Most
-positive lung cancers will develop ALK-independent resistance after treatment with next-generation ALK inhibitors.
amplification has been described in patients progressing on ALK inhibitors, ...but frequency of this event has not been comprehensively assessed.
We performed FISH and/or next-generation sequencing on 207 posttreatment tissue (
= 101) or plasma (
= 106) specimens from patients with ALK-positive lung cancer to detect
genetic alterations. We evaluated ALK inhibitor sensitivity in cell lines with
alterations and assessed antitumor activity of ALK/MET blockade in ALK-positive cell lines and 2 patients with MET-driven resistance.
amplification was detected in 15% of tumor biopsies from patients relapsing on next-generation ALK inhibitors, including 12% and 22% of biopsies from patients progressing on second-generation inhibitors or lorlatinib, respectively. Patients treated with a second-generation ALK inhibitor in the first-line setting were more likely to develop
amplification than those who had received next-generation ALK inhibitors after crizotinib (
= 0.019). Two tumor specimens harbored an identical
rearrangement, one of which had concurrent
amplification. Expressing
in the sensitive H3122 ALK-positive cell line induced resistance to ALK inhibitors that was reversed with dual ALK/MET inhibition. MET inhibition resensitized a patient-derived cell line harboring both
and
amplification to ALK inhibitors. Two patients with ALK-positive lung cancer and acquired
alterations achieved rapid responses to ALK/MET combination therapy.
Treatment with next-generation ALK inhibitors, particularly in the first-line setting, may lead to MET-driven resistance. Patients with acquired
alterations may derive clinical benefit from therapies that target both ALK and MET.
Inactivation of SMARCA4/BRG1, the core ATPase subunit of mammalian SWI/SNF complexes, occurs at very high frequencies in non-small cell lung cancers (NSCLC). There are no targeted therapies for this ...subset of lung cancers, nor is it known how mutations in
contribute to lung cancer progression. Using a combination of gain- and loss-of-function approaches, we demonstrate that deletion of BRG1 in lung cancer leads to activation of replication stress responses. Single-molecule assessment of replication fork dynamics in BRG1-deficient cells revealed increased origin firing mediated by the prelicensing protein, CDC6. Quantitative mass spectrometry and coimmunoprecipitation assays showed that BRG1-containing SWI/SNF complexes interact with RPA complexes. Finally, BRG1-deficient lung cancers were sensitive to pharmacologic inhibition of ATR. These findings provide novel mechanistic insight into BRG1-mutant lung cancers and suggest that their dependency on ATR can be leveraged therapeutically and potentially expanded to BRG1-mutant cancers in other tissues. SIGNIFICANCE: These findings indicate that inhibition of ATR is a promising therapy for the 10% of non-small cell lung cancer patients harboring mutations in SMARCA4/BRG1. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/18/3841/F1.large.jpg.
Histologic transformation from non-small cell to small cell lung cancer has been reported as a resistance mechanism to targeted therapy in
-mutant and
fusion-positive lung cancers. Whether small cell ...transformation occurs in other oncogene-driven lung cancers remains unknown. Here we analyzed the genomic landscape of two pre-mortem and 11 post-mortem metastatic tumors collected from an advanced,
fusion-positive lung cancer patient, who had received sequential ROS1 inhibitors. Evidence of small cell transformation was observed in all metastatic sites at autopsy, with inactivation of
and
, and loss of
fusion expression. Whole-exome sequencing revealed minimal mutational and copy number heterogeneity, suggestive of "hard" clonal sweep. Patient-derived models generated from autopsy retained features consistent with small cell lung cancer and demonstrated resistance to ROS1 inhibitors. This case supports small cell transformation as a recurring resistance mechanism, and underscores the importance of elucidating its biology to expand therapeutic opportunities.
Abstract
Over 30% of lung adenocarcinomas harbor KRAS mutation. Despite progress in targeted therapies for specific genetic subsets of lung cancer (e.g. EGFR or ALK), there are no clinically ...effective targeted therapies for KRAS non-small cell lung cancer (NSCLC). Interpatient heterogeneity of KRAS mutant lung cancers, with large variability in co-occurring mutations, may contribute to its refractory phenotype. Specifically, KRAS mutant lung cancers with concurrent loss of the tumor suppressor LKB1 (KRAS-LKB1) have increased metastatic frequency and resistance to chemotherapy in pre-clinical models, and these patients have poor responses to immune checkpoint inhibitor. Thus, there is a critical need to develop novel therapeutic approaches for this subset of NSCLC. Intrinsic resistance to apoptosis limits the efficacy of therapies targeting KRAS signaling. Previous studies reported BCL-2/BCL-XL + MEK inhibition can increase apoptotic responses of some KRAS mutant cancers. Recently, potent and selective inhibitors of MCL-1 have been developed, creating additional possibilities for targeting apoptotic machinery for cancers that dependent upon MCL-1 for survival.
