Optimizing KRAS Therapeutics for Non-Small Cell Lung Cancer.

Background: VS-6766 is a unique small molecule inhibitor of both RAF and MEK. In contrast to several other MEK inhibitors available, VS-6766 blocks both MEK kinase activity and RAF phosphorylation of MEK. This sequential blockade mechanism enables VS-6766 to block MEK signaling more consistently without the compensatory and paradoxical activation of MEK that reduces the efficacy of other small molecule inhibitors of MEK and RAF. Defactinib (VS-6063), an orally active small molecule, is a potent adenosine 5'- triphosphate (ATP) competitive, reversible inhibitor of focal adhesion kinase (FAK). Defactinib has shown synergistic activity with BRAF and MEK inhibitors in both in vitro and in vivo preclinical solid tumor models. Specifically, in several human tumor cell lines with mutations in RAS or BRAF or wildtype RAS and BRAF, defactinib has shown synergy with MEK inhibitors or VS-6766. In mouse models of KRAS mutant ovarian cancer, BRAF-mutant melanoma or uveal melanoma, FAK inhibition has been shown to induce tumor regression when combined with RAF, MEK or RAF/MEK inhibitors, while the single agents have only induced tumor stasis. The combination of VS-6766 and defactinib is currently being evaluated in the Investigator Sponsored FRAME study. Preliminary efficacy results are available for a small number of subjects with KRAS mutated NSCLC treated with a combination of VS-6766 and defactinib. Of the 10 subjects with NSCLC, 1 subject with KRAS-G12V- mutated cancer achieved PR while 3/10 subjects received treatment for ≥24 weeks. In an updated analysis of response in 15 subjects with KRAS-mutant NSCLC, there were 2 PRs and 7 SDs (ORR: 13%). The two subjects with KRAS G12V- mutated NSCLC both achieved PR (ORR: 100%). Methods: This is an adaptive, two-part, Phase 2, multicenter, parallel cohort, randomized, open label study designed to evaluate the efficacy and safety of VS-6766 versus VS-6766 in combination with defactinib in subjects with recurrent NSCLC(NCT04620330). The study will be conducted in two parts. Part A will determine the optimal regimen, either VS-6766 monotherapy or VS-6766 in combination with defactinib based on confirmed overall response rate as assessed by independent radiology review. Part A will consist of three arms in KRAS mutated NSCLC Arm 1 monotherapy of VS-6766, Arm 2 the combination of VS-6766 and defactinib in KRAS G12V mutated and Arm 3 the combination in other KRAS mutated. Part B will determine the efficacy of the optimal regimen identified in Part A in KRAS mutated NSCLC. Subjects enrolled must have histologic or cytologic evidence of NSCLC, measurable disease according to RECIST V1.1 and known KRAS mutation. The study will enroll up to 377 subjects globally with 57 subjects (32 KRAS G12V & 25 KRAS other) in Part A and an additional 144 KRAS G12V and 176 KRAS other in Part B. This study is open for enrollment. Citation Format: D. Ross Camidge, Jonathan Pachter, Andrew Koustenis, Gloria Patrick, David R. Spigel. A phase 2 study of VS-6766 (dual RAF/MEK inhibitor) RAMP 202, as a single agent and in combination with defactinib (FAK inhibitor) in recurrent KRAS-mutant (KRAS-MT) non-small cell lung cancer (NSCLC) [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P048.

KRAS-mutant non-small cell lung cancer (NSCLC) is a major lung cancer subtype that leads to many cancer-related deaths worldwide. Although numerous studies on KRAS-mutant type NSCLC have been conducted, new oncogenic or tumor suppressive genes need to be detected because a large proportion of NSCLC patients does not respond to currently used therapeutics. Here, we show the tumor-promoting function of a cell cycle-related protein, PIERCE1, in KRAS-mutant NSCLC. Mechanistically, PIERCE1 depletion inhibits cell growth and AKT phosphorylation (pAKT) at S473, which is particularly observed in KRAS-mutant lung cancers. Analyses of AKT-related genes using microarray, immunoblotting, and real-time quantitative PCR indicated that PIERCE1 negatively regulates the gene expression of the AKT suppressor, TRIB3, through the CHOP pathway, which is a key regulatory pathway for TRIB3 expression. Similarly, in vivo analyses of PIERCE1 depletion in the KRAS mutation-related lung cancer mouse models revealed the suppressive effect of PIERCE1 knockout in urethane- and KRASG12D-induced lung tumorigenesis with decreased pAKT levels observed in the tumors. Tissue microarrays of human lung cancers indicated the expression of PIERCE1 in 83% of lung cancers and its correlation with pAKT expression. Thus, we illustrate how PIERCE1 depletion may serve as a therapeutic strategy against KRAS-mutant NSCLC and propose the clinical benefit of PIERCE1.

