Hyperthermia-induced injury and adaptation in adrenocortical carcinoma cells

Adrenocortical carcinoma (ACC) is a rare and aggressive malignancy that presents with limited therapeutic options, particularly for the 50% of patients who are diagnosed at late stages and are inoperable. While hyperthermia-based approaches (e.g. thermal ablation, heated intraperitoneal chemotherapy) have demonstrated promise against various solid tumours, the precise mechanisms by which hyperthermia induces ACC cell death, the effects on steroidogenesis, and whether cells develop thermal resistance, remain unclear. This thesis investigates the impact of hyperthermia on ACC cells using in vitro models (H295R, HAC15) alongside the non-tumorigenic endothelial cells (HUVEC). Lethal exposures (≥48°C) were found to markedly reduce ACC cell viability, primarily through caspase-independent pathways consistent with necrosis and necroptosis, rather than classical apoptosis. In contrast, sublethal exposures (42–45°C) caused minimal cell death where most ACC cells survived, regained typical morphology, and eventually resumed proliferation. These surviving cells did not manifest robust thermotolerance upon rechallenge, indicating a limited capacity for long-term hyperthermia resistance. Sublethal hyperthermia also transiently suppressed steroidogenesis, specifically Cortisol and Aldosterone biosynthesis, immediately following hyperthermia exposure; however, cells that recovered from sublethal hyperthermia regained their functional ability to synthesise steroids. Western blotting, live-cell imaging, and flow cytometry showed the involvement of necroptosis-related proteins like mixed lineage kinase domain-like protein (MLKL), and increased intracellular calcium flux upon hyperthermia exposure. Furthermore, scanning electron microscopy revealed classic morphological changes indicative of necrotic damage at lethal temperatures, alongside profound structural alterations that subsequently normalised in sub-lethally heated cells. Collectively, these findings provide a comprehensive evaluation of hyperthermia- induced cytotoxicity in ACC cells. By delineating the thermal thresholds that govern cell death, clarifying a dominant cell-death pathway, and assessing the functional resilience of surviving cells, this thesis sets the groundwork for refining hyperthermia- based interventions.

Funder

Science Foundation Ireland under Grant number [20/US/ 3676], the National Institutes of Health Grant number [R01EB028848]

Publisher

University of Galway

Rights

CC BY-NC-ND

Collections

University of Galway Theses (PhD Theses)