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Tyrosine kinase inhibitors (TKI) directed against MET have been recently approved to treat advanced non-small cell lung cancer (NSCLC) harbouring activating MET mutations. This success is the consequence of a long characterization of MET mutations in cancers, which we propose to outline in this review. MET, a receptor tyrosine kinase (RTK), displays in a broad panel of cancers many deregulations liable to promote tumour progression. The first MET mutation was discovered in 1997, in hereditary papillary renal cancer (HPRC), providing the first direct link between MET mutations and cancer development. As in other RTKs, these mutations are located in the kinase domain, leading in most cases to ligand-independent MET activation. In 2014, novel MET mutations were identified in several advanced cancers, including lung cancers. These mutations alter splice sites of exon 14, causing in-frame exon 14 skipping and deletion of a regulatory domain. Because these mutations are not located in the kinase domain, they are original and their mode of action has yet to be fully elucidated. Less than five years after the discovery of such mutations, the efficacy of a MET TKI was evidenced in NSCLC patients displaying MET exon 14 skipping. Yet its use led to a resistance mechanism involving acquisition of novel and already characterized MET mutations. Furthermore, novel somatic MET mutations are constantly being discovered. The challenge is no longer to identify them but to characterize them in order to predict their transforming activity and their sensitivity or resistance to MET TKIs, in order to adapt treatment.

Targeted therapies against receptor tyrosine kinases (RTKs) are currently used with success on a small proportion of patients displaying clear oncogene activation. Lung cancers with a mutated EGFR provide a good illustration. The efficacy of targeted treatments relies on oncogene addiction, a situation in which the growth or survival of the cancer cells depends on a single deregulated oncogene. MET, a member of the RTK family, is a promising target because it displays many deregulations in a broad panel of cancers. Although clinical trials having evaluated MET inhibitors in large populations have yielded disappointing results, many recent case reports suggest that MET inhibition may be effective in a subset of patients with unambiguous MET activation and thus, most probably, oncogene addiction. Interestingly, preclinical studies have revealed a particularity of MET addiction: it can arise through several mechanisms, and the mechanism involved can differ according to the cancer type. The present review describes the different mechanisms of MET addiction and their consequences for diagnosis and therapeutic strategies. Although in each cancer type MET addiction affects a restricted number of patients, pooling of these patients across all cancer types yields a targetable population liable to benefit from addiction-targeting therapies.

Background The ETS transcription factor ETV4 is involved in the main steps of organogenesis and is also a significant mediator of tumorigenesis and metastasis, such as in breast cancer. Indeed, ETV4 is overexpressed in breast tumors and is associated with distant metastasis and poor prognosis. However, the cellular and molecular events regulated by this factor are still misunderstood. In mammary epithelial cells, ETV4 controls the expression of many genes, MMP13 among them. The aim of this study was to understand the function of MMP13 during ETV4-driven tumorigenesis. Methods Different constructs of the MMP13 gene promoter were used to study the direct regulation of MMP13 by ETV4. Moreover, cell proliferation, migration, invasion, anchorage-independent growth, and in vivo tumorigenicity were assayed using models of mammary epithelial and cancer cells in which the expression of MMP13 and/or ETV4 is modulated. Importantly, the expression of MMP13 and ETV4 messenger RNA was characterized in 456 breast cancer samples. Results Our results revealed that ETV4 promotes proliferation, migration, invasion, and anchorage-independent growth of the MMT mouse mammary tumorigenic cell line. By investigating molecular events downstream of ETV4, we found that MMP13, an extracellular metalloprotease, was an ETV4 target gene. By overexpressing or repressing MMP13, we showed that this metalloprotease contributes to proliferation, migration, and anchorage-independent clonogenicity. Furthermore, we demonstrated that MMP13 inhibition disturbs proliferation, migration, and invasion induced by ETV4 and participates to ETV4-induced tumor formation in immunodeficient mice. Finally, ETV4 and MMP13 co-overexpression is associated with poor prognosis in breast cancer. Conclusion MMP13 potentiates the effects of the ETV4 oncogene during breast cancer genesis and progression.

Nonsense mutations cause about 10% of genetic disease cases, and no treatments are available. Nonsense mutations can be corrected by molecules with nonsense mutation readthrough activity. An extract of the mushroom Lepista inversa has recently shown high-efficiency correction of UGA and UAA nonsense mutations. One active constituent of this extract is 2,6-diaminopurine (DAP). In Calu-6 cancer cells, in which TP53 gene has a UGA nonsense mutation, DAP treatment increases p53 level. It also decreases the growth of tumors arising from Calu-6 cells injected into immunodeficient nude mice. DAP acts by interfering with the activity of a tRNA-specific 2′-O-methyltransferase (FTSJ1) responsible for cytosine 34 modification in tRNA Trp . Low-toxicity and high-efficiency UGA nonsense mutation correction make DAP a good candidate for the development of treatments for genetic diseases caused by nonsense mutations. Nonsense mutations can be corrected by several molecules that activate readthrough of premature termination codon. Here, the authors report that 2,6-diaminopurine efficiently corrects UGA nonsense mutations with no significant toxicity.

