Proteomics at the heart

of multiomics studies
Contents
Chapter 1 ............................ 4
Finding proteins that cause
disease: pQTLs are the key

Chapter 2 ............................ 9
How proteomics enriches the
multiomic study of disease

Chapter 3 .......................... 16
The future of proteomics in
multiomics research

2
Introduction
Achieving the goal of precision medicine and more targeted therapeutics will require the
use of systems biology approaches to understand the molecular mechanisms at work
within the human body. This trend is already apparent in scientific research, with more
scientists beginning to use multiomics studies to better understand disease and to help
develop the drugs needed to treat them. To this end, the integration of data from high-
throughput proteomics technologies is essential, as proteins best represent individual
phenotypes and the effects of environmental and lifestyle factors. In other words, proteins
best reflect our real-time biology, which is key in developing precision medicine.

This ebook aims to show how the integration of proteomics data in multiomics studies
is advancing disease research and drug development, and to consider what lies ahead
in terms of trends and challenges in multiomics research. Each chapter comprises a
curation of excerpts from academic and industry key opinion leaders on three central
themes: How combining genetic data with proteomics help researchers identify proteins
that cause disease, how proteomics adds value to multiomics studies on complex
diseases, and the future of proteomics in multiomics research.

Read on to learn how combining multiple omics technologies, with proteomics at the
heart of such strategies, can give a much clearer and comprehensive picture of real-time
biology, and how this will drive future healthcare development.

3
Finding proteins that cause
disease: pQTLs are the key
The ‘DNA revolution’, prompted by the sequencing of the human genome, has transformed many
aspects of our biological understanding of disease through explosive growth of our knowledge of
genetic variation. DNA testing has been effective in the areas of diagnosis for monogenic disorders
and risk susceptibility. For example, in oncology, geneticists have identified cancer susceptibility genes,
notably for breast and ovarian cancers (BRCA1 and BRCA2), colorectal cancer (MLH1 and MSH2) and
melanoma (CDKN2A). Through genome-wide association studies (GWAS), polygenic risk scores have
been used widely in research and show promise for clinical implementation to assist in diagnosis, inform
treatment options and individualize screening frequency.

Several biopharmaceutical companies have focused on RNA for its therapeutic potential because of
its role as a messenger of genetic information; RNA was perceived to be a closer reflection of real-time
health. Certain RNA-based therapeutics such as anti-sense oligonucleotides (ASOs) or short interfering
RNAs (siRNA) have been approved as drug treatments. With the coronavirus pandemic the usefulness
of RNA-based vaccines has been demonstrated. Yet, despite these ongoing advances in using DNA and
RNA to improve drug development, there is still room for improving drug development.

Approximately 95% of all current approved drugs target proteins. Proteins provide the final link in
the central dogma between genetics and disease. Until now, the wealth of information that proteins
could provide was limited by insufficient scalability, efficiency, and throughput of protein analysis
methods. However, much like with DNA sequencing technology, over the last several years proteomics
technologies have undergone somewhat of a revolution. It is now feasible to conduct proteomics
analyses at the scale and throughput achieved by DNA and RNA sequencing methods, making it
possible to include protein data as part of large multiomics analyses.

Large scale studies integrating proteomics, genomics, and clinical data are ushering in a new paradigm
to identify potential therapeutic targets and biomarkers for the development of clinical diagnostics,
as well as proteins that may be useful for clinical trial treatment stratification. This has evolved into
an emerging field called proteogenomics, where researchers are using protein quantitative trait loci
(pQTLs) to link genetic variation, proteins, and disease.

4
What are pQTLs?
Protein quantitative trait loci are locations in the genome that contain
one or more genetic variants associated with circulating protein levels.
Genetic variants that are close to, or within the same gene that makes
the protein are called cis-pQTLs, and those that are in a different
location, and therefore indirectly affect the protein-making gene, are
called trans-pQTLs (Figure 1).

Of particular interest to disease researchers are the genetic variants

that are connected to both protein levels and disease. In a podcast with
Drug Discovery News, Dr. Anders Mälarstig, Director of Human Genetics
at Pfizer, explains pQTLs in more detail, and how they benefit the drug
development process (Podcast snippet 1.1). Podcast 1
Dr. Anders Mälarstig, Director of
Target sciences at Pfizer Worldwide
Research & Development and
Affiliate Researcher at Karolinska
Institute, talks about how
proteogenomics can improve drug
discovery and development.

Listen to the podcast

Figure 1 Schematic representation of a cis- and trans-pQTL, and how they affect protein levels. A cis-pQTLs is a genetic variant (typically a
SNP) that lies very close to (<1 Mb) or within the gene that codes for the protein (Protein 1) being measured. Trans-pQTLs are located more
than 1 Mb away from the gene encoding the affected protein and may be on a different chromosome altogether (chromosome B above).
Trans-pQTLs are assumed to regulate the protein being studied indirectly and can reveal new mechanisms of how Gene B could be related
functionally to Gene A.