A large fraction of non-small cell lung cancers (NSCLC) have mutant KRAS, which is associated with poor response to current cytotoxic therapy and a poor prognosis. Although the KRAS signaling pathway has been well characterized, no current therapies target this critical oncogene. Several studies have demonstrated that bypass of senescence in Kras-mediated adenocarcinoma mouse models is essential for tumorigenesis. Therefore, activation of senescence in KRAS mutant NSCLC may be an effective therapeutic strategy. We recently demonstrated that the basic helix loop helix transcription factor Twist1 cooperates with mutant Kras to induce lung adenocarcinoma in transgenic mouse models and that inhibition of Twist1 in these models led to activation of Kras-induced senescence and tumor stasis. In the current study, we examine the role of TWIST1 in KRAS mutant human NSCLC. Silencing of TWIST1 in multiple KRAS mutant NSCLC cell lines resulted in dramatic growth inhibition and either reactivation of oncogene-induced senescence or in some cases, apoptosis. Similar effects were also observed in four KRAS wild type lines, including cell lines with key driver mutations including a cell line with an activating EGFR mutation and a cell line with c-Met amplification. Gene set enrichment analysis of NSCLC cell lines after silencing of TWIST1 revealed a striking cell cycle arrest gene signature. Growth inhibition by silencing of TWIST1 was independent of p53 or Rb/p16 mutational status. Furthermore, activation of oncogene-induced senescence by TWIST1 silencing did not require previously defined mediators of senescence, p21 and p27, nor could this phenotype be rescued by overexpression of SKP2. To extend these observations in vivo, TWIST1 was silenced in both KRAS mutant and wildtype cell lines and these cells were implanted in NOD-SCID mice to assess tumor formation. Interestingly, silencing of TWIST1 in xenograft models preferentially inhibited KRAS mutant tumor formation suggesting that TWIST1 plays a critical in mediating KRAS tumorigenesis. Finally, inducible silencing of TWIST1 resulted in significant growth inhibition of established xenograft KRAS mutant tumors. Together these findings suggest TWIST1 is essential for the establishment and maintenance of KRAS mutant NSCLC tumors and silencing of TWIST1 in KRAS mutant NSCLC represents a novel and promising therapeutic strategy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2954. doi:1538-7445.AM2012-2954

Efficacy of Immune Checkpoint Inhibitors in KRAS-Mutant Non-Small Cell Lung Cancer (NSCLC)

An integrative pharmacogenomics analysis identifies therapeutic targets in KRAS-mutant lung cancer.

