Abstract
Despite the great success of small molecule inhibitors in the treatment of patients with BRAFV600E mutated melanoma, the response to these drugs remains transient and patients eventually relapse within a few months, highlighting the need to develop novel combination therapies based on the understanding of the molecular changes induced by BRAFV600E inhibitors. The acute inhibition of oncogenic signaling can rewire entire cellular signaling pathways and thereby create novel cancer cell vulnerabilities. Here, we demonstrate that inhibition of BRAFV600E oncogenic signaling in melanoma cell lines leads to destabilization of the large subunit of RNA polymerase II POLR2A (polymerase RNA II DNA-directed polypeptide A), thereby preventing its binding to the unconventional prefoldin RPB5 interactor (URI1) chaperone complex and the successful assembly of RNA polymerase II holoenzymes. Furthermore, in melanoma cell lines treated with mitogen-activated protein kinase (MAPK) inhibitors, α-amanitin, a specific and irreversible inhibitor of RNA polymerase II, induced massive apoptosis. Pre-treatment of melanoma cell lines with MAPK inhibitors significantly reduced IC50 values to α-amanitin, creating a state of collateral vulnerability similar to POLR2A hemizygous deletions. Thus, the development of melanoma specific α-amanitin antibody-drug conjugates could represent an interesting therapeutic approach for combination therapies with BRAFV600E inhibitors.
Highlights
Melanoma is an aggressive tumor with poor survival when disseminated to distant organs
We performed a mass spectrometry analysis to investigate whether BRAFV600E oncogenic signaling might influence interactions of the URI1 chaperone complex with client proteins in melanoma cell lines
Acute inhibition of BRAFV600E did not affect the composition of the URI1 core-chaperone complex nor its association with transcription regulatory proteins
Summary
Melanoma is an aggressive tumor with poor survival when disseminated to distant organs. Several compounds inhibiting BRAFV600E and the downstream mitogen-activated protein kinase (MAPK) signal transduction pathway were developed and are successfully used in the clinic[2]. These therapies are effective in only a subset of patients, approximately 50% with tumors harboring the gain-of-function BRAFV600E mutation, and drug resistance inevitably develops and most patients relapse within a few months[1,2]. Www.nature.com/scientificreports phosphorylation target of p70-S6 kinase 1 (S6K1) This URI1 phosphorylation is controlling its interaction with the phosphatase PP1γ (protein phosphatase 1, catalytic subunit, gamma isoform) and thereby PP1γ-mediated de-phosphorylation of S6K18. Together with its co-members of the URI1/‘prefoldin-like’ and the R2TP [RUVBL1, RUVBL2, RPAP3 (in yeast known as Tah1), PIH1D1] complex it is tightly involved in the cytoplasmic assembly of all three RNA polymerase complexes[13,15,16,17]
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