CD4 Th1 Cytokines and HER-2/HER-3 Blockade Induces Tumor Senescence in Breast Cancer

ROSEMBLIT C; CZERNIECKI BJ

San Antonio, Texas

Simposio; San Antonio Breast Cancer Symposium; 2014

American Association for Cancer Research (AACR)

Background: HER-2 a molecular oncodriver in breast tumorigenesis is over expressed in 25% of human breast cancers, and its expression correlates with enhanced tumor aggressiveness. While targeted therapies have improved outcomes many patients become resistant or recur. We have recently established a progressive loss in anti-HER-2 CD4 Th1 responses during disease progression and is associated with outcomes. The pleiotropic Th1 cytokines IFN-γ and TNF-α have diverse effects on tumor epithelial cells. In this study we sought to determine whether these Th1 cytokines induce senescence of HER-2 expressing breast cancer cells and assess the impact of IFN-γ and TNF-α with simultaneous HER-2 and HER-3 blockade. Results: All breast cancer cell lines tested express IFN-γ and TNF-α receptors measured by western blot analysis. The high and intermediate HER-2 expressing cells are sensitive to tumor senescence induction when treated with combinations of IFN-γ and TNF-α in a dose dependent manner. Low HER-2 expressing cells were less sensitive to senescence induction as measured by positive b-galactosidase activity and the expression of p15INK4b and p16INK4a by western blot. Addition of IFN-γ and TNF-α treatment to HER-2-depleted cells by RNAi resulted in an increase senescent phenotype and was increased further when the cells were double HER-2- and HER-3-depleted.  To determine whether CD4 Th1 mediated effects on high HER-2 human breast cancer cell lines, we co-cultured increasing number of HER-2 antigen-primed CD4+ T cells with high HER-2 human breast cancer cells in a transwell cell culture system. This resulted in a dose-dependent senescence of breast cancer cells compared with CD4+ T cells primed either with immature dendritic cells (DC) or mature DC plus irrelevant (Class II BRAF) peptide.  In addition, SK-BR-3 breast cancer cells incubated with the supernatant of the CD4+ T cells-immature DC or mature DC co-culture demonstrated similar results. CD4+ Th1-elaborated cytokines IFN-γ and TNF-α in the supernatants were confirmed using ELISA. Blocking antibodies against IFN-γ and TNF-α demonstrated reduced senescence induction. Conclusions: Our results establish a role for IFN-γ and TNF-α in inducing tumor senescence in breast cancer. An effective CD4 Th1 responses combined with HER-2 and HER-3 blockade can significantly drive tumor senescence in breast cancer that can be explored as treatment to effectively eliminate residual breast cancer cells and prevent recurrence.