Christopher M. Overall (Co-Chair C-HPP), Bill Hancock (Past Chair C-HPP) and contributions modified from several relevant papers on which we are authors.
What is the mission of the Chromosome-centric Human Proteome Project (C-HPP?)
We have started with an easy question to answer. The Human Proteome Project (HPP) of the Human Proteome Organization (HUPO) aims to find high-stringency evidence for all proteins encoded by the human genome. Indeed, one cannot do the best biology and/or medical research without a complete understanding of the parts of the human proteome. Conversely, the international C-HPP teams need the input of biology/disease (B/D) teams to understand the biological context of the parts list. It is a question of focus. A focus on the parts list promotes a measurement- and detail-focused analytical mindset, while a focus on the biological context is focused on outcomes and may ignore individual parts in the context of the overall picture. In conclusion, both mindsets are indivisible parts of the larger HPP initiative.
What is a Missing Protein?
Amazingly, as of 11th January 2019, 2,129 proteins representing 9.3% of all 19,823 human proteins recognized by neXtProt, are currently defined as ‘missing’ without compelling mass spectrometry (MS) or other evidence of protein existence (PE). These proteins have classification as PE2 – 4. Moreover, a surprising 1,057 PE1 proteins have only been identified by protein chemistry or other techniques, and still lack convincing evidence at the MS level with ≥ 2 proteotypic, non-nested peptides, ≥ 9 amino acids in length. Such PE1 entries that are missing MS evidence are designated as MS-Evi PE1 proteins.
Why do some proteins elude detection by mass spectrometry?
Various factors influence the ability of MS-based technologies to unambiguously detect missing and MS-Evi proteins. Conventional bottom-up workflows utilize trypsin to digest samples prior to LC-MS/MS analysis. Thus, a typical proteomics experiment is unlikely to detect proteins that are not amenable to trypsin digestion, or proteins which yield tryptic peptides that are difficult to ionize, fall outside of the range of typical mass detection ranges, or are not proteotypic (i.e. unique to a particular protein). Furthermore, proteins that are extensively crosslinked, insoluble or expressed in very low amounts or in a very limited temporal-spatial manner or found in challenging tissues to extract for analysis, for example mineralized tissues (bone, dental enamel, dentine and cementum), also present challenges for detection by MS. Other factors are limitations in mass spectrometry instrumentation and search algorithms—despite overlap, various search engines will preferentially identify different subsets of peptides.
Are missing proteins missing because they are unimportant?
No. The olfactory receptors are a compelling example of the lack of our knowledge in important areas of biology. These receptors represent a large class of proteins whose coding regions appear to be distributed over almost all the chromosomes. The proteins are extensively expressed in nasal tissue but there is also unconfirmed evidence at the protein level that olfactory receptors may also be expressed in other tissues, ranging from reproductive organs to tooth dentine to the brain, and they appear to have important interactions with the immune system. Another huge example are the hundreds of ORF genes for which tissue expression of mRNA (PE2) has been demonstrated. MPs are also expressed in less well studied cells and tissues that may provide phenotypic differences to these cell populations and tissues, but are difficult to sample or access.
Have we finished the C-HPP once we have found all the missing proteins?
No. Finding all MPS is just Phase I of the C-HPP. Next comes functionalization of the PE1 proteins with no known or predicted function, or homology or orthologues with known function. Extensive goals such as ASV and PTM characterizations follow; thus, each team has many proteomic adventures ahead. Our objective across the HPP will be to identify and characterize the dynamics, functions, associations and turnover of their numerous isoforms and variants, their evolution, and their roles in pathways, networks, embryogenesis, growth and development, and disease processes.
How is proteomic data captured and used for the C-HPP?
While a quality review is the foundation of a scientific study, the acceptance of a proteomic study for publication, even in high impact journals, does not guarantee the use of its dataset. Many experimental details and lab-to-lab variations make very difficult the comparison of the datasets from different research groups, especially in a global collaboration like the C-HPP. For the C-HPP to succeed, the data must be curated and accepted at either PeptideAtlas (http://www.peptideatlas.org) and/or now at MassIVE (https://massive.ucsd.edu/ProteoSAFe/static/massive.jsp) and integrated into the databases. Furthermore, our gold standard for the C-HPP is acceptance with gold-level evidence in neXtProt (https://www.nextprot.org).