Non-small cell lung cancers (NSCLCs) treated with tyrosine kinase inhibitors (TKIs) of the epidermal growth factor receptor (EGFR) almost invariably relapse in the long term, due to the emergence of subpopulations of resistant cells. Through a DNA barcoding approach, we show that the clinically approved drug sorafenib specifically abolishes the selective advantage of EGFR-TKI-resistant cells, while preserving the response of EGFR-TKI-sensitive cells. Sorafenib is active against multiple mechanisms of resistance/tolerance to EGFR-TKIs and its effects depend on early inhibition of MAPK-interacting kinase (MKNK) activity and signal transducer and activator of transcription 3 (STAT3) phosphorylation, and later down-regulation of MCL1 and EGFR. Using different xenograft and allograft models, we show that the sorafenib-EGFR-TKI combination can delay tumor growth and promote the recruitment of inflammatory cells. Together, our findings indicate that sorafenib can prolong the response to EGFR-TKIs by targeting NSCLC capacity to adapt to treatment through the emergence of resistant cells. The emergence of resistant subpopulations often underlies the development of resistance to cancer therapy. Here, using a DNA barcoding approach, the authors demonstrate EGFR TKI treatment in non-small cell lung cancer enriches for resistant subpopulation which can be prevented by treatment with the multikinase inhibitor sorafenib via inhibition of MKNK, STAT3 and MCL1.

The MET exon 14 skipping mutation (named METex14Del) described in lung cancer leads to prolonged activation of signaling pathways and aberrant cell responses, but the link between HGF signaling and cell responses remains unclear. A putative lung cancer regulatory network of influential transcription factors was constructed from the transcriptomes of lung cancer cell lines. Transcriptomic data from METex14Del-expressing cells, stimulated or not by HGF, were mapped onto this lung cancer reference network and revealed activation of a major regulatory node composed mainly by the highly influential transcription factors ETS1, FOSL1 and SMAD3. HGF activation of METex14Del receptor induced the expression and phosphorylation of these three master regulators and the expression of their predicted target genes involved in migration and invasion. All these molecular and biological effects were inhibited by trametinib, a MEK inhibitor, which was potentiated by combination with capmatinib, a MET inhibitor. New mapping with transcriptomic data from trametinib-treated METex14Del cells validated the key role of the RAS-ERK pathway signaling in the activation of ETS1, FOSL1 and SMAD3 regulators and the induction of their target genes in HGF-activated METex14Del receptor. Thus, we report an original and powerful strategy to uncover key regulators, including transcription factors that have not been widely described in METex14Del signaling, such as SMAD3. These factors are activated by specific signaling pathways and could provide a novel therapeutic strategy involving a combination of receptor and signaling inhibitors.

Prostate cancer, the most common malignancy in men, has a relatively favourable prognosis. However, when it spreads to the bone, the survival rate drops dramatically. The development of bone metastases leaves patients with aggressive prostate cancer, the leading cause of death in men. Moreover, bone metastases are incurable and very painful. Hepatocyte growth factor receptor (MET) and fusion of genes encoding E26 transformation‐specific (ETS) transcription factors are both involved in the progression of the disease. ETS gene fusions, in particular, have the ability to induce the migratory and invasive properties of prostate cancer cells, whereas MET receptor, through its signalling cascades, is able to activate transcription factor expression. MET signalling and ETS gene fusions are intimately linked to high‐grade prostate cancer. However, the collaboration of these factors in prostate cancer progression has not yet been investigated. Here, we show, using cell models of advanced prostate cancer, that ETS translocation variant 1 (ETV1) and transcriptional regulator ERG (ERG) transcription factors (members of the ETS family) promote tumour properties, and that activation of MET signalling enhances these effects. By using a specific MET tyrosine kinase inhibitor in a humanised hepatocyte growth factor (HGF) mouse model, we also establish that MET activity is required for ETV1/ERG‐mediated tumour growth. Finally, by performing a comparative transcriptomic analysis, we identify target genes that could play a relevant role in these cellular processes. Thus, our results demonstrate for the first time in prostate cancer models a functional interaction between ETS transcription factors (ETV1 and ERG) and MET signalling that confers more aggressive properties and highlight a molecular signature characteristic of this combined action. The MET receptor activation pathway is intrinsically linked to the activity of the transcription factors ETV1 and ERG (ETS family) to promote tumorigenesis in prostate cancer cells. Here we focus on the transcriptional programme leading to this partnership and have identified 5 commonly regulated target genes that may represent a potential signature of prostate cancer progression at advanced stages.