5
pQTLs uncover novel drug
target candidates in the drug
development process
pQTLs are at the heart of proteogenomic studies
Proteins are the main targets for most on-market drugs today. Exploring the blood proteome with
high-throughput proteomics technologies has been of great value to scientists in the drug discovery
and development field. By combining genomics and proteomics through the study of pQTLs, research
conducted by the SCALLOP consortium not only advances our understanding of diseases such as
cardiovascular disease but also uncovers causal proteins in disease which may serve as novel drug
targets for future medicines.

The SCALLOP consortium was begun in 2017 by Pfizer’s Dr. Anders Mälarstig with the aim to bring
together researchers with an interest in proteogenomics to explore opportunities that collaboration
and combining their datasets could offer. The consortium now has 35 principal investigators from
all over the world and Olink proteomic data from over 45 different human cohorts, representing
over 65,000 samples. The resulting statistical power from such large amounts of protein data has
led to amazing findings in disease research, as evidenced by a landmark paper published in Nature
Metabolism in 2020 titled ‘Genomic and drug target evaluation of 90 cardiovascular proteins in 30,931
individuals.’ This study exemplifies the value of pQTLs in drug development, using a pQTL-based
framework to map protein regulatory pathways, identify new drug targets, repurpose known drugs,
and stratify patients according to treatment response.

pQTLs help us understand the relationship between genes, proteins, and disease
The main advantage of using pQTLs in drug development studies is that they help researchers
understand the interplay between genes, proteins, and disease. In this case, researchers used pQTLs
to establish which proteins play a causal role in disease and may therefore be promising drug targets.
This is done using pQTLs and a method called Mendelian randomization. Mendelian randomization is
a technique that uses genetic variants that lead to pQTLs, as a natural experiment to investigate the
causal relationship between certain risk factors and disease outcomes in observational data. Cis-pQTLs
are particularly powerful here, as their random allocation at conception as well as their direct influence
on protein levels limit the effects of any confounding factors like health-related behaviors or post-
translational protein modifications. As such, Mendelian randomization is less likely to be affected by
confounding or reverse causation than observational studies.

Publication highlight
Folkersen L et al. Genomic and drug target evaluation of 90 cardiovascular proteins in 30,931 individuals.
Nature Metabolism (2020).

Read publication

6
By comparing the effect of that genetic variant on disease against its
effect on protein levels, researchers can determine whether altered protein
levels cause, or are the consequence of, the disease. In her talk as part of a
webinar on SCALLOP research titled ‘Driving Next-Generation Medicine
Discovery with Proteogenomics’, Erin Macdonald-Dunlop (University of
Edinburgh) describes Mendelian randomization as part of her research
in investigating the relationship between plasma proteins and different
diseases (Webinar snippet 1.1). Her study involved a genome-wide meta-
analysis of 184 proteins from the Olink® Target Cardiovascular I and II
panels to collect pQTL data, and Mendelian randomization to find evidence
for causal relationships between these proteins and disease.

The study found strong evidence for causal associations for 24 proteins,
14 of which are already targets for drugs in various stages of development.
However, this research may be used to investigate whether these drugs Webinar 1
target the disease for which they found evidence for a causal relationship;
in other cases the evidence may point to potential opportunities for drug Erin Macdonald-Dunlop, M. Sc.
repositioning and repurposing. The other 10 proteins were not current is the lead analyst for the SCALLOP
Consortium’s large-scale, genome-
drug targets and may be further investigated as potential drug target
wide meta-analysis of 184
candidates for the indications of interest. cardiovascular-related plasma proteins.
Here she talks about how to drive
Mendelian randomization was also carried out across 35 complex diseases Driving next-generation medicine
for the 90 cardiovascular proteins of the Nature Metabolism study to find discovery with proteogenomics.
causal links between proteins and disease. The authors then compared this
data with different drugs on the market or still under development, which Watch the webinar
raised some concerns regarding drug safety.

An example that Dr. Mälarstig discusses in the webinar on SCALLOP

research is the relationship between the protein CD40, rheumatoid
arthritis, and stroke. The study found evidence for a positive causal
relationship between CD40 and rheumatoid arthritis, but the opposite
between CD40 levels and stroke. Therefore, a CD40 inhibitor drug that
decreases the effects of rheumatoid arthritis may also increase the risk
of stroke, raising concerns about the safety of such drugs. Dr. Mälarstig
also discusses another example in a clip (Webinar snippet 1.2) from the
same webinar.

Webinar 1
Dr. Anders Mälarstig, Director of
Target sciences at Pfizer Worldwide
Research & Development and
Affiliate Researcher at Karolinska
Institute, talks about how
proteogenomics can improve drug
discovery and development.

