Direct interaction of TrkA/CD44v3 is essential for NGF-promoted aggressiveness of breast cancer cells

Background CD44 is a multifunctional membrane glycoprotein. Through its heparan sulfate chain, CD44 presents growth factors to their receptors. We have shown that CD44 and Tropomyosin kinase A (TrkA) form a complex following nerve growth factor (NGF) induction. Our study aimed to understand how CD44 and TrkA interact and the consequences of inhibiting this interaction regarding the pro-tumoral effect of NGF in breast cancer. Methods After determining which CD44 isoforms (variants) are involved in forming the TrkA/CD44 complex using proximity ligation assays, we investigated the molecular determinants of this interaction. By molecular modeling, we isolated the amino acids involved and confirmed their involvement using mutations. A CD44v3 mimetic peptide was then synthesized to block the TrkA/CD44v3 interaction. The effects of this peptide on the growth, migration and invasion of xenografted triple-negative breast cancer cells were assessed. Finally, we investigated the correlations between the expression of the TrkA/CD44v3 complex in tumors and histo-pronostic parameters. Results We demonstrated that isoform v3 (CD44v3), but not v6, binds to TrkA in response to NGF stimulation. The final 10 amino acids of exon v3 and the TrkA H112 residue are necessary for the association of CD44v3 with TrkA. Functionally, the CD44v3 mimetic peptide impairs not only NGF-induced RhoA activation, clonogenicity, and migration/invasion of breast cancer cells in vitro but also tumor growth and metastasis in a xenograft mouse model. We also detected TrkA/CD44v3 only in cancerous cells, not in normal adjacent tissues. Conclusion Collectively, our results suggest that blocking the CD44v3/TrkA interaction can be a new therapeutic option for triple-negative breast cancers

Rapid response to the M_w 4.9 earthquake of November 11, 2019 in Le Teil, Lower Rhône Valley, France

On November 11, 2019, a Mw 4.9 earthquake hit the region close to Montelimar (lower Rhône Valley, France), on the eastern margin of the Massif Central close to the external part of the Alps. Occuring in a moderate seismicity area, this earthquake is remarkable for its very shallow focal depth (between 1 and 3 km), its magnitude, and the moderate to large damages it produced in several villages. InSAR interferograms indicated a shallow rupture about 4 km long reaching the surface and the reactivation of the ancient NE-SW La Rouviere normal fault in reverse faulting in agreement with the present-day E-W compressional tectonics. The peculiarity of this earthquake together with a poor coverage of the epicentral region by permanent seismological and geodetic stations triggered the mobilisation of the French post-seismic unit and the broad French scientific community from various institutions, with the deployment of geophysical instruments (seismological and geodesic stations), geological field surveys, and field evaluation of the intensity of the earthquake. Within 7 days after the mainshock, 47 seismological stations were deployed in the epicentral area to improve the Le Teil aftershocks locations relative to the French permanent seismological network (RESIF), monitor the temporal and spatial evolution of microearthquakes close to the fault plane and temporal evolution of the seismic response of 3 damaged historical buildings, and to study suspected site effects and their influence in the distribution of seismic damage. This seismological dataset, completed by data owned by different institutions, was integrated in a homogeneous archive and distributed through FDSN web services by the RESIF data center. This dataset, together with observations of surface rupture evidences, geologic, geodetic and satellite data, will help to unravel the causes and rupture mechanism of this earthquake, and contribute to account in seismic hazard assessment for earthquakes along the major regional Cévenne fault system in a context of present-day compressional tectonics

Involvement of membrane receptors networks in NGF and proNGF signaling in breast cancer cells

Notre laboratoire a montré que le NGF et les neurotrophines sont des facteurs déterminants de la cancérogenèse mammaire, et plus particulièrement de la croissance et de l’invasion des cellules cancéreuses de sein. Les effets du NGF dépendent, entre autres, de l’activation du récepteur à activité tyrosine kinase TrkA. Récemment, nous avons décrit que le précurseur du NGF, le proNGF, est également produit et sécrété par les cellules cancéreuses de sein pour induire leur invasion via la formation d’un complexe sortiline/TrkA. Néanmoins, les différentes signalisations induites par le NGF et le proNGF ne sont pas encore parfaitement décryptées dans le cancer du sein. Au cours de ma thèse, j’ai démontré que le NGF et le proNGF induisent des complexes de récepteurs distincts. Ainsi, alors que le NGF conduit à la formation d’un complexe membranaire TrkA/CD44, le proNGF recrute quant à lui la sortiline, TrkA et EphA2. En outre, les études in vivo, réalisées dans un modèle de souris SCID, montrent que la croissance tumorale est dépendante de la formation de ces complexes. Ainsi, l’administration conjointe d’un inhibiteur de l’activité kinase de TrkA (CEP-701) et d’un ARN interférent dirigé soit contre CD44, soit contre EphA2, diminue significativement le volume tumoral en comparaison à l’utilisation en monothérapie du CEP-701 ou d’un siEphA2 ou du siCD44. L’ensemble de mes résultats a permis d’approfondir les mécanismes d’action du NGF et de son précurseur dans les cellules cancéreuses mammaires et suggère l’existence de mécanismes de résistance à travers les coopérations TrkA/CD44 et TrkA/EphA2, ouvrant ainsi l’opportunité à de nouvelles stratégies thérapeutiques dans les cancers.We have shown that autocrine stimulation of breast tumor cells by NGF increased both cell growth and migration/invasion. These effects are mediated by TrkA, a receptor tyrosine kinases. Recently, it has been described that proNGF, the NGF precursor, is also produced and secreted by breast cancer cells to induce invasion through activation of sortilin/TrkA complex. Nevertheless, NGF and proNGF signaling are still unclear in breast cancer. During my thesis, I have demonstrated that the pro-invasive signaling of (pro)NGF requires the formation of NGF/TrkA/CD44 and proNGF/sortilin/TrkA/EphA2 complexes. I have showed that each of these receptor complexes leads to the induction of two separate transduction pathways, one dependent on TrkA phosphorylation, and the other independently. Furthermore, by using a SCID mouse xenograft model, we have demonstrated the involvement of TrkA/CD44 and TrkA/EphA2 complexes in tumor growth. Interestingly, the co-administration of an inhibitor of the TrkA kinase activity (CEP-701) and a siRNA directed either against CD44 or against EphA2, decreases significantly the tumor burden in comparison to the use of CEP-701, siEphA2 or siCD44 alone. Thus, our findings have improved the knowledge of NGF and proNGF signaling in breast cancer cells and suggest the presence of resistance mechanisms mediated by TrkA/CD44 and TrkA/EphA2 interactions, thereby giving the opportunity for new therapeutic modulations in breast cancer

