Abstract No. 164 ▪ FEATURED ABSTRACT Natural Killer Cell-Based Immunotherapy Maintains Cytotoxicity After Temperature-Controlled Ablation in Poorly and Well-Differentiated Hepatocellular Carcinoma

Image-guided thermal ablation is considered the standard of care for patients with unresectable early-stage HCC. The peripheral zone of the ablation exposes tumor cells to non-lethal hyperthermic temperatures. Histopathologic grade of HCC has been correlated with post-ablation outcomes, with poorly differentiated HCCs being associated with higher rates of disease progression. This study evaluated the feasibility of using NK-cell based immunotherapy after sub-lethal ablation for treating both well- and poorly differentiated HCC. HCC cell lines HepG2 (well-differentiated) and Snu-423 (poorly differentiated) were incubated in 37, 43, and 47°C water baths for 10 minutes before cooling to 25°C for 20 minutes. HCC cells were then exposed to NK effector cells (primary NK (pNK) cells, IL-2 stimulated NK (pNK+IL-2) cells, and supercharged NK (sNK) cells). HCC cell percent cytolysis was analyzed on the live cell imaging platform for 72 hours. HCC cell cytotoxicity was also analyzed via chromium release assay, using a gamma counter to quantify cell death. IL-2 stimulated and sNK cell therapies demonstrated increased cytotoxicity with higher percent cytolysis after exposure to non-lethal hyperthermia. There was significantly higher percent cytolysis in both well- and poorly differentiated HCC exposed to 47°C compared to cells exposed to 37°C, although there was not a significant difference in cytotoxicity between the well and poorly differentiated HCC cell lines. This result was recapitulated in the chromium assay where there was a significantly higher LU30 in the IL-2 stimulated and supercharged NK cells at 47°C compared to 37°C (Table 164.1). Our in-vitro results confirm that non-lethal hyperthermia improves NK cell cytotoxicity to both well- and poorly differentiated HCC cell lines. While thermal ablation currently provides excellent outcomes for early-stage HCC, improvements may be achieved by augmenting thermal ablation with NK-cell based immunotherapy. Additional validation via in-vivo studies is required to confirm the safety of NK-cell based immunotherapies.