Acquired and intrinsic resistance to vemurafenib in BRAFV600E-driven melanoma brain metastases

BRAFV600-mutated melanoma brain metastases (MBMs) are responsive to BRAF inhibitors, but responses are generally less durable than those of extracranial metastases. We tested the hypothesis that the drug efflux transporters P-glycoprotein (P-gp; ABCB1) and breast cancer resistance protein (BCRP; ABCG2) expressed at the blood–brain barrier (BBB) offer MBMs protection from therapy. We intracranially implanted A375 melanoma cells in wild-type (WT) and Abcb1a/b;Abcg2−/− mice, characterized the tumor BBB, analyzed drug levels in plasma and brain lesions after oral vemurafenib administration, and determined the efficacy against brain metastases and subcutaneous lesions. Although contrast-enhanced MRI demonstrated that the integrity of the BBB is disrupted in A375 MBMs, vemurafenib achieved greater antitumor efficacy against MBMs in Abcb1a/b;Abcg2−/− mice compared with WT mice. Concordantly, P-gp and BCRP are expressed in MBM-associated brain endothelium both in patients and in A375 xenografts and expression of these transporters limited vemurafenib penetration into A375 MBMs. Although initially responsive, A375 MBMs rapidly developed therapy resistance, even in Abcb1a/b;Abcg2−/− mice, and this was unrelated to pharmacokinetic or target inhibition issues. Taken together, we demonstrate that both intrinsic and acquired resistance can play a role in MBMs

Caffeine as a tool for investigating the integration of Cdc25 phosphorylation, activity and ubiquitin-dependent degradation in Schizosaccharomyces pombe

The evolutionarily conserved Cdc25 phosphatase is an essential protein that removes inhibitory phosphorylation moieties on the mitotic regulator Cdc2. Together with the Wee1 kinase, a negative regulator of Cdc2 activity, Cdc25 is thus a central regulator of cell cycle progression in Schizosaccharomyces pombe. The expression and activity of Cdc25 is dependent on the activity of the Target of Rapamycin Complex 1 (TORC1). TORC1 inhibition leads to the activation of Cdc25 and repression of Wee1, leading to advanced entry into mitosis. Withdrawal of nitrogen leads to rapid Cdc25 degradation via the ubiquitin- dependent degradation pathway by the Pub1 E3- ligase. Caffeine is believed to mediate the override of DNA damage checkpoint signalling, by inhibiting the activity of the ataxia telangiectasia mutated (ATM)/Rad3 homologues. This model remains controversial, as TORC1 appears to be the preferred target of caffeine in vivo. Recent studies suggest that caffeine induces DNA damage checkpoint override by inducing the nuclear accumulation of Cdc25 in S. pombe. Caffeine may thus modulate Cdc25 activity and stability via inhibition of TORC1. A clearer understanding of the mechanisms by which caffeine stabilises Cdc25, may provide novel insights into how TORC1 and DNA damage signalling is integrated