We recently reported that novel MCL-1 inhibitor AMG-176 combined with MEK inhibitor trametinib can induce tumor regression in subsets of KRAS mutant NSCLC pre-clinical tumor models. In addition to commercially available cell lines, we established 20 patient-derived cell line/xenograft mouse model and showed that KRAS-LKB1 cell lines are particularly sensitive to MEK + MCL1 inhibition with high synergy score. Moreover, restoring LKB1 expression in LKB1-/- cell lines hampers the synergy and blocks mitochondrial depolarization and apoptosis. BH3-profiling reveals high dependency on MCL-1 in KRAS-LKB1 cell lines. We reported that trametinib increase intercellular Bim (pro-apoptotic protein) and subsequent loading of Bim onto pro-survival proteins BCL-XL and/or MCL1. MCL1 inhibitor releases Bim from MCL1 and initiates the apoptotic cascade. Interestingly, trametinib induces preferential sequestration of Bim by MCL-1 in KRAS-LKB1 models. Restoration of LKB1 in LKB1-/- cell lines reduces the trametinib-induced Bim:MCL-1 protein-protein interactions. Combined inhibition of MEK + MCL1 caused dramatic tumor regression in LKB1-deficient xenograft mouse models compared to LKB1-restored models.
Our results reveal fundamental insights into a novel role for LKB1 in the regulation of mitochondrial apoptosis and will lay a solid pre-clinical foundation for the clinical investigation of the MEK + MCL-1 inhibitor combination. Our data may also support using LKB1 as a genetic biomarker for guiding the selection of BH3 mimetics in targeting KRAS mutant NSCLC.
Citation Format: Chendi Li, Audris Oh, Varuna Nangia, Aaron N. Hata. LKB1 regulates BH3-mimetics vulnerability of KRAS mutant non-small cell lung cancer by alternating mitochondrial apoptotic protein interactions abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2049.
Abstract
Background: There are currently no approved targeted therapies for KRAS-mutant non-small cell lung cancers (NSCLC), which represent 25-30% of lung adenocarcinomas. The development of ...mutant-specific covalent inhibitors of KRAS G12C has invigorated hope that clinically effective KRAS-targeted therapies are within reach. While these agents have shown activity in early phase clinical trials, identification of specific vulnerabilities conferred by common co-occurring mutations in KRAS-mutant NSCLC may enable development of combination therapies with enhanced activity in distinct subsets of patients.
Results: We screened a panel of KRAS-mutant NSCLC cell lines and observed that loss of the tumor suppressor STK11/LKB1 is associated with increased MCL-1 dependence and sensitivity to combined MAPK (either MEK inhibitor or KRAS G12C inhibitor AMG 510) and MCL-1 inhibition (AMG 176). Restoration of LKB1 expression in LKB1-deficient cell lines and mouse xenograft tumors blunted the apoptotic response to MAPK + MCL-1 inhibition; conversely, deletion of LKB1 in LKB1 wild-type models restored the sensitivity. Mechanistically, LKB1 deficiency is associated with an altered phosphoproteome and increased MCL-1-dependent apoptotic priming. LKB1 loss increased cellular stress leading to hyperactivation of JNK1, phosphorylation and stabilization of MCL-1 protein, and increased BIM sequestration by MCL-1. Upon suppression of MAPK signaling, LKB1-deficient cells exhibited greater levels of BIM bound to MCL-1 that could be liberated by AMG 176 to induce apoptosis. Consistent with these results, ex vivo treatment of tumor tissue from a KRAS-LKB1 mutant NSCLC patient with MEK inhibitor or AMG 510 increased MCL-1 dependent priming.
Conclusion: These results uncover a novel link between LKB1, cellular stress, and the regulation of MCL-1. LKB1 loss confers a dependency on MCL-1 that can be exploited therapeutically. Moreover, our study provides preclinical rationale for the exploration of combined KRAS G12C + MCL-1 inhibitors, particularly for KRAS-LKB1 mutant patients who respond poorly to standard-of-care checkpoint inhibitor therapy.
Citation Format: Chendi Li, Yi Shen, Mohammed Usman Syed, Audris Oh, Cameron Fraser, Johannes Kreuzer, Kaitlyn Webster, Robert Morris, Sean Caenepeel, Anne Y. Saiki, Karen Rex, James Lipford, Wilhelm Hass, Kristopher Sarosiek, Paul E. Hughes, Aaron N. Hata. LKB1 loss rewires stress signaling-induced apoptotic protein dynamics and sensitizes KRAS-mutant non-small cell lung cancers to combined MAPK + MCL-1 blockade abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 982.
