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347 Development of optimized CAR T cells for therapy of glioblastoma
  1. Jonathan Khan1,
  2. Winson Cai1,
  3. Yanhong Yang1,
  4. Lauren Dong1,
  5. Kelsey Hopland1,
  6. Kenny Kwok-Hei Yu1,
  7. Ruby Freeman1,
  8. Terence Purdon2,
  9. Viviane Tabar1,
  10. Taha Merghoub1,
  11. Jedd Wolchok1 and
  12. Renier Brentjens2
  1. 1Memorial Sloan Kettering Cancer Center, Floral Park, NY, USA
  2. 2Roswell Park Comprehensive Cancer Center, Buffalo, NY, USA


Background Glioblastoma (GBM) is the most lethal form of primary brain tumor in adults, with a 95% five-year mortality rate. Current therapy consists of surgical resection, chemotherapy, and radiotherapy. However, there remains an urgent need for novel therapies as recent trials have failed to improve overall survival.1 A new approach to target GBM is the administration of Chimeric Antigen Receptor (CAR) T cells to treat relapsed/refractory disease.2 To date, multiple clinical trials employing CAR T cells targeting disparate antigens in the context of GBM have been completed; and analysis of data accrued from these trials identified antigen loss and the upregulation of T cell inhibitory pathways to be major limitations of CAR T cell antitumor efficacy.3,4 Accordingly, we have identified B7-H3 as a potential target antigen for CAR T cell therapy of GBM, and the Programmed Death-1/Programmed Death Ligand-1 (PD-1/PD-L1) axis as an inhibitory pathway suitable for combinatorial immunotherapy with CAR T cell therapy of GBM.

Methods B7-H3 expression on U251, U87, and 4 primary GBM cells lines was evaluated by flow cytometry, while patient tumor samples were analyzed by immunofluorescence. B7-H3 targeting CAR T cells were engineered by transducing primary human T cells with a retroviral vector encoding either a CD28-based CAR, or a CD28-based CAR containing a PD-1 blocking scFv. In vitro luciferase-based killing assays, and Luminex-based quantification of cytokine release were used to characterize CAR function. NSG mice were engrafted with U251 GBM cell line and treated with a single peripheral infusion of CAR T cells. Tumors were harvested at various time points following CAR T cell treatment and characterized for T cell infiltrate, T cell phenotype, and expression of checkpoint pathways.

Results We demonstrate that B7-H3 is a suitable antigen for CAR T cell therapy of GBM, and that B7-H3-targeting CAR T cells are able to safely, and efficiently control disease progression in a disease-relevant xenograft model. We also show that GBM tumor cells upregulate PD-L1 in direct response to CAR T cell activity, and that CAR T cells upregulate PD-1 following activation through the CAR. We subsequently demonstrate that concomitant PD-1 blockade augments the antitumor capabilities of B7-H3-targeting CAR T cells through increased T cell engraftment and improved effector function to confer durable, long-term remissions in a disease-relevant preclinical model.

Conclusions These data demonstrate the concomitant PD-1 blockade is safe and efficient method for improving the antitumor capabilities of B7-H3 targeting CAR T cells in the context of GBM.


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  2. Bagley SJ, Desai AS, Linette GP, June CH, O’Rourke, DM. CAR T-cell therapy for glioblastoma: recent clinical advances and future challenges. Neuro Oncol. 2018; 20: 1429–1438.

  3. Brown CE, Badie B, Barish ME, Weng L, Ostberg JR, Chang W-C, Naranjo A, Starr R, Wagner J, Wright C, Zhai Y, Bading JR, Ressler JA, Porgnow J, D’Apuzzo M, Forman SJ, Jensen MC. Bioactivity and safety of IL13R 2-redirected chimeric antigen receptor CD8+ T cells in patients with recurrent glioblastoma. Clin Cancer Res. 2015; 21: 4062–4072.

  4. O’Rourke DM, Nasrallah MP, Desai A, Melenhorst JJ, Mansfield K, Morrissette JJD, Martinez-Lage M, Brem S, Maloney E, Shen A, Isaacs R, Mohan S, Plesa G, Lacey SF, Navenot J-M, Zheng Z, Levine BL, Okada H, June CH, Brogdon JL, Maus MV. A single dose of peripherally infused EGFRvIII-directed CAR T cells mediates antigen loss and induces adaptive resistance in patients with recurrent glioblastoma. Sci Transl Med. 2017; 9: eaaa0984.

Ethics Approval The human biospecimen analyses were approved by Memorial Sloan Kettering Cancer Center IRB #09-156.

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