Using RNAseq, Li et al. determined that exon 3 of glypican 2 (GPC2) is highly expressed in neuroblastoma (NB) and the least expressed exon in normal tissue. They generated a monoclonal antibody (CT3) with high affinity for a tumor-associated GPC2 epitope expressed on 95% of NB tissue and encoded by both exon 3 and exon 10. There was little to no binding of CT3 to normal tissue or other glypicans. CT3 CAR T cells killed GPC2+ tumor cells in vitro. In NOD/SCID/IL-2Rgcnull (immuno-incompetent) mice bearing a human NB tumor cell line, CT3 CAR T cells were selectively expanded and caused tumor regression.
Contributed by Margot O’Toole
ABSTRACT: Targeting solid tumors must overcome several major obstacles, in particular, the identification of elusive tumor-specific antigens. Here, we devise a strategy to help identify tumor-specific epitopes. Glypican 2 (GPC2) is overexpressed in neuroblastoma. Using RNA sequencing (RNA-seq) analysis, we show that exon 3 and exons 7–10 of GPC2 are expressed in cancer but are minimally expressed in normal tissues. Accordingly, we discover a monoclonal antibody (CT3) that binds exons 3 and 10 and visualize the complex structure of CT3 and GPC2 by electron microscopy. The potential of this approach is exemplified by designing CT3-derived chimeric antigen receptor (CAR) T cells that regress neuroblastoma in mice. Genomic sequencing of T cells recovered from mice reveals the CAR integration sites that may contribute to CAR T cell proliferation and persistence. These studies demonstrate how RNA-seq data can be exploited to help identify tumor-associated exons that can be targeted by CAR T cell therapies.