Abstract No. 330 ▪ FEATURED ABSTRACT Advanced Biphasic Pulsed Electric Field (PEF) Stimulates a More Favorable Immune Response Profile versus Irreversible Electroporation (IRE) Upon Ablation of Murine Breast Cancer

Aliya Pulsed Electric Fields (PEF) is an ablation modality that does not rely on thermal processes to kill cells, providing a means to destroy tissue without damaging the extracellular matrix and potentially promoting an immune response. The Aliya PEF system uses an advanced waveform design compared with irreversible electroporation (IRE) that makes it is simpler to deliver and generates less mechanical trauma than IRE waveforms. This study evaluates whether these improvements to PEF technology provide differentially favorable immune response. Female Balb/c mice were orthotopically inoculated in the mammary fat pad with 200,000 cells from the EMT6 murine breast cancer cell line. When the tumors reached 5-7 mm in size (10 days), the mice were randomly assigned to Sham treatment (n=4), IRE (n=6), or Aliya PEF (n=6), where the ablation technologies were titrated to target matched-size ablations for approximately 80% of the tumor volume. Four days later, the mice were euthanized to collect the treated tumors as well as peripheral blood. The tumor samples were lysed to extract total proteins to quantify 32 cytokines by multiplex laser bead technology. Flow cytometry on blood samples was used to enumerate dendritic and natural killer cells, T and B lymphocytes, macrophage (M1 and M2), and myeloid derived suppressor cells (mMDSC). The intratumoral cytokine profiles revealed unique tumor microenvironments, where Aliya PEF-treated tumors had higher concentration of LIX, MIP2, and MIP1A and lower concentrations of IL-13, Il-2, Eotaxin, Il-12, IL-7, RANTES and GM-CSF compared to both Sham and IRE treatment. These cytokines were unchanged between Sham and IRE. Flow cytometric analyses revealed that Aliya PEF-treated tumors four days post-ablation had a higher percentage of T-cells (CD3+), B-cells, dendritic, and natural killer cells compared to Sham and IRE. Moreover, Aliya PEF-treated tumors had fewer immunosuppressive mMDSC and M2 macrophages, while the level of M1 was unchanged between groups. The IRE treatment did not promote any statistical change in the recruitment of these immune populations. The proprietary Aliya PEF ablation modality resulted in important differences in post-ablation intratumoral cytokines that indicate distinct effects in the tumor microenvironment between the ablation technologies. This resulted in increased intratumoral immune cell populations for Aliya PEF relative to Sham or IRE. These data suggest that Aliya PEF generates a uniquely favorable immunostimulatory profile versus IRE in a murine model.