ADAM protease inhibition overcomes resistance of breast cancer stem-like cells to γδ T cell immunotherapy

Abstract Breast cancer stem cells (BCSC) are highly resistant to current therapies, and are responsible for metastatic burden and relapse. Gamma delta T cells (γδTc) are immunosurveillance cells with tremendous anti-tumoral activity, and a growing number of clinical trials have confirmed the safety of γδTc immunotherapy for various malignancies. Herein, we demonstrate that γδTc can kill BCSC, but to a lesser extent than non-cancer stem cells (NSC). Immune evasion was orchestrated by several mechanisms. The BCSC secretome rendered γδTc hypo-responsive by reducing proliferation, cytotoxicity and IFN-γ production, while increasing expression of co-inhibitory receptors on γδTc. BCSC and target cells surviving γδTc cytotoxicity had higher PD-L1 co-inhibitory ligand expression, and blocking PD-1 on γδTc significantly overcame BCSC resistance to γδTc killing. Fas/FasL signaling was dysfunctional in BCSC due to upregulation of the anti-apoptotic protein MCL-1, which could be partially overcome using dMCL1-2, an MCL-1 degrader. Moreover, the BCSC fraction shed higher levels of the NKG2D ligand MICA compared to NSC. Inhibiting MICA shedding using the ADAM inhibitor GW280264X overcame BCSC resistance to γδTc killing, rendering BCSC as sensitive to γδTc cytotoxicity as NSC. Collectively, our data unravel multiple mechanisms exploited by BCSC to evade γδTc killing, which may also come into play in BCSC resistance to other cytotoxic lymphocytes. Developing strategies to overcome this resistance will increase the efficacy of cancer immunotherapy and lead to improved outcomes for cancer patients. One Sentence SummaryBreast cancer stem-like cells are resistant to γδ T cell targeting, which can be overcome by inhibiting ADAM proteases that facilitate MICA/B shedding.