Targeting CDK4 overcomes EMT-mediated tumor heterogeneity and therapeutic resistance in KRAS-mutant lung cancer.
Ontology highlight
ABSTRACT:
SUBMITTER: Padhye A
PROVIDER: S-EPMC8492319 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Evolved resistance to tyrosine kinase inhibitor (TKI)-targeted therapies remains a major clinical challenge. In epidermal growth factor receptor (EGFR) mutant non-small-cell lung cancer (NSCLC), failure of EGFR TKIs can result from both genetic and epigenetic mechanisms of acquired drug resistance. Widespread reports of histologic and gene expression changes consistent with an epithelial-to-mesenchymal transition (EMT) have been associated with initially surviving drug-tolerant persister cells, which can seed bona fide genetic mechanisms of resistance to EGFR TKIs. While therapeutic approaches targeting fully resistant cells, such as those harboring an EGFRT790M mutation, have been developed, a clinical strategy for preventing the emergence of persister cells remains elusive. Using mesenchymal cell lines derived from biopsies of patients who progressed on EGFR TKI as surrogates for persister populations, we performed whole-genome CRISPR screening and identified fibroblast growth factor receptor 1 (FGFR1) as the top target promoting survival of mesenchymal EGFR mutant cancers. Although numerous previous reports of FGFR signaling contributing to EGFR TKI resistance in vitro exist, the data have not yet been sufficiently compelling to instigate a clinical trial testing this hypothesis, nor has the role of FGFR in promoting the survival of persister cells been elucidated. In this study, we find that combining EGFR and FGFR inhibitors inhibited the survival and expansion of EGFR mutant drug-tolerant cells over long time periods, preventing the development of fully resistant cancers in multiple vitro models and in vivo. These results suggest that dual EGFR and FGFR blockade may be a promising clinical strategy for both preventing and overcoming EMT-associated acquired drug resistance and provide motivation for the clinical study of combined EGFR and FGFR inhibition in EGFR-mutated NSCLCs.
| S-EPMC6742540 | biostudies-literature
Project description:KRAS mutant lung adenocarcinomas remain intractable for targeted therapies. Genetic interrogation of KRAS downstream effectors, including the MAPK pathway and the interphase CDKs, identified CDK4 and RAF1 as the only targets whose genetic inactivation induces therapeutic responses without causing unacceptable toxicities. Concomitant CDK4 inactivation and RAF1 ablation prevented tumor progression and induced complete regression in 25% of KRAS/p53-driven advanced lung tumors, yet a significant percentage of those tumors that underwent partial regression retained a population of CDK4/RAF1-resistant cells. Characterization of these cells revealed two independent resistance mechanisms implicating hypermethylation of several tumor suppressors and increased PI3K activity. Importantly, these CDK4/RAF1-resistant cells can be pharmacologically controlled. These studies open the door to new therapeutic strategies to treat KRAS mutant lung cancer, including resistant tumors.
| S-EPMC7533841 | biostudies-literature
Project description:Non-small cell lung cancers (NSCLCs) that harbor an oncogenic KRAS mutation are often associated with resistance to targeted therapies. The MUC1-C transmembrane protein is aberrantly overexpressed in NSCLCs and confers a poor outcome; however, the functional role for MUC1-C in mutant KRAS NSCLC cells has remained unclear. The present studies demonstrate that silencing MUC1-C in A549/KRAS(G12S) and H460/KRAS(Q61H) NSCLC cells is associated with downregulation of AKT signaling and inhibition of growth. Overexpression of a MUC1-C(CQC?AQA) mutant, which inhibits MUC1-C homodimerization and function, suppressed both AKT and MEK activation. Moreover, treatment with GO-203, an inhibitor of MUC1-C homodimerization, blocked AKT and MEK signaling and decreased cell survival. The results further demonstrate that targeting MUC1-C suppresses expression of the ZEB1 transcriptional repressor by an AKT-mediated mechanism, and in turn induces miR-200c. In concert with these effects on the ZEB1/miR-200c regulatory loop, targeting MUC1-C was associated with reversal of the epithelial-mesenchymal transition (EMT) and inhibition of self-renewal capacity. Loss of MUC1-C function also attenuated KRAS independence and inhibited growth of KRAS mutant NSCLC cells as tumors in mice. These findings support a model in which targeting MUC1-C inhibits mutant KRAS signaling in NSCLC cells and thereby reverses the EMT phenotype and decreases self-renewal.
