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Pre-clinical validation of a humanized anti-EGFR variant III chimeric antigen receptor and phase I trial of CART-EGFRvIII in glioblastoma
  1. Laura A Johnson1,
  2. John Scholler2,
  3. Takayuki Ohkuri3,
  4. Akemi Kosaka3,
  5. Prachi R Patel2,
  6. Shannon E McGettigan4,
  7. Arben Nace5,
  8. Pramod Thekkat6,
  9. Andreas Loew7,
  10. Taylor J Chen2,
  11. Joseph A Fraietta1,
  12. Avery D Posey2,
  13. Alina C Boesteanu8,
  14. Alexandria P Cogdill2,
  15. Boris Engels7,
  16. Reshma Singh7,
  17. Tucker R Ezell7,
  18. Neeraja Idamakanti9,
  19. Gabriela Plesa10,
  20. John Seykora2,
  21. Hideho Okada11,
  22. Carl June2,
  23. Jennifer Brogdon7 and
  24. Marcela Maus12
  1. Aff1 grid.25879.310000000419368972Translational Research ProgramUniversity of Pennsylvania Perelman School of Medicine Philadelphia PA USA
  2. Aff2 grid.25879.310000000419368972University of Pennsylvania Philadelphia PA USA
  3. Aff3 grid.21925.3d0000000419369000University of Pittsburgh Pittsburgh PA USA
  4. Aff4 grid.25879.310000000419368972University of Pennsylvania Hatboro PA USA
  5. Aff5 grid.25879.310000000419368972University of Pennsylvania Landenberg PA USA
  6. Aff6 Novartis Institutes of BioMedical Research Inc Quincy MA USA
  7. Aff7 grid.418424.f0000000404392056Novartis Institutes for Biomedical Research Inc Cambridge MA USA
  8. Aff8 University of Pennsylvania Abramson Cancer Center Wilow Grove PA USA
  9. Aff9 grid.418424.f0000 0004 0439 2056Novartis Burlington VT USA
  10. Aff10 grid.25879.310000000419368972University of Pennsylvania Blue Bell PA USA
  11. Aff11 grid.266102.10000000122976811University of California San Francisco San Francisco CA USA
  12. Aff12 grid.25879.310000000419368972Abramson Cancer Center, Dept. of MedicineUniversity of Pennsylvania Perelman School of Medicine Bryn Mawr PA USA

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Meeting abstracts

Chimeric antigen receptors are synthetic molecules designed to re-direct T cells to specific surface antigens; CAR-modified T cells can mediate long-term durable remissions in B cell malignancies, but expanding this platform to solid tumors requires the discovery of novel surface targets with limited expression. The variant III mutation of the epidermal growth factor receptor (EGFR variant III) is the most common variant of the EGF receptor observed in human tumors, and results from an in-frame deletion of a portion of the extracellular domain. In glioblastoma, the EGFRvIII mutation is oncogenic, portends a poor prognosis, and is thought to be enriched in glioblastoma stem cells. However, because the neoepitope of EGFR variant III is based on a small peptide sequence, an antibody or single-chain variable fragment (scFv) directed to this epitope must be rigorously tested to confirm lack of cross-reactivity to the ubiquitously expressed normal EGFR. Having selected a candidate murine scFv directed to EGFRvIII and a vector backbone encoding a second generation CAR, we generated a panel of humanized scFv's and tested their specificity and function as soluble proteins and in the form of CAR-transduced T cells. The lead candidate scFv was tested in vitro for its ability to direct CAR-transduced T cells to kill antigen-bearing targets effectively, and proliferate and secrete cytokines specifically in response to antigen. We further evaluated the specificity of the lead candidate CAR by comparing it to a cetuximab-based CAR which does not discriminate between EGFR and EGFR variant III; the two CARs, along with negative controls, were tested in vitro against primary cells derived from a panel of normal tissues, and in vivo in immunodeficient mice grafted with normal human skin, which naturally expresses EGFR. CAR-T cells were also able to control tumor growth in xenogeneic subcutaneous and orthotopic models of human EGFR variant III+ glioblastoma. We have designed a Phase I clinical study of CAR T cells transduced with humanized scFv directed to EGFR variant III in patients with glioblastoma.