Upregulation of ERBB3 by FOXD3, an adaptive response to RAF and MEK inhibitors
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ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Ertel Adam
PROVIDER: S-ECPF-GEOD-43962 | biostudies-other |
REPOSITORIES: biostudies-other
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Project description:To understand the transcriptional impact of FOXD3 in melanoma cells, we utilized a microarray approach. We collected RNA from three unrelated mutant B-RAF melanoma cell lines (WM115, WM793, and A375) that were engineered to inducibly express FOXD3 or the control gene, ?-galactosidase (LacZ), after 5 days of transgene induction. This time point was chosen based on maximal phenotypic changes previously observed. Comparison of gene signatures between the 3 cell lines produced approximately 2,600 common genes differentially regulated by FOXD3-expressing cells compared to the LacZ controls. Three unrelated mutant B-RAF melanoma cell lines (WM115, WM793, and A375) were induced to express FOXD3 and compared against the same cell lines expressing the control gene, ?-galactosidase (LacZ).
2013-03-31 | E-GEOD-43962 | biostudies-arrayexpress
Project description:To understand the transcriptional impact of FOXD3 in melanoma cells, we utilized a microarray approach. We collected RNA from three unrelated mutant B-RAF melanoma cell lines (WM115, WM793, and A375) that were engineered to inducibly express FOXD3 or the control gene, β-galactosidase (LacZ), after 5 days of transgene induction. This time point was chosen based on maximal phenotypic changes previously observed. Comparison of gene signatures between the 3 cell lines produced approximately 2,600 common genes differentially regulated by FOXD3-expressing cells compared to the LacZ controls.
2013-03-31 | GSE43962 | GEO
Project description:The mechanisms underlying adaptive resistance of melanoma to targeted therapies remain unclear. By combining ChIP sequencing with microarray-based gene profiling, we determined that ERBB3 is upregulated by FOXD3, a transcription factor that promotes resistance to RAF inhibitors in melanoma. Enhanced ERBB3 signaling promoted resistance to RAF pathway inhibitors in cultured melanoma cell lines and in mouse xenograft models. ERBB3 signaling was dependent on ERBB2; targeting ERBB2 with lapatinib in combination with the RAF inhibitor PLX4720 reduced tumor burden and extended latency of tumor regrowth in vivo versus PLX4720 alone. These results suggest that enhanced ERBB3 signaling may serve as a mechanism of adaptive resistance to RAF and MEK inhibitors in melanoma and that cotargeting this pathway may enhance the clinical efficacy and extend the therapeutic duration of RAF inhibitors.
| S-EPMC3635724 | biostudies-literature
Project description:ERBB3/HER3 expression and signaling are upregulated in mutant BRAF melanoma as an adaptive, prosurvival response to FDA-approved RAF inhibitors. Because compensatory ERBB3 signaling counteracts the effects of RAF inhibitors, cotargeting ERBB3 may increase the efficacy of RAF inhibitors in mutant BRAF models of melanoma. Here, we corroborate this concept by showing that the ERBB3 function-blocking monoclonal antibody huHER3-8 can inhibit neuregulin-1 activation of ERBB3 and downstream signaling in RAF-inhibited melanoma cells. Targeting mutant BRAF in combination with huHER3-8 decreased cell proliferation and increased cell death in vitro, and decreased tumor burden in vivo, compared with targeting either mutant BRAF or ERBB3 alone. Furthermore, the likelihood of a durable tumor response in vivo was increased when huHER3-8 was combined with RAF inhibitor PLX4720. Together, these results offer a preclinical proof of concept for the application of ERBB3-neutralizing antibodies to enhance the efficacy of RAF inhibitors in melanoma to delay or prevent tumor regrowth. As ERBB3 is often upregulated in response to other kinase-targeted therapeutics, these findings may have implications for other cancers as well.
