by ubiquitous alloantigens (foreign/host and tumor antigens) and excess B Cell Activating Factor (BAFF). Both alloantigen activation of the BCR and BAFF promote survival of B cells that potentially produce polyreactive antibodies. Thus, we hypothesize early after HCT, B cells are primed for production of antibodies with tumor-binding potential. In Aim 1, we will employ HCT- related factors that promote BCR-activated B cells in order to clone cells that would otherwise die via immune tolerance death mechanisms. In Aim 2, we will establish a tumor identification strategy using pooled CRISPR knockdown in a flow cytometry-based assay. Our immediate objective in this R21 proposal is to understand and overcome barriers by harnessing human tumor antigen-responsive, antibody-producing B cells and methods that identify tumor binding receptors (scFV) to engineering novel cancer-eradicating CARs. We will capitalize on what we’ve learned about B cell tolerance after HCT, along with our collaborators at Duke, to establish a pipeline for B cell for rapid cloning of key anti-tumor antibody fragments, scFv, for CAR production.