Abstract 6638: Pulsed electric fields combined with anti-PD1 prolongs survival and triggers an adaptive immune response in an IO-non-responsive orthotopic mouse model
Authors: Nafie Ebtesam , Wagh Mukta , Pastori Chiara , Trikha Mo , Neal Robert 
Published: June 2023

Abstract
Background: Although anti-PD1 therapy is standard of care in multiple tumor types, unmet need remains for those non-responsive to immunotherapy. Pulsed Electric Field (PEF) treatment may alter the tumor micro-environment, converting an immune “cold” tumor into an immune “hot” tumor, triggering an adaptive immune response and abscopal effect. We evaluated PEF in combination with anti-PD1 in a highly aggressive orthotopic triple negative breast tumor mouse model.
Methods: Murine mammary 4T1 cells (2 × 105) non-responsive to anti-PD1 therapy were orthotopically implanted in mammary fat of 48 female Balb/C mice. Once tumors were established (5 mm diameter), mice were randomized into six groups: Sham+IgG controls, aPD-1-only, PEF (1X), PEF (2X), aPD-1 + PEF (1X), and aPD-1 + PEF (2X). Groups 4 and 6 received two PEF treatments, five days apart. Anti-PD1 and isotype matched control IgG was administered (200 ug, intraperitoneal) weekly, starting 3 days after PEF treatment. Tumor volumes were recorded 3x/week and standard survival analyses were conducted per IACUC protocol. In the cohort of tumors collected 30 days after PEF, flow cytometry (FC) evaluated T-cell immune infiltrates and associated markers. Blood was collected at different time points to evaluate systemic immunological response. Tumor-specific cytotoxic lymphocytes (CTL) were isolated from tumor-infiltrating and circulating lymphocytes by gp70 MHC tetramer formation assay.
Results: Single PEF reduced tumor growth compared to IgG and aPD-1-only groups by 65% and 62%, respectively. Double PEF treatment reduced tumor growth by 74% and prolonged survival. FC demonstrated circulating helper T-cells and CTL frequency was increased two weeks post-PEF compared to the control group. The tetramer detected substantial increases in antigen-specific T-cell populations in both single and double PEF treatment groups. PEF groups had significant tumor and spleen infiltration of CD8 and CD4 T-cells, including increased tetramer levels. Anti-PD1-only animals showed no meaningful reductions in tumor growth, prolonged survival, or significant changes in CD8, CD4 T cells or in the tetramer assay.
Conclusions: PEF treatments prior to initiation of anti-PD1 in orthotopic 4T1 mouse model triggers a tumor-specific adaptive immune response in an immune “cold” tumor model, irrespective of anti-PD-1 therapy. PEF increases circulating and tumor-infiltrating T-cells and antigen-specific T-cells, with these effects observed after a single or two treatments with PEF. Collectively, our results demonstrate that PEF treatment prior to initiation of anti-PD1 treatment can reduce tumor growth, prolong survival, and activate an adaptive immune response in an immune “cold” tumor. These findings suggest that PEF treatment merits further evaluation in IO-non-responsive cancer patients.
Citation Format: Ebtesam Nafie, Mukta Wagh, Chiara Pastori, Mo Trikha, Robert Neal. Pulsed electric fields combined with anti-PD1 prolongs survival and triggers an adaptive immune response in an IO-non-responsive orthotopic mouse model. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6638.