CD16-158-valine chimeric receptor T cells overcome the resistance of KRAS-mutated colorectal carcinoma cells to cetuximab

KRAS mutations hinder therapeutic efficacy of epidermal growth factor receptor (EGFR)-specific monoclonal antibodies cetuximab and panitumumab-based immunotherapy of EGFR+ cancers. Although cetuximab inhibits KRAS-mutated cancer cell growth in vitro by natural killer (NK) cell-mediated antibody-dependent cellular cytotoxicity (ADCC), KRAS-mutated colorectal carcinoma (CRC) cells escape NK cell immunosurveillance in vivo. To overcome this limitation, we used cetuximab and panitumumab to redirect Fcγ chimeric receptor (CR) T cells against KRAS-mutated HCT116 colorectal cancer (CRC) cells. We compared four polymorphic Fcγ-CR constructs including CD16158F-CR, CD16158V-CR, CD32131H-CR, and CD32131R-CR transduced into T cells by retroviral vectors. Percentages of transduced T cells expressing CD32131H-CR (83.5 ± 9.5) and CD32131R-CR (77.7 ± 13.2) were significantly higher than those expressing with CD16158F-CR (30.3 ± 10.2) and CD16158V-CR (51.7 ± 13.7) (p < 0.003). CD32131R-CR T cells specifically bound soluble cetuximab and panitumumab. However, only CD16158V-CR T cells released high levels of interferon gamma (IFNγ = 1,145.5 pg/ml ±16.5 pg/ml, p < 0.001) and tumor necrosis factor alpha (TNFα = 614 pg/ml ± 21 pg/ml, p < 0.001) upon incubation with cetuximab-opsonized HCT116 cells. Moreover, only CD16158V-CR T cells combined with cetuximab killed HCT116 cells and A549 KRAS-mutated cells in vitro. CD16158V-CR T cells also effectively controlled subcutaneous growth of HCT116 cells in CB17-SCID mice in vivo. Thus, CD16158V-CR T cells combined with cetuximab represent useful reagents to develop innovative EGFR+KRAS-mutated CRC immunotherapies

Generation and characterization of cytotoxic activity against tumor cell lines in human peripheral blood mononuclear cells stimulated "in vitro" by a glucomannan-protein preparation of Candida albicans.

Human peripheral blood mononuclear cells (PBMC) proliferated and generated non-specific cell-mediated cytotoxicity (CMC) after stimulation with a cell-wall glucomannan-protein (GMP) fraction of Candida albicans or chemically-inactivated intact microrganism. No effects were observed using other fungal cell wall components such as glucan or alkali-acid treated glucomannan. The highest CMC level was detected after 7-10 days of PBMC incubation in the presence of 50 micrograms/ml of whole Candida cells and the cytotoxic immunoeffectors elicited by these antigenic stimulations equally affected NK-susceptible (K562) and NK-resistant (Raji, Daudi and Jurkat) tumor cell lines. Both Interleukin-2 (IL-2) and gamma interferon (IFN-gamma) were produced by GMP-stimulated PBMC, the IL-2 peak production constantly preceding that of IFN production. GMP-induced generation of natural CMC was potentiated by the addition of IFN-gamma and a monospecific anti IFN-gamma serum totally abrogated both IFN activity and CMC generation. The cytolytic effectors were shown to be OKT3-, OKT8- and HLA-DR-. They did not possess typical NK markers (e.g. Leu-7 and AB8.28) but were partially recognized by A10, a IgG2a monoclonal antibody reacting to PBMC-NK lymphocytes and activated T cells. These results suggest that the antitumor cytolytic effectors generated in PBMC cultures exposed to Candida material belong either to a discrete subset of natural effectors lacking classical NK markers or to other lymphokine-activated cells. This study also suggests that the human indigenous microrganisms C.albicans may play a role in raising nonspecific antitumor effects in normal host

Induction of potent antitumor CTL responses by recombinant vaccinia encoding a melan-A peptide analogue.

There is considerable interest in the development of vaccination strategies that would elicit strong tumor-specific CTL responses in cancer patients. One strategy consists of using recombinant viruses encoding amino acid sequences corresponding to natural CTL-defined peptide from tumor Ags as immunogens. However, studies with synthetic tumor antigenic peptides have demonstrated that introduction of single amino acid substitutions may dramatically increase their immunogenicity. In this study we have used a well-defined human melanoma tumor Ag system to test the possibility of translating the immunological potency of synthetic tumor antigenic peptide analogues into recombinant vaccinia viruses carrying constructs with the appropriate nucleotide substitutions. Our results indicate that the use of a mutated minigene construct directing the expression of a modified melanoma tumor Ag leads to improved Ag recognition and, more importantly, to enhanced immunogenicity. Thus, recombinant vaccinia viruses containing mutated minigene sequences may lead to new strategies for the induction of strong tumor-specific CTL responses in cancer patients