Inhibition of autophagy and MEK promotes ferroptosis in Lkb1-deficient Kras-driven lung tumors

Abstract

LKB1 and KRAS are the third most frequent co-mutations detected in non-small cell lung cancer (NSCLC) and cause aggressive tumor growth. Unfortunately, treatment with RAS-RAF-MEK-ERK pathway inhibitors has minimal therapeutic efficacy in LKB1-mutant KRAS-driven NSCLC. Autophagy, an intracellular nutrient scavenging pathway, compensates for Lkb1 loss to support Kras-driven lung tumor growth. Here we preclinically evaluate the possibility of autophagy inhibition together with MEK inhibition as a treatment for Kras-driven lung tumors. We found that the combination of the autophagy inhibitor hydroxychloroquine (HCQ) and the MEK inhibitor Trametinib displays synergistic anti-proliferative activity in KrasG12D/+;Lkb1-/- (KL) lung cancer cells, but not in KrasG12D/+;p53-/- (KP) lung cancer cells. In vivo studies using tumor allografts, genetically engineered mouse models (GEMMs) and patient-derived xenografts (PDXs) showed anti-tumor activity of the combination of HCQ and Trametinib on KL but not KP tumors. We further found that the combination treatment significantly reduced mitochondrial membrane potential, basal respiration, and ATP production, while also increasing lipid peroxidation, indicative of ferroptosis, in KL tumor-derived cell lines (TDCLs) and KL tumors compared to treatment with single agents. Moreover, the reduced tumor growth by the combination treatment was rescued by ferroptosis inhibitor. Taken together, we demonstrate that autophagy upregulation in KL tumors causes resistance to Trametinib by inhibiting ferroptosis. Therefore, a combination of autophagy and MEK inhibition could be a novel therapeutic strategy to specifically treat NSCLC bearing co-mutations of LKB1 and KRAS.

Inhibition of ULK1/2 and KRASG12Ccontrols tumor growth in preclinical models of lung cancer

“…Our data suggest that combined inhibition of KRAS G12C plus autophagy may have superior activity against KRAS G12C -driven lung cancer. These data are in accord with a recent study in which HCQ-mediated inhibition of lysosome function sensitized a KRAS G12D /LKB1 Null -driven GEM model of lung cancer to MEK1/2 inhibition 11 . An interesting aspect of these results is the apparent dispensability of LKB1 for the induction of autophagy in response to inhibition of KRAS signaling.…”

“…Cancer cells exhibit increased autophagy in response to a variety of cellular stresses including nutrient deprivation or the application of pathway-targeted therapy, thereby allowing cancer cells to recycle a wide variety of macromolecules and organelles to promote cell viability. 8, 11 Moreover, GEM models of lung or pancreatic cancer indicate that cancer cells employ autophagy - both cancer cell autonomous and from the mouse as a whole - at a several stages of cancer progression 46, 49 . Recently, we and others have demonstrated that many RAS-driven cancer cells display increased autophagy in response to blockade of RAS>RAF>MEK>ERK signaling 8, 9, 11 .…”

“…8, 11 Moreover, GEM models of lung or pancreatic cancer indicate that cancer cells employ autophagy - both cancer cell autonomous and from the mouse as a whole - at a several stages of cancer progression 46, 49 . Recently, we and others have demonstrated that many RAS-driven cancer cells display increased autophagy in response to blockade of RAS>RAF>MEK>ERK signaling 8, 9, 11 . Moreover, in our work, combined inhibition of MEK1/2 (with trametinib) plus inhibition of lysosome function (with CQ or HCQ) displayed synergistic anti-proliferative activity in vitro , superior tumor control (compared to the single agents or standard of care) in c ell line- or p atient- d erived x enografts (CDX or PDX) models of pancreatic cancer and, in one patient with advanced pancreatic cancer, there was evidence of anti-tumor activity of the combination of trametinib plus hydroxychloroquine 8 .…”

“…Liver kinase B1 (LKB1) activates AMP kinases in response to cellular nutrient changes to maintain homeostasis 10 . However, induction of autophagy has also been reported in response to MEK inhibition in models of KRAS G12D -driven lung cancer with loss of LKB1 expression 11 , suggesting that LKB1 is dispensable for increases in autophagy in lung cancer. LKB1 silencing is frequently observed in patients with KRAS-driven lung cancer, and such patients exhibit decreased overall survival and are more refractory to current therapeutic treatments than in LKB1 proficient KRAS-driven lung cancers 12, 13 .…”

Liver Kinase B1 Mediates Its Anti-Tumor Function by Binding to the N-Terminus of Malic Enzyme 3

“…Liver kinase B1 (LKB1), also known as the serine/threonine kinase 11 (STK11) gene, is a widely expressed tumor suppressor gene that is commonly associated with Peutz-Jeghers syndrome (PJS) ( Avilés-Salas et al ., 2023 ; Omori et al ., 2023 ). It plays a crucial role in regulating multiple cellular processes, including tumor initiation and progression ( Gao et al ., 2010 ; Hardie, 2013 ), invasion, and adhesion ( Zagórska et al ., 2010 ; Konen et al ., 2016 ), energy metabolism ( Bhatt et al ., 2023 ), cell cycle ( Gurumurthy et al ., 2010 ), cell migration, and embryonic development ( Men et al ., 2015 ). LKB1 functions as a master regulator by directly phosphorylating the AMP-activated protein kinase (AMPK) family of proteins, thereby modulating their activity ( Shackelford and Shaw, 2009 ; Zhang et al ., 2013 ).…”