Multiple Injections of Electroporated Autologous T Cells Expressing a Chimeric Antigen Receptor Mediate Regression of Human Disseminated Tumor

Abstract

Redirecting T lymphocyte antigen specificity by gene transfer can provide large numbers of tumor-reactive T lymphocytes for adoptive immunotherapy. However, safety concerns associated with viral vector production have limited clinical application of T cells expressing chimeric antigen receptors (CAR). T lymphocytes can be gene modified by RNA electroporation without integration-associated safety concerns. To establish a safe platform for adoptive immunotherapy, we first optimized the vector backbone for RNA in vitro transcription to achieve high-level transgene expression. CAR expression and function of RNA-electroporated T cells could be detected up to a week after electroporation. Multiple injections of RNA CAR-electroporated T cells mediated regression of large vascularized flank mesothelioma tumors in NOD/scid/γc(−/−) mice. Dramatic tumor reduction also occurred when the preexisting intraperitoneal human-derived tumors, which had been growing in vivo for >50 days, were treated by multiple injections of autologous human T cells electroporated with anti-mesothelin CAR mRNA. This is the first report using matched patient tumor and lymphocytes showing that autologous T cells from cancer patients can be engineered to provide an effective therapy for a disseminated tumor in a robust preclinical model. Multiple injections of RNA-engineered T cells are a novel approach for adoptive cell transfer, providing flexible platform for the treatment of cancer that may complement the use of retroviral and lentiviral engineered T cells. This approach may increase the therapeutic index of T cells engineered to express powerful activation domains without the associated safety concerns of integrating viral vectors. Cancer Res; 70(22); 9053-61. ©2010 AACR.

Chimeric antigen receptor T cells are vulnerable to immunosuppressive mechanisms present within the tumor microenvironment

“…4 In contrast to prior studies with a different 5,6 tumor outgrowth was delayed but no regressions were seen. One explanation for this finding could be that the tumors underwent a process of "immunoediting" in which they downregulated or lost mesothelin expression allowing for the outgrowth of antigennegative tumor variants.…”

Strength of PD-1 signaling differentially affects T-cell effector functions

“…Before electroporation, T cells were washed three times with Opti-MEM (Invitrogen) and were resuspended in Opti-MEM to a final concentration of 1-2 × 10 8 /mL. Subsequently, 0.1 mL of the cells was mixed with 10 μg (or as otherwise indicated) in vitro-transcribed RNA and was electroporated in a 2-mm cuvette using BTX EM830 (Harvard Apparatus BTX) as previously described (56).…”

The Flipside of the Power of Engineered T Cells: Observed and Potential Toxicities of Genetically Modified T Cells as Therapy

“…16 However, other studies targeting Her2 with lower doses and different CAR constructs, based on either a lower-affinity scFv or less potent signaling domains, show that Her2-targeted CAR T cells may be safe. [17][18][19] Some antigens that are expressed on normal tissues may still be targetable by CARs without causing dose-limiting toxicity, such as mesothelin and prostate-specific membrane antigen (PSMA) (S.F. Slovin et al, 2013, J Clin Oncol., abstract); 20 however, neither of these targets has brought about dramatic clinical efficacy, and it is possible that these two outcomes are inexorably linked.…”