Dual Targeting Of MEK Research Articles

Dual targeting of MEK and PI3K effectively controls the proliferation of human EGFR-TKI resistant non-small cell lung carcinoma cell lines with different genetic backgrounds

BackgroundMolecular targeted therapy for non-small cell lung carcinoma (NSCLC) is restricted due to resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs). This study evaluated the effects of dual targeting of MEK and PI3K in human EGFR-TKI resistant NSCLC cell lines.MethodsEGFR-TKI resistant NSCLC cell lines H1975, H460, and A549, with different mutation and amplification status in EGFR, K-RAS, PIK3CA, and MET genes, were treated with a MEK162 (MEK inhibitor) and BKM120 (PI3K inhibitor) combination or a BIBW2992 (EGFR inhibitor) and ARQ197 (MET inhibitor) combination and assayed for cell proliferation, apoptosis, and cell cycle distribution.ResultsDual targeting of MEK and PI3K efficiently inhibited the cell proliferation, induced apoptosis and the G0/G1 cell cycle, and decreased the phosphorylation of ERK1/2, AKT, S6, and 4E-BP1. H460 cells with K-RAS and PIK3CA mutation were most sensitive to MEK162 and BKM120 combinations. H1975 cells with EGFR and PIK3CA mutation and MET amplification were sensitive to BIBW2992 and ARQ197 combinations.ConclusionDual targeting regulated the proliferation of EGFR-TKI-resistant NSCLC cells, especially mutants in K-RAS and PIK3CA that are promising for EGFR-TKI-resistant NSCLC therapeutics.

Blocking Y-Box Binding Protein-1 through Simultaneous Targeting of PI3K and MAPK in Triple Negative Breast Cancers.

Simple SummaryTriple-negative breast cancer (TNBC) is associated with the high rates of relapse and metastasis and poor survival. YB-1 is overexpressed in TNBC tumor tissues. In the present study, we demonstrated that S102 phosphorylation of YB-1 in TNBC cell lines depend on the mutation status of the components of the MAPK/ERK and PI3K/Akt pathways. Simultaneous targeting of MEK and PI3K was found to be the most effective approach to block YB-1 phosphorylation and to inhibit YB-1 dependent cell proliferation. YBX1 knockout was sufficient to block TNBC tumor growth.The multifunctional protein Y-box binding protein-1 (YB-1) regulates all the so far described cancer hallmarks including cell proliferation and survival. The MAPK/ERK and PI3K/Akt pathways are also the major pathways involved in cell growth, proliferation, and survival, and are the frequently hyperactivated pathways in human cancers. A gain of function mutation in KRAS mainly leads to the constitutive activation of the MAPK pathway, while the activation of the PI3K/Akt pathway occurs either through the loss of PTEN or a gain of function mutation of the catalytic subunit alpha of PI3K (PIK3CA). In this study, we investigated the underlying signaling pathway involved in YB-1 phosphorylation at serine 102 (S102) in KRAS(G13D)-mutated triple-negative breast cancer (TNBC) MDA-MB-231 cells versus PIK3CA(H1047R)/PTEN(E307K) mutated TNBC MDA-MB-453 cells. Our data demonstrate that S102 phosphorylation of YB-1 in KRAS-mutated cells is mainly dependent on the MAPK/ERK pathway, while in PIK3CA/PTEN-mutated cells, YB-1 S102 phosphorylation is entirely dependent on the PI3K/Akt pathway. Independent of the individual dominant pathway regulating YB-1 phosphorylation, dual targeting of MEK and PI3K efficiently inhibited YB-1 phosphorylation and blocked cell proliferation. This represents functional crosstalk between the two pathways. Our data obtained from the experiments, applying pharmacological inhibitors and genetic approaches, shows that YB-1 is a key player in cell proliferation, clonogenic activity, and tumor growth of TNBC cells through the MAPK and PI3K pathways. Therefore, dual inhibition of these two pathways or single targeting of YB-1 may be an effective strategy to treat TNBC.

Dual-targeting of MEK and CDK4/6 in KRAS-mutated Non-Small Cell Lung Cancer

Discovering new ways to modulate the responsiveness of non-small cell lung cancer (NSCLC) to treatment represents one of the forefronts of cancer research. KRAS mutations, present in roughly 30% of NSCLCs, lead to increased levels of cellular proliferation and decreased responsiveness to treatment modalities. The present study examined the RB/p16/CDK4 pathway as a way to modulate the responsiveness of these cell lines. The diverse downstream effects of these pathways suggest that molecular profiling of NSCLCs may assist clinicians and researchers to predict the effectiveness in the application of particular therapeutics.