Watch the webinar

7
pQTLs connect layers of biological information to understand disease
The value of protein quantitative trait loci is especially beneficial in the study of complex diseases.
The previous research examples combined proteomics with genomics to do this, but how much
more may be understood by adding other omics data?

A study resulting in a paper published in 2022 in Journal of Crohn’s and Colitis titled The Effect of
Phenotype and Genotype on the Plasma Proteome in Patients with Inflammatory Bowel Disease,
is the largest pQTL study to date using high-resolution genotype data to understand inflammatory
bowel disease (IBD). IBD is a collective term for complex immune-mediated diseases of the GI
tract, is difficult to treat, and includes Crohn’s disease and ulcerative colitis. Its etiology is not
well understood, but involves a complex interplay between host genes, a dysregulated immune
response, environmental factors, and the gut microbiome. In this study, researchers used the Olink®
Target 96 Inflammation protein panel to gather protein data for 1028 patients with IBD as well
as 148 healthy controls. They combined the protein data with extensive genotype data, RNA-seq
data from intestinal biopsies, as well as metagenomic data assessing the gut microbiome in fecal
samples, and their analysis of pQTLs connected these layers of biological data together.

From the protein data alone, the researchers identified unique plasma protein profiles for both
Crohn’s disease and ulcerative colitis, and overall, 32 proteins were significantly different between
the IBD cohort and healthy controls after adjustment for age, sex, and BMI. Overlapping this data
with genetic data revealed 23 cis-pQTLs that were associated with the levels of 21 proteins, 13 of
which were novel.

However, integration of these results with RNA-seq data from intestinal biopsies revealed how the
cis-pQTLs affected protein levels. The authors discovered that some of the cis-pQTLs were also
expression quantitative trait loci (eQTLs) for relevant proteins, such as CCL25, where the cis-pQTL
variant rs3745387 was also an eQTL for CCL25 gene expression. The CCL25 protein is connected
to immune regulation, which is compromised in patients with IBD. Understanding the genotype
association with plasma protein levels of CCL25 could aid in functional studies.

By integrating multiple omics data, the authors uncovered a complex interplay between host
genotype and the plasma proteome, uncovering a variety of inflammatory pathways that define
IBD. This may serve as a basis for future research to disentangle these pathways to give further
insight into the disease. The identified pQTLs may aid clinicians in patient stratification, drug
selection and validation and drug repurposing, to help them find therapies and treatments for
this disease that so far has remained elusive.

Publication highlight
Bourgonje AR et al. The Effect of Phenotype and Genotype on the Plasma Proteome in Patients with
Inflammatory Bowel Disease. J Crohn’s Colitis (2022).÷

Read publication

8
How proteomics enriches the
multiomic study of disease
A deeper understanding of the biology underlying pathophysiological processes is essential in any
efforts to better predict, prevent, manage, or treat disease. It is particularly important for more
effective and personalized medicine. Diseases frequently represent multi-layered changes in complex
underlying biology, which may itself be only partially understood at best. Pathophysiology may result
from a combination of genetic and environmental factors that trigger multiple interactions between
cells and biomolecules, with subsequent changes that occur both locally and systemically.

The development and combination of large-scale omics methodologies are critical to these efforts.
Proteomics, genomics, transcriptomics, epigenomics, metabolomics, lipidomics, mass cytometry etc.
are increasingly combined in comprehensive “systems biology” studies, with proteomics now taking
an ever more central role. Combining different omics datasets answer questions that cannot be fully
addressed by using one approach alone. Genetics can help define and quantify disease risk and
enables stratification of targeted therapies for some specific cancer mutations, but sequencing data is
usually unable to provide dynamic information on disease progression and outcomes. Transcriptomics
provides a valuable proxy for gene action, but RNA levels often correlate quite poorly (or even anti-
correlate) with protein levels, especially systemically. Proteins provide the closest and most dynamic
link to complex phenotypes and constitute almost all drug targets but may be difficult to analyze at
scale and may require additional data to establish causality.

9
In this chapter, we will look at several recent examples where proteomics has made crucial
contributions in multiomic studies in several key disease areas:

• Early Detection and Treatment of Cancer – A combination of proteomics and transcriptomics has
been used to determine biomarkers of lung cancer years before conventional diagnosis.

• Cancer Immunotherapy – Baseline predictive plasma protein biomarkers combined with the
presence of tertiary lymphoid structures were reported as a powerful predictor of response to
immunotherapy among patients with a variety of cancers.

• COVID-19 – Proteomics, transcriptomics, mass cytometry and other omics have been used in systems
biology approaches to address COVID-19 disease severity, comorbidities, and immune response.

• Inflammatory Bowel Disease – A study from the Crohn’s & Colitis Foundation determined
prognostic biomarker signatures to predict the future onset of complications as well as treatment
response in juvenile Crohn’s Disease, using tissue-specific RNA-seq and plasma proteomics in
combination with machine learning data analysis.

