ABSTRACT
Background: Boiling histotripsy (BH), a mechanical focused ultrasound ablation strategy, can elicit intriguing signatures of anti-tumor immunity. However, the influence of BH on dendritic cell function is unknown, compromising our ability to optimally combine BH with immunotherapies to control metastatic disease. Methods: BH was applied using a sparse scan (1 mm spacing between sonications) protocol to B16F10-ZsGreen melanoma in bilateral and unilateral settings. Ipsilateral and contralateral tumor growth was measured. Flow cytometry was used to track ZsGreen antigen and assess how BH drives dendritic cell behavior. Results: BH monotherapy elicited ipsilateral and abscopal tumor control in this highly aggressive model. Tumor antigen presence in immune cells in the tumor-draining lymph nodes (TDLNs) was ~3-fold greater at 24h after BH, but this abated by 96h. B cells, macrophages, monocytes, granulocytes, and both conventional dendritic cell subsets (i.e. cDC1s and cDC2s) acquired markedly more antigen with BH. BH drove activation of both cDC subsets, with activation being dependent upon tumor antigen acquisition. Our data also suggest that BH-liberated tumor antigen is complexed with damage-associated molecular patterns (DAMPs) and that cDCs do not traffic to the TDLN with antigen. Rather, they acquire antigen as it flows through afferent lymph vessels into the TDLN. Conclusion: When applied with a sparse scan protocol, BH monotherapy elicits abscopal melanoma control and shapes dendritic cell function through several previously unappreciated mechanisms. These results offer new insight into how to best combine BH with immunotherapies for the treatment of metastatic melanoma.
Subject(s)
High-Intensity Focused Ultrasound Ablation , Melanoma , Humans , Melanoma/therapy , High-Intensity Focused Ultrasound Ablation/methods , Antigens, Neoplasm , Dendritic CellsABSTRACT
Background: Boiling histotripsy (BH), a mechanical focused ultrasound ablation strategy, can elicit intriguing signatures of anti-tumor immunity. However, the influence of BH on dendritic cell function is unknown, compromising our ability to optimally combine BH with immunotherapies to control metastatic disease. Methods: BH was applied using a sparse scan (1 mm spacing between sonications) protocol to B16F10-ZsGreen melanoma in bilateral and unilateral settings. Ipsilateral and contralateral tumor growth was measured. Flow cytometry was used to track ZsGreen antigen and assess how BH drives dendritic cell behavior. Results: BH monotherapy elicited ipsilateral and abscopal tumor control in this highly aggressive model. Tumor antigen presence in immune cells in the tumor-draining lymph nodes (TDLNs) was ~3-fold greater at 24h after BH, but this abated by 96h. B cells, macrophages, monocytes, granulocytes, and both conventional dendritic cell subsets (i.e. cDC1s and cDC2s) acquired markedly more antigen with BH. BH drove activation of both cDC subsets, with activation being dependent upon tumor antigen acquisition. Our data also suggest that BH-liberated tumor antigen is complexed with damage-associated molecular patterns (DAMPs) and that cDCs do not traffic to the TDLN with antigen. Rather, they acquire antigen as it flows through afferent lymph vessels into the TDLN. Conclusion: When applied with a sparse scan protocol, BH monotherapy elicits abscopal melanoma control and shapes dendritic cell function through several previously unappreciated mechanisms. These results offer new insight into how to best combine BH with immunotherapies for the treatment of metastatic melanoma.
ABSTRACT
BACKGROUND: Triple-negative breast cancer (TNBC) remains recalcitrant to most targeted therapy approaches. However, recent clinical studies suggest that inducing tumor damage can render TNBC responsive to immunotherapy. We therefore tested a strategy for immune sensitization of murine TNBC (4T1 tumors) through combination of focused ultrasound (FUS) thermal ablation and a chemotherapy, gemcitabine (GEM), known to attenuate myeloid-derived suppressor cells (MDSCs). METHODS: We applied a sparse-scan thermally ablative FUS regimen at the tumor site in combination with systemically administered GEM. We used flow cytometry analysis to investigate the roles of monotherapy and combinatorial therapy in mediating local and systemic immunity. We also tested this combination in Rag1-/- mice or T cell-depleted wild-type mice to determine the essentiality of adaptive immunity. Further, we layered Programmed cell death protein 1 (PD-1) blockade onto this combination to evaluate its impact on tumor outgrowth and survival. RESULTS: The immune-modulatory effect of FUS monotherapy was insufficient to promote a robust T cell response against 4T1, consistent with the dominant MDSC-driven immunosuppression evident in this model. The combination of FUS+GEM significantly constrained primary TNBC tumor outgrowth and extended overall survival of mice. Tumor control correlated with increased circulating antigen-experienced T cells and was entirely dependent on T cell-mediated immunity. The ability of FUS+GEM to control primary tumor outgrowth was moderately enhanced by either neoadjuvant or adjuvant treatment with anti-PD-1. CONCLUSION: Thermally ablative FUS in combination with GEM restricts primary tumor outgrowth, improves survival and enhances immunogenicity in a murine metastatic TNBC model. This treatment strategy promises a novel option for potentiating the role of FUS in immunotherapy of metastatic TNBC and is worthy of future clinical evaluation. TRIAL REGISTRATION NUMBERS: NCT03237572 and NCT04116320.