Phenformin enhances the therapeutic effect of selumetinib in KRAS-mutant non-small cell lung cancer irrespective of LKB1 status.

MEK inhibition is potentially valuable in targeting KRAS-mutant non-small cell lung cancer (NSCLC). Here, we analyzed whether concomitant LKB1 mutation alters sensitivity to the MEK inhibitor selumetinib, and whether the metabolism drug phenformin can enhance the therapeutic effect of selumetinib in isogenic cell lines with different LKB1 status. Isogenic pairs of KRAS-mutant NSCLC cell lines A549, H460 and H157, each with wild-type and null LKB1, as well as genetically engineered mouse-derived cell lines 634 (krasG12D/wt/p53-/-/lkb1wt/wt) and t2 (krasG12D/wt/p53-/-/lkb1-/-) were used in vitro to analyze the activities of selumetinib, phenformin and their combination. Synergy was measured and potential mechanisms investigated. The in vitro findings were then confirmed in vivo using xenograft models. The re-expression of wild type LKB1 increased phospho-ERK level, suggesting that restored dependency on MEK->ERK->MAPK signaling might have contributed to the enhanced sensitivity to selumetinib. In contrast, the loss of LKB1 sensitized cells to phenformin. At certain combination ratios, phenformin and selumetinib showed synergistic activity regardless of LKB1 status. Their combination reduced phospho-ERK and S6 levels and induced potent apoptosis, but was likely through different mechanisms in cells with different LKB1 status. Finally, in xenograft models bearing isogenic A549 cells, we confirmed that loss of LKB1 confers resistance to selumetinib, and phenformin significantly enhances the therapeutic effect of selumetinib. Irrespective of LKB1 status, phenformin may enhance the anti-tumor effect of selumetinib in KRAS-mutant NSCLC. The dual targeting of MEK and cancer metabolism may provide a useful strategy to treat this subset of lung cancer.

Abstract 3018: Cotargeting MEK and CDK4/6 to treat pancreatic adenocarcinoma

Abstract Hyperactivation of KRAS and inactivation of CDKN2A [p16] play a prominent role in tumor initiation and progression in a broad spectrum of human cancers. In pancreatic adenocarcinoma, KRAS mutation occurs in 90-95% of cases and is often coupled with inactivation of CDKN2A (>90% incidence), typically by homozygous deletion. Based on the high incidence of these genetic events, the present study addresses the hypothesis that dual targeting of MEK and CDK4/6 represents a viable therapeutic strategy for the treatment of pancreatic adenocarcinoma. A panel of primary and high passage xenograft models was screened for in vitro synergy to the antiproliferative effects of the MEK inhibitor trametinib and the CDK4/6 inhibitor palbociclib. Two models that emerged as highly sensitive to this combination treatment strategy, L3.6pl and UM59, both exhibited G1 cell cycle arrest that was significantly more pronounced in response to the combination compared to either single agent. In vivo studies were subsequently carried out to evaluate the efficacy of this combination in tumor-bearing mice. Mice implanted subcutaneously with L3.6pl or UM59 cells were treated daily by oral gavage for ten days with trametinib (3 mg/kg), palbociclib (100 mg/kg), or the combination of these two agents at these doses. Consistent with in vitro synergy observations, both models proved to be exceptional responders in vivo to this combination treatment strategy, showing a robust response to the combination not seen with either single agent. These results suggest that the combined inhibition of MEK and CDK4/6 may offer a valuable therapeutic strategy for the treatment of pancreatic adenocarcinoma. Studies are ongoing to identify molecular determinants of response in pancreatic tumors that are highly sensitive to this combination approach to guide future patient enrichment strategies. Citation Format: Joel D. Maust, Diane Simeone, Judith Sebolt-Leopold. Cotargeting MEK and CDK4/6 to treat pancreatic adenocarcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3018.

