BRAF Isoforms and Resistance to MAPK Inhibitors in Melanoma

Authors: Francisco Aya, Pablo Lanuza-Gracia, Abel González-Pérez, Sophie Bonnal, Estefania Mancini, Nuria López-Bigas, Ana Arance, Juan Valcárcel

Published in Cell Reports on April 23, 2024       .

Introduction:

Resistance to MAPK inhibitors (MAPKi), such as BRAF and MEK inhibitors, is a major challenge in treating BRAF-mutant melanoma. The production of alternative BRAF mRNA isoforms (altBRAFs) has been associated with up to 30% of cases of resistance to BRAF inhibitor monotherapy.

Previously, altBRAFs were thought to be generated by alternative splicing, leading to proposals for splicing modulation as a therapeutic strategy. This study challenges this view and provides evidence that altBRAFs are primarily caused by genomic deletions in the BRAF gene.

Design/Methods:

The researchers utilized different transcriptome datasets, including RNA-seq data from melanoma cell lines and patient tumors, to identify and quantify the expression of altBRAFs.

They employed whole-genome sequencing (WGS) and PCR amplification followed by Sanger sequencing to detect genomic deletions in the BRAF locus. In vitro models of altBRAF-mediated melanoma resistance were used to study the mechanisms of altBRAF generation.

Key Findings:

• altBRAFs were detected not only in BRAF-mutant and MAPKi-resistant samples but also in treatment-naive and BRAF wild-type melanoma samples, suggesting that altBRAFs are not solely associated with MAPKi resistance.
• In resistant melanoma cell lines, altBRAFs were exclusively expressed from the BRAF V600E allele, indicating a cis-acting genetic alteration rather than a trans-acting splicing factor alteration.
• Whole-genome sequencing and PCR amplification revealed cell line-specific genomic deletions in the BRAF locus, precisely encompassing the skipped exons responsible for generating different altBRAF isoforms.
• Evidence of genomic rearrangements was also found in patient tumor samples, which was consistent with the observed altBRAF expression profiles.

Key Highlights:

• altBRAFs are generated through genomic deletions, not alternative splicing.
• This challenges the prevailing paradigm of altBRAFs being produced by competition between alternative splice sites.
• Genomic deletions were detected in resistant cell lines and patient tumors, explaining the generation of specific altBRAF isoforms.
• altBRAFs were found in treatment-naive and BRAF wild-type tumors, suggesting a broader relevance beyond MAPKi resistance.

Key Takeaway Messages:

• The generation of altBRAFs in melanoma is primarily driven by genomic deletions in the BRAF locus rather than alternative splicing events.
• This finding has significant implications for understanding the resistance mechanisms to MAPKi and challenges the proposed therapeutic strategy of using splicing modulators to overcome resistance.
• The detection of altBRAFs in treatment-naive and wild-type BRAF tumors expands the potential clinical relevance of these BRAF isoforms beyond MAPKi resistance.
• Further investigation is warranted to explore the effects of type II RAF inhibitors in tumours harbouring dimer-enhancing genomic deletions in BRAF, regardless of BRAF mutational status.

Summary by Amalya Sargsyan, MD

Genomic deletions explain the generation of alternative BRAF isoforms conferring resistance to MAPK inhibitors in melanoma