KRAS gene mutation is linked to poor prognosis and resistance to oncology therapeutics in Non Small Cell Lung Cancer (NSCLC). We have explored the possibility of exploiting inherent differences in KRAS mutant cell metabolism to enhance the efficacy of treatment. We have identified a greater dependency on purine biosynthesis and related pathways in KRAS mutant compared to KRAS wild type NSCLC cell lines. Purine synthesis requires factors generated from other metabolic reactions including ribose-5-phosphate from the pentose phosphate pathway, THF cofactors from folate metabolism and glycine / amide nitrogen groups from glutamine and aspartate metabolism. In this study, microarray gene expression and biological pathway analysis identified higher expression of purine synthesis and accessory pathways such as folate metabolism, 5-aminoimidazole ribonucleotide biosynthesis and glycine synthesis pathways in KRAS mutant NSCLC cells compared to wildtype counterparts. KRAS knockdown and overexpression studies demonstrated the ability of KRAS to regulate expression of genes that comprise purine synthesis and folate metabolism pathways. Moreover, pathway analysis and knockdown studies suggest a role for MYC, an oncogene previously associated with KRAS mutant tumors, in the regulation of these pathways in KRAS mutant NSCLC cells. Proliferation studies demonstrated higher responsiveness to antifolates such as methotrexate and pemetrexed in KRAS mutant NSCLC cells, both of which may interfere indirectly with purine biosynthesis. In vivo analysis of NSCLC tumorgraft models in nude mice also identified an association between KRAS mutant tumor status and response to pemetrexed. The expression of a KRAS driven folate/purine synthesis gene, Methylenetetrahydrofolate Dehydrogenase 2 (MTHFD2), was also correlated with antifolate activity suggesting its use as a possible biomarker of response to antifolates. We propose that KRAS mutation drives increased purine synthesis activity and as a result an elevated dependency on the factors needed to feed this biosynthetic pathway such as those generated by folate metabolism. Thus, antifolates can indirectly inhibit purine synthesis through the depletion of folate cofactors which may account for the stronger response to these agents in KRAS mutant cells. We are currently expanding this study to examine alternative inhibitors of purine synthesis as possible therapeutics in KRAS mutant NSCLC and other cancers. Collectively, our findings highlight that a better understanding of the molecular mechanisms underlying the dependency of cancer cells on specific metabolic pathways may result in more effective metabolic targeting and new approaches in treating specific cancers. Citation Format: Diarmuid M. Moran, Patricia B. Trusk, Karen Pry, David Sidransky, Keren Paz, Sarah S. Bacus. KRAS mutation status is associated with enhanced dependency on purine biosynthesis and related pathways in non small cell lung cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4410. doi:10.1158/1538-7445.AM2014-4410

KRAS gene mutation is linked to poor prognosis and resistance to oncology therapeutics in Non Small Cell Lung Cancer (NSCLC). We have explored the possibility of exploiting inherent differences in KRAS mutant cell metabolism to enhance the efficacy of treatment. We have identified a greater dependency on purine biosynthesis and related pathways in KRAS mutant compared to KRAS wild type NSCLC cell lines. Purine synthesis requires factors generated from other metabolic reactions including ribose-5-phosphate from the pentose phosphate pathway, THF cofactors from folate metabolism and glycine / amide nitrogen groups from glutamine and aspartate metabolism. In this study, microarray gene expression and biological pathway analysis identified higher expression of purine synthesis and accessory pathways such as folate metabolism, 5-aminoimidazole ribonucleotide biosynthesis and glycine synthesis pathways in KRAS mutant NSCLC cells compared to wildtype counterparts. KRAS knockdown and overexpression studies demonstrated the ability of KRAS to regulate expression of genes that comprise purine synthesis and folate metabolism pathways. Moreover, pathway analysis and knockdown studies suggest a role for MYC, an oncogene previously recognized to be associated with KRAS mutant tumors, in the regulation of these pathways in KRAS mutant NSCLC cells. Proliferation studies demonstrated higher responsiveness to antifolates such as methotrexate and pemetrexed in KRAS mutant NSCLC cells, both of which may interfere indirectly with purine biosynthesis. In vivo analysis of NSCLC tumorgraft models in nude mice also identified an association between KRAS mutant tumor status and response to pemetrexed. The expression of a KRAS driven folate/purine synthesis gene, Methylenetetrahydrofolate Dehydrogenase 2 (MTHFD2), was also correlated with antifolate activity suggesting its use as a possible biomarker of response to antifolates. We propose that KRAS mutation drives increased purine synthesis activity and as a result an elevated dependency on the factors needed to feed this biosynthetic pathway such as those generated by folate metabolism. Thus, antifolates can indirectly inhibit purine synthesis through the depletion of folate cofactors which may account for the stronger response to these agents in KRAS mutant cells. We are currently expanding this study to examine alternative inhibitors of purine synthesis as possible therapeutics in KRAS mutant NSCLC and other cancers. Collectively, our findings highlight that a better understanding of the molecular mechanisms underlying the dependency of cancer cells on specific metabolic pathways may result in more effective metabolic targeting and new approaches in treating specific cancers. Citation Format: Diarmuid M. Moran, Patricia B. Trusk, Karen Pry, Keren Paz, David Sidransky, Sarah S. Bacus. KRAS mutation status is associated with enhanced dependency on purine biosynthesis and related pathways in non small cell lung cancer cells. [abstract]. In: Proceedings of the AACR Special Conference on RAS Oncogenes: From Biology to Therapy; Feb 24-27, 2014; Lake Buena Vista, FL. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(12 Suppl):Abstract nr B25. doi: 10.1158/1557-3125.RASONC14-B25