Abstract 4695: Dual targeting of MEK and PI3K pathways can act via tumor-intrinsic mechanisms to overcome resistance and tumor-extrinsic mechanisms to modulate immunity and limit cancer cachexia

Abstract Prior studies by our group have shown that single-agent MEK inhibitors have clinical activity in cholangiocarcinoma (CC), including objective responses. Yet no relationship was found between clinical benefit, and oncogenic driver mutations such as KRAS or BRAF. Strategies to expand upon MEKi have focused on combinations with agents targeting resistance, such as the PI3K/Akt pathway. CC patients treated with MEKi also had reduced pro-inflammatory cytokines and weight gain with restoration of muscle mass. These findings are clinically significant, and led us to hypothesize that MEKi act via tumor-extrinsic mechanisms to modulate immunity and limit cancer cachexia, while overcoming tumor-intrinsic resistance. We used the colon-26 adenocarcinoma model to evaluate the effect of single and combined treatment with MEK162 (30mg/kg) and buparlisib (25mg/kg) on muscle wasting, tumor growth, and immune modulation. This murine model depends on interleukin-6 (IL-6) and recapitulates the cancer cachexia syndrome. Single agent MEK162 inhibited growth initially, but after 14 days, tumor volume was comparable between MEKi and vehicle treated mice, possibly from acquired MEK162 resistance. Despite these data, reduced serum IL-6 and splenic Gr1+CD11b+ cells were evident compared to control mice (mean = 358 pg/mL to 43 pg/mL, and 13.8% to 8.4%, respectively), while body weight from MEK162 treated mice was spared (mean = 24.0g to 20.5g in control mice). In addition, markers of muscle catabolism, including the E3 ubiquitin ligases Atrogin-1 (from 1 to 0.043 in combo) and MuRF1 (from 1 to 0.03 in combo), and the autophagy gene, Bnip3, were all reduced in tibialis anterior muscles from tumor-bearing mice treated with MEK162. Similar to our clinical data, these results suggest MEK162 modulates immune biomarkers, and acts as an anti-cachexia agent. Since cross talk between MAPK and PI3K/AKT pathways promotes resistance to monotherapy, we next investigated the effects of MEK162 combined with buparlisib in the colon-26 model. As before, single agent MEK162 had a modest growth inhibitory effect, yet rescued body weight. Consistent with in vitro studies, single agent buparlisib was cytostatic, but not as effective as MEK162 in rescuing weight loss. However, dual treatment with MEK162 + buparlisib significantly inhibited tumor growth. This combination also significantly reduced splenic MDSC (mean = 37% to 9% in control mice) and increased CD4+ and CD8+ (mean = 24% to 6%, and 7.7% to 2%, respectively) T cells. These results suggest that dual inhibition of MEK and PI3K has efficacy, while MEKi may have under-appreciated tumor-extrinsic mechanisms of activity that can be leveraged to benefit patients with advanced malignancy. This treatment combination will be evaluated in CC patients in the setting of a Phase II clinical trial. Citation Format: Jennifer Yang, Erin Talbert, Omar Elnaggar, Priyani Rajasekera, Thomas Mace, Matthew Farren, Zheng Che, Benjamin Swanson, Gregory Young, Ericka Haverick, Cynthia Timmers, Mark Bloomston, Tanios Bekaii-Saab, Denis Guttridge, Gregory Lesinski. Dual targeting of MEK and PI3K pathways can act via tumor-intrinsic mechanisms to overcome resistance and tumor-extrinsic mechanisms to modulate immunity and limit cancer cachexia. [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 4695. doi:10.1158/1538-7445.AM2015-4695

Dual Targeting of MEK and PI3K Pathways Attenuates Established and Progressive Pulmonary Fibrosis

Pulmonary fibrosis is often triggered by an epithelial injury resulting in the formation of fibrotic lesions in the lung, which progress to impair gas exchange and ultimately cause death. Recent clinical trials using drugs that target either inflammation or a specific molecule have failed, suggesting that multiple pathways and cellular processes need to be attenuated for effective reversal of established and progressive fibrosis. Although activation of MAPK and PI3K pathways have been detected in human fibrotic lung samples, the therapeutic benefits of in vivo modulation of the MAPK and PI3K pathways in combination are unknown. Overexpression of TGFα in the lung epithelium of transgenic mice results in the formation of fibrotic lesions similar to those found in human pulmonary fibrosis, and previous work from our group shows that inhibitors of either the MAPK or PI3K pathway can alter the progression of fibrosis. In this study, we sought to determine whether simultaneous inhibition of the MAPK and PI3K signaling pathways is a more effective therapeutic strategy for established and progressive pulmonary fibrosis. Our results showed that inhibiting both pathways had additive effects compared to inhibiting either pathway alone in reducing fibrotic burden, including reducing lung weight, pleural thickness, and total collagen in the lungs of TGFα mice. This study demonstrates that inhibiting MEK and PI3K in combination abolishes proliferative changes associated with fibrosis and myfibroblast accumulation and thus may serve as a therapeutic option in the treatment of human fibrotic lung disease where these pathways play a role.