Once a dataset has been deposited in ProteomeXchange, is the task complete?
No. The authors must also provide the PXD identifiers in the publication, preferably also in the abstract, and then provide permission for the dataset to be made public. To facilitate the dataset’s use by other teams, additional steps should be considered to guide each team to access the C-HPP Wiki (https://c-hpp.web.rug.nl/tiki-index.php?page=HomePage) and distribute the produced dataset with biological and other details. MissingProteinPedia (http://www.missingproteins.org/) also contains orthogonal MP evidence.
How is a Missing Protein identified for the HPP?
Peptides and proteins are identified in proteomic data sets, but only those reaching very high stringency in spectral quality and sequence coverage are considered as unequivocal evidence for the promotion of a PE2-4 protein (missing protein) to a PE1 protein (i.e. found). Full details of the criteria used to designate a missing protein as found are those meeting the current HPP Guidelines (v 3.0 in press) (https://hupo.org/Guidelines, and for v2.1 DOI: 10.1021/acs.jproteome.6b00392, J. Proteome Res. 2016, 15, 3961−3970).
Notably, while all peptides that pass thresholds are visible in PeptideAtlas and neXtProt, only the proteins with two uniquely-mapping non-nested proteotypic peptides with length 9AA or greater, as called by neXtProt, are deemed to have sufficient evidence to be labeled as confidently detected by MS methods. Therefore, neXtProt is the final arbiter to designate if a PE2 – 4 protein in UniProtKB is now deemed PE1 in neXtProt for the purposes of the HPP.
What are the details of the HPP Data Processing Pipeline?
The current basic process by which the HPP manages the process of reducing the number of missing proteins of the human proteome begins with the collection of mass spectrometry data sets and deposition in one of the ProteomeXchange repositories after registration with ProteomeXchange. From their ProteomeXchange deposition, PeptideAtlas collects the raw data files and reprocesses those data using the tools of the Trans-Proteomic Pipeline (TPP). In 2019 the C-HPP is now also utilizing the MassIVE database. Thresholds are set extremely high in PeptideAtlas in order to obtain a 1% protein-level FDR across the ensemble of all datasets. In November each year, PeptideAtlas stops processing new datasets and creates a build reflecting the current state of the human proteome with MS evidence. In December the final peptide list is transferred to neXtProt for integration into its next build, usually in January or February each year. The official numbers to determine progress on the HPP are taken from the January/February neXtprot release, even though neXtprot is updated several times a year, which also change the numbers of proteins in each of the PE categories.
Does the C-HPP study biology and/or disease?
Virtually all proteomic experiments are performed in a chromosome agnostic manner. Further, most of the chromosome teams conduct at least some of their search for the missing proteins (PE2 – 4) in samples relevant to biology or disease that allows the observation of unusual protein expression patterns that will provide unique insights for other biological studies. Many teams employ clinical studies to search for ‘rare’ proteins that are observed only in the unusual context of a disease state or rare human cells and tissues. Examples of teams supplying samples and data between groups include the US and ANZ teams on chromosome 17 and 7 and the Chinese teams (Chr 1, 8, 20) and Korean teams (Chr 9, 11, 13).
Does each chromosome team study only one chromosome?
Because proteomics experiments study proteins expressed potentially form every gene, the data generated are relevant to all the chromosome teams and thus must be shared with all the groups. Furthermore, a team’s B/D studies will involve all the chromosomes, and these insights will be shared across teams. A focus of the C-HPP teams can also be on the parts list and biology of co-expression or cis-regulation of genes on that particular chromosome. Such biological features, exemplified by the HER2/neu (ERBB2) amplicon at chromosome 17q12, reinforces the scientific rationale for chromosome-by-chromosome studies of the proteome.
Is the value of a team defined by the number of missing proteins that they discover?
