Exploring the effects of combining KRAS inhibitor MRTX1133 and glutaminase inhibitor CB-839 in inhibiting pancreatic ductal adenocarcinoma cell proliferation

Pancreatic ductal adenocarcinoma (PDAC) is the third leading cause of cancer deaths in the United States with nearly all patients (~95%) containing a point mutation in the KRAS oncogene. Mutated KRAS promotes cancer cell proliferation primarily through constitutive activation of effector signaling pathways that deregulate genes that support cancer growth. The RAF-MEK-ERK mitogen-activated protein kinase (MAPK) cascade is the predominant signaling pathway downstream of KRAS driving cell growth. To prevent such oncogenic proliferation, there is a critical need for the development of potent KRAS inhibitors that block effector signaling. Recently, the non-covalent, KRASG12D-selective inhibitor, clinical candidate MRTX1133 (G12Di) suppressed activation of the RAF-MEK-ERK pathway and potently suppressed the growth of PDAC cancer cell lines in vitro and in vivo. Because the G12D mutation accounts for 42% of all KRAS-mutant PDAC cases, KRASG12D inhibitors show great potential as an efficacious treatment for a significant proportion of PDAC patients. However, recent data of the clinically approved KRASG12C inhibitors sotorasib and adagrasib for non-small cell lung cancer (NSCLC) demonstrated anti-tumor activity that was limited by onset of acquired treatment-associated resistance. Thus, it is highly possible that resistance could develop to G12Di treatment in PDAC. In this study, we focus on determining the mechanisms behind G12Di resistance in PDAC. To identify genes involved in G12Di sensitivity, we performed a loss-of-function CRISPR screen using the druggable genome library in PDAC cancer cells. One identified gene linked to resistance was KEAP1. KEAP1 is a negative regulator of NRF2, a transcription factor involved in the adaptation of cellular stress response and expression of pro-cell survival genes. To assess the impact of KEAP1 on PDAC G12Di resistance, we employed CRISPR to eliminate KEAP1 from the genome. As a result, we discovered that KEAP1 knockout PDAC cell lines showed decreased sensitivity to G12Di to inhibit proliferation in vitro, and additionally developed a dependence on glutamine metabolism. Through cell proliferation assays, we further discovered that the combination of a glutaminase inhibitor (CB-839) and G12Di improved G12D-mutant growth inhibition in KEAP1 knockout cell lines and in parental cell lines. These findings point to glutaminase inhibition as a potential strategy to address NRF2-driven resistance and make progress toward a more effective treatment of PDAC.