Apoptosis of human melanoma cells induced by inhibition of B-RAFV600E involves preferential splicing of bimS

Jiang, Chen Chen; Lai, Fritz; Tay, Kwang Hong; Croft, Amanda; Rizos, Helen; Becker, Therese M.; Yang, Fan; Liu, H; Thorne, Rick F.; Hersey, Peter; Zhang, Xu Dong

doi:10.1038/cddis.2010.48

Cited by 86 publications

(92 citation statements)

References 36 publications

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“…The small RNA interference (siRNA) constructs used were obtained as the siGENOME SMARTpool reagents (Dharmacon), MEK1 siGENOME SMARTpool (M-003571-01-0005), C-RAF siGENOME SMARTpool (M-003601-02-0010), ERK1 siGENOME SMARTpool (M-003592-03-0010), ERK2 siGENOME SMARTpool (M-003555-04-0010), Akt1 siGENOME SMARTpool (M-003000-03-0010), Akt2 siGENOME SMARTpool (M-003001-02-0010), Akt3 siGENOME SMARTpool (M-003002-02-0010), and siGENOME Nontargeting siRNA pool (D-001206- [13][14][15][16][17][18][19][20]. Transfection of siRNA pools was carried out as described previously (20,21).…”

Section: Small Rna Interferencementioning

confidence: 99%

“…Cell viability assays (MTS assays) were performed as reported previously (20). In brief, cells were seeded at 5,000 cells/well onto flat-bottomed 96-well culture plates and allowed to grow for 24 hours followed by the desired treatment.…”

Section: Cell Viability Assaysmentioning

confidence: 99%

“…Human melanoma cell lines Mel-RMu and Mel-CV have been described previously (20). They were cultured in Dulbecco's Modified Eagle Medium (DMEM) containing 5% fetal calf serum (FCS) (Commonwealth Serum Laboratories).…”

Section: Cell Culture and Reagentsmentioning

confidence: 99%

“…Western blot analysis was carried out as described previously (20,21). Labeled bands were detected by ImmunStar HRP Chemiluminescent Kit, and images were captured and the intensity of the bands was quantitated with the BioRad VersaDoc image system (Bio-Rad).…”

Section: Western Blot Analysismentioning

confidence: 99%

“…Quantitation of apoptotic cells was carried out by measurement of sub-G1 DNA content using propidium iodide (PI) on a flow cytometer as described elsewhere (20,21).…”

Section: Apoptosismentioning

confidence: 99%

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MEK-Independent Survival of B-RAFV600E Melanoma Cells Selected for Resistance to Apoptosis Induced by the RAF Inhibitor PLX4720

Jiang

Lai

Thorne

et al. 2011

Clinical Cancer Research

105

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Purpose: To examine mechanisms that determine long-term responses of B-RAF V600E melanoma cells to B-RAF inhibitors. Experimental Design: B-RAF V600E melanoma cells were exposed to the B-RAF inhibitor PLX4720 for prolonged periods to select for cells resistant to apoptosis induced by the inhibitor. The resultant cells were analyzed for activation of extracellular signal regulated kinase (ERK), MAP/ERK kinase (MEK), and Akt, and related signals. Their roles in survival of the cells were also examined.Results: B-RAF V600E melanoma cells selected for resistant to PLX4720-induced apoptosis retained the V600E mutation in B-RAF, and proliferated steadily in the presence of the inhibitor, albeit with slow growth rate. These cells displayed high levels of ERK activation, that is, at least in part, independent of the conventional RAF/MEK/ERK pathway, as MEK activation was low and inhibition of MEK did not significantly block activation of ERK. In contrast, extracellular signals appeared involved. This was associated with elevated activation of the phosphoinositide 3-kinase (PI3k)/Akt pathway and could be inhibited by serum starvation and inhibition of PI3k/Akt. Inhibition of MEK did not impact on survival of these cells, whereas serum starvation, inhibition of PI3K/Akt, and inhibition of ERK1/2 reduced their viability.Conclusions: These results indicate that sensitivity to induction of apoptosis may be a major determinant of long-term responses of B-RAF V600E melanomas to specific inhibitors and suggest that rebound melanoma growth after initial treatment with the inhibitors may not be responsive to MEK inhibitors, but may be susceptible to inhibition of the PI3k/Akt pathway.