While the number of proteins discovered is an important metric, the data are only valuable once deposited and accepted by Peptide Atlas, MassIVE, and neXtProt. Now that C-HPP is well established, there are new opportunities to collaborate with the B/D groups to begin to understand the function of previously missing parts of the proteome. This is particularly relevant for functionalization of proteins, both MPs and PE1s, that have no known or predicted function, termed uPE1 proteins. Such flexibility will promote additional interactions between chromosome teams and B/D-HPP teams and allow for differences in priorities for various national and international funding agencies.
How does the C-HPP facilitate interactions and collaborations?
To facilitate productive and mutually beneficial interactions and collaborations within the C-HPP and between the C-HPP teams and those in B/D-HPP and the four support pillars of the HPP, in 2013 the C-HPP:
Dear HUPO Members,
The Human Proteome Project (HPP) was launched in 2010 under the aegis of HUPO. The HPP was built upon a matrix structure, incorporating existing HUPO scientific initiatives (e.g., plasma, brain, liver proteome, etc), the gene/chromosomal-centric (started 2012 and abbreviated as C-HPP) and protein-biology-disease-centric approaches (also started 2012 and called B/D-HPP). These are now underpinned by 4 resource pillars (affinity/antibody reagents, mass spectrometry, knowledgebase and pathology). The vision of HUPO is that HPP activities will collectively and ultimately lead to breakthroughs in medicine, biotechnology and the life sciences, thereby leaving a legacy of human proteome research.
The HPP continues to make progress, addressing two specific challenges;
In January 2019, Mark Baker (previous HUPO President), took over as Chair of the HPP to carry on the mantle from Gil Omenn (HUPO co-Founder and Chair of the HPP since its inception) in driving the current and future initiatives of the Human Proteome Project. With the expanding goals of the HPP and aggressive timelines to drive the numerous initiatives under the HPP umbrella of activities, we seek to engage a vibrant, well-organized, and goal-oriented proteomics researcher to the newly created HPP Co-Chair position to support and assist the HPP Chair.
The HPP Co-Chair position is a 2-year term and will commence January 2020. Responsibilities of the HPP Co-Chair include:
The HPP is seeking a strong, strategic, vibrant, enthusiastic and collegial leader who would be a suitable candidate to support and represent the HPP. HUPO is keen to ensure regional, gender and early career scientist equity across its management structures. This position is honorary and in line with the many organizational positions within the HUPO executive committee.
Applications will be reviewed and voted on by the HUPO and HPP Executive Committees, and the successful candidate will need to be ratified by HUPO Council at their HUPO2019 Meeting in Adelaide.
To apply, please submit a brief (<1 page) vision statement outlining why you are a suitable candidate for this position. Email vision statement to email@example.com before August 23, 2019.
Marta del Campo Milan, Charlotte Teunissen
We are glad to announce the 29th HUPO Brain Proteome Project (HBPP) Workshop will be held on 27th and 28th of May 2019 in Amsterdam, The Netherlands.
HBPP Spring Workshops cover a wide range of topics relevant to neuroproteomics research. The program is highly dynamic and interactive, and all participants are given the opportunity to give an oral presentation on their research or updates of novel techniques and analytical approaches used in their labs. Workshops have included, for example, sessions on myelin proteomics, autoimmunity, and bioinformatics.
HBPP is committed to a strong clinical and translational focus. Hence, sessions in last meetings were dedicated to proteomics of psychiatric disorders, movement disorders, spinal cord injury & trauma, dementia, and cancers of the CNS. On top of that, there will be a very nice evening program that will allow all the delegates to have relax interactions with a unique local atmosphere.We encourage every researcher interested in brain proteomics, including junior scientists to get in contact and join this event. For more information about the workshop and abstract submission please visit https://www.hbpp2019.com/.
We would like to ask you to extent and share this information between all the researchers and/or in your mailing list. It would be also very helpful if you could include our flyer in your main webpage, which is attached within this email.
Thank you in advance for your collaboration!
