Periodic Reporting for period 2 - EPIC-XS (European Proteomics Infrastructure Consortium providing Access)
Okres sprawozdawczy: 2020-07-01 do 2021-12-31
The large-scale European proteomics infrastructure EPIC-XS is fundamental in ensuring Europe’s leading role in proteomics by ensuring access for all researchers to prominent European proteomics facilities. Access is provided to the latest generations of mass spectrometry platforms, as well as advanced peptide separation methods, the newest targeted analysis approaches, and state-of-the-art bioinformatics tools. Moreover, EPIC-XS provides access to experts in the field allowing efficient dissemination to first-time users of proteomics technologies. Research studies can yield information spanning the whole life science area including developments towards potential disease biomarkers that can ultimately be used fordetection, treatment, and in some cases, disease prevention.
The main results of the EPIC-XS project, since the beginning of the project are listed below. The project website (epic-xs.eu) provides comprehensive information about the project's goals, publications, protocols, and partners involved.
Progress from the four collaborative cutting-edge Joint Research Activities.
JRA1: Computational proteomics and cross-omics integration
This JRA manages and improves processes and data manipulation for datasets generated by all other JRAs.
Progress:
• Improved algorithms for cross-linked peptide identifications include OpenPepXL, OpenNuXL, Ms-Annika.
• Software controller for enhanced MS data-dependent acquisition FLASHIda and FLASHDeconv includes novel identification tools for top-down proteomics.
• Innovative ionbot search engine evaluated in JRA3 identified several novel histone glycations and glycation derivatives, validated on clinical samples from breast cancer patients.
• Novel sensitive protein-protein cross-linking identification algorithms called OpenPepXL and XlinkX.
• Identification method for protein-DNA cross-links and integrated improved algorithms into OpenNuXL.
• Algorithm capable of extracting information on positional isomers (multiple modified proteins) from recorded fragmentation spectra.
JRA2: Future and emerging proteomics technologies
Progress:
• Ongoing development of a microfluidics approach for handling and lysing single cells on chips for further MS-based proteomics.
• Discovery that hints towards a multi-peptide vaccination approach against highly conserved tumor suppressors may be a viable option in patients with low tumor-mutational burden.
• Development of new rapid DIA methods and novel software tools for the MS/MS analysis of intact proteins.
• Development, optimization, and application of cutting-edge MS methodologies which facilitates
- The study of emergent and obscure post-translational modifications, including SUMOylation, ADP-ribosylation and Citrullination, for which no reliable contemporary MS methods exist.
- Revealing circulating IgG1 repertoire in human plasma is dominated by a limited number of clones in healthy donors and septic patients.
- MS analysis as a suitable technique for analyzing fast photochemical oxidation of proteins (FPOP)
• Improved workflows for the discrimination of bacterial pathogens.
JRA3: Translational proteomics
Improved analysis of clinical and patient-derived proteomes.
Progress:
• Improved analysis of clinical and patient-derived proteomes.
• Identification of more than 1,400 proteins from cerebral spinal fluid samples.
• Optimization of the proteomic characterization of exosomes and other small extracellular vesicles in a panel of cancer cell lines.
• Robust and reproducible pipeline for plasma proteomics of larger cohorts of patient samples
• Identification of panels of plasma protein biomarkers for a variety of diseases, including COVID-19
• Robust and reproducible phosphoproteomics workflow applied to cancer biopsies for diseases stratification
• Novel protocol for in-gel digestion of histones.
JRA4: Proteome organization: structural and spatial proteomics
Progress:
• Novel crosslinking reagent (Phox) commercialized through industrial partners of UU, Bruker Daltonics and PIERCE, available with optimized protocols for any laboratory.
• Chemical cross-linking and hydrogen-deuterium exchange (HDX) to study protein-ds DNA complexes.
• Improvement and extension of the synthetic peptide library for benchmarking cross-linking mass spectrometry search engines for proteins and protein complexes.
• Improved application and method devleopment of limited proteolysis-coupled mass spectrometry (LiP-MS).
• Localization by Reduction and Reconstruction (LoRRe), based on LOPIT protocol but subcellular fractions are partitioned into protein and RNA, separately quantified by quantitative MS and RNA-seq respectively.
• hyperLOPIT technology applied to THP-1 human leukemia cells to examine re-localization of proteins upon treatment with lipopolysaccharide.
•Structural characterization of human aSyn,implicated in the development of Parkinson’s disease.
•Analysis of the plasma higher-order structural proteome.
Transnational Access provision
Since the start of the project, EPIC-XS has provided access for 233 projects involving 210 different users. Proposal submission has exceeded our expectations, indicating a healthy and growing proteomics field. EPIC-XS users are mostly academic, although interest from non-academic parties has also been experienced, mainly to be informed on high-end proteomics technology, but also to participate in EPIC-XS TNA opportunities. Hands-on training helps researchers to develop best practice workflows and aid the dissemination of proteomics data into publicly available databases, thereby broadening the expertise of experienced scientists and those new to the field of proteomics. Also, standard operating procedures and new protocols are rapidly exchanged in between the access sites, to broaden and strengthen the services that can be offered. During the pandemic, many sites found it difficult to keep up with sample analysis because of limited or no access to mass spectrometry equipment as well as restrictions and access to laboratories for parts of 2020 and 2021. The temporary closure of the submission system was necessary to ensure proper functioning of TNA activity. The system will reopen in April 2022.
