Abstract 1800A: STK11 and KEAP1 co-mutation cooperatively promotes rapid tumor growth in lung adenocarcinoma
Authors: Wohlhieter Corrin A. , Richards Allison L. , Poirier John T. , Uddin Fathema , Hulton Christopher , De Stanchina Elisa , Papagiannakopoulos Thales , Sen Triparna , Rudin Charles M. 
Published: September 2020

Abstract
Background: Genomic profiling and targeted therapies have significantly improved outcomes for oncogene-addicted subgroups of lung adenocarcinoma (LUAD) patients (e.g., EGFR mutations or rearrangements of ALK, ROS1 and RET). However, we continue to lack effective targeted therapies for some of the common mutationally defined subsets, including those associated with poor outcome in patients with lung adenocarcinoma. A recent study from our group has shown that concomitant loss-of-function mutations in STK11/LKB1 and KEAP1 (LK) defines a cohort of LUAD patients with shortest overall survivals (Shen et al., 2019). Patients whose tumors harbor LK mutations across all stages of disease show a median overall survival of 11.8 months, compared to KEAP1 only (34.5 months), STK11 only (34.2 months), or neither (40.2 months). The biology and therapeutic vulnerabilities of LUAD subtype with concurrent loss of LKB1 and KEAP1 (either with or without KRAS mutation) is particularly understudied.
Methods: We analyzed genomic characteristics of LK lung adenocarcinomas from patients (n=292) at Memorial Sloan Kettering. We performed CRISPR/Cas9 mediated knockout of LKB1, KEAP1, or both to develop matched isogenic clones for three adenocarcinoma cell lines to investigate phenotypic and molecular changes imparted by each single mutation and the co-mutation. We investigated growth in genetically manipulated isogenic groups of LUAD human cell lines (n=8).To investigate the gene expression changes and therapeutic vulnerabilities in samples with concurrent loss of LKB1/KEAP1 we performed bulk RNA-seq and druggable genome CRISPR screening respectively across derivatives of two of these lines, NCI-H358 and NCI-H292.
Results: Study of variant allele frequency (VAF) on patient samples with mutations in both STK11and KEAP1 demonstrated a high correlation between VAF of the two genes suggesting clonal expansion of LK tumor cells without defining a preferred order in mutational timing. We show that loss of both genes increases cell proliferation and invasion in vitro and faster/enhanced tumor growth in vivo in all models independent of their carrier mutation status. RNA-sequencing of our isogenic cell lines showed that LK tumors have a distinct gene expression signature different from either of the single mutants with high expression of transcripts involved in cell division and cell survival.
Conclusions: Our data suggests that co-mutation of STK11 and KEAP1 cooperate to enhance cell proliferation and tumor growth in human models of lung adenocarcinoma. This is in agreement with clinical observations demonstrating aggressiveness of this subpopulation. Furthermore, identified hits from the CRISPR gene dependency screening have clear translational potential and pave the way for the development of rational therapeutic strategies to improve treatment of LKB1/KEAP1-deficient tumors.
Citation Format: Corrin A. Wohlhieter, Allison L. Richards, John T. Poirier, Fathema Uddin, Christopher Hulton, Elisa De Stanchina, Thales Papagiannakopoulos, Triparna Sen, Charles M. Rudin. STK11 and KEAP1 co-mutation cooperatively promotes rapid tumor growth in lung adenocarcinoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1800A.