Abstract 1058: Inhibition of nuclear transport protein XPO1 potentiates the effect of KRASG12C inhibitors

Introduction: The discovery of compounds that can bind covalently to KRASG12C and lock it in its inactive GDP-bound conformation has opened a window of opportunity to selectively target KRASG12C. Such agents have shown promising results in preclinical tumor models and recently in clinical trials as well. However, factors such as feedback reactivation or bypass of KRAS dependence are known to limit antitumor activity of KRAS inhibitors. This necessitates the need for combination approaches that can potentially sensitize tumors to KRAS inhibitors when co-targeted. In this study, we have tested two potent KRASG12C inhibitors, MRTX1257 and AMG510, on PDAC and NSCLC cell lines both as single agents and also in combination with nuclear transport inhibitor, Selinexor (KPT-330/XPOVIO). Methods: Cell growth inhibition was determined by MTT assay. Drug synergy analysis was performed and the isobolograms were generated using CalcuSyn 2.1 software (Biosoft, Cambridge, UK). Spheroid disintegration assay, clonogenic assay, immunofluorescence assay, and Western blotting were performed by standard methods. Results: MRTX1257 and AMG510, when combined with KPT-330, synergistically inhibited (Combination Index values less than 1) the proliferation of KRASG12C mutant cancer cell lines, namely MiaPaCa-2 (PDAC) and NCI-H2122 (NSCLC). Such an effect was observed neither in KRASWT (NCI-H1650) nor KRASG12D mutant (Panc-1) cells, suggesting that the synergistic effect on cell viability was specific to KRASG12C. Combined treatment of KPT-330 with either MRTX1257 or AMG510 resulted in enhanced disintegration of MiaPaCa-2 and NCI-H2122 spheroids, indicating the efficacy of the drug combinations in a 3-D cell growth model. In addition, similar synergistic action was also seen in a clonogenic assay, where MRTX1257 and AMG510 reduced the number and size of colonies formed by MiaPaCa-2 cells and KPT-330 further augmented this effect. KPT-330 also potentiated the MRTX1257- or AMG510-induced inhibition of KRAS-dependent signaling targets such as ERK1/2 phosphorylation. Mechanistically, we observed that KPT-330 caused nuclear retention of ETS repressor factor (ERF), a transcriptional repressor that is exported upon phosphorylation by ERK. Given that XPO1 transports hundreds of proteins, we are currently evaluating the global changes in RAS pathway proteins using high throughput approaches. Such studies are anticipated to unveil the molecular underpinnings of the effectiveness of this co-targ