| S-EPMC4253405 | biostudies-literature
Project description:CDK4/cyclin D kinase constitutes an attractive pharmacological target for development of anticancer therapeutics, in particular in KRAS-mutant lung cancer patients, who have a poor prognosis and no targeted therapy available yet. Although several ATP-competitive inhibitors of CDK4 have been developed for anticancer therapeutics, they suffer from limited specificity and efficacy. Methods: As an alternative to ATP-competitive inhibitors we have designed a stapled peptide to target the main interface between CDK4 and cyclin D, and have characterized its physico-chemical properties and affinity to bind cyclin D1. Results: We have validated a positive correlation between CDK4/cyclin D level and KRAS mutation in lung cancer patients. The stapled peptide enters cells rapidly and efficiently, and inhibits CDK4 kinase activity and proliferation in lung cancer cells. Its intrapulmonary administration in mice enables its retention in orthotopic lung tumours and complete inhibition of their growth when co-administered with Abemaciclib. Conclusion: The stapled peptide targeting the main interface between CDK4 and cyclin D provides promising therapeutic perspectives for patients with lung cancer.
| S-EPMC7019173 | biostudies-literature
Project description:In non-small-cell lung cancer (NSCLC), concurrent mutations in the oncogene KRAS and tumor suppressor STK11 (also known as LKB1) confer an aggressive malignant phenotype, an unfavourability towards immunotherapy, and overall poor prognoses in patients. In a previous study, we showed that murine KRAS/LKB1 co-mutant tumors and human co-mutant cancer cells have an enhanced dependence on glutamine-fructose-6-phosphate transaminase 2 (GFPT2), a rate-limiting enzyme in the hexosamine biosynthesis pathway (HBP), which could be targeted to reduce survival of KRAS/LKB1 co-mutants. Here, we found that KRAS/LKB1 co-mutant cells also exhibit an increased dependence on N-acetylglucosamine-phosphate mutase 3 (PGM3), an enzyme downstream of GFPT2. Genetic or pharmacologic suppression of PGM3 reduced KRAS/LKB1 co-mutant tumor growth in both in vitro and in vivo settings. Our results define an additional metabolic vulnerability in KRAS/LKB1 co-mutant tumors to the HBP and provide a rationale for targeting PGM3 in this aggressive subtype of NSCLC.