This study aimed to confirm whether Kirsten rat sarcoma viral oncogene (KRAS) mutations affect the therapeutic efficacy of non-small cell lung cancer (NSCLC) and, if so, to explore what the possible mechanisms might be. We retrospectively analyzed the efficacy of immunochemotherapy in KRAS-mutant NSCLC patients compared to driver-negative patients. Online data platforms were used to find immunotherapy cases, and survival analysis compared treatments' efficacy. Cytotoxicity assays measured chemosensitivity in KRAS-mutant versus wild-type NSCLC to drugs like paclitaxel, carboplatin, and pemetrexed. Bioinformatics confirmed the KRAS-SLC7A11 link and cell experiments tested SLC7A11's role in chemoresistance. Animal studies verified the antitumor effects of SLC7A11 inhibitors with chemotherapy. Patients with KRAS-mutated NSCLC have a shorter therapeutic effectiveness duration with immunochemotherapy than patients with driver gene-negative status. The efficacy of immunotherapy alone is similar between the two groups. The KRAS mutation can enhance chemoresistance by upregulating SLC7A11, and inhibiting SLC7A11 can increase the sensitivity of KRAS-mutated NSCLC to chemotherapy. This study suggests that KRAS-mutant NSCLC can enhance its acquired chemoresistance by overexpressing SLC7A11, leading to poorer therapeutic outcomes. Targeting the KRAS-SLC7A11 axis could increase sensitivity to chemotherapeutic drugs, providing theoretical support for future treatment directions.

Immune-Related Adverse Events and Outcomes in Patients with Advanced Non–Small Cell Lung Cancer: A Predictive Marker of Efficacy?

East Asian, including Thailand, lung cancer population may have a relatively lower prevalence of KRAS mutations than Caucasians. We investigated the prevalence and clinical characteristics of KRAS-driven non-small cell lung cancer (NSCLC) patients and the expression of cyclin D1, one of the KRAS downstream targets. Lung cancer patients who received treatment at the King Chulalongkorn Memorial Hospital between January 2015 and July 2017 were enrolled. We identified KRAS mutations using allele specific PCR KRAS mutation testing. Cyclin D1 expression was determined using immunohistochemistry. After excluding 376 EGFR mutations and inadequate samples, we enrolled 95 patients eligible for KRAS mutation testing. KRAS mutations were identified in 28 out of 95 patients. There were 26 patients with KRAS codon 12/13 and 2 patients with KRAS codon 61 mutations. The prevalence of KRAS mutations among informative samples was 28 out of 357 (7.8%) which was relatively lower than that reported in Caucasian population. Smoking and male were significantly associated with KRAS mutations. The prognosis of KRAS-mutant NSCLC patients in particular codon 61 mutations was worse than that found in KRAS- and EGFR-wild-type (KRAS WT/EGFR WT) NSCLC patients (P=0.048). The levels of cyclin D1 expression in KRAS-mutant NSCLC were significantly higher than those in KRAS WT/EGFR WT NSCLC (P=0.02). A better prognosis of KRAS-mutant NSCLC patients with low cyclin D1 expression was observed when compared with those with high cyclin D1 expression (median overall survival 41.7 vs. 3.5 months, P=0.037). We found a moderate prevalence of KRAS mutations in lung cancer in Thailand. Clinical characteristics were similar to those of Caucasian population. Most KRAS-mutant NSCLC had high cyclin D1 expression. Cyclin D1 expression may serve as a useful prognostic biomarker in KRAS-mutant lung cancer. Validation of this finding in larger cohort is required.

