QUEST-MSI : Quantitative Highly-sensitive MALDI MS Imaging of Proteins
The motivation of QUEST MSI is to create a spatially-resolved highly-sensitive analytical platform which can generate valuable proteomic information while avoiding ion suppression and to use these data for image reconstruction. To achieve this challenging goal, we propose a new spatially-resolved analytical technique where images are reconstructed based on data coming from identification/quantification analysis. This will be achieved by rostering the tissue section using liquid micro-extraction system and performing identification and relative quantification LC ESI MS to analyse the extracts from all pixels. Based on our results, we believe that our spatially-resolved analytical technique will be extremely useful to reveal fundamental roles of proteins in physio-pathological processes by getting access to the deep proteome and the ghost proteome. Thus, the overarching goal of the QUEST MSI project is to develop a new experimental and computational strategy to generate MS images based on identification/quantification data for proteins highlighting spatial abundance of identified proteins.
Tag-Mass : New kit for multiplexing and quantifying transcriptome, proteome, glycome, metabolome by targeting MALD-MSI
Support : SATT Nord
Partners : Nucleosyn
GHOST : Alternative proteins
Proteogenomic associated to MALDI MSI for targeting alternative proteins involved in oncological processes
Support : Siric OncoLille, La Ligue Contre Le Cancer, Canceropole Nord-Ouest
Partners : Oncovet Clinical Research (OCR), Sherbooke Univ, Pr. X. Roucou)
With the recent advances of high throughput sequencing in the field of genomics and mass spectrometry (MS) based proteomics, it has been shown that traditional computational genome annotation algorithms have underestimated the number of open reading frames (ORFs). Recent evidence has shown that small ORFs (smORFs) are present in RNAs including non-coding RNAs. Recently, PRISM has shown that many microproteins translated from smORFs can be detected in cancer including ovarian cancer and glioma. Even if their existence has been widely proven, their biological functions remain to be elucidated. Thus GHOST project will respond to questions focusing on the role of microproteins and therefore their involvement in the glioma. Novel potential therapeutic targets can be identified and may not lead to tumor resistance or secondary effects of treatment. Additionally, large scale proteomics analysis combining microproteins and canonical proteins will provide more insights about patient profiles in response to specific treatments.
Delcourt V, Staskevicius A, Salzet M, Fournier I, Roucou X (2017) Small Proteins Encoded by Unannotated ORFs are Rising Stars of the Proteome, Confirming Shortcomings in Genome Annotations and Current Vision of an mRNA Proteomics. 2017 Jun 19. doi: 10.1002/pmic.201700058 Download PDF
Delcourt V, Brunelle M, Roy AV, Jacques JF, Salzet M, Fournier I, Roucou X First evidence that a protein coded by a short ORF, not by the annotated CDS is the main gene product Molecular and Cellualr Proteomics. (in press)
Delcourt V, Franck J, Quanico J, Gimeno JP, Wisztorski M, Raffo Romero A, Kobeissy F, Roucou X, Salzet M, Fournier I (2017) Top-down microproteomics bridged to MALDI MS imaging reveals the molecular physiome of brain regions Molecular & cellular proteomics Protemics 2018 17(2):357-372 Download PDF
Delcourt V, Franck J, Leblanc E, Narducci F, Robin YM, Gimeno JP, Quanico J, Wisztorski M, Kobeissy F, Jacques JF, Roucou X, Salzet M, Fournier I (2017) Combined Mass Spectrometry Imaging and Top-down Microproteomics Reveals Evidence of a Hidden Proteome in Ovarian Cancer EBioMedicine 21, 55-64 Download PDF
Mouilleron H, Delcourt V, Roucou X (2016) Death of a dogma: eukaryotic mRNAs can code for more than one protein Nucleic acids research 44,14-23 Download PDF
Vanderperre B, Lucier JF, Bissonnette C, Motard J, Tremblay G, Vanderperre S, Wisztorski M, Salzet M, Boisvert FM, Roucou X (2013) Direct detection of alternative open reading frames translation products in human significantly expands the proteome PloS one 8, e70698 Download PDF
TBI : Imaging Mass Spectrometry in 3D for brain Trauma
Support : Clic-Imaging
Partners : Univ. Florida (Pr. F. Kobeissy), Univ. Lebanon (Pr. K. Zibara)
Traumatic brain injury (TBI) is a major cause of death and disability worldwide affecting almost 100 000 people per year. A primary mechanical impact due to an external force is followed by a cascade of events involving the recruitment of different cell types, specifically immune system cells. A need for visualization and trafficking of changes post injury at different molecular levels such as lipids, metabolites, and peptides has emerged, rather than relying on conventional methods such as immunofluorescence or western blot to target a specific protein and monitor changes at the level of single cellular class/family. In our project, we track lipid changes post-TBI in a 3-dimensional manner after performing matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-MSI) on rat brain subjected to TBI Lipid mass spectrometry images of injured brain tissue cryosectioned in a coronal view were acquired using a high-resolution mass spectrometer (MALDI LTQ orbitrap XL). Our result reveals a unique lipid signature for the injured cortical tissue which further differentiates into two subgroups of injury. In addition, certain m/z peaks show unique expression to both the injured area and the ventricular system within the brain.