Proteomics as part of a multiomics

approach in the early detection and
treatment of cancer
A better understanding of cancer biology along with advances in screening and targeted and immune
therapies are transforming the ways in which we prevent, diagnose, treat, and ultimately survive
cancer. Development of personalized targeted therapies and immunotherapy approaches have made
major strides in the past decade; however further developments in new technologies and multiomic
strategies will be needed for actionable insights into cancer biology and improved targeted treatments.

Advances in RNA sequencing technologies, DNA methylation profiling and high-plex proteomics are
now being used to enable multiomic studies that can improve our understanding of early malignant
transformation and therapeutic outcomes. Here we describe key recent findings and provide new insights
generated by the high-plex proteomic analyses in early cancer detection and cancer immunotherapy.

Publication highlight
Dagnino S et al. Prospective Identification of Elevated Circulating CDCP1 in Patients Years before Onset
of Lung Cancer. Cancer Research (2021).

Read publication

10
The potential exists for circulatory plasma protein biomarkers to enable early cancer detection.
However, due to the lack of currently available tests, ~50% of cancers are diagnosed only after
diagnosis of symptoms that indicate an advanced stage of disease, when treatment options are
limited, and prognosis is poor. While ctDNA has been extensively researched and is currently under
development as a liquid biopsy biomarker for cancer detection, there is a lack of both predictability in
its utility and an understanding of the pathophysiology of ctDNA release. Not surprisingly, there is a
trend back toward examining protein biomarkers for early detection diagnostic test development.

Previous studies have indicated that protein biomarkers in combination with ctDNA can enhance
the accuracy of cancer detection. In the study highlighted below, Marc Chadeau-Hyam’s group
from Imperial College London used a combination of proteomics and transcriptomics to identify
predictive biomarkers for lung cancer years ahead of clinical diagnosis, as well as provide deeper
pathophysiological insights into the disease.

Analyzing plasma proteomics with an Olink panel for 92 inflammation-related proteins in 325 female
lung cancer patients and 325 healthy controls from two prospective cohorts identified 12 proteins
(CDCP1, HGF, IL6, OSM, MCP1, IL8, VEGFA, CD6, CD5, SCF, TWEAK & IL12B) associated with increased
incidence of lung cancer. After statistical adjustment for smoking history, CDCP1 (a protein involved
in CD6 T-cell migration and immune function) remained as a smoking-independent predictive risk
marker for development of lung cancer. These findings were then confirmed using 450 additional
subjects from an independent validation cohort. CDCP1 therefore shows promise as an early predictive
biomarker for lung cancer since the levels of this protein were high even in individuals who did not
smoke, well before cancer diagnosis.

The team then combined the CDCP1 protein data with RNA-seq analysis of over 11 000 transcripts
and identified significant associations between CDCP1 levels and the expression of LRRN3 and SEM1
mRNA. Principal Component Analysis (PCA) of reactome pathway associations indicated that CDCP1
may be involved in molecular pathways related to WNT/ß-catenin signaling, a canonical signaling
pathway associated with carcinogenesis.

The integration of proteomic and transcriptomic data has potential to improve early diagnosis of
lung cancer by enabling an in-depth pathway analyses of underlying malignant transformation
mechanisms. Moreover, an effective pre-symptomatic, population-wide early detection strategy could
shift the burden of disease to an earlier stage, where treatment may be more effective or potentially
curative. A more detailed study synopsis is available in this blog post.

Visit the Olink to Science blog

This Olink to Science blog post includes a detailed study synopsis of the S. Dagnino et al. Prospective
Identification of Elevated Circulating CDCP1 in Patients Years before Onset of Lung Cancer study.

Read the blog post

11
Proteomics as part of a multiomics
approach in cancer immunotherapy
Immune checkpoint blockade (ICB) treatment has revolutionized the way we approach cancer and
continues transforming the practice of medical oncology across different tumor types for a small but
significant proportion of patients. The growing need to better predict which patients will benefit from
ICB treatment is driving the demand for minimally invasive predictive biomarkers. Such research will
continue to drive the future of precision immunotherapy, an example of which is presented below.

Antoine Italiano and colleagues from INSERM used the Olink® Explore platform to interrogate ~1500
plasma proteins in two independent prospective cohorts of cancer patients treated with immune
checkpoint inhibitors. Correlations between protein plasma levels, clinical benefit, progression-
free survival, and overall survival were then examined, showing that elevated baseline serum level
of leukemia inhibitory factor (LIF) was associated with a poor clinical outcome in patients. This
association was independent of any other prognostic factors (such as PD-L1 expression or tumor
mutational burden) and was replicated in two additional cohorts.