| S-EPMC8750012 | biostudies-literature
Project description:Patients with EGFR-mutant non-small-cell lung cancer (NSCLC) have significantly benefited from the use of EGFR tyrosine kinase inhibitors (TKIs). However, long-term efficacy of these therapies is limited due to de novo resistance (~30%) as well as acquired resistance. Epithelial-mesenchymal transition transcription factors (EMT-TFs), have been identified as drivers of EMT-mediated resistance to EGFR TKIs, however, strategies to target EMT-TFs are lacking. As the third generation EGFR TKI, osimertinib, has now been adopted in the first-line setting, the frequency of T790M mutations will significantly decrease in the acquired resistance setting. Previously less common mechanisms of acquired resistance to first generation EGFR TKIs including EMT are now being observed at an increased frequency after osimertinib. Importantly, there are no other FDA approved targeted therapies after progression on osimertinib. Here, we investigated a novel strategy to overcome EGFR TKI resistance through targeting the EMT-TF, TWIST1, in EGFR-mutant NSCLC. We demonstrated that genetic silencing of TWIST1 or treatment with the TWIST1 inhibitor, harmine, resulted in growth inhibition and apoptosis in EGFR-mutant NSCLC. TWIST1 overexpression resulted in erlotinib and osimertinib resistance in EGFR-mutant NSCLC cells. Conversely, genetic and pharmacological inhibition of TWIST1 in EGFR TKI-resistant EGFR-mutant cells increased sensitivity to EGFR TKIs. TWIST1-mediated EGFR TKI resistance was due in part to TWIST1 suppression of transcription of the pro-apoptotic BH3-only gene, BCL2L11 (BIM), by directly binding to BCL2L11 intronic regions and promoter. As such, pan-BCL2 inhibitor treatment overcame TWIST1-mediated EGFR TKI resistance and were more effective in the setting of TWIST1 overexpression. Finally, in a mouse model of autochthonous EGFR-mutant lung cancer, Twist1 overexpression resulted in erlotinib resistance and suppression of erlotinib-induced apoptosis. These studies establish TWIST1 as a driver of resistance to EGFR TKIs and provide rationale for use of TWIST1 inhibitors or BCL2 inhibitors as means to overcome EMT-mediated resistance to EGFR TKIs.
| S-EPMC6358506 | biostudies-literature
Project description:KRAS is the most frequently mutated oncogene in human tumors, and its activating mutations represent important therapeutic targets. The combination of Cas9 and guide RNA from the CRISPR-Cas system recognizes a specific DNA sequence and makes a double-strand break, which enables editing of the relevant genes. Here, we harnessed CRISPR to specifically target mutant KRAS alleles in cancer cells. We screened guide RNAs using a reporter system and validated them in cancer cells after lentiviral delivery of Cas9 and guide RNA. The survival, proliferation, and tumorigenicity of cancer cells in vitro and the growth of tumors in vivo were determined after delivery of Cas9 and guide RNA. We identified guide RNAs that efficiently target mutant KRAS without significant alterations of the wild-type allele. Doxycycline-inducible expression of this guide RNA in KRAS-mutant cancer cells transduced with a lentiviral vector encoding Cas9 disrupted the mutant KRAS gene, leading to inhibition of cancer cell proliferation both in vitro and in vivo. Intra-tumoral injection of lentivirus and adeno-associated virus expressing Cas9 and sgRNA suppressed tumor growth in vivo, albeit incompletely, in immunodeficient mice. Expression of Cas9 and the guide RNA in cells containing wild-type KRAS did not alter cell survival or proliferation either in vitro and in vivo. Our study provides a proof-of-concept that CRISPR can be utilized to target driver mutations of cancers in vitro and in vivo.
| S-EPMC5848616 | biostudies-literature
Project description:Acquired drug resistance to tyrosine kinase inhibitor (TKI) targeted therapies remains a major clinical challenge. In EGFR mutant non-small cell lung cancer (NSCLC), therapeutic failure of EGFR TKIs can result from both genetic and epigenetic mechanisms of acquired drug resistance. Histologic and gene expression changes consistent with an epithelial-to-mesenchymal transition (EMT) have been associated with resistance to EGFR TKIs in both experimental models and in patients, and may coincide with genetic mechanisms of resistance such as the EGFRT790M gatekeeper mutation. While therapeutic approaches targeting EGFRT790M have been developed, a strategy for overcoming EMT-related resistance remains unclear. We performed whole-genome CRISPR screening on patient-derived, mesenchymal EGFRT790M-positive cell lines and identified FGFR1 as a critical gene promoting resistance to third generation EGFR TKIs. The FGFR1-3 inhibitor, BGJ398 (infigratinib), resensitized resistant mesenchymal-like cell lines to EGFR inhibition in a synergistic manner. Combining EGFR + FGFR inhibitors also inhibited the in vitro survival and expansion of EGFR mutant drug tolerant cells with mesenchymal-like features prior to the development of drug resistance, and delayed the development of in vivo resistance in EGFR mutant NSCLC xenograft tumors. These results suggest that dual EGFR + FGFR blockade may be a promising clinical strategy for preventing and overcoming EMT-associated acquired drug resistance in EGFR mutant NSCLC.
