Anti-MAPK1 monoclonal antibody, clone 2C4C0

The concept of polypharmacology involves the interaction of drug molecules with multiple molecular targets. It provides a unique opportunity for the repurposing of already-approved drugs to target key factors involved in human diseases. Herein, we used anin silicotarget prediction algorithm to investigate the mechanism of action of mebendazole, an antihelminthic drug, currently repurposed in the treatment of brain tumors. First, we confirmed that mebendazole decreased the viability of glioblastoma cellsin vitro(IC(50)values ranging from 288 nmto 2.1 mu m). Ourin silicoapproach unveiled 21 putative molecular targets for mebendazole, including 12 proteins significantly upregulated at the gene level in glioblastoma as compared to normal brain tissue (fold change > 1.5;P < 0.0001). Validation experiments were performed on three major kinases involved in cancer biology: ABL1, MAPK1/ERK2, and MAPK14/p38 alpha. Mebendazole could inhibit the activity of these kinasesin vitroin a dose-dependent manner, with a high potency against MAPK14 (IC50 = 104 +/- 46 nm). Its direct binding to MAPK14 was further validatedin vitro, and inhibition of MAPK14 kinase activity was confirmed in live glioblastoma cells. Consistent with biophysical data, molecular modeling suggested that mebendazole was able to bind to the catalytic site of MAPK14. Finally, gene silencing demonstrated that MAPK14 is involved in glioblastoma tumor spheroid growth and response to mebendazole treatment. This study thus highlighted the role of MAPK14 in the anticancer mechanism of action of mebendazole and provides further rationale for the pharmacological targeting of MAPK14 in brain tumors. It also opens new avenues for the development of novel MAPK14/p38 alpha inhibitors to treat human diseases.

Hypoxia inducible factor-1 (HIF-1) is the main transcriptional activator of the cellular response to hypoxia and an important target of anticancer therapy. Phosphorylation by ERK1 and/or ERK2 (MAPK3 and MAPK1, respectively; hereafter ERK) stimulates the transcriptional activity of HIF-1 alpha by inhibiting its CRM1 (XPO1)-dependent nuclear export. Here, we demonstrate that phosphorylation by ERK also regulates the association of HIF-1 alpha with a so-far-unknown interaction partner identified as mortalin (also known as GRP75 and HSPA9), which mediates non-genomic involvement of HIF- 1 alpha in apoptosis. Mortalin binds specifically to HIF- 1a that lacks modification by ERK, and the HIF-1 alpha-mortalin complex is localized outside the nucleus. Under hypoxia, mortalin mediates targeting of unmodified HIF-1 alpha to the outer mitochondrial membrane, as well as association with VDAC1 and hexokinase II, which promotes production of a C-terminally truncated active form of VDAC1, denoted VDAC1-Delta C, and protection from apoptosis when ERK is inactivated. Under normoxia, transcriptionally inactive forms of unmodified HIF-1 alpha or its C-terminal domain alone are also targeted to mitochondria, stimulate production of VDAC1-Delta C and increase resistance to etoposide-or doxorubicin-induced apoptosis. These findings reveal an ERK-controlled, unconventional and anti-apoptotic function of HIF-1 alpha that might serve as an early protective mechanism upon oxygen limitation and promote cancer cell resistance to chemotherapy.