The relevance of LIF in the tumor microenvironment was then explored, using RNA-seq to profile the
immune cell composition in patients with high or low plasma levels of the LIF protein. LIF levels were
shown to be inversely correlated with both B cells and helper T cells, which are major components
of tertiary lymphoid structures (TLS). The presence of TLS features has been linked to improved
responses in cancer immunotherapy. Multiplex immunohistochemistry of tumor biopsies confirmed
that the presence of TLS was significantly more frequent in patients with low levels of plasma LIF, in
line with the behavior of LIF as a biomarker for poorer prognosis in cancer patients being treated with
immunotherapy.

Recently Dr. Italiano gave a presentation (available here) discussing the results of this important work.

“ “In the wake of genomics, the study of proteins is now emerging as the new frontier
for understanding real-time human biology”.
Loriot et al. Annals of Oncology (2021)

Publication highlight Publication highlight

Loriot Y et al. Plasma proteomics identifies leukemia inhibitory Arunachalam P et al. Systems biological assessment of
factor (LIF) as a novel predictive biomarker of immune- immunity to mild versus severe COVID-19 infection in
checkpoint blockade resistance. Annals of Oncology (2021). humans. Science (2020).

Read publication Read publication

12
Addressing the COVID-19 pandemic
through multiomics
The global pandemic caused by the rapid spread of the SARS-CoV-2 coronavirus has had an enormous
impact on global health and society, while presenting unprecedented challenges to both the scientific
community and the pharmaceutical industry. It became clear early on that patient immune responses
were critical in determining the broad spectrum of disease severity and clinical outcomes observed,
requiring comprehensive strategies to map the complexity of the immune system. Here we highlight
two outstanding COVID-19 studies that demonstrate the value of a multiomics strategy with proteins
as a central focus.

Prabhu Arunachalam and colleagues from Bali Pulendran’s group at Stanford University undertook
a comprehensive multiomics study of immune responses in 76 COVID-19 patients and 69 healthy
individuals, using a combination of single-cell transcriptomics, plasma proteomics and mass cytometry.
Thus, cellular and molecular immune system responses could be assessed at both a local and a
systemic level. Results revealed a spatial dichotomy in the innate immune response, with suppression
of peripheral innate immunity in the face of a pro-inflammatory response in the lung, which develops
into a systemic pro-inflammatory response. Using a multiplexed Olink panel to assess 92 inflammation
biomarkers in circulation, this work identified EN-RAGE, OSM and TNFSF14 as potential protein
biomarkers associated with disease severity.

This group also produced a multiomics study of immune responses to the anti-SARS-CoV-2 mRNA
vaccine from Pfizer (Arunachalam P, Scott M, Hagan T, et al. Systems vaccinology of the BNT162b2
mRNA vaccine in humans. (2021) Nature, DOI: 10.1038/s41586-021-03791-x).

Figure 2 Illustrative summary of the main findings from Arunachalam et al. The process on the left represents the innate immune response
in patients with mild COVID-19, while the process on the right represents the innate immune response in patients with severe COVID-19.
ISGs = IFN-stimulated genes, pDCs = Plasmacytoid dendritic cells. Image courtesy of Dr. Prabhu Arunachalam.

13
Dr. Arunachalam explains why a multiomics approach is so important to
understand complex immune system responses, in the short video clip
(Webinar snippet 2.1).

“ ”Using a multiomics approach is essential to provide

a comprehensive picture of immune response, not
just to this vaccine, but to any vaccine”.
Dr. Prabhu,Arunachalam, Stanford University

A team from Dr. Petter Brodin’s group at the Karolinska Institute used a
combination of autoantibody screening, flow cytometry and Olink-based
plasma proteomics to gain a systems biology level understanding of a rare Webinar 2
but severe complication in children with COVID-19.
Dr. Prabhu Arunachalam, Senior
Multisystem Inflammatory Syndrome in Children (MIS-C) presents as high research scientist at the Institute for
fever, organ dysfunction and strongly elevated levels of inflammatory Immunity, Transplantation, and
Infection Operations at Stanford
proteins, and exhibits several overlapping features with Kawasaki Disease
University, explains why a multiomics
(an autoimmune condition accompanied by vascular inflammation of approach is so important to understand
small- to medium-sized arteries in young children). By taking a multiomics complex immune system responses.
approach to study MIS-C, they were able to show that the inflammatory
response in MIS-C differs from that seen in severe acute COVID-19 in adults Watch the webinar
and shares several features with Kawasaki Disease. Crucially, however,
they also found important differences between MIS-C and Kawasaki
Disease, particularly with respect to the T-cell subsets involved, expression
of IL-17A and biomarkers associated with arterial damage. These findings
have significant implications for the understanding and possible treatment
strategies for the two diseases. Additionally, while treatments targeting
TNF-alpha and IL-6 have been suggested for severe acute COVID-19 in
adults, neither of these proteins were seen to be significantly elevated in
children with MIS-C.

More details of this study and an additional investigation into severe adult
COVID-19 can be found by viewing a webinar presented by Dr. Brodin
(Webinar snippet 3.1).