KRAS mutations are one of the most common driver mutations in non-small-cell lung cancer (NSCLC) and finding druggable target molecules to inhibit oncogenic KRAS signaling is a significant challenge in NSCLC therapy. We recently identified epiregulin (EREG) as one of several putative transcriptional targets of oncogenic KRAS signaling in both KRAS-mutant NSCLC cells and immortalized bronchial epithelial cells expressing ectopic mutant KRAS. In the current study, we found that EREG is overexpressed in NSCLCs harboring KRAS, BRAF or EGFR mutations compared with NSCLCs with wild-type KRAS/BRAF/EGFR. Small interfering RNAs (siRNAs) targeting mutant KRAS, but not an siRNA targeting wild-type KRAS, significantly reduced EREG expression in KRAS-mutant and EREG-overexpressing NSCLC cell lines. In these cell lines, EREG expression was downregulated by MEK and ERK inhibitors. Importantly, EREG expression significantly correlated with KRAS expression or KRAS copy number in KRAS-mutant NSCLC cell lines. Further expression analysis using 89 NSCLC specimens showed that EREG was predominantly expressed in NSCLCs with pleural involvement, lymphatic permeation or vascular invasion and in KRAS-mutant adenocarcinomas. In addition, multivariate analysis revealed that EREG expression is an independent prognostic marker and EREG overexpression in combination with KRAS mutations was associated with an unfavorable prognosis for lung adenocarcinoma patients. In KRAS-mutant and EREG overexpressing NSCLC cells, siRNA-mediated EREG silencing inhibited anchorage-dependent and -independent growth and induced apoptosis. Our findings suggest that oncogenic KRAS-induced EREG overexpression contributes to an aggressive phenotype and could be a promising therapeutic target in oncogenic KRAS-driven NSCLC.

Background: KRAS mutations are frequently found in non-small cell lung cancer (NSCLC). The KRAS mutations could be predictive of resistance to targeted therapy like epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) or a prognostic factor in NSCLC. No successful targeted therapy currently exists to treat patients with KRAS mutant NSCLC, and novel therapeutic strategies are needed to prolong survival in patients with KRAS mutant NSCLC. Nicotinamide phosphoribosyltransferase (nampt) is a rate-limiting enzyme in NAD+ salvage, and we previously reported that nampt is a potent therapeutic target in NSCLC. Based on the analysis result of GEO database, we found that expression of nampt is associated with RAS-MAPK signaling pathway in NSCLC, and we therefore, conducted an in vitro study to evaluate a role of nampt in growth of KRAS mutant NSCLC. Methods: We used a panel of KRAS mutant NSCLC cell lines (A427, A549, H157, H23, H1792, H2009, H358, H441, Calu6, Calu1). The cells were treated with a nampt inhibitor, FK866, and acid phosphatase (APH) assay (Thermo SCIENTIFIC) was used to evaluate cell viability of the KRAS mutant cells. Changes in expression of MAPK signaling pathway and BCL2 family proteins (antibodies from Cell Signaling and Santa Cruz) were evaluated by western blot. Apoptosis of the cells treated with FK866 was detected using an Annexin V FITC apoptosis detection kit (BD Biosciences). FACS analysis was performed using a BD FACSAria II cytometer (BD Bioscience). Autophagic proteolysis was studied by Cyto-ID autophagy detection kit (Cosmo Bio), and the expression of the fluorescent signal was visualized by a FV1000 confocal microscope (OLYMPUS JAPAN). Results: We found that the sensitivity to FK866 was different among the KRAS mutant cell lines, and for instance, A427 cells were sensitive while H157 cells were resistant. FK866 suppressed growth of KRAS mutant NSCLC and decreased expression of p-ERK expression in the cell line sensitive to FK866. We will report alteration of apoptosis and autophagy in the KRAS mutant NSCLC treated with FK866. Conclusion: Nampt could be a potent therapeutic target in KRAS mutant NSCLC. Citation Format: Shunsuke Okumura, Takaaki Sasaki, Yoshinobu Ohsaki. A role of nicotinamide phosphoribosyltransferase in growth of KRAS mutant non-small cell lung cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1770. doi:10.1158/1538-7445.AM2015-1770

