Critical Role for HSF2 and HSP110 axis in Regulation of Pre-mRNA Splicing and Radiation Response

ABSTRACT Pancreatic ductal adenocarcinoma (PDAC) has a poor 5-year survival outcome with a limited therapeutic opportunity. A major barrier to the durable clinical responses is the tumor’s ability to adapt to therapeutic pressures (such as cytotoxic chemotherapy, radiotherapy, targeted agents, and immunotherapy) through transcriptional and post-transcriptional mechanisms that promote cell survival, enhance DNA repair, and suppress immune surveillance. Disrupting transcriptional fidelity can simultaneously impair DNA repair, oncogenic signaling, and enhance anti-tumor immune responses, offering broad therapeutic leverage. This proposal aims to address these critical scientific and clinical unmet needs. Our preliminary results show that Heat Shock Transcription Factor 2 (HSF2) and its downstream transcriptional target HSP110 are upregulated in cells in response to x-irradiation (IR) exposure. Loss of HSF2 or HSP110 increases transcription-associated DNA damage in cells, and their loss leads to suppression of RNA polymerase II (RNAPII) C-terminal domain (CTD) phosphorylation at serine 7 (RNAPIICTD pS7). RNAPIICTD pS7 regulates transcription of small nuclear RNAs (snRNAs), which are essential components of pre-mRNA splicing and spliceosomes. Consequently, reduction in pS7 in HSF2- or HSP110-deficient cells reduces snRNA levels, resulting in altered alternative splicing and potentially generating novel transcripts. Furthermore, loss of HSF2 or HSP110 impairs RNAPII processivity, resulting in altered RNAPII occupancy dynamics, reduced transcriptional fidelity, and transcription-associated genome instability. Under genotoxic stress induced by IR, loss of HSF2 or HSP110 reduces the expression of DNA repair genes, leading to cellular sensitivity to IR exposure, which is consistent with a significant increase in DNA damage, cell death, and infiltration of CD8+ T cells, thereby significantly inhibiting PDAC tumor growth in mice. Based on these observations, we hypothesize that: Disruption of the HSF2-HSP110 axis impairs adaptive transcriptional responses, sensitizes tumor cells to genotoxic therapies, and enhances anti-tumor immunity, revealing a novel and actionable vulnerability in cancer treatment. To determine the therapeutic vulnerability of tumors following HSF2 or HSP110 loss, we propose the following Aims: (1). Determine the role of HSF2 or HSP110 in regulating RNAPII processivity that culminates in DNA damage and x-irradiation sensitivity and altered splicing providing multiple avenues of PDAC-specific cell killing. (2). Determine the therapeutic significance of the HSF2–HSP110 axis in genotoxic stress response and identify HSF2/HSP110 inhibitors as candidate therapeutics. We predict that the successful completion of this study will establish novel therapeutic strategies for treating PDAC through simultaneous induction of transcription stalling, splicing impairment, immunomodulation, and tumor cell-specific radiation sensitivity.