Progress from the four collaborative cutting-edge Joint Research Activities.
JRA1: Computational proteomics and cross-omics integration
This JRA manages and improves processes and data manipulation for datasets generated by all other JRAs.
Progress:
• Improved algorithms for cross-linked peptide identifications include OpenPepXL, OpenNuXL, Ms-Annika.
• Software controller for enhanced MS data-dependent acquisition FLASHIda and FLASHDeconv includes novel identification tools for top-down proteomics.
• Innovative ionbot search engine evaluated in JRA3 identified several novel histone glycations and glycation derivatives, validated on clinical samples from breast cancer patients.
• Novel sensitive protein-protein cross-linking identification algorithms called OpenPepXL and XlinkX.
• Identification method for protein-DNA cross-links and integrated improved algorithms into OpenNuXL.
• Algorithm capable of extracting information on positional isomers (multiple modified proteins) from recorded fragmentation spectra.
JRA2: Future and emerging proteomics technologies
Progress:
• Ongoing development of a microfluidics approach for handling and lysing single cells on chips for further MS-based proteomics.
• Discovery that hints towards a multi-peptide vaccination approach against highly conserved tumor suppressors may be a viable option in patients with low tumor-mutational burden.
• Development of new rapid DIA methods and novel software tools for the MS/MS analysis of intact proteins.
• Development, optimization, and application of cutting-edge MS methodologies which facilitates
- The study of emergent and obscure post-translational modifications, including SUMOylation, ADP-ribosylation and Citrullination, for which no reliable contemporary MS methods exist.
- Revealing circulating IgG1 repertoire in human plasma is dominated by a limited number of clones in healthy donors and septic patients.
- MS analysis as a suitable technique for analyzing fast photochemical oxidation of proteins (FPOP)
• Improved workflows for the discrimination of bacterial pathogens.
JRA3: Translational proteomics
Improved analysis of clinical and patient-derived proteomes.
Progress:
• Improved analysis of clinical and patient-derived proteomes.
• Identification of more than 1,400 proteins from cerebral spinal fluid samples.
• Optimization of the proteomic characterization of exosomes and other small extracellular vesicles in a panel of cancer cell lines.
• Robust and reproducible pipeline for plasma proteomics of larger cohorts of patient samples
• Identification of panels of plasma protein biomarkers for a variety of diseases, including COVID-19
• Robust and reproducible phosphoproteomics workflow applied to cancer biopsies for diseases stratification
• Novel protocol for in-gel digestion of histones.
JRA4: Proteome organization: structural and spatial proteomics
Progress:
• Novel crosslinking reagent (Phox) commercialized through industrial partners of UU, Bruker Daltonics and PIERCE, available with optimized protocols for any laboratory.
• Chemical cross-linking and hydrogen-deuterium exchange (HDX) to study protein-ds DNA complexes.
• Improvement and extension of the synthetic peptide library for benchmarking cross-linking mass spectrometry search engines for proteins and protein complexes.
• Improved application and method devleopment of limited proteolysis-coupled mass spectrometry (LiP-MS).
• Localization by Reduction and Reconstruction (LoRRe), based on LOPIT protocol but subcellular fractions are partitioned into protein and RNA, separately quantified by quantitative MS and RNA-seq respectively.
• hyperLOPIT technology applied to THP-1 human leukemia cells to examine re-localization of proteins upon treatment with lipopolysaccharide.
•Structural characterization of human aSyn,implicated in the development of Parkinson’s disease.
•Analysis of the plasma higher-order structural proteome.
Transnational Access provision
Since the start of the project, EPIC-XS has provided access for 233 projects involving 210 different users. Proposal submission has exceeded our expectations, indicating a healthy and growing proteomics field. EPIC-XS users are mostly academic, although interest from non-academic parties has also been experienced, mainly to be informed on high-end proteomics technology, but also to participate in EPIC-XS TNA opportunities. Hands-on training helps researchers to develop best practice workflows and aid the dissemination of proteomics data into publicly available databases, thereby broadening the expertise of experienced scientists and those new to the field of proteomics. Also, standard operating procedures and new protocols are rapidly exchanged in between the access sites, to broaden and strengthen the services that can be offered. During the pandemic, many sites found it difficult to keep up with sample analysis because of limited or no access to mass spectrometry equipment as well as restrictions and access to laboratories for parts of 2020 and 2021. The temporary closure of the submission system was necessary to ensure proper functioning of TNA activity. The system will reopen in April 2022.
EPIC-XS heralds democratization of the proteomics field as leading European experts pool together their state of the art technologies, expertise and data sharing. Resources aren’t just limited to MS-based proteomics as access is also offered for antibody-based imaging technologies. The success of the EPIC-XS infrastructure in its availability to all Life Science genres has resulted in over 290 applications, showing the need for a proteomics initiative like this one cannot be underestimated. We will continue our extensive discussions with the management committee, closely monitoring trans-national access, networking, dissemination and other issues the consortium may have. The constructive cooperation of all partners involved and input from its Scientific Advisory Board, User, Review, and Industrial panels will ensure the continued success of EPIC-XS.