Péter Horvatovich, University of Groningen, The Netherlands
The Journal of Proteome Research Special Issue 2018 (Associate Editor: Christopher M. Overall, Guest Editors: Paik, YK, Eric Deutsch, Fernando Corrales, Lydie Lane and Gil Omenn) was published on December 7, 2018 (Volume 17, Issue 12). In this issue, a total of 32 papers covered 4 major research topics: (i) missing proteins (MPs), (ii) uPE1 proteins, (iii) bioinformatics tool development and (iv) biology/disease proteomes. According to the article summarising the progress on identification status and metrics of the Human Proteome Project, the number of missing proteins (PE1+2+3+4) decreased from 2579 to 2186 and the number of proteins with sufficient evidence at proteome level (PE1) reached 17470, which represents 89% of the human proteome, while the number of dubious proteins is 574. From the 17470 PE1 proteins there are mass spectrometry evidence in the Peptide Atlas for 15798 proteins. The launch of the neXt-CP50 pilot project to find at least one function of well identified 1260 PE1 proteins with unknown function (uPE1 proteins) using state-of-the-art gene editing technologies such as CRISPCas or gene silence with miRNA is discussed in Young-Ki Paik et al. Deutsch et al discusses all aspects of the use of spectral libraries and spectral library search in proteomics workflows including quality at library, spectra and peak (fragment ion) levels the used spectral similarity methods, construction of consensus spectra and merging different spectral library following the discussion at the 2017 Dagstuhl Seminar on Computational Proteomics. The work of Macron et al. describe identification of missing proteins in human cerebrospinal fluid following immunodepletion and TMT labeling. This work identified 12 missing proteins candidates from which 8 proteins were identified based on 2 to 6 uniquely mapping peptides and 4 matched a new peptide with a complementary “stranded” single peptide in PeptideAtlas from previous CSF studies. Sun et al. presents a study on human testis, using multiple proteases and high and low pH deep proteomics analysis and identified 14 PE2 MPs after spectrum quality analysis, isobaric post-translational modification, and single amino acid variant filtering, and synthesized peptide matching, from which 3 was testis specific. The study by He et al. used LysargiNase, the trypsin “mirror protease” that cuts before lysine and arginine equally as efficiently as trypsin that cuts after the basic residues, to identify low molecular weight missing proteins and validated 2 MPs from 7MPs candidates. Pullman et al. presented the tool ProteinExplorer, which allows to explore the large amount of reanalysed public proteomics data available in MASSIVE, which allowed to build a spectral library containing 2.1 million precursors matching to 1 million unique peptides and 19000 proteins. The use of ProteinExplorer allowed to validate HPP-compliant evidence for 107 MPs (PE2, PE3, and PE4) and 23 dubious (PE5) proteins.
Michelle Hill, QIMR Berghofer Medical Research Institute, and The University of Queensland, Australia
The 2018 post-HUPO congress HPP workshop kicked off with a new interactive session moderated by Rob Moritz, where selected senior and young investigators each had <5 minutes to share what they found most exciting during the entire HUPO congress. All speakers strove to beat Rob’s musical interlude, most were successful. The floor was then opened to all participants, with great diversity of inputs and interests. This new format was a refreshing and productive start to the workshop.
The HPP Town Hall Meeting session led by Fernando Corrales and Young-Ki Paik outlined the activities and goals for the B/D- and C- HPP, respectively. Mark Baker then presented his vision and strategic plan for the NextGen HPP, with input and discussions from the senior Scientific Advisory Board (SAB) members, Cathy Costello, John Yates and Naoyuki Taniguchi. Although the B/D- and C- HPP branches were organised for logistics in the first phase of HPP, synergistic efforts are being established between various B/D- and C- HPP teams. The SAB further challenged the NextGen HPP leadership team to aim for HPP integration in the coming years.
With the establishment of the Pathology pillar, HPP is inviting external interdisciplinary partners towards clinical translation. Steve Pennington and Henry Rodriguez shared their views and experiences on the strategies for successful international translational research. Finally, Mike Synder delivered insightful grand challenges for the entire field of proteomics.
The post-congress workshop ended on a high note with celebration of Gil Omenn’s leadership of HPP, with refreshments. The well-researched presentation by Mark Baker provided a rare insight to the talents and achievements of Gil.