2019-04-30 | GSE114647 | GEO
Project description:The exact influence of statins on gefitinib resistance in human non-small cell lung cancer (NSCLC) cells with KRAS mutation alone or KRAS/PIK3CA and KRAS/PTEN comutations remains unclear. This work found that transfection of mutant KRAS plasmids significantly suppressed the gefitinib cytotoxicity in Calu3 cells (wild-type KRAS). Gefitinib disrupted the Kras/PI3K and Kras/Raf complexes in Calu3 cells, whereas not in Calu3 KRAS mutant cells. These trends were corresponding to the expression of pAKT and pERK in gefitinib treatment. Atorvastatin (1??M) plus gefitinib treatment inhibited proliferation, promoted cell apoptosis, and reduced the AKT activity in KRAS mutant NSCLC cells compared with gefitinib alone. Atorvastatin (5??M) further enhanced the gefitinib cytotoxicity through concomitant inhibition of AKT and ERK activity. Atorvastatin could interrupt Kras/PI3K and Kras/Raf complexes, leading to suppression of AKT and ERK activity. Similar results were also obtained in comutant KRAS/PTEN or KRAS/PIK3CA NSCLC cells. Furthermore, mevalonate administration reversed the effects of atorvastatin on the Kras/Raf and Kras/PI3K complexes, as well as AKT and ERK activity in both A549 and Calu1 cells. The in vivo results were similar to those obtained in vitro. Therefore, mutant KRAS-mediated gefitinib insensitivity is mainly derived from failure to disrupt the Kras/Raf and Kras/PI3K complexes in KRAS mutant NSCLC cells. Atorvastatin overcomes gefitinib resistance in KRAS mutant NSCLC cells irrespective of PIK3CA and PTEN statuses through inhibition of HMG-CoA reductase-dependent disruption of the Kras/Raf and Kras/PI3K complexes.
| S-EPMC3789171 | biostudies-literature
Project description:The KRAS gain-of-function mutation confers intrinsic resistance to targeted anti-cancer drugs and cytotoxic chemotherapeutic agents, ultimately leading to treatment failure. KRAS mutation frequency in lung adenocarcinoma is ~15-30%. Novel therapeutic strategies should be developed to improve clinical outcomes in these cases. Deregulation of the p16/cyclin-dependent kinase (CDK) 4/retinoblastoma (Rb) pathway is frequently observed in various cancers and it represents an attractive therapeutic target. We compared the anti-tumor efficacy of genetically knocked-down CDK4 and a pharmacological inhibitor of CDK4/6, CINK4, in KRAS mutation-positive lung adenocarcinoma cells. We also investigated changes in anti-proliferative activity and downstream molecules with these treatments in combination with paclitaxel. CDK4 short interfering RNA (siRNA) significantly increased paclitaxel sensitivity in KRAS mutation-positive H23 cells. CINK4 demonstrated concentration- and time-dependent anti-proliferative activity in 5 adenocarcinoma lines. CINK4 induced G 1 arrest by downregulating the p16/cyclin D1/Rb pathway, resulting in apoptotic induction via increased expression of cleaved caspase3, cleaved PARP and Bax. Combined CINK4 and paclitaxel produced synergistic anti-proliferative activity and increased apoptosis through reduced cyclin D1 and Bcl-2 in KRAS mutation-positive cancer cells. These data suggest CDK4 is a promising target for development of anti-cancer drugs and CINK4 combined with paclitaxel may be an effective therapeutic strategy for enhancing anti-tumor efficacy in KRAS mutation-positive lung adenocarcinoma.
| S-EPMC3742489 | biostudies-literature