| S-EPMC4120074 | biostudies-literature
Project description:BackgroundTreatment of advanced melanoma has been improved with the advent of the BRAF inhibitors. However, a limitation to such treatment is the occurrence of resistance. Several mechanisms have been identified to be responsible for the development of resistance, either MEK-dependent or MEK-independent. In order to overcome resistance due to reactivation of MEK signaling, MEK inhibitors are being clinically developed with promising results. However, also in this case resistance inevitably occurs. It has been recently reported that ErbB3, a member of the EGFR receptor family, may be involved in the establishment of drug resistance.MethodsThree melanoma cell lines were tested: LOX IMVI (BRAF V600E), MST-L (BRAF V600R) and WM266 (BRAF V600D). Phosphorylation of Receptor Tyrosine Kinases (RTKs) was assessed by an RTK array. Western blot analysis was performed on total protein extracts using anti-ErbB3, anti-AKT and anti-ERK 1/2 antibodies. The expression of neuregulin after vemurafenib treatment was assessed by Real Time PCR and Western blotting. The growth inhibitory effects of vemurafenib, GSK1120212b and/or anti-ErbB3 mAbs were evaluated by in vitro colony formation assays.ResultsIn the present study we demonstrate that ErbB3 is the main RTK undergoing rapidly hyperphosphorylation upon either treatment with a BRAF inhibitor or with a MEK inhibitor in a panel of melanoma cell lines harboring a variety of V600BRAF mutations and that this results in a strong activation of phospho-AKT. Importantly, ErbB3 activation is fully abrogated by the simultaneous use of anti-ErbB3 monoclonal antibodies, which are also shown to potently synergize with BRAF inhibitors in the inactivation of both AKT and ERK pathways and in the inhibition of melanoma cell growth. We show that upregulation of phospho-ErbB3 is due to an autocrine loop involving increased transcription and production of neuregulin by melanoma cells.ConclusionsOn the basis of these results, we propose that initial co-treatment with BRAF and/or MEK inhibitors and anti-ErbB3 antibodies should be pursued as a strategy to reduce the ErbB3-dependent feedback survival mechanism and enhance duration of clinical response.
| S-EPMC3729364 | biostudies-literature
Project description:RAF inhibitor therapy yields significant reductions in tumour burden in the majority of V600E-positive melanoma patients; however, resistance occurs within 2-18 months. Here we demonstrate that the mixed lineage kinases (MLK1-4) are MEK kinases that reactivate the MEK/ERK pathway in the presence of RAF inhibitors. Expression of MLK1-4 mediates resistance to RAF inhibitors and promotes survival in V600E-positive melanoma cell lines. Furthermore, we observe upregulation of the MLKs in 9 of 21 melanoma patients with acquired drug resistance. Consistent with this observation, MLKs promote resistance to RAF inhibitors in mouse models and contribute to acquired resistance in a cell line model. Lastly, we observe that a majority of MLK1 mutations identified in patients are gain-of-function mutations. In summary, our data demonstrate a role for MLKs as direct activators of the MEK/ERK pathway with implications for melanomagenesis and resistance to RAF inhibitors.
| S-EPMC4046110 | biostudies-literature
Project description:The discovery of activating mutations in BRAF at high frequency in cutaneous melanoma opened the door to new treatment options, which have resulted in significantly better patient outcomes. Treatments such as the FDA-approved RAF inhibitor vemurafenib and the more recently approved dabrafenib and trametinib combination therapy are designed to target the ERK1/2 pathway. Initial success in targeting this pathway is evidenced by the high percentage of melanoma patients who undergo tumor remission. However, the beneficial effects of these targeted therapies are usually short-lived due to the development of resistance, which leads to disease progression. As a result, studies have focused on the acquired forms of resistance that develop following continued exposure to therapy. Conversely, far fewer studies have investigated the adaptive forms of resistance, which activate rapidly, promote cell survival, and may underlie the development of acquired resistance by providing melanoma cells the time to develop additional mutations. We provide a detailed review of the known mechanisms of adaptive resistance in melanoma and relate them to similar responses to targeted therapies in other tumor types.
