Abstract 6328: Preclinical development of CAR T cells with antigen-inducible IL-18 enforcement to treat GD2-positive solid cancers
Authors: Fischer-Riepe Lena , Kailayangiri Sareetha , Zimmermann Katharina , Pfeifer Rita , Aigner Michael , Altvater Bianca , Kretschmann Sascha , Voelkl Simon , Hartley Jordan , Dreger Celine , Petry Katja , Bosio Andreas , von Doellen Angelika , Hartmann Wolfgang , Lode Holger , Goerlich Dennis , Mackensen Andreas , Jungblut Melanie , Schambach Axel , Abken Hinrich , Rossig Claudia 
Published: March 2024

Cytokine-engineering is a promising strategy to overcome limitations of CAR T cell therapy in solid tumors. We have developed a novel investigational medicinal product, consisting of CAR T cells directed against ganglioside GD2 along with CAR-inducible secretion of IL-18, to enforce their activation response and effector functions in the microenvironment of solid cancers. Our lentiviral all-in-one vector combines constitutive expression of a 14.G2a antibody-derived GD2-specific, 4-1BB-costimulated CAR with inducible nuclear factor of activated T cells (NFAT)-driven expression of the human IL-18 transgene. A control construct contains EGFP in the place of IL-18 (GD2EGFPCART). Lentiviral transduction of T cells from various individual donors with both preclinical and clinical batches of virus resulted in CAR surface expression in &gt;40% of T cells. GD2IL18CART contained effector memory and central memory T cell phenotypes at similar frequencies as the control. CAR engagement by immobilized anti-idiotype antibody or by GD2-positive tumor cells selectively induced IL-18 release, with production of IL-18 strictly depending on GD2 antigen engagement. GD2IL18CART responded to interaction with GD2-positive tumor cells with superior IFN-γ, TNF-α and IL-2 cytokine release and superior target cytolysis compared to the control CAR T cells without inducible IL-18, whereas IL-6 was secreted at similar concentrations. In an NSG xenograft model of disseminated GD2-positive tumors, GD2IL18CART had superior in vivo anti-tumor activity than GD2EGFPCART, with eradication of GD2-positive tumor xenografts as detected by bioluminescence imaging while the control achieved only limited extension of progression-free survival. Finally, we established GMP-compliant manufacturing of GD2IL18CART that is feasible and efficient at clinical scale, using an automated protocol and device. Three validation runs confirmed robust expansion of CAR+ T cells to final cell numbers of 3.5x109 CAR+ cells (median; range 3.4 to 3.7x109), with consistent transduction efficiencies of &gt;54% at the in-process control time-point and at harvest. GD2IL18CART from clinical-scale manufacturing runs responded to interaction with GD2 with strictly antigen-dependent secretion of both IL-18 and IFN-γ and efficiently lysed various GD2-positive tumor targets. In conclusion, we assembled all prerequisites for clinical evaluation of an IL-18 cytokine enhanced, GD2-specific CAR T cell product for the treatment of patients with a variety of solid tumors, recruited on the basis of GD2 immunofluorescence pre-screening (EU CT 2022-501725-21-00). Besides addressing the safety of GD2IL18CART therapy in pediatric and adult patients as the primary endpoint, the study will allow to gain substantial knowledge on the biological effects of IL-18 in the TME of the individual solid tumors.

Citation Format: Lena Fischer-Riepe, Sareetha Kailayangiri, Katharina Zimmermann, Rita Pfeifer, Michael Aigner, Bianca Altvater, Sascha Kretschmann, Simon Voelkl, Jordan Hartley, Celine Dreger, Katja Petry, Andreas Bosio, Angelika von Doellen, Wolfgang Hartmann, Holger Lode, Dennis Goerlich, Andreas Mackensen, Melanie Jungblut, Axel Schambach, Hinrich Abken, Claudia Rossig. Preclinical development of CAR T cells with antigen-inducible IL-18 enforcement to treat GD2-positive solid cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6328.