BackgroundThe KRAS gene is a member of the Ras protein family. KRAS mutations are prevalent oncogenic drivers in non-small cell lung cancer (NSCLC), even in the Chinese populations, where they account for 10–15% of cases, and are correlated with aggressive disease and poor clinical outcomes. Despite recent breakthroughs in direct KRASG12C inhibitors (e.g., sotorasib and adagrasib), the therapeutic efficacy of these inhibitors remains limited; the median progression-free survival of KRAS-mutated NSCLC patients rarely exceeds 6 months, and actionable therapies for non-G12C variants are lacking. Based on this foundation, the study aimed to identify novel potential targeted therapeutic strategies for patients with KRAS-mutant NSCLC through systematic target screening.MethodsThe following three-step process and selection criteria were employed to identify the KRAS function-related genes: (I) the genes were differentially expressed in the NSCLC tissues compared to the normal tissues; (II) the differentially expressed genes were highly expressed in the KRAS-mutated tissues compared to the KRAS-wild-type NSCLC tissues in The Cancer Genome Atlas (TCGA) cohort; and (III) the genes had a dependency score of >0.9 in the KRAS-mutated NSCLC cell lines from the Cancer Cell Line Encyclopedia based on the dataset. We then examined the prognostic value of the KRAS function-related genes in TCGA cohort (504 lung adenocarcinoma samples) and validated their prognostic value in the GSE72094 dataset (398 lung adenocarcinoma samples). Additionally, we conducted a gene set enrichment analysis (GSEA) to investigate the potential mechanisms by which the candidate genes affected prognosis, and performed a drug sensitivity analysis to identify compounds exhibiting sensitivity to candidate gene expression levels in the KRAS-mutated NSCLC cell lines.Results We identified four KRAS function-related genes and showed that ATR expression was significantly associated with overall survival (P=0.008). After adjusting for age, gender and TNM stage (Tumor-Node-Metastasis stage), high ATR expression remained an independent predictor of a worse prognosis in KRAS-mutated NSCLC [hazard ratio (HR) =2.192; P=0.01] in the TCGA cohort, and this prognostic significance was validated in the GSE72094 dataset (HR =2.06; P=0.02). The GSEA results showed that the enriched genes in the ATR high-expression group were significantly associated with ubiquitin mediated proteolysis, pathways in cancer, and the mitogen-activated protein kinase signaling pathway compared to the ATR low-expression group. Additionally, the drug sensitivity analysis identified two compounds (i.e., AZ20 and AZD6738) that were sensitive to ATR expression.ConclusionsATR holds promise as both a prognostic marker and therapeutic target for KRAS-mutated NSCLC. Our findings may assist in the prediction of prognosis and the development of novel targeted therapies for this disease.

BackgroundKirsten rat sarcoma viral oncogene homolog (KRAS) mutation seemingly suffered less effective therapeutic regimens in the absence of widely-accepted targeted drugs compared with other mutation types in non-small cell lung cancer (NSCLC). However, whether these non-selective therapy schedules for KRAS mutation matters is still under debate. Correspondingly, we aimed to compare the long term expectancy of indicated therapeutic regimes and further explore the optimal schemes of KRAS mutated NSCLC in the absence of targeted drugs in this retrospective study cohort.MethodsWe conducted a single-center retrospective analysis among 66 patients diagnosed with KRAS-mutant advanced NSCLC from November 2018 to December 2020. These enrolled cases were divided into different subgroups in light of mutant isotypes, pathological characteristics, and therapeutic regimes to uncover indicated long-term survival benefits. Additionally, clinical outcomes of treatment schedules and interventional lines to KRAS-mutant NSCLC were described in detail.ResultsThis cohort enrolled 8 patients with stage IIIB (12.1%) and 58 patients with stage IV (87.9%) with the median age 62 years, ranging from 32 to 91 years old. Genetically, G12C conducted as the most common KRAS mutation type, accounting for 30.3%. Pemetrexed combined with platinum chemotherapy seemed to be a priority (72.7%), and chemotherapy combined with immunotherapy became an alternative (15.2%) in clinic. Performing further analysis of long-term survival of patients receiving different treatment methods indicated that the median overall survival (mOS) in first-line therapy with antiangiogenesis or untreated was 13 and 12 months, respectively (P=0.79). In the first-line regimen, median survival was 17 months for patients who received combined immune checkpoint inhibitors and 12 months for those who did not (P=0.34). The mOS was 20 months for those who had used immune checkpoint inhibitors and 12 months for those who had not (P=0.11). Survival analysis results of NSCLC patients with different KRAS mutation types showed the median survival time of patients with G12C mutation type and patients without with nonG12C mutation type was 19 and 12 months, respectively (P=0.37).ConclusionsIn the absence of KRAS targeted drugs, available treatment plans failed to benefit KRAS mutant sufferers regardless of isotypes, making the KRAS-targeted drugs urgent.