| S-EPMC4212018 | biostudies-literature
Project description:RAF and MEK inhibitors are effective in BRAF-mutant melanoma but not in BRAF-mutant colorectal cancer. To gain additional insights into this difference, we performed a genome-scale pooled shRNA enhancer screen in a BRAF-mutant, RAF inhibitor-resistant colorectal cancer cell line exposed to the selective RAF inhibitor PLX4720. We identified multiple genes along the receptor tyrosine kinase (RTK)/mitogen-activated protein kinase (MAPK) signaling axis that, when suppressed, either genetically or pharmacologically, sensitized cells to the selective RAF inhibitor through sustained inhibition of MAPK signaling. Strikingly, CRAF was a key mediator of resistance that could be overcome by the use of pan-RAF inhibitors in combination with a MEK inhibitor. Furthermore, the combination of pan-RAF and MEK inhibitors displayed strong synergy in melanoma and colorectal cancer cell lines with RAS-activating events such as RTK activation, KRAS mutation, or NF1 loss-of-function mutations. Combinations of selective RAF inhibitors, such as PLX4720 or dabrafenib, with MEK inhibitors did not incur such profound synergy, suggesting that inhibition of CRAF by pan-RAF inhibitors plays a key role in determining cellular response. Importantly, in contrast to the modest activity seen with single-agent treatment, dual pan-RAF and MEK inhibition results in the induction of apoptosis, greatly enhancing efficacy. Notably, combined pan-RAF and MEK inhibition can overcome intrinsic and acquired resistance to single-agent RAF/MEK inhibition, supporting dual pan-RAF and MEK inhibition as a novel therapeutic strategy for BRAF- and KRAS-mutant cancers.
| S-EPMC4674359 | biostudies-other
Project description:BackgroundBRAF-mutant melanoma patients respond to BRAF inhibitors and MEK inhibitors (BRAFi/MEKi), but drug-tolerant cells persist, which may seed disease progression. Adaptive activation of receptor tyrosine kinases (RTKs) has been associated with melanoma cell drug tolerance following targeted therapy. While co-targeting individual RTKs can enhance the efficacy of BRAFi/MEKi effects, it remains unclear how to broadly target multiple RTKs to achieve more durable tumour growth inhibition.MethodsThe blockage of adaptive RTK responses by the new BET inhibitor (BETi), PLX51107, was measured by RPPA and Western blot. Melanoma growth was evaluated in vitro by colony assay and EdU staining, as well as in skin reconstructs, xenografts and PDX models following BRAFi, MEKi and/or PLX51107 treatment.ResultsTreatment with PLX51107 limited BRAFi/MEKi upregulation of ErbB3 and PDGFR-β expression levels. Similar effects were observed following BRD2/4 depletion. In stage III melanoma patients, expression of BRD2/4 was strongly correlated with ErbB3. PLX51107 enhanced the effects of BRAFi/MEKi on inhibiting melanoma growth in vitro, in human skin reconstructs and in xenografts in vivo. Continuous triple drug combination treatment resulted in significant weight loss in mice, but intermittent BETi combined with continuous BRAFi/MEKi treatment was tolerable and improved durable tumour inhibition outcomes.ConclusionsTogether, our data suggest that intermittent inhibition of BET proteins may improve the duration of responses following BRAFi/MEKi treatment in BRAF-mutant melanoma.
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Project description:Substituting a carbon atom with a nitrogen atom (nitrogen substitution) on an aromatic ring in our leads 11a and 13g by applying nitrogen scanning afforded a set of compounds that improved not only the solubility but also the metabolic stability. The impact after nitrogen substitution on interactions between a derivative and its on- and off-target proteins (Raf/MEK, CYPs, and hERG channel) was also detected, most of them contributing to weaker interactions. After identifying the positions that kept inhibitory activity on HCT116 cell growth and Raf/MEK, compound 1 (CH5126766/RO5126766) was selected as a clinical compound. A phase I clinical trial is ongoing for solid cancers.
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