Abstract LB-078: Effect of a combined immune checkpoint inhibitor and mechanical focused ultrasound treatment on intratumoral immune response in a MC38 preclinical model
Authors: Fant Cécile , Grasselly Chloé , Denis Morgane , Mathé Doriane , Choffour Pierre-Antoine , Daunizeau Loic , Mestas Jean-Louis , Lafon Cyril , Caux Christophe , Depil Stéphane , Dumontet Charles , Padilla Frédéric 
Published: September 2020

Abstract
Recent preclinical and clinical data suggest that Focused ultrasound (FUS) can enhance host antitumor immune response. FUS is a minimally invasive therapy focusing beams of high power ultrasound to locally induce tissue damage such as thermal necrosis or apoptosis, approved for treatment of some solid malignancies. As a result, there is an interest in combining FUS and immune checkpoint inhibitors (ICI) to enhance ICI response. However, the effect of such combined treatment on the immune response remains unclear. We have evaluated the combination of anti-PD1 and FUS in the MC38 preclinical colon cancer model. FUS was delivered using a confocal device generating and monitoring mechanical cavitational ultrasound (peak negative pressure -20.5MPa). Mice were treated each week with FUS three days before injections of anti-PD1 antibodies. A longitudinal follow-up of tumor growth and overall survival was performed. Lymphoid and myeloid intratumoral immune cells were assessed by flow cytometry on days one, three and six after the start of the combined treatment, including expression levels of co-inhibitory and co-stimulatory molecules such as PD1, PDL-1, TIGIT, LAG3, TIM3 and ICOS. Combined treatment of FUS and ICI led to significant increased survival (up to three-fold for the combination compared to control) and improved tumor growth control. Flow cytometry revealed various changes in lymphoid as well as myeloid populations. The CD8+ TNFa+ / CD4+ FoxP3+ cells ratio was increased for combined group compared to non-treated, the fraction of CD11b+ cells in treated groups was increased and could be explained by an increase in the granulocyte population. Moreover, ICI and FUS combination may lead to increased antigen priming as increased CD11c+ CMHIIhigh conventional dendritic cells (DC), activated CD8- and Sirpa-CD24- subpopulations were found. As for co-inhibitory and co-stimulatory molecules expression at cell surface, flow cytometry showed increased PD1 and LAG3 expression by CD8+ cells on day 1 (respectively two- and three-fold increases for combined group compared to control); an increased TIM3 expression by CD4+ cells for combined group on day 3;an increased PD1 expression by CD4+ cells for FUS-treated groups and by tumor cells for combined group on day 6; increased PDL1 and TIGIT expression by CD8+ cells on day 6. Combination of therapeutic ultrasound and anti PD1 results in a significant improvement in tumor growth control and survival, possibly through significant changes in immune infiltrate such as increased activated CD8+ cells / inhibitory CD4+ cells ratio, an important clinical biomarker of response, increased granulocyte cell populations and an increased DC-mediated response. However, the combined treatment led to increased PD1 expression, as well as of other co-inhibitory molecules, by CD4+, CD8+ and tumor cells possibly explaining the transient effect of the combination. Our observation carries important implications for future FUS/ICI therapies at clinical and mechanistic levels.
Citation Format: Cécile Fant, Chloé Grasselly, Morgane Denis, Doriane Mathé, Pierre-Antoine Choffour, Loic Daunizeau, Jean-Louis Mestas, Cyril Lafon, Christophe Caux, Stéphane Depil, Charles Dumontet, Frédéric Padilla. Effect of a combined immune checkpoint inhibitor and mechanical focused ultrasound treatment on intratumoral immune response in a MC38 preclinical model [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr LB-078.