Péter Horvatovich, University of Groningen, The Netherlands
With a good attendance at the Thursday HPP workshop, talks on the Knowledge Base Pillar progress and outlook were well presented by Eric Deutsch, Lydie Lane and Henning Hermjakob. They highlighted the raw MS data exchange between ProteomeXchange and partners such as Pride, iProx, JPOST, MassIVE and Panorama, which serves to collect all public proteomics data generated by the scientific community, protein evidence status in PeptideAtlas and neXtProt according to the latest release. The report also gave updates on the peptide uniqueness checker and introduction of Universal Peptide Identifier that aims to identify specific MS/MS spectra in a ProteomeXchange dataset.
Measurement of intact proteoforms with top-down approach to map the proteome of human cell types (a Cell-Based Human Proteome Project) was presented by Neil Kelleher. In discussions with the Chairs of the C-HPP a collaboration has been formalised to prepare a proposal for the C-HPP and HPP ECs titled “The Human Cell Proteome Atlas”. This is an extension of the current rare cells and tissues project of the C-HPP led by Chris Overall. In the HCPA the current bottom up rare cells and tissues proteomics projects of the C-HPP would be complemented by the a top-down proteomics approach led by Neil to produce an atlas of proteins and proteoforms making up each of the hundreds of different cells of the human being.
On emerging technology topics, Bonghee Lee (Korea) presented CRISP/cas9 technology used to modify human iPSC and differentiated cells for Hemophilia B treatment . Janne Lethiö (Sweden) presented proteogenomics pipeline that can be used to process high-resolution iso-electric focusing fractioned peptides to better annotate protein coding region of the human genome and identify variants in human blood. Mike Snyder presented ‘Grand Challenges for the Entire Field of Proteomics’ where he addressed an outlook and plans on the directions of the proteomics technology development and those key issues that the HPP will address in the future. Henry Rodrigez showed an overview on the international collaboration within Cancer Human Proteome Project and within the Cancer Moonshot international collaboration. Mark Baker, incoming HPP Chair (Australia) outlined his strategic plans on the key HPP components, C-HPP and B/D-HPP in the next 2 years. The Antibody Validation Initiative showed their progress to develop validated antibody reagents for the scientific community. The September/October neXt-MP50 report from the Chromosome Teams was presented to the HUPO council. The chromosome teams reported the identification of ~401 missing proteins identified according to the C-HPP guidelines in 2018. These were presented in 26 J Proteome Res Special Issue papers in December 2017 and will be reported in 34 J Proteome Res papers in the upcoming December 2018 Special Issue. In addition, the Chromosome Teams reported HPP findings in 63 other papers in 2018, many of which reported missing proteins or uPE1 functionalization. At the end of the first quarter 2019 the next builds of the Peptide Atlas and NeXtProt will have parsed through these and other Proteome Exchange data sets for confirmation of found MPs.
The C-HPP welcomes the leadership of Dr. Rob Moritz, Institute of Systems Biology, Seattle, WA as the new leader of the Chromosome 6 Team, that has passed from Christoph Borchers, University of Victoria. The experimentalists, coders and data-base curators at ISB are a welcome addition to the C-HPP and join active labs at the University of British Columba, Vancouver, in a “Cascadia Chromosome Collaboration”.
Plans are moving forward for the 21st C-HPP Project Symposium, to be held May 12-14, 2019 in Saint Malo, France. Focussing on “Illuminating the Dark Proteome” the symposium will be an important milestone for the uPE1 project and will include speakers from the B/D-HPP on functionalizing the dark proteome. What is also certain, the cuisine at this meeting will not be dark or need illumination thanks to Charles Pineau, Chair of the workshop.
Miguel Marcilla, Spanish National Biotechnology Centre, Spain
A total of 57 attendees –including 41 academics and 16 delegates from the industry– from 19 different countries gathered together at the Spanish National Biotechnology Centre in Madrid for the 1st HIPP Summer School on Immunopeptidomics. To provide students with a critical understanding of the challenges and experimental strategies in immunopeptidomics, the contents of the Summer School were structured around three fundamental topics: sample preparation, LC-MS/MS analysis and bioinformatics and data sharing. The course encompassed keynote talks by recognized experts in the field, training lectures and poster presentations by the participants. Additionally, it also included a dedicated session to promote networking with representatives from companies actively working in the area of immunopeptidomics.
