3P - Metabolic reprogramming induced by KEAP1 mutation in NSCLC

Background

Recently, KRAS G12C inhibitors (KRAS G12Ci) have been approved for the treatment of advanced KRAS G12C-mutant NSCLC. However, the group of patients with KEAP1 and LKB1 co-mutations did not achieve the expected clinical benefit of KRASG12Ci monotherapy. The independent impact of these alterations and the molecular mechanisms contributing to resistance are poorly understood. In this work, we investigated the impact of these co-mutations in vitro/in vivo in response to KRAS G12Ci and characterized transcriptomic and metabolic profiles.

Methods

We established a murine model of KRAS G12C NSCLC using CRISPR/Cas9 gene editing to create knockouts of LKB1(393p KL) and LKB1/KEAP1 (393p KLK). Several human KRAS G12C- mutant NSCLCs were used to confirm our findings (H2030, HCC44). Sotorasib (AMG 510) was used as a KRAS G12C inhibitor. RNA-seq, Seahorse Mito Stress test, metabolome analysis and 13C metabolic flux analysis were conducted. For animal studies, 8-week-old female 129S2/SvPasCrl mice were used.

Results

KEAP1 mutation drives resistance to KRAS G12Ci in cell viability assay and syngeneic murine model. Despite differences in response to KRAS G12Ci in vitro/in vivo, the MAPK pathway in KL and KLK was inhibited at the same level. RNA seq results revealed the enrichment of multiple metabolic and oxidative stress transcriptional programs in KLK cells. To gain insights into metabolic reprogramming in cells with KEAP1 and LKB1 alterations, metabolome and uniformly 13C-labeled glucose tracer assay were performed. In KLK cells central carbon metabolism including glutamine metabolism was greatly changed. Moreover, we observed an increase in several intermediates of the tricarboxylic acid cycle. Hence, this metabolic reprogramming caused by KEAP1 mutation can provide a survival advantage to KLK cells.

Conclusions

Clinically known resistance to KRAS G12Ci in patients with KEAP1 mutation was proved in vitro/in vivo. The presence of co-mutations in LKB1 (without additional alterations in KEAP1) does not affect the resistance to KRAS G12Ci. KEAP1 mutation induces metabolic reprogramming to fulfil the increased needs for energy and anabolism.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

The authors.

Funding

Ministry of Education, Culture, Sports, Science and Technology.

Disclosure

All authors have declared no conflicts of interest.