IMMU-04. DEVELOPMENT OF GPC2-DIRECTED CHIMERIC ANTIGEN RECEPTOR THERAPY FOR PEDIATRIC BRAIN TUMORS WITH IN VITRO TRANSCRIBED mRNA
Year: 2019
Type: conference-abstract
Abstract: The recent identification of glypican-2 (GPC2) as a cell surface oncoprotein in neuroblastoma, high grade glioma (HGG), and medulloblastoma provides the opportunity for the development of targeted immunotherapy. We hypothesized that chimeric antigen receptor (CAR) T cell therapy directed against GPC2 could be achieved using in vitro transcribed mRNA, allowing for rapid and safe iterative trials to test novel CAR constructs. We created mRNA encoding four different CAR proteins using a GPC2-specific scFv paired with 4-1BB and CD3ζ co-stimulatory domains, with varied heavy and light chain orientation and linker length between chains. After electroporation of CAR encoding mRNA into human T cells, we evaluated for CAR persistence, markers of T cell exhaustion, and cellular cytotoxicity against four primary and two isogenic neuroblastoma cell lines and three primary HGG cell lines. All four constructs showed >80% CAR expression and GPC2-specific binding by flow cytometry. CAR molecules in the light-to-heavy (VL-VH) configuration showed persistence on the surface over seven days and increased cytotoxicity compared to heavy-to-light (VH-VL) configuration. VH-VL configuration with a long linker provided the weakest cytotoxic effect, and evaluation of negative checkpoint regulators revealed the highest expression of PD1 and Lag3 (62% vs. 17–40% in other constructs, p<0.0001). Based on the in vitro data, the two VL-VH CAR constructs were chosen for testing in murine flank models of neuroblastoma treated with IV GPC2 CAR T cells once weekly for three doses. At day 14, animals treated with both VL-VH CAR constructs had reduced tumor burden compared to CD19 CAR controls (p<0.01), with several animals showing complete response. Studies are currently underway evaluating efficacy in orthotopic models of pediatric HGG using local delivery. These data highlight the utility of using mRNA to efficiently design and test novel CAR T cells, providing a platform for clinical testing for proof of efficacy and screening for toxicity.
Source: Neuro-Oncology
Institutions: Children's Hospital of Philadelphia, University of Pittsburgh
Language: English
Topic: CAR-T cell therapy research
Subfield: Oncology
Field: Medicine
Domain: Health Sciences
Open Access status: bronze
OpenAlex ID: W2942087142