Memory T cells are uniquely resistant to melanoma-induced suppression.

We have previously observed that in vivo exposure to growing melanoma tumors fundamentally alters activated T cell homeostasis by suppressing the ability of naïve T cells to undergo antigen-driven proliferative expansion. We hypothesized that exposure of T cells in later stages of differentiation to melanoma would have similar suppressive consequences. C57BL/6 mice were inoculated with media or syngeneic B16F10 melanoma tumors 8 or 60 days after infection with lymphocytic choriomeningitis virus (LCMV), and splenic populations of LCMV-specific T cells were quantified using flow cytometry 18 days after tumor inoculation. Inoculation with melanoma on post-infection day 8 potentiated the contraction of previously activated T cells. This enhanced contraction was associated with increased apoptotic susceptibility among T cells from tumor-bearing mice. In contrast, inoculation with melanoma on post-infection day 60 did not affect the ability of previously established memory T cells to maintain themselves in stable numbers. In addition, the ability of previously established memory T cells to respond to LCMV challenge was unaffected by melanoma. Following adoptive transfer into melanoma-bearing mice, tumor-specific memory T cells were significantly more effective at controlling melanoma growth than equivalent numbers of tumor-specific effector T cells. These observations suggest that memory T cells are uniquely resistant to suppressive influences exerted by melanoma on activated T cell homeostasis; these findings may have implications for T cell-based cancer immunotherapy.

Melanoma-induced suppression of tumor antigen-specific T cell expansion is comparable to suppression of global T cell expansion.

We have observed that in vivo interaction between melanoma and resting T cells promotes suppression of antigen-driven proliferative T cell expansion. We hypothesized that this suppression would affect tumor antigen-specific T cell populations more potently than tumor-unrelated T cell populations. A B16F10 cell line was stably transfected to express low levels of the lymphocytic choriomeningitis virus (LCMV) glycoprotein GP33 (B16GP33). Mice bearing B16F10 or B16GP33 tumors were infected with LCMV, and proliferative expansion of LCMV epitope-specific T cell populations was quantified. In vitro and in vivo assays confirmed low levels of antigenic GP33 expression by B16GP33 tumors. Suppressed expansion of GP33-specific T cells was equivalent between mice bearing B16F10 and B16GP33 tumors. These observations suggest that the ability of growing melanoma tumors to impair antigen-driven proliferative expansion of activated T cells is global and not antigen-specific, and provide further insight into the influence of cancer on activated T cell homeostasis.

Suppression of T-cell expansion by melanoma is exerted on resting cells.

BACKGROUND: Immunotherapeutic cancer protocols often rely on the ability to promote proliferative expansion of tumor-specific T-cell, but the influence of cancer on in vivo T-cell expansion remains largely undefined. METHODS: The ability of control and B16F10 melanoma-bearing C57BL/6 mice to expand lymphocytic choriomeningitis virus antigen-specific T-cell populations in response to acute viral infection was compared by using flow cytometric assays of splenocytes. RESULTS: The ability to expand virus-specific CD8+ and CD4+ T-cells was globally and markedly suppressed in tumor-bearing mice. Expanded cytotoxic T lymphocytes (CTLs) retained in vivo and in vitro functionality, suggesting that melanoma growth did not induce T-cell anergy. The magnitude of suppressed proliferative expansion was proportional to the extent of tumor burden. Melanoma-induced suppression of CTL expansion was correlated with upregulated apoptotic activity and hampered the induction of memory precursor effector cells. Adoptive transfer of resting LCMV antigen-specific T-cells before or after tumor establishment demonstrated that a critical period of in vivo exposure of resting T-cells to growing melanoma was responsible for the induction of suppressed expansion. This suppression was durable; surgical resection of melanoma after in vivo exposure to resting T-cells but before antigenic stimulation did not restore full expansion. CONCLUSIONS: These data suggest that growing melanoma tumors exert a global, antigen-independent influence on resting T-cells that fundamentally reprograms their ability to undergo proliferative expansion in response to subsequent antigenic stimulation. This finding may have direct implications for T-cell-based immunotherapeutic strategies.

Down-regulation of Notch1 signaling inhibits tumor growth in human hepatocellular carcinoma.

In some human cancers, the Notch1 receptor and signaling pathway is upregulated, which increases the oncogenic potential of the cell through prevention of differentiation and inhibition of apoptosis. We sought to evaluate the role of Notch1 in hepatocellular cancer (HCC), and evaluate the therapeutic efficacy of curcumin, a known Notch1 inhibitor. Human liver tumors were compared to normal liver to evaluate for Notch1 and Notch1 Intracellular Domain (NICD). Three human HCC cell lines were exposed to curcumin and evaluated for downstream effectors by Western blot. In addition, standard MTT assays were performed to assess the effect of curcumin in vitro. Finally, a nude mouse xenograft model was utilized to assess the response to curcumin in vivo. High levels of NICD were present in the majority of human HCC samples and all three HCC cell lines. Treatment with curcumin led to a dose-dependent decrease in the expression of NICD associated with the induction of cleaved poly ADP-ribose polymerase (PARP), the degradation of cyclin D1 and increase in cyclin-dependent kinase p21. Curcumin inhibited HCC cell proliferation in vitro. Importantly, transfection of Notch1 small-interfering RNA (siRNA) into HCC cells resulted in cell growth inhibition and apoptosis, recapitulating the effects of curcumin. Finally, treatment with curcumin resulted in a 40% decrease in tumor growth in vivo. These results suggest for the first time that down-regulation of Notch1 signaling with curcumin is an attractive new therapeutic strategy for the treatment of patients with HCC.