Boiling histotripsy and in-situ CD40 stimulation improve the checkpoint blockade therapy of poorly immunogenic tumors

Abstract

Background: Advanced stage cancers with a suppressive tumor microenvironment (TME) are often refractory to immune checkpoint inhibitor (ICI) therapy. Recent studies have shown that focused ultrasound (FUS) TME-modulation can synergize ICI therapy, but enhancing survival outcomes in poorly immunogenic tumors remains challenging. Here, we investigated the role of focused ultrasound based boiling histotripsy (HT) and in-situ anti-CD40 agonist antibody (αCD40) combinatorial therapy in enhancing therapeutic efficacy against ICI refractory murine melanoma. Methods: Unilateral and bilateral large (~330-400 mm 3 ) poorly immunogenic B16F10 melanoma tumors were established in the flank regions of mice. Tumors were exposed to single local HT followed by an in-situ administration of αCD40 (HT+ αCD40: HT40). Inflammatory signatures post treatment were assessed using pan-cancer immune profiling and flow cytometry. The ability of HT40 ± ICI to enhance local and systemic effects was determined by immunological characterization of the harvested tissues, and by tumor growth delay of local and distant untreated tumors 4-6 weeks post treatment. Results: Immune profiling revealed that HT40 upregulated a variety of inflammatory markers in the tumors. Immunologically, HT40 treated tumors showed an increased population of granzyme B+ expressing functional CD8+ T cells (~4-fold) as well as an increased M1 to M2 macrophage ratio (~2-3-fold) and CD8+ T: regulatory T cell ratio (~5-fold) compared to the untreated control. Systemically, the proliferation rates of the melanoma-specific memory T cell population were significantly enhanced by HT40 treatment. Finally, the combination of HT40 and ICI therapy (anti-CTLA-4 and anti-PD-L1) caused superior inhibition of distant untreated tumors, and prolonged survival rates compared to the control. Conclusions: Data suggest that HT40 reprograms immunologically cold tumors and sensitizes them to ICI therapy. This approach may be clinically useful for treating advanced stage melanoma cancers.

Pulsed focused ultrasound can improve the anti-cancer effects of immune checkpoint inhibitors in murine pancreatic cancer

“…These data suggested that pHIFU exposures designed to induce mechanical damage in tissue in the absence of significant thermal change may be better equipped to create a pro-inflammatory environment than thermally ablative continuous HIFU treatments which are likely to heat fix target tissue, thus limiting intratumoral antigen release [ 16 , 19 ]. In order to facilitate clinical translation, we have used lower peak negative pressures and acoustic powers than those used in histotripsy studies [ 19 , 20 ]. Here ‘histotripsy-like’ pHIFU parameters ( P − = 17 MPa, d.c. = 1%, 10 ms exposures) that were considered ‘safe’ were chosen to avoid the induction of non-specific inflammation since treatment with higher pulse negative pressures ( P − ≥ 22 MPa), and attempts to ablate the whole tumour resulted in some skin and spleen damage.…”

MRI Guided Focused Ultrasound-Mediated Delivery of Therapeutic Cells to the Brain: A Review of the State-of-the-Art Methodology and Future Applications

“…MRgFUS technology is gaining substantial interest for its ability to provide controlled, non-invasive, and targeted therapeutic ultrasound energy, which can be adjusted to create a variety of beneficial biological effects for treatments in the brain (52). These specific effects include destructive thermal ablation (53,54), radiosensitization (55), immune activation (56,57), BBB opening for therapeutic delivery (58,59), and stem cell homing (60,61).…”

Therapeutic Ultrasound-Enhanced Immune Checkpoint Inhibitor Therapy

“…Other recent studies found that histotripsy stimulated more potent intratumoral CD8 T cells and antigen presentation than HIFU thermal ablation in a murine breast cancer model [15] and melanoma model [16]. One recent study showed that combining histotripsy with intratumor anti-CD40 agonist antibody, αCTLA-4, and anti-PD-L1 antibody (αPD-L1) significantly improved the therapeutic efficacy against ICI refractory murine melanoma [17].…”

“…As a promising therapeutic technology, FUS has the unique combined advantages of being noninvasive, nonionizing, nonpharmaceutical, spatially targeted, and deeply penetrating the body. Since 2017, five FUS modalities, including highintensity focused ultrasound (HIFU) thermal ablation [7][8][9][10][11], HIFU hyperthermia [12], HIFU mechanical ablation [13][14][15][16][17], ultrasound-targeted microbubble destruction (UTMD) [18][19][20], and sonodynamic therapy (SDT) [21,22], have been investigated in combination with ICIs for treating solid tumors in mouse models. The enhancement of antitumor immune responses by these FUS modalities demonstrated the great promise of FUS as a transformative cancer treatment modality to improve ICI therapy.…”