Immunohistochemical application of a highly sensitive and specific murine monoclonal antibody recognising the extracellular domain of the human hepatocyte growth factor receptor (MET).

AIMS: Development of novel targeted therapies directed against hepatocyte growth factor (HGF) or its receptor (MET) necessitates the availability of quality diagnostics to facilitate their safe and effective use. Limitations of some commercially available anti-MET antibodies have prompted development of the highly sensitive and specific clone A2H2-3. Here we report its analytical properties when applied by an automated immunohistochemistry method. METHODS AND RESULTS: Excellent antibody specificity was demonstrated by immunoblot, ELISA, and IHC evaluation of characterised cell lines including NIH3T3 overexpressing the related kinase MST1R (RON). Sensitivity was confirmed by measurements of MET in cell lines or characterised tissues. IHC correlated well with FISH and quantitative RT-PCR assessments of MET (P < 0.001). Good total agreement (89%) was observed with the anti-MET antibody clone SP44 using whole-tissue sections, but poor positive agreement (21-47%) was seen in tissue microarray cores. Multiple lots displayed appropriate reproducibility (R(2)  > 0.9). Prevalence of MET positivity by IHC was higher in non-squamous cell NSCLC, MET or EGFR amplified cases, and in tumours harbouring abnormalities in EGFR exon 19 or 21. CONCLUSIONS: The anti-MET antibody clone A2H2-3 displays excellent specificity and sensitivity. These properties make it suitable for clinical trial investigations and development as a potential companion diagnostic.

Cotargeting MNK and MEK kinases induces the regression of NF1-mutant cancers.

Neurofibromin 1-mutant (NF1-mutant) cancers are driven by excessive Ras signaling; however, there are currently no effective therapies for these or other Ras-dependent tumors. While combined MEK and mTORC1 suppression causes regression of NF1-deficient malignancies in animal models, the potential toxicity of cotargeting these 2 major signaling pathways in humans may necessitate the identification of more refined, cancer-specific signaling nodes. Here, we have provided evidence that MAPK-interacting kinases (MNKs), which converge on the mTORC1 effector eIF4E, are therapeutic targets in NF1-deficient malignancies. Specifically, we evaluated primary human NF1-deficient peripheral nervous system tumors and found that MNKs are activated in the majority of tumors tested. Genetic and chemical suppression of MNKs in NF1-deficient murine tumor models and human cell lines potently cooperated with MEK inhibitors to kill these cancers through effects on eIF4E. We also demonstrated that MNK kinases are important and direct targets of cabozantinib. Accordingly, coadministration of cabozantinib and MEK inhibitors triggered dramatic regression in an aggressive genetically engineered tumor model. The cytotoxicity of this combination required the suppression of MNK-induced eIF4E phosphorylation and was not recapitulated by suppressing other cabozantinib targets. Collectively, these studies demonstrate that combined MNK and MEK suppression represents a promising therapeutic strategy for these incurable Ras-driven tumors and highlight the utility of developing selective MNK inhibitors for these and possibly other malignancies.