Expression of miR-17-92 enhances anti-tumor activity of T-cells transduced with the anti-EGFRvIII chimeric antigen receptor in mice bearing human GBM xenografts

Abstract

BackgroundExpression of miR-17-92 enhances T-cell survival and interferon (IFN)-γ production. We previously reported that miR-17-92 is down-regulated in T-cells derived from glioblastoma (GBM) patients. We hypothesized that transgene-derived co-expression of miR17-92 and chimeric antigen receptor (CAR) in T-cells would improve the efficacy of adoptive transfer therapy against GBM.MethodsWe constructed novel lentiviral vectors for miR-17-92 (FG12-EF1a-miR-17/92) and a CAR consisting of an epidermal growth factor receptor variant III (EGFRvIII)-specific, single-chain variable fragment (scFv) coupled to the T-cell receptor CD3ζ chain signaling module and co-stimulatory motifs of CD137 (4-1BB) and CD28 in tandem (pELNS-3C10-CAR). Human T-cells were transduced with these lentiviral vectors, and their anti-tumor effects were evaluated both in vitro and in vivo.ResultsCAR-transduced T-cells (CAR-T-cells) exhibited potent, antigen-specific, cytotoxic activity against U87 GBM cells that stably express EGFRvIII (U87-EGFRvIII) and, when co-transduced with miR-17-92, exhibited improved survival in the presence of temozolomide (TMZ) compared with CAR-T-cells without miR-17-92 co-transduction. In mice bearing intracranial U87-EGFRvIII xenografts, CAR-T-cells with or without transgene-derived miR-17-92 expression demonstrated similar levels of therapeutic effect without demonstrating any uncontrolled growth of CAR-T-cells. However, when these mice were re-challenged with U87-EGFRvIII cells in their brains, mice receiving co-transduced CAR-T-cells exhibited improved protection compared with mice treated with CAR-T-cells without miR-17-92 co-transduction.ConclusionThese results warrant the development of novel CAR-T-cell strategies that incorporate miR-17-92 to improve therapeutic potency, especially in patients with GBM.

Lenalidomide enhances the function of chimeric antigen receptor T cells against the epidermal growth factor receptor variant III by enhancing immune synapses

“…Gene sequence data for the third-generation CAR was kindly provided by Dr Carl June (University of Pennsylvania, Philadelphia, PA, USA). 21 Using this gene sequence data, 3C10-CAR cDNA was generated by gene synthesis (Genscript, Piscataway, NJ, USA). The mock vector was designed to harbor the scrambled sequence of the scFv portion that has shown no functional activity against glioma, breast cancer, colon cancer and pancreatic cancer cell lines.…”

“…18,19 Further, we constructed human T cells that expressed chimeric antigen receptor (CAR) targeting the EGFRvIII antigen (3C10-CAR). 20,21 CARs are genetically constructed from scFvs connected by a transmembrane hinge region, the T-cell signaling complex, and costimulatory signaling domains (e.g. CD3ζ, CD28, 4-1BB and/or OX-40).…”

Immunotherapy in glioblastoma: emerging options in precision medicine

“…Typically, it can take approximately 3-5 weeks to manufacture clinical-grade modified CAR T cells [78][79][80], depending on the genetic modification method used. The EGFRvIII and IL13Rα2 CAR T-cell therapies have shown efficacy in murine model systems of glioma and CNS melanoma [81][82][83][84], and these strategies, as well as HER2-CAR T cell therapy, are currently being investigated in GBM Phase I clinical trials (NCT02209376, NCT01454596, NCT02208362, NCT02442297, NCT01109095). However, this approach will require a portfolio of CARs and may also have treatment failures owing to antigenic loss/clonotypic selection.…”

Regulation of Memory CD8+ T Cell Differentiation by MicroRNAs

“…At present, it was found that miR-153 inhibits IDO1 expression in cancer cells, and which is a tumor-suppressive miRNA that enhances CAR-T cell immunotherapy for solid tumors [86]. More importantly, the over-expression of the miR-17-92 cluster in CAR-transduced T cells results in prolonged protection against tumor re-challenge in a xenograft model of glioma, indicating the potential of engineered T cells with miRNAs for the treatment of cancer [87]. There are also studies demonstrating better antitumor responses and long-term persistence when both CD4 + and CD8 + CAR-T cells are transferred [88], and it has been proven that one critical role for miRNAs in CD4 + T cells is to help maintain the functional phenotypes of distinct T cell lineages [58].…”