ERK1/2, a key downstream effector of RAS mutations, is involved in the signaling network which drives cell proliferation, survival, metastasis and cancer resistance to drug treatment (including MEK and BRAF inhibitors). Lung cancer is a leading cause of cancer death worldwide. KRAS mutation present in up to 30% of NSCLC patients is associated with a poor prognosis and represents an unmet medical need. In KRAS mutant NSCLC, enhanced ERK activation cooperates with dysregulation of the cell cycle checkpoint (e.g., cyclin D, CDK4 and CDK6 complex), and contributes to tumor progression; thus, the simultaneous inhibition of ERK and the CDK4/6 pathway is hypothesized to augment tumor growth inhibition. LY3214996, a novel and highly selective small molecule inhibitor of ERK1 and ERK2, is currently in phase I clinical trial and has been shown to inhibit cell proliferation in RAS or BRAF mutant tumor cells in vitro and xenograft tumor growth in vivo. Abemaciclib, a CDK4 and CDK6-selective inhibitor is currently in phase III studies for ER positive breast cancer and KRAS mutant NSCLC. In this study we explore the potential efficacy of combined inhibition of ERK1/2 and CDK4 and CDK6 in KRAS mutant NSCLC. The combination of LY3214996 and abemaciclib synergistically inhibited cell proliferation in 85% of KRAS mutant cells in an unbiased NSCLC panel. Combination treatment with LY3214996 and abemaciclib significantly decreased levels of phospho- p90RSK, phospho-Rb, phospho-S6 and Ki67; and synergistically inhibited cell proliferation and survival in KRAS mutant NSCLC cell lines including NCI-H2122 (G-12C), A549 (G-12S) and NCI-H441 (G-12V). Subsequent in vivo studies showed that the combination treatment with LY3214996 and abemaciclib was well tolerated and led to more robust tumor growth inhibition or regression in all KRAS mutant NSCLC xenograft models (H2122, A549 and H441) compared with either single agent treatment (p≤0.002). Furthermore, in xenograft tumors the combination of LY3214996 and abemaciclib resulted in more significant reduction of phospho-p90RSK, phospho-Rb, phospho-S6 and Ki67 in H2122 tumors compared with either single agent. Overall, the combined inhibition of ERK1/2 and CDK4 and CDK6 was tolerated and enhanced antitumor efficacy in several KRAS mutant NSCLC preclinical models. These data support the feasibility of combining ERK inhibitor LY3214996 with CDK4 and CDK6 inhibitor abemaciclib as a promising strategy for the treatment of KRAS mutant NSCLC patients, and provides the rationale for the combination study in the on-going phase I LY3214996 clinic trial (NCT02857270). Citation Format: Wenjuan Wu, Shripad V. Bhagwat, Constance King, Susan Pratt, Xueqian Gong, Julie Stewart, Bonita Jones, Robert Flack, Richard Beckman, Beverly Falcon, Jason Manro, William T. McMillen, Ramon V. Tiu, Sheng-Bin Peng, Christoph Reinhard, Sajan Joseph, Sean Buchanan. Combination of a novel ERK1/2 inhibitor (LY3214996) with CDK4 and CDK6 inhibitor (abemaciclib) enhances antitumor efficacy in KRAS mutant non-small cell lung cancer (NSCLC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 317. doi:10.1158/1538-7445.AM2017-317