Organizing Committee and Sponsors
The 1st Summer School on Immunopeptidomics was organized by the following members of the HIPP initiative: Nicola Ternette (University of Oxford, Oxford, UK), Stefan Tenzer (University of Mainz, Mainz, Germany), Miguel Marcilla (Spanish National Biotechnology Centre, Madrid, Spain), Fabio Marino and Michal Bassani-Sternberg (both from the Ludwig Institute, Lausanne, Switzerland). In addition, the organization of this event was possible thanks to the financial support of Agenus, Genentech, Bioinformatics Solutions, the Spanish Proteomics Network (ProteoRed) and the HIPP.
Final Remarks and Future Editions
This Summer School proved to be an ideal setting for discussion and exchange of ideas allowing young researchers to meet renowned experts as well as to interact with delegates from companies. Based on the results of a survey completed by the participants, the overall perception of the quality of the course was high and it fulfilled the expectations of both the attendees and the organizers. Considering its success, HUPO-HIPP is promoting the organization of a new edition of the Summer School to be held in one or two years´ time.
The annual meeting of the C-HPP and C-HPP PIs (PIC) was held at HUPO 2018 in Orlando, Florida. The meeting was very well attended by members and PIs. A progress report on the next-MP50 was prepared before the meeting and will be updated when all reports have been submitted and uploaded to the C-HPP Wiki. Several important decisions were made after extensive discussions. Following Young-Ki Paik’s Paik (Yonsei University) proposal to rotate the leadership of C-HPP. Chris Overall (University of British Columbia) was unanimously elected as C-HPP Chair. Young-Ki was then elected as the new Co-Chair with Lydie Lane continuing her current position as shared Co-Chair of C-HPP Executive Committee (EC).
Collaborations with B/D-HPP. To further the collaborations with the B/D-HPP, the membership of the C-HPP agreed that the uPE1 project was ideal to combine with the talents and resources of the B/D-HPP, and in discussions with the B/D-HPP leadership this too was agreed (see Figure, noting that for clarity the Pillars etc. are not shown). Relevant papers in the JPR SI can be found in PMID 30269496.
Human Cell Proteome Atlas. As part of the rare cells and tissues strategy to find MPs, a parallel value-added project to annotate the proteins of specific normal human cell types received considerable support with the view of creating a Human Cell Proteome Atlas that would be done in collaboration with B/D-HPP. The aim of the HCPA is to annotate the proteomes of normal human cells (fresh prepared or low passage number primary culture) so as to provide an authorative and convenient single lookup data base that would complement the other resources based on antibodies (Human Protein Atlas), Cellosaurus, and various genetic based resources. During HUPO the C-HPP initiated discussions with Dr. Neil Keller to bring this proposal to the HUPO EC for discussion and approval.
C-HPP 2.0. Deep discussions took place on a proposed “chromosome-rearrangement” of the working nucleus of the C-HPP, which had been floated to the membership at the Santiago C-HPP workshop on 23rd June. The proposed new structure, as part of “C-HPP 2.0”, would collapse the number of national Chromosome teams by approximately half based on their activities. Freed-up C-HPP team members would remain as members at large or nucleate bottom up to form new annotation C-HPP annotation teams based on protein families, cell types, or new technologies. Such teams would have equal standing with those teams wishing to remain chromosome based.
Following further discussion, the membership clearly expressed their wish and desire to pursue their national projects under the current chromosome-organised structure. Points brought up included that Chromosome teams very much liked having a recognised national “brand” as a platform to nucleate like-minded labs and centers to work collaboratively together, importantly a Chromosome “face” was valuable to interface more effectively with Universities, Funding Agencies and Governments. It was also expressed that this structure was the most effective to support the current projects, many of which are in fact chromosome orientated, i.e. finding missing proteins (neXt-MP50), discovering uPE1 protein functions (neXt-CP50), and to maintain funding and salaries by some teams that have secured significant operating funds. Members of the Italian Mt team proposed an innovative cross matrix solution that has been drawn in the figure, that was greeted with great enthusiasm as meeting the new needs and scientific objectives of the C-HPP under the HPP umbrella.