Abstract
The recent approval of the KRAS G12C inhibitor sotorasib (AMG 510) for non-small cell lung cancer (NSCLC) marked a milestone in the development of targeted therapies for KRAS mutant cancers. ...While sotorasib and other KRAS G12C inhibitors have demonstrated rapid and durable responses in the clinic, some patients do not achieve responses. The identification of specific vulnerabilities conferred by recurrent co-occurring mutations may enable the development of biomarker-driven combination therapies with enhanced activity in distinct subsets of patients. We screened a panel of KRAS-mutant NSCLC cell lines as well as patient-derived xenograft (PDX) mouse models and observed that loss of the tumor suppressor STK11/LKB1 is associated with increased sensitivity to combined MAPK (either the KRAS G12C inhibitor sotorasib or MEK inhibitor trametinib) and MCL-1 inhibition (AMG 176). Restoration of LKB1 expression in LKB1-deficient cell lines and PDX tumors blunted the apoptotic response to MAPK + MCL-1 inhibition; conversely, deletion of LKB1 in LKB1 wild-type models increased sensitivity. Mitochondrial apoptotic cell death is regulated by interactions between pro- (e.g., BIM) and anti-apoptotic (e.g., MCL-1, BCL-XL) BCL-2 family members. MAPK inhibition increases BIM, while MCL-1 inhibition prevents BIM sequestration by MCL-1, resulting in apoptosis. LKB1 deficient cells exhibit increased association of BIM and MCL-1 upon MAPK inhibition, effectively priming cells for death upon inhibition of MCL-1. Mechanistically, LKB1 deficiency and associated loss of NUAK phosphorylation leads to hyperactivation of the JNK phospho-kinase network. JNK phosphorylates MCL-1 at S64 and T163, which enhances BIM: MCL-1 protein-protein interaction. Conversely, JNK phosphorylates BCL-XL at S62 and prevents sequestration of BIM. This series of phosphorylation events increases MCL-1 dependence and creates a specific vulnerability of KRAS-LKB1 tumors to MAPK + MCL-1 inhibition. Consistent with this mechanism, ex vivo treatment of tumor tissue from a KRAS-LKB1 mutant NSCLC patient with sotorasib or trametinib increased MCL-1 dependent priming. These results reveal a novel link between LKB1 and the regulation of BCL-2 family proteins and provide preclinical rationale for evaluation of combined KRAS G12C + MCL-1 inhibitors for KRAS-LKB1 mutant NSCLC.
Citation Format: Chendi Li, Mohammed Usman Syed, Yi Shen, Audris Oh, Cameron Fraser, Johannes Kreuzer, Christopher Nabel, Kaitlyn Webster, Robert Morris, Sean Caenepeel, Anne Y. Saiki, Karen Rex, J. Russell Lipford, Wilhelm Hass, Kristopher Sarosiek, Paul E. Hughes, Aaron Hata. LKB1 loss rewires JNK-induced apoptotic protein dynamics through NUAKs and sensitizes KRAS-mutant non-small cell lung cancers to combined KRAS G12C + MCL-1 blockade abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2150.
Histologic transformation from non-small cell to small cell lung cancer has been reported as a resistance mechanism to targeted therapy in EGFR-mutant and ALK fusion-positive lung cancers. Whether ...small cell transformation occurs in other oncogene-driven lung cancers remains unknown. Here we analyzed the genomic landscape of two pre-mortem and 11 post-mortem metastatic tumors collected from an advanced, ROS1 fusion-positive lung cancer patient, who had received sequential ROS1 inhibitors. Evidence of small cell transformation was observed in all metastatic sites at autopsy, with inactivation of RB1 and TP53, and loss of ROS1 fusion expression. Whole-exome sequencing revealed minimal mutational and copy number heterogeneity, suggestive of "hard" clonal sweep. Patient-derived models generated from autopsy retained features consistent with small cell lung cancer and demonstrated resistance to ROS1 inhibitors. This case supports small cell transformation as a recurring resistance mechanism, and underscores the importance of elucidating its biology to expand therapeutic opportunities.
Histologic transformation from non-small cell to small cell lung cancer has been reported as a resistance mechanism to targeted therapy in EGFR-mutant and ALK fusion-positive lung cancers. Whether ...small cell transformation occurs in other oncogene-driven lung cancers remains unknown. Here we analyzed the genomic landscape of two pre-mortem and 11 post-mortem metastatic tumors collected from an advanced, ROS1 fusion-positive lung cancer patient, who had received sequential ROS1 inhibitors. Evidence of small cell transformation was observed in all metastatic sites at autopsy, with inactivation of RB1 and TP53, and loss of ROS1 fusion expression. Whole-exome sequencing revealed minimal mutational and copy number heterogeneity, suggestive of "hard" clonal sweep. Patient-derived models generated from autopsy retained features consistent with small cell lung cancer and demonstrated resistance to ROS1 inhibitors. This case supports small cell transformation as a recurring resistance mechanism, and underscores the importance of elucidating its biology to expand therapeutic opportunities.