Fimbria : Molecular histology guided Spatially resolved large scale microproteomic coupled to HR MALDI MSI in tubal serous carcinoma and precancerous lesions
Support : I-Site, ULille, SATT Nord
Partners : Comprehensive Care Center Oscar Lambret, OCR
Whereas the ovarian cancer is only at the 7th position in women’s cancer, it is by far the most lethal (only 30-40% survival rate at 5 years). Amongst the different classes, the High Grade Serous Carcinomas (HGSC) is the most represented (>75%) and within this class, 10% of the patients are mutated for BRCA1/2 or presents a Lynch syndrome. They showed an increased risk of 20-40% to present this HGSC from 30-35 years old (versus 1.4% in the global population). In such women, without efficient detection method, the recommended action is the prophylactic adnexectomies (ovary and tubal ablations). Histopathological studies on the excised pieces showed that these HGSC are taking their origin not in the ovary but in the fimbrial part of the fallopian tube from precursor lesions called STICs (Serous tubal intraepithelial carcinoma). In this project, prophylactic adnexectomies, tissues were prepared following the See-FIM protocol (Serial Extended Examination of the FIMbria) and ImunoHistoChemistry (IHC) against p53 and Ki-67 allowed the pathologist to precisely explore the tubal extremities. Lesion of interest (“p53 lesion”, STIC, STOUT, and HGSC) were analyze by HR MALDI MSI coupled to spatially resolved tissue micro-proteomic guided by molecular histology in FFPE tissue.
SentiRad : New approaches to sentinel nodes in ovarian Cancer
Support : INCA (PHRC), H2020
Partners : Comprehensive Care Center Oscar Lambret, ESGO, Chieti University (Pr. S. Albertini)
Since the last 15 years, ovarian cancer pathology (OVC) has been investigated transversally in PRISM by MALDI-MSI coupled to spatially-resolved tissue micro-proteomic. This apaproach was used on ovarian cancer tissues for hunting specific biomarkers and for research on cancer specific markers in OVC and in their sentinel nodes. Novel set of biomarkers involve in immune response inhibition and tolerance have been discovered and their etiology investigated. Knowledge of the origin of the different OVC permits well-fitted prophylactic measures. Moreover, the description of molecular specificity of the proteins of each subtype will permit definition of early biomarkers, useful for early diagnosis. Thus, molecular insights will identify therapeutic targets and clinical trials should no longer aggregate the various ovarian types. Two markers reg-Alpha and Trop-2 are now under multicenter investigation as novel diagnosis markers for OVC. Based on these results, therapeutic target has been investigated and the prohormone convertases PACE4 seems a promising one (See the Therapeutic Axis). OVC project is now in two directions i.e. validating MALDI MS molecular signature on large cohort for clinical diagnosis and prognosis in conjunction with the pathologist and macrophages M1 phenotype reactivation in tumor though PC1/3 inhibition (MacBeth Project).
EMT : Lipidomic approaches for investigating epithelial-mesenchymal transition in breast cancer and investigation of β-catenin in driven metabolic reprogramming of EMT in cancer
Support : INCA (PHRC), H2020
Partners : Comprehensive Care Center Oscar Lambret, ESGO, Univ. Lecce (D. Vergara, Pr. M. Maffia)
Several studies have identified a specific metabolic reprogramming that is associated with the process of epithelial-mesenchymal transition (EMT). Whereas much is known about the association between glucose metabolism and EMT, the contribution of lipid metabolism is not still completely understood. In the present project, we want to investigate epithelial and mesenchymal breast cancer cells by proteomic and lipidomic approaches and identified significant differences that characterized these models concerning specific metabolic enzymes and metabolites including fatty acids and phospholipids. Higher levels of monounsaturated fatty acids together with increased expression of de novo fatty acid enzymes is the distinct signature of epithelial with respect to mesenchymal cells that, on the contrary, show reduced lipogenesis, higher polyunsaturated fatty acids level and increased expression of genes involved in the triglyceride (TAG) synthesis and lipid droplet formation. In mesenchymal model, the diacylglycerol acyltransferase (DGAT)-1 appears to be the major enzyme accounting for TAG synthesis. Inhibition of DGAT1 but not DGAT2 drastically reduces, in mesenchymal cells, the incorporation of labelled palmitate into TAG. Moreover, we recently investigated by knockdown experiments the role of β-catenin and its pathways in the regulation of metabolism and energy homeostasis in breast cancer cells