Webinar 3
Dr. Petter Brodin, Professor in
Paediatric Immunology at
Karolinska Institute, talks about
their multiomics study on the rare
but severe complication MIS-C in
Publication highlight children with COVID-19 and an
additional investigation into
Consiglio CR et al. The Immunology of Multisystem Inflammatory Syndrome
servere adult COVID-19.
in Children with COVID-19. Cell (2020).

Read publication Watch the webinar

14
Prognostic and predictive
biomarkers for complications
in juvenile Crohn’s disease
The Crohn’s & Colitis Foundation have recently run a major study to find
prognostic markers that can determine if children with the autoimmune
inflammatory bowel condition, Crohn’s disease, will develop serious
complications such as fibrosis and/or fistulas, and predictive markers that
foretell whether they are likely to respond to more aggressive therapy with
anti-TNFα biologics. In a large-scale project, they analyzed RNA-seq data from Webinar 4
mucosal biopsy samples taken via colonoscopy and used machine learning
algorithms to identify a high-performance, prognostic 13-gene expression Dr. Andres Hurtado-Lorenzo, VP
Translational Research at the Crohn’s
signature for Crohn’s complications. The information of key genes active locally
& Colitis Foundation, describes
at the site of disease also provided valuable pathophysiological insights and the results of the study where they
potential future drug targets. tried to find prognostic markers
that could determine if children
The sampling method is highly invasive, however, and does not lend itself to with Crohn’s disease would develop
routine clinical use with paediatric patients. To address this issue, they also serious complications.
measured ~1000 proteins in the plasma of these patients using Olink, again
Watch the webinar
applying machine learning, to identify a 14-protein prognostic signature that
predicts those individuals who will go on to develop fibrosis/fistulas with an
accuracy of 84%, as well as a 3-protein signature that predicts responders to
anti-TNFα therapy with an accuracy of 90%. These data provided the basis
to develop a custom panel comprising the proteins identified during the
discovery phase, which was subsequently tested and verified and offers a
potential prognostic test for future management of paediatric Crohn’s disease.

In Webinar snippet 4.1, Dr. Hurtado-Lorenzo describes the results of the

study in more detail.

“ ”I am glad to report that we were able to identify a plasma

protein signature that can predict at diagnosis, the likelihood
of a child with Crohn’s disease to develop complications
within 5 years”.
Dr. Andrés Hurtado-Lorenzo, Ph.D. (VP, Translational Research,
Crohn’s & Colitis Foundation)

15
The future of proteomics
in multiomics research
With the implementation of high-throughput proteomics technologies like the Olink Proximity
Extension Assay (PEA) platform, proteomics is fast becoming a core omic in multiomics studies. Studies
incorporating protein biomarkers to monitor real-time biology enrich genomics and single-cell insights
to rapidly advance our knowledge of human health and disease and to drive development of next
generation therapies.
What will the future hold for multiomics strategies with proteomics at its center? Below we share some
key trends that are already becoming apparent in multiomics research and perspectives from industry
experts in both pharmaceutical and academic context on what can be expected over the next decade.

16
Key trends and future
perspectives
Proteomics at population scale will lead to breakthroughs
in disease research
With continued development of high-throughput proteomics technologies,
researchers will combine protein data with other omics in ever larger sample
cohorts, with clear repercussions on drug development. Dr. Cindy Lawley
(Global Director for Population Health Olink Proteomics) highlights a few key
projects using the Olink platform:

“
Webinar 5
”This approach is being deployed by the United Kingdom’s UK Biobank
Pharma Proteomics Project (UKB-PPP), a pre-competitive collaboration of Dr. Claudia Langenberg, professor
over 10 biopharmaceutical companies to build a framework and systematic of Computational Medicine in
approach to therapeutic target identification. They are confident of the results the Berlin Institute of Health at
because they have seen a powerful precursor to the UKBB-PPP in the SCALLOP Charité and MRC Investigator and
(Systematic and Combined AnaLysis of Olink Proteins) consortium study. In Programme Leader at the MRC
a milestone paper, Folkersen et al (2020) deployed such an approach in over Epidemiology Unit at the University
30,000 samples with genetic data, clinical data for 38 diseases and protein data of Cambridge, talks about exciting
for 90 proteins. They identified over 450 pQTLs (protein Quantitative Trait Loci) developments in the multiomics field.
with 25 of these suggesting causality through Mendelian Randomization. Of
the 25 causal markers, 14 of these were recapitulating others’ findings, 11 were Watch the webinar
novel and another 18 protein-disease relationships suggested repurposing
opportunities with existing drugs for new disease indications”.

Dr. Cindy Lawley (Global Director for Population

Health Olink Proteomics)

Dr. Claudia Langenberg, a professor of Computational Medicine in the Berlin

Institute of Health at Charité and MRC Investigator and Programme Leader
at the MRC Epidemiology Unit at the University of Cambridge, sees much
potential in the use of blood biomarker studies and broad capture proteomics
in large populations to give us more mechanistic insights into unclassified
and complex diseases. Listen to her talk about these exciting developments in
the multiomics field in Webinar snippet 5.1.