In hereditary papillary renal cell carcinoma (HPRCC), the hepatocyte growth factor receptor (MET) receptor tyrosine kinase (RTK) mutations recorded to date are located in the kinase domain and lead to constitutive MET activation. This contrasts with MET mutations identified in non‐small‐cell lung cancer (NSCLC), which lead to exon 14 skipping and deletion of a regulatory domain: In this latter case, the mutated receptor still requires ligand stimulation. Sequencing of MET in samples from 158 HPRCC and 2808 NSCLC patients revealed 10 uncharacterized mutations. Four of these, all found in HPRCC and leading to amino acid substitutions in the N‐lobe of the MET kinase, proved able to induce cell transformation, which was further enhanced by hepatocyte growth factor (HGF) stimulation: His1086Leu, Ile1102Thr, Leu1130Ser, and Cis1125Gly. Similar to the variant resulting in MET exon 14 skipping, the two N‐lobe MET variants His1086Leu and Ile1102Thr were found to require stimulation by HGF in order to strongly activate downstream signaling pathways and epithelial cell motility. The Ile1102Thr mutation also displayed transforming potential, promoting tumor growth in a xenograft model. In addition, the N‐lobe‐mutated MET variants were found to trigger a common HGF‐stimulation‐dependent transcriptional program, consistent with an observed increase in cell motility and invasion. Altogether, this functional characterization revealed that N‐lobe variants still require ligand stimulation, in contrast to other RTK variants. This suggests that HGF expression in the tumor microenvironment is important for tumor growth. The sensitivity of these variants to MET inhibitors opens the way for use of targeted therapies for patients harboring the corresponding mutations. MET variants in the N‐lobe of the kinase domain, found in hereditary papillary renal cell carcinoma, require ligand stimulation to promote cell transformation, in contrast to other RTK variants. This suggests that HGF expression in the microenvironment is important for tumor growth in such patients. Their sensitivity to MET inhibitors opens the way for use of targeted therapies in these patients.

About 10% of patients with a genetic disease carry a nonsense mutation causing their pathology. A strategy for correcting nonsense mutations is premature termination codon (PTC) readthrough, i.e. incorporation of an amino acid at the PTC position during translation. PTC-readthrough-activating molecules appear as promising therapeutic tools for these patients. Unfortunately, the molecules shown to induce PTC readthrough show low efficacy, probably because the mRNAs carrying a nonsense mutation are scarce, as they are also substrates of the quality control mechanism called nonsense-mediated mRNA decay (NMD). The screening systems previously developed to identify readthrough-promoting molecules used cDNA constructs encoding mRNAs immune to NMD. As the molecules identified were not selected for the ability to correct nonsense mutations on NMD-prone PTC-mRNAs, they could be unsuitable for the context of nonsense-mutation-linked human pathologies. Here, a screening system based on an NMD-prone mRNA is described. It should be suitable for identifying molecules capable of efficiently rescuing the expression of human genes harboring a nonsense mutation. This system should favor the discovery of candidate drugs for treating genetic diseases caused by nonsense mutations. One hit selected with this screening system is presented and validated on cells from three cystic fibrosis patients.

Exon skipping mutations of the MET receptor tyrosine kinase (METex14), increasingly reported in cancers, occur in 3–4% of non–small‐cell lung cancer (NSCLC). Only 50% of patients have a beneficial response to treatment with MET‐tyrosine kinase inhibitors (TKIs), underlying the need to understand the mechanism of METex14 oncogenicity and sensitivity to TKIs. Whether METex14 is a driver mutation and whether it requires hepatocyte growth factor (HGF) for its oncogenicity in a range of in vitro functions and in vivo has not been fully elucidated from previous preclinical models. Using CRISPR/Cas9, we developed a METex14/WT isogenic model in nontransformed human lung cells and report that the METex14 single alteration was sufficient to drive MET‐dependent in vitro anchorage‐independent survival and motility and in vivo tumorigenesis, sensitising tumours to MET‐TKIs. However, we also show that human HGF (hHGF) is required, as demonstrated in vivo using a humanised HGF knock‐in strain of mice and further detected in tumour cells of METex14 NSCLC patient samples. Our results also suggest that METex14 oncogenicity is not a consequence of an escape from degradation in our cell model. Thus, we developed a valuable model for preclinical studies and present results that have potential clinical implication.