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.
Author Info: (1) Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892,
USA (2) Genetics Branch, Center for C
Author Info: (1) Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892,
USA (2) Genetics Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA (3) Mouse Cancer Genetics Program, Center for Cancer Research, National Cancer Institute, Frederick, MD 21702, USA (4) Pediatric Oncology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA (5) Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA (6) Cancer Research Technology Program, Leidos Biomedical Research, Inc., Frederick, MD 21702, USA (7) Data Science and Information Technology Program, Leidos Biomedical Research, Frederick, MD 21702, USA (8) Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA (9) Lead contact.