Targeting copper metabolism to defeat KRAS-driven colorectal cancer.

KRAS-driven cancers acquire profound metabolic dependencies that are intimately linked to tumor growth. Our work revealed that colorectal cancers that harbor KRAS mutations are addicted to copper metabolism. This adaptation renders tumor cells critically dependent on the copper transporter ATP7A, which reveals copper metabolism as a promising therapeutic target for KRAS-driven colorectal cancers

Targeting copper metabolism to defeat KRAS-driven colorectal cancer

KRAS-driven cancers acquire profound metabolic dependencies that are intimately linked to tumor growth. Our work revealed that colorectal cancers that harbor KRAS mutations are addicted to copper metabolism. This adaptation renders tumor cells critically dependent on the copper transporter ATP7A, which reveals copper metabolism as a promising therapeutic target for KRAS-driven colorectal cancers

A Molecular Description of the Cell Surface Proteome in HER2-transformed cells

Despite continuous efforts, breast cancer remains the leading cause of cancer-related death in women. Occurring respectively in 20-25% and 40% of breast tumors, ERBB2 amplification and PIK3CA activating mutations (E545K, H1047R) are among the most common genomic aberrations in human breast cancer. However, significant advances in therapies targeting HER2 and PI3K oncoproteins fail to demonstrate good efficacy, mainly due to tumor heterogeneity and the development of acquired resistance. To expand the treatment options for breast cancer, new targets are desperately needed. In this respect, the characterization of cell surface proteome (surfaceome) changes occurring in transformed cells is essential to identify novel targets for cancer therapy and diagnosis. Insights into the complexity of the surfaceome have been yet limited by the lack of suitable methodologies. Herein, we have optimized a state-of-the-art proteomics approach based on the labeling of cell surface proteins with biotin reagents, their subsequent purification with avidin chromatography, and quantification using label-free quantitative proteomics with liquid chromatography-tandem mass spectrometry (LC-MS/MS). We have employed this proteomics approach to identify secreted and plasma membrane proteins that are differentially expressed on the cell surface of several MCF-10A human mammary epithelial cell lines that reflect the initiation of breast cancer induced either by HER2 overexpression or PIK3CA mutations. Interestingly, our LC-MS/MS analyses identified over 200 cell surface proteins in MCF-10A overexpressing HER2 from which 35% were significantly upregulated compared with isogenic MCF-10A cells. This molecular description of the surface of ERBB2-transformed cells will allow to characterize new putative targets for breast tumors

The MYC oncogene induces specific surfaceome changes associated with TGF-β hypersensitivity in mammary cells.

The c-Myc oncoprotein belongs to a family of “super-transcription factors” that regulates the transcription of several downstream effectors, thereby orchestrating a broad range of biological responses. c-Myc has long been considered the Higgs boson of anticancer drug targets because of its high expression levels and functional deregulation in many human cancers. Although c-Myc inhibition would be a powerful approach for the treatment of many cancers, direct targeting of c-Myc has been challenging owing to its “undruggable” protein structure. Hence, alternatives to c-Myc direct blockade have been widely explored to achieve desirable anti-tumor effects, including c-Myc/Max complex disruption, c-Myc destabilization, as well as the targeting of synthetic lethality proteins. Nevertheless, this enthusiasm has been mitigated by the recognition that c-Myc is almost universally involved in normal physiological processes, thereby raising concerns that c-Myc inhibition would lead to unacceptable toxicities. Intriguingly, recent transcriptomic data from our lab predict that most of the changes associated with c-Myc overexpression involved transcripts coding for cell surface proteins. Elucidating how oncogenic c-Myc modifies the cell surface proteome (surfaceome) could help understand its complex mechanism of action, and possibly identify new “druggable” targets and/or tumor-specific biomarkers. Herein, we have optimized a cutting-edge chemoproteomic approach based on the labeling of cell surface proteins with biotin reagents, their subsequent purification with avidin chromatography, and quantification using label-free quantitative proteomics with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Using mammary epithelial cells overexpressing c-Myc as model system, our LC-MS/MS analyses allowed the identification of >500 cell surface molecules from which 32% and 21% were significantly upregulated and downregulated in c-Myc-overexpressing cells, respectively. Interestingly, we found that c-Myc modulates the surface expression of a large network of proteins associated with TGF-β-induced Epithelial-Mesenchymal Transition (EMT). These results were confirmed using several in vitro assays, which highlighted the existence of a TGF-β autocrine loop induced specifically by c-Myc overexpression. Taken together, these results indicate that oncogenic c-Myc leads to a massive reprogramming of the epithelial cell surface, and suggest new options for targeting c-Myc-dependent cancers