Dataset Information

Downregulation of the Ubiquitin Ligase RNF125 Underlies Resistance of Melanoma Cells to BRAF Inhibitors via JAK1 Deregulation.

ABSTRACT: Despite the remarkable clinical response of melanoma harboring BRAF mutations to BRAF inhibitors (BRAFi), most tumors become resistant. Here, we identified the downregulation of the ubiquitin ligase RNF125 in BRAFi-resistant melanomas and demonstrated its role in intrinsic and adaptive resistance to BRAFi in cultures as well as its association with resistance in tumor specimens. Sox10/MITF expression correlated with and contributed to RNF125 transcription. Reduced RNF125 was associated with elevated expression of receptor tyrosine kinases (RTKs), including EGFR. Notably, RNF125 altered RTK expression through JAK1, which we identified as an RNF125 substrate. RNF125 bound to and ubiquitinated JAK1, prompting its degradation and suppressing RTK expression. Inhibition of JAK1 and EGFR signaling overcame BRAFi resistance in melanoma with reduced RNF125 expression, as shown in culture and in in vivo xenografts. Our findings suggest that combination therapies targeting both JAK1 and EGFR could be effective against BRAFi-resistant tumors with de novo low RNF125 expression.

SUBMITTER: Kim H

PROVIDER: S-EPMC4681438 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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Downregulation of the Ubiquitin Ligase RNF125 Underlies Resistance of Melanoma Cells to BRAF Inhibitors via JAK1 Deregulation.

Kim Hyungsoo H Frederick Dennie T DT Levesque Mitchell P MP Cooper Zachary A ZA Feng Yongmei Y Krepler Clemens C Brill Laurence L Samuels Yardena Y Hayward Nicholas K NK Perlina Ally A Piris Adriano A Zhang Tongwu T Halaban Ruth R Herlyn Meenhard M MM Brown Kevin M KM Wargo Jennifer A JA Dummer Reinhard R Flaherty Keith T KT Ronai Ze'ev A ZA

Cell reports 20150528 9

Despite the remarkable clinical response of melanoma harboring BRAF mutations to BRAF inhibitors (BRAFi), most tumors become resistant. Here, we identified the downregulation of the ubiquitin ligase RNF125 in BRAFi-resistant melanomas and demonstrated its role in intrinsic and adaptive resistance to BRAFi in cultures as well as its association with resistance in tumor specimens. Sox10/MITF expression correlated with and contributed to RNF125 transcription. Reduced RNF125 was associated with elev ...[more]

PMID: 26027934

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Project description:BackgroundMost mutations in melanoma affect one critical amino acid on BRAF gene, resulting in the V600E substitution. Patient management is often based on the use of specific inhibitors targeting this mutation.MethodsDNA and RNA mutation status was assessed in 15 melanoma cell lines by Sanger sequencing and RNA-seq. We tested the cell lines responsiveness to BRAF inhibitors (vemurafenib and PLX4720, BRAF-specific and sorafenib, BRAF non-specific). Cell proliferation was assessed by MTT colorimetric assay. BRAF V600E RNA expression was assessed by qPCR. Expression level of phosphorylated-ERK protein was assessed by Western Blotting as marker of BRAF activation.ResultsThree cell lines were discordant in the mutation detection (BRAF V600E at DNA level/Sanger sequencing and BRAF WT on RNA-seq). We initially postulated that those cell lines may express only the WT allele at the RNA level although mutated at the DNA level. A more careful analysis showed that they express low level of BRAF RNA and the expression may be in favor of the WT allele. We tested whether the discordant cell lines responded differently to BRAF-specific inhibitors. Their proliferation rate decreased after treatment with vemurafenib and PLX4720 but was not affected by sorafenib, suggesting a BRAF V600E biological behavior. Yet, responsiveness to the BRAF specific inhibitors was lower as compared to the control. Western Blot analysis revealed a decreased expression of p-ERK protein in the BRAF V600E control cell line and in the discordant cell lines upon treatment with BRAF-specific inhibitors. The discordant cell lines showed a lower responsiveness to BRAF inhibitors when compared to the BRAF V600E control cell line. The results obtained from the inhibition experiment and molecular analyses were also confirmed in three additional cell lines.ConclusionCell lines carrying V600E mutation at the DNA level may respond differently to BRAF targeted treatment potentially due to a lower V600E RNA expression.

Dataset Information

Downregulation of the Ubiquitin Ligase RNF125 Underlies Resistance of Melanoma Cells to BRAF Inhibitors via JAK1 Deregulation.

Publications

Downregulation of the Ubiquitin Ligase RNF125 Underlies Resistance of Melanoma Cells to BRAF Inhibitors via JAK1 Deregulation.

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