A motion to retain the existing Chromosome Structure was seconded and put to the vote, which was carried unanimously.
Bioinformatics Hub. In other activities at the bioinformatics hub shared great interactions between the leaders of the main HPP resources and tools (ProteomExchange, PetideAtlas, MASSIVE, neXtProt and HPA) in order to make the most of the available data and complete the annotation of the human proteome parts list.
Annual C-HPP Workshop. The annual C-HPP workshop entitled: “Illuminating the Dark Proteome” has a major goal to advance uPE1 functionalization in the next-CP50 project. The workshop is organized by Charles Pineau in collaboration with Chris Overall and Fernando Corrales. The workshop will take place in Saint-Malo, France, a historic city on the Brittany coast (France), from 12 to 14 May 2019, with the hotels within the actual historic citadel and featuring excellent French cuisine and wines.
HPP Scientific Terms, Definitions & Abbreviations
Definition of most often used terms such as Human Proteome Project (HPP), Chromosome-Centric Human Proteome Project (C-HPP), Biology/Disease Human Proteome Project (B/D-HPP), Protein Evidence (PE), Uncharacterized PE1 proteins (uPE1s), Missing Proteins (MPs), HPP Guidelines, Dark Proteome, Proteoforms, neXt-MP50, neXt-CP50 and Popular proteins. Exact definitions of these terms with explanation and link to scientific article as supporting reference, to additional resources such as list of MPs or uPE1s is provided in PDF document at the HUPO webpage, at the C-HPP Wiki, and as wiki page with live links at C-HPP Wiki. We encourage the scientific community to discuss further these terms and complete it with additional ones during the HUPO 2018 Orlando congress.
C-HPP Wiki update
The program of C-HPP activities at HUPO 2018 (Orlando, US, September 30-october 4, 2018) are summarized at C-HPP Wiki, showing pre and post congress C-HPP programs, Bioinformatics Hub and C-HPP sessions. Many presentations provided by authors are already available at C-HPP Wiki and others will be made public upon availability in the next few weeks. We encourage the C-HPP Chromosome members to share news, presentations and highlights on their work themselves on the C-HPP Wiki page.
Launch of the HPP Pathology Pillar at HUPO2018 was met with great enthusiasm, particularly with the clinical scientist and pathologist travel grant winners. “I was very excited to learn about the establishment of the new Pathology Pillar of the Human Proteome Project”, said Kun-Hsing Yu (Department of Pathology, Brigham and Women’s Hospital and Department of Biomedical Informatics, Harvard Medical School, USA). Peter Stewart’s (Royal Prince Alfred Hospital Sydney) advice to tackle the “low-hanging fruit”, as a strategy forward particularly struck a chord with Yunki Yau (Gastroenterology Department, Concord Repatriation General Hospital, AUSTRALIA): ‘By improving on current clinical tests that aren’t satisfactory with proteomic techniques and technology, we can foster confidence in our workflows in the wider clinical community.”
The seven travel grant winners, the pathology pillar leaders and the Biology Disease Human Proteome Project Executive Committee members had a productive roundtable meeting in Orlando. HUPO Young Gun, ECR member and clinician-scientist Ferdinando Cerciello (Department of Oncology, University Hospital Zurich, Switzerland) moderated the discussion.
In-depth discussions ensued around the topic “Can proteomics speed up drug discovery?””, posed by Ferdinando. Danni Li (Director of Clinical Chemistry, Department of Laboratory Medicine and Pathology, University of Minnesota) posed the thought-provoking question, what is proteomics? “To me, proteomics is to measure proteins. In this definition, proteomic technologies have been used for drug discovery for a long time.”, said Danni. Yunki Yau agrees. “Are we fundamentally interested in making MS-based protein assays a clinical instrument? Or do we essentially want clinical legitimacy for protein biomarkers and drug targets, no matter how we do it?” Jesper Kers (Department of Pathology, University of Amsterdam & VU University Amsterdam, The Netherlands) also voiced the point “Clinical proteomics should be driven by clinician’s unmet needs, because they are the most important potential end-users of the technique.”