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Section: Small Rna Interferencementioning

confidence: 99%

Section: Cell Viability Assaysmentioning

confidence: 99%

Section: Cell Culture and Reagentsmentioning

confidence: 99%

Section: Western Blot Analysismentioning

confidence: 99%

“…Quantitation of apoptotic cells was carried out by measurement of sub-G1 DNA content using propidium iodide (PI) on a flow cytometer as described elsewhere (20,21).…”

Section: Apoptosismentioning

confidence: 99%

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MEK-Independent Survival of B-RAFV600E Melanoma Cells Selected for Resistance to Apoptosis Induced by the RAF Inhibitor PLX4720

Jiang

Lai

Thorne

et al. 2011

Clinical Cancer Research

105

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HDAC inhibitors restore BRAF‐inhibitor sensitivity by altering PI3K and survival signalling in a subset of melanoma

Gallagher

Gunatilake

Beaumont

et al. 2017

Intl Journal of Cancer

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Mutations in BRAF activate oncogenic MAPK signalling in almost half of cutaneous melanomas. Inhibitors of BRAF (BRAFi) and its target MEK are widely used to treat melanoma patients with BRAF mutations but unfortunately acquired resistance occurs in the majority of patients. Resistance results from mutations or non-genomic changes that either reactivate MAPK signalling or activate other pathways that provide alternate survival and growth signalling. Here, we show the histone deacetylase inhibitor (HDACi) panobinostat overcomes BRAFi resistance in melanoma, but this is dependent on the resistant cells showing a partial response to BRAFi treatment. Using patient- and in vivo-derived melanoma cell lines with acquired BRAFi resistance, we show that combined treatment with the BRAFi encorafenib and HDACi panobinostat in 2D and 3D culture systems synergistically induced caspase-dependent apoptotic cell death. Key changes induced by HDAC inhibition included decreased PI3K pathway activity associated with a reduction in the protein level of a number of receptor tyrosine kinases, and cell line dependent upregulation of pro-apoptotic BIM or NOXA together with reduced expression of anti-apoptotic proteins. Independent of these changes, panobinostat reduced c-Myc and pre-treatment of cells with siRNA against c-Myc reduced BRAFi/HDACi drug-induced cell death. These results suggest that a combination of HDAC and MAPK inhibitors may play a role in treatment of melanoma where the resistance mechanisms are due to activation of MAPK-independent pathways.

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The splicing factor SRSF6 is amplified and is an oncoprotein in lung and colon cancers

Cohen-Eliav¹,

Golan-Gerstl²,

Siegfried³

et al. 2013

The Journal of Pathology

129

123

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An increasing body of evidence connects alterations in the process of alternative splicing with cancer development and progression. However, a direct role of splicing factors as drivers of cancer development is mostly unknown. We analysed the gene copy number of several splicing factors in colon and lung tumours, and found that the gene encoding for the splicing factor SRSF6 is amplified and over-expressed in these cancers. Moreover, over-expression of SRSF6 in immortal lung epithelial cells enhanced proliferation, protected them from chemotherapy-induced cell death and converted them to be tumourigenic in mice. In contrast, knock-down of SRSF6 in lung and colon cancer cell lines inhibited their tumourigenic abilities. SRSF6 up- or down-regulation altered the splicing of several tumour suppressors and oncogenes to generate the oncogenic isoforms and reduce the tumour-suppressive isoforms. Our data suggest that the splicing factor SRSF6 is an oncoprotein that regulates the proliferation and survival of lung and colon cancer cells.

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Apoptosis of human melanoma cells induced by inhibition of B-RAFV600E involves preferential splicing of bimS

Cited by 86 publications

References 36 publications

MEK-Independent Survival of B-RAFV600E Melanoma Cells Selected for Resistance to Apoptosis Induced by the RAF Inhibitor PLX4720

MEK-Independent Survival of B-RAFV600E Melanoma Cells Selected for Resistance to Apoptosis Induced by the RAF Inhibitor PLX4720

HDAC inhibitors restore BRAF‐inhibitor sensitivity by altering PI3K and survival signalling in a subset of melanoma

The splicing factor SRSF6 is amplified and is an oncoprotein in lung and colon cancers

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