Adding protein data to multiomics strategies will change

the way we classify disease
Results from protein biomarker research combined with genomics and often
transcriptomics are changing the way we study disease. In a pharmaceutical Webinar 6
roundtable discussion three pharmaceutical research scientists discussed the
opportunities and challenges in using biomarker data in drug discovery and Dr. Timothy Radstake MD, Ph.D,
development. Dr. Timothy Radstake, MD, Ph.D. (Executive Director Immunology Executive Director Immunology
Discovery, Head of Transformational and Translational Immunology Discovery, Discovery, Head of Transformational
and Translational Immunology
AbbVie) emphasized how drug discovery based upon a diagnostic approach is
Discovery, AbbVie talks about
wrong. Biomarker research shows that in fact, different diseases exhibit shared how drug discovery based upon a
disease drivers and biological pathways. This will lead to a reclassification diagnostic approach is wrong.
of medicine, where new types of clinical trials may be conducted based on
biomarkers and protein profiles, rather than by disease. Listen to Dr. Timothy Watch the webinar
Radstake’s perspective in Webinar snippet 6.1.

17
Prof. Langenberg has already found evidence of this “shared-ness across
diseases” in her research combining genetics and proteomics. In a roundtable
discussion on how proteomics can complement genomics, Prof. Langenberg
describes how protein data have enhanced their ability to connect different
diseases (Webinar snippet 5.2).

Her perspective was echoed by another roundtable panelist, Dr. Janne Lehtiö,
Professor of Clinical Proteomics (Karolinska Institute, Stockholm, Sweden),
who describes how the use of proteomics in clinical trials is changing their
perspective on how to treat complex disease (Webinar snippet 5.3).

One inevitable consequence of this growing trend in increasingly large-scale,

multiomic studies is a logarithmic growth in the volume and complexity of
the data being generated. Webinar 5
Dr. Claudia Langenberg, professor
of Computational Medicine in
the Berlin Institute of Health at
Protein data visualization tools help integrate protein data Charité and MRC Investigator and
Programme Leader at the MRC
into multiomics datasets Epidemiology Unit at the University
As the biomedical research community continues to strive for precision of Cambridge, describes how protein
medicine and personalized therapy, proteomics will become ever more data have enhanced their ability to
connect different diseases.
integral to reaching that goal. For proteomics data to become even more
informative and useful in multiomics strategies, we must reach a higher
Watch the webinar
resolution in our knowledge of the human proteome. This will involve
new ways of analyzing protein data and the use of open-source protein
databases to further our understanding of protein pathways.

Efforts such as the Human Protein Atlas and the Reactome Project aim to
support proteomics researchers by organizing the ever growing wealth
of human circulating biomarker data into protein maps and biological
pathways, as well as offering tools to enable more effective analysis of
protein data. These initiatives reflect ongoing trends in multiomics research
analysis that will only continue to grow in the future.

The Human Protein Atlas is an open-access resource combining data from

protein and RNA expression to map all human proteins across organs,
tissues, and cells. The latest version of the atlas (released in November
2021) incorporates single cell transcriptomics data to increase the resolution
of the protein map in different types of tissues such as blood and brain
cells. As a result, they have introduced four new sections to the atlas which
includes proteins from single cells and specific cell lines. Combining these
different omics is no small feat, and the consortium continues to make use Webinar 5
of bioinformatics tools such as machine learning and AI to produce updated
versions of the protein atlas. Dr. Janne Lehtiö, Professor of Clinical
Proteomics (Karolinska Institute,
Stockholm, Sweden) describes how
the use of proteomics in clinical trials
is changing their perspective on how
to treat complex disease.

Watch the webinar

18
Listen to Podcast snippet 2 by Prof. Mattias Uhlén (Head of the Division of
Systems Biology at the KTH Royal Institute of Technology Sweden and head of
the Protein Atlas consortium) discuss the Human Protein Atlas and their
plans on how to develop the atlas in the future to support proteomics studies.

“ “I think more effort should be made in visualizing the [protein

data] results better. We have gone beyond massive Excel
spreadsheets that we could sit at and separate. Now we need
to create innovative ways of integrating the different biological
domains and bring them to our results from proteogenomics.“
Dr. Claudia Langenberg, Computational Medicine, Berlin Institute
of Health, Charité.
Podcast 2
Such innovation will also enable more effective analysis of protein and multiomics Dr. Mathias Uhlén, Head of the
data, exemplified through the development of better bioinformatics tools by Division of Systems Biology at the
Olink. The Reactome project maps the human proteome into biological pathways, KTH Royal Institute of Technology
as well as develops tools to handle large protein data sets. This biological Sweden and head of the Protein
pathway map is featured in Olink’s new cloud-based bioinformatics tool, Atlas consortium discusses the
Human Protein Atlas and their plans
Olink® Insight Beta, a platform for protein biomarker data discovery. In
on how to develop the atlas in the
addition to visualizing Reactome pathway connections from lists of your future to support proteomics studies.
selected proteins or genes, the platform also allows you to search, save and
share relevant information from millions of online publication abstracts
and explore our interactive Data Stories, including hands-on analysis of a Listen to the podcast
COVID-19 data set and a study of the variability of ~3000 proteins in the
”normal” adult population (available here). See Figure 3 below.