In addition to the current strategy of engaging clinicians & pathologists, there was agreement on the importance of engaging the pharmaceutical companies in future HUPO meetings. Michael Roehrl (Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, USA) suggested that “we all need to engage directly with biotech and pharma to make functional proteomics a key strategy for the next generation of companion diagnostics.”
Several clinician scientists and pathologists strongly agree with the need for standardization and automation, which was highlighted by several speakers during the HPP track session: Partnering with Pathology Towards Precision Medicine. “I think the HUPO pathology pillar can play a crucial role in standardization of proteomics assays among laboratories in order to assure high quality and reproducible results.”, says Jesper Kers.
Ferdinando’s second discussion point “Can next-gen proteomics biomarkers be as good or better than current imaging/diagnostics?” brought additional discussions around reimbursement and funding of clinical tests. There was general agreement that proteomics can and should add value to the existing tests, with opportunities and strategies to cross barriers discussed in detail. Jesper Kers suggests that “An example where I think mass spectrometry can have an additional value over DNA sequencing is holistic profiling of immune responses, in both the setting of oncology and immunometabolic diseases.”
As 90 minutes quickly passes, Jennifer Van Eyk (Cedars Sinai Medical Center) suggests the preparation and publication of a white paper on the key points and conclusions, as a way forward. With support from the group, Michael Roehrl graciously agrees to lead this endeavour. “We need to work as a community to firmly establish proteomics as the new frontier of theragnostic molecular testing in pathology”, says Michael. From the roundtable and HUPO congress, Margaret Simonian (University of California Los Angeles) was inspired to collaborate between all omics, clinicians and scientist “to advance treatments and diagnostics in clinical medicine and precision medicine.”
The Human Proteome Project (HPP) MS-Pillar Phosphopeptide challenge resource continues to collect new data and invites new participants to also contribute to this knowledgebase. Participants are invited to contribute to both phases of phosphopeptide identification methods development by analyzing a set of phosphopeptides by their favorite method and follow up with a method of affinity purification step using the popular ReSyn HUPO affinity kit to see improvements in their methods. Together with our partners, SynPeptide Co. Ltd in Shanghai (www.synpeptide.com) and Resyn Biosciences Pty Ltd in South Africa (www.resynbio.com), the HPP MS-Pillar will provide the SynPeptide-HUPO phosphopeptide mixtures as well as a comprehensive ReSyn phosphopeptide purification kit to use as the affinity method for the HUPO phosphopeptide mixtures. The SynPeptide phospho peptides and the Resyn MagReSyn® kit including the magnet separator are valued at over US$1000 each are provided free to all interested HUPO members.
The MS-Pillar Phosphopeptide challenge samples are a complex set of human phosphopeptides (Ser, Thr or Tyr) singly and multiply phosphorylated) and their unphosphorylated counterparts that can be used for method development and verification for phosphopeptide enrichment, sequence analysis by mass spectrometry and bioinformatic evaluation.
The first stage of this initiative is for interested members of the HUPO community to obtain a set of the peptides and apply their own methods and bioinformatic analysis to fully characterize the peptides as a neat mixture and in a tryptic digest background. Samples are still available by contacting the email below.
The second stage is now to compare your methods used with the Resyn purification kit for phospho-enrichment and peptide clean-up. The kit contains 2mL of each MagReSyn® Ti-IMAC, Zr-IMAC, and HILIC, and 4-place magnetic separator. As a result of this collaborative endeavor, multiple purification schemes, analytical protocols and data processing strategies will be evaluated, making it possible to determine the approach(es) that provide the highest coverage of phosphopeptides in the mixture.
By partnering with Resyn and SynPeptide, the HPP MS-pillar has combined the peptide sets and affinity purification kit free-of-charge to all HUPO members, with the understanding that the analytical and data processing methods and results will be returned to the HPP MS Resource Pillar committee, so they can be collated, combined with the HUPO 2018 data, and reported at the HUPO 2019 meeting.
The HUPO phosphopeptide mixtures Peptides and iMac kits can be picked up from SynPeptide and Resyn at their booth at HUPO 2018 Orlando.
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