CXCR4 Inhibition Counteracts Immunosuppressive Properties of Metastatic NSCLC Stem Cells

Abstract

Cancer stem cells (CSCs) are functionally defined as the cell subset with greater potential to initiate and propagate tumors. Within the heterogeneous population of lung CSCs, we previously identified highly disseminating CD133+CXCR4+ cells able to initiate distant metastasis (metastasis initiating cells-MICs) and to resist conventional chemotherapy. The establishment of an immunosuppressive microenvironment by tumor cells is crucial to sustain and foster metastasis formation, and CSCs deeply interfere with immune responses against tumors. How lung MICs can elude and educate immune cells surveillance to efficiently complete the metastasis cascade is, however, currently unknown. We show here in primary tumors from non-small cell lung cancer (NSCLC) patients that MICs express higher levels of immunoregulatory molecules compared to tumor bulk, namely PD-L1 and CD73, an ectoenzyme that catalyzes the production of immunosuppressive adenosine, suggesting an enhanced ability of MICs to escape immune responses. To investigate in vitro the immunosuppressive ability of MICs, we derived lung spheroids from cultures of adherent lung cancer cell lines, showing enrichment in CD133+CXCR4+MICs, and increased expression of CD73 and CD38, an enzyme that also concurs in adenosine production. MICs-enriched spheroids release high levels of adenosine and express the immunosuppressive cytokine IL-10, undetectable in an adherent cell counterpart. To prevent dissemination of MICs, we tested peptide R, a novel CXCR4 inhibitor that effectively controls in vitro lung tumor cell migration/invasion. Notably, we observed a decreased expression of CD73, CD38, and IL-10 following CXCR4 inhibition. We also functionally proved that conditioned medium from MICs-enriched spheroids compared to adherent cells has an enhanced ability to suppress CD8+ T cell activity, increase Treg population, and induce the polarization of tumor-associated macrophages (TAMs), which participate in suppression of T cells. Treatment of spheroids with anti-CXCR4 rescued T cell cytotoxic activity and prevented TAM polarization, likely by causing the decrease of adenosine and IL-10 production. Overall, we provide evidence that the subset of lung MICs shows high potential to escape immune control and that inhibition of CXCR4 can impair both MICs dissemination and their immunosuppressive activity, therefore potentially providing a novel therapeutic target in combination therapies to improve efficacy of NSCLC treatment.

The intrinsic role and mechanism of tumor expressed-CD38 on lung adenocarcinoma progression

“…Orazio Fortunato identified that CD133+CXCR4+ lung cancer stem cells were able to initiate distant metastasis and led to a resistance for chemotherapy. They also found that CD133+CXCR4+ cancer stem cells evaded immune surveillance via increasing expression of CD38 and CD73 [ 18 ]. Understanding the essential mechanisms of CD38 employed by tumor cells in tumor microenvironment represents an attractive therapeutic opportunity to selectively target tumor cells.…”

Dual Role of p73 in Cancer Microenvironment and DNA Damage Response

“…The later stages of cancer are associated with well-known cancer hallmarks [ 86 ] including induction of angiogenesis to feed the growth of cancer cells [ 87 , 88 ], activation of metastasis driven by the epithelial-to-mesenchymal transition (EMT) [ 89 , 90 , 91 ], evasion from the immune surveillance, and formation of the tumor microenvironment [ 92 , 93 , 94 , 95 , 96 , 97 ].…”

Tumor-Associated Macrophages Regulate PD-1/PD-L1 Immunosuppression

“…BL-8040 (motixafortide), one of the CXCR4 antagonists, plus the anti-PD-1 pembrolizumab in the COMBAT trial contributes to the improvement found in pancreatic ductal adenocarcinoma (PDAC) patients ( 95 ). After using a new CXCR4 inhibitor peptide R, the expression of CD73, CD38, and IL-10 in non-small cell lung cancer is reduced, which can rescue the cytotoxic activity of T cells and prevent TAM polarization ( 96 ). Another CXCR4 antagonist, Pep R, demonstrated efficacy in combination with nivolumab in melanoma.…”