P-07.05 Overcoming endothelial cell anergy by VEGFR2 inhibition to enhance CAR T cell response in aggressive B cell lymphoma

<h3>Background</h3> CAR T cell therapy has greatly improved outcomes for patients suffering from relapsed and/or refractory B cell lymphoma.¹ However, resistance and CAR T cell failure occurs, leading to decreased treatment efficacy and non-durable response.² Resistance against CAR T cell therapy can occur due to endothelial cell anergy - a tumor induced phenomenon mediated by VEGF-A - VEGFR2 interaction, in which permeability of the endothelium is drastically decreased. Anergic endothelial cells form a physical barrier between tumor cells and circulating CAR T cells, keeping them confined to the vascular system and thereby, leading to reduced efficacy of CAR T cells.^{3 4} To overcome endothelial cell anergy and thereby, resistance against CAR T cell therapy, we hypothesized that combined VEGFR2 inhibition through PTK787 and CAR T cells displays synergistic treatment effects. <h3>Materials and Methods</h3> To elucidate mechanisms that are associated with CAR T-cell resistance in patients with aggressive B-cell lymphoma, we applied multi-omic analyses to pre- and post CAR-T-cell treatment lymphoma specimens. In our <i>in vitro</i> studies, we used methods such as flow cytometry, adhesion assays and IF staining. For in vivo experiments we worked with an autochthonous DLBCL mouse model, using VEGFR2 inhibitor PTK787 and CD19 CAR T cells. After harvesting the mice, we performed immunohistochemistry and IF stainings on spleens. <h3>Results</h3> Upon performing bulk-RNA sequencing on biopsies of lymphoma patients before undergoing CAR T cell therapy, we observed that high VEGF-A levels in pre-CAR T cell specimens associate with non-durable response to CAR T cell therapy and that markers of endothelial cell anergy correlate with worse prognosis. Our <i>in vitro</i> data revealed an upregulation of endothelial adhesion molecules upon VEGFR2 inhibition, leading to increased CAR T cell adhesion. Strikingly, in an autochthonous aggressive B cell lymphoma mouse model we demonstrate a significantly prolonged overall and progression free survival of mice treated with VEGFR2 inhibition in combination with murine CD19-CAR T cells. <h3>Conclusions</h3> To conclude, our <i>in vivo</i> results strongly suggest that combining CD19 directed CAR T cells and VEGFR2 inhibition is a promising strategy to improve treatment efficacy of CAR T cell therapy, potentially increasing outcome of patients with aggressive B cell lymphoma. <h3>References</h3> Locke FL, Miklos DB, Jacobson CA, Perales MA, Kersten MJ, Oluwole OO, <i>et al</i>. Axicabtagene ciloleucel as second-line therapy for large b-cell lymphoma. <i>N Engl J Med.</i> 2022 Feb 17;<b>386</b>(7):640–54. Neelapu SS, Jacobson CA, Ghobadi A, Miklos DB, Lekakis LJ, Oluwole OO, <i>et al</i>. Five-year follow-up of ZUMA-1 supports the curative potential of axicabtagene ciloleucel in refractory large B-cell lymphoma. <i>Blood</i> 2023 May 11;<b>141</b>(19):2307–15. Huinen ZR, Huijbers EJM, van Beijnum JR, Nowak-Sliwinska P, Griffioen AW. Anti-angiogenic agents — overcoming tumour endothelial cell anergy and improving immunotherapy outcomes. <i>Nat Rev Clin Oncol.</i> 2021 Aug;<b>18</b>(8):527–40. Flati V, Pastore LI, Griffioen AW, Satijn S, Toniato E, D’Alimonte I, <i>et al</i>. Endothelial cell anergy is mediated by bFGF through the sustained activation of p38-MAPK and NF-kappaB inhibition. <i>Int J Immunopathol Pharmacol.</i> 2006 Oct-Dx;<b>19</b>(4):761–73. L. Schlözer: None. N. Neumaier: None. D. Stahl: None. T. Riet: None. M. Nill: None. R. Flümann: None. C. Reinhardt: None. N. Abedpour: None. J. Brägelmann: None. M. Müller: None. A. Florin: None. P. Borchmann: None. R. Ullrich: None.