Figure 3 A snapshot of the pathway browser tool in Olink® Insight, which is based upon the Reactome project. Each cluster represents
pathways involved in different biological systems, with nodes representing individual pathways. The colors of each pathway represent the
Olink® panels that may be used to investigate each pathway.

19
The challenge of big data
Beyond the development of data visualization tools there remains much work to be done to more
effectively integrate data from multiple omics to generate more actionable insights. The era of ‘big
data’ is well and truly upon us, and this greatly impacts the multiomics research space.

“ “A general challenge for those working in this field is that comprehensive drug analysis
also creates big data, so we need to have proper data mining tools in place which is a
challenge for everyone. Multiomics is in the field, and all around us, so we need to look
at this challenge.“
Dr. Jurgen Braunger - Head of Cellular and Protein Biomarkers Boehringer Ingelheim.

“ “This is where the bottleneck is at the moment, as we’re gathering so much data
in terms of different layers, time points, different sample types [serum vs tissues].
The integration of all that data is really challenging and is the limiting step in truly
understanding mechanistic biomarkers and identifying phenotypes in patient disease.“
Dr. Timothy Radstake, Executive Director Immunology Discovery, Distinguished Research
Fellow Head Transformational and Translational Immunology Discovery, AbbVie

We conclude this chapter with some thoughts by Prof. Jochen Schwenk (Professor of Translational
Proteomics KTH Royal Institute of Technology).

“ “The future utility of capturing the components of the blood proteome will continue to
rely on combining proteome insights with other types of data. Starting from the list of
proteins that can be measured by the current technologies, the ultimate goal to analyze
even more, if not all 20,000 protein-encoding genes has remained unchanged. But
efforts to measure all human proteins in every sample pose a challenge for proteomics
technologies, and moving forward, there will be a continuous need to improve the
limits for detecting the circulating proteins at the lowest levels. We must also continue
to improve our understanding of how protein modifications and interactions influence
the assembly and analyses of the circulating proteome. When combining proteomics
and genomics, more data, preferably from multiple technology platforms might be
required to fully understand how genetic variation influences the circulating proteome.
In the future, prospective longitudinal investigations with complete phenotype
information can provide an important opportunity for proteomics in health and in the
development of disease. It is through such studies that proteomics can contribute to
a better understanding of intermediate phenotypes, can offer time-resolved insights
into health or disease states, and thereby transform single snapshots of data into
projections of human health.”
Dr. Jochen Schwenk, Professor of Translational Proteomics
at KTH Royal Institute of Technology

20
This challenge is compounded by just how different these omics data sets
are from one another. Dr. Janne Lehtiö explains how multiomics studies
utilize data from systems that are regulated differently, have different
dynamic ranges, and temporal/spatial regulation, and calls for a better way
to marry biological understanding with bioinformatics to make sense of
multiomics data. Listen to his explanation in Webinar snippet 5.4.

The incorporation of high-throughput proteomics technologies into

multiomics studies provides a clear and comprehensive picture of real-
time biology that will translate breakthrough findings in the lab to the
implementation of precision medicine in the clinic. Developments in
technologies and data science will continue to enable fully integrated
multiomic studies at an ever-increasing scale, deepening our understanding
of complex disease biology and driving future healthcare.
Webinar 5
Dr. Janne Lehtiö, Professor of
Clinical Proteomics (Karolinska
Institute, Stockholm, Sweden)
discusses ways to marry biological
understanding with bioinformatics
to make sense of multiomics data.

Watch the webinar

We would like to thank all the experts that contributed to this ebook:

Dr. Anders Mälarstig, Karolinska Institute/Pfizer
Dr. Mathias Uhlén, KTH Royal Institute of Technology
Erin McDonald-Dunlop, University of Edinburgh
Dr. Andres Hurtado-Lorenzo, Crohn's & Colitis Foundation
Dr. Prabhu Arunachala, Stanford University
Prof. Petter Brodin, Karolinska Institute
Prof. Claudia Langenberg, Berlin Institute of Health
Prof. Janne Lehtiö, Karolinska Institute
Dr. Timothy Radstake, AbbVie
Prof. Jochen Schwenk, KTH Royal Institute of Technology
Dr. Marijana Rucevic, Olink Proteomics
Dr. Cindy Lawley, Olink Proteomics

21
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