Newly combined digital pathology, artificial intelligence (AI), and omics technologies are providing anatomic pathologists and medical laboratory scientists with powerful diagnostic tools
Add “spatial transcriptomics” to the growing list of “omics” that have the potential to deliver biomarkers which can be used for earlier and more accurate diagnoses of diseases and health conditions. As with other types of omics, spatial transcriptomics might be a new tool for surgical pathologists once further studies support its use in clinical care.
Among this spectrum of omics is spatial transcriptomics, or ST for short.
Spatial Transcriptomics is a groundbreaking and powerful molecular profiling method used to measure all gene activity within a tissue sample. The technology is already leading to discoveries that are helping researchers gain valuable information about neurological diseases and breast cancer.
Marriage of Genetic Imaging and Sequencing
Spatial transcriptomics is a term used to describe a variety of methods designed to assign cell types that have been isolated and identified by messenger RNA (mRNA), to their locations in a histological section. The technology can determine subcellular localization of mRNA molecules and can quantify gene expression within anatomic pathology samples.
In “Spatial: The Next Omics Frontier,” Genetic Engineering and Biotechnology News (GEN) wrote, “Spatial transcriptomics gives a rich, spatial context to gene expression. By marrying imaging and sequencing, spatial transcriptomics can map where particular transcripts exist on the tissue, indicating where particular genes are expressed.”
In an interview with Technology Networks, George Emanuel, PhD, co-founder of life-science genomics company Vizgen, said, “Spatial transcriptomic profiling provides the genomic information of single cells as they are intricately spatially organized within their native tissue environment.
“With techniques such as single-cell sequencing, researchers can learn about cell type composition; however, these techniques isolate individual cells in droplets and do not preserve the tissue structure that is a fundamental component of every biological organism,” he added.
“Direct spatial profiling the cellular composition of the tissue allows you to better understand why certain cell types are observed there and how variations in cell state might be a consequence of the unique microenvironment within the tissue,” he continued. “In this way, spatial transcriptomics allows us to measure the complexity of biological systems along the axes that are most relevant to their function.”
“Although spatial genomics is a nascent field, we are already seeing broad interest among the community and excitement across a range of questions, all the way from plant biology to improving our understanding of the complex interactions of the tumor microenvironment,” George Emanuel, PhD (above), told Technology Networks. Oncologists, anatomic pathologists, and medical laboratory scientists my soon see diagnostics that take advantage of spatial genomics technologies. (Photo copyright: Vizgen.)
According to 10x Genomics, “spatial transcriptomics utilizes spotted arrays of specialized mRNA-capturing probes on the surface of glass slides. Each spot contains capture probes with a spatial barcode unique to that spot.
“When tissue is attached to the slide, the capture probes bind RNA from the adjacent point in the tissue. A reverse transcription reaction, while the tissue is still in place, generates a cDNA [complementary DNA] library that incorporates the spatial barcodes and preserves spatial information.
“Each spot contains approximately 200 million capture probes and all of the probes in an individual spot share a barcode that is specific to that spot.”
“The highly multiplexed transcriptomic readout reveals the complexity that arises from the very large number of genes in the genome, while high spatial resolution captures the exact locations where each transcript is being expressed,” Emanuel told Technology Networks.
Spatial Transcriptomics for Breast Cancer and Neurological Diagnostics
In that paper, the authors wrote “we envision that in the coming years we will see simplification, further standardization, and reduced pricing for the ST protocol leading to extensive ST sequencing of samples of various cancer types.”
Spatial transcriptomics is also being used to research neurological conditions and neurodegenerative diseases. ST has been proven as an effective tool to hunt for marker genes for these conditions as well as help medical professionals study drug therapies for the brain.
“You can actually map out where the target is in the brain, for example, and not only the approximate location inside the organ, but also in what type of cells,” Malte Kühnemund, PhD, Director of Research and Development at 10x Genomics, told Labiotech.eu. “You actually now know what type of cells you are targeting. That’s completely new information for them and it might help them to understand side effects and so on.”
The field of spatial transcriptomics is rapidly moving and changing as it branches out into more areas of healthcare. New discoveries within ST methodologies are making it possible to combine it with other technologies, such as Artificial Intelligence (AI), which could lead to powerful new ways oncologists and anatomic pathologists diagnose disease.
“I think it’s going to be tricky for pathologists to look at that data,” Kühnemund said. “I think this will go hand in hand with the digital pathology revolution where computers are doing the analysis and they spit out an answer. That’s a lot more precise than what any doctor could possibly do.”
Spatial transcriptomics certainly is a new and innovative way to look at tissue biology. However, the technology is still in its early stages and more research is needed to validate its development and results.
Nevertheless, this is an opportunity for companies developing artificial intelligence tools for analyzing digital pathology images to investigate how their AI technologies might be used with spatial transcriptomics to give anatomic pathologists a new and useful diagnostic tool.
Proteins in human saliva make up its proteome and may be the key to new, precision medicine diagnostics that would give clinical pathologists new capabilities to identify disease
Clinical pathologists may soon have an array of new precision medicine diagnostic tools based on peoples’ saliva. There are an increasing number of “–omes” that can be the source of useful diagnostic biomarkers for developing clinical laboratory tests. The latest is the world’s first saliva protein biome wiki.
Called the Human Salivary Proteome Wiki (HSP Wiki), the “public data platform,” which was created by researchers at the University of Buffalo, is the “first of its kind,” according to Labroots, and “contains data on the many thousands of proteins present in saliva.”
The HSP Wiki brings together data from independent studies on proteins present in human saliva. One of the researchers’ goals is to speed up the development of saliva-based diagnostics and personalized medicine tools.
In “The Human Salivary Proteome Wiki: A Community-Driven Research Platform,” published in the Journal of Dental Research, the researchers wrote, “Saliva has become an attractive body fluid for on-site, remote, and real-time monitoring of oral and systemic health. At the same time, the scientific community needs a saliva-centered information platform that keeps pace with the rapid accumulation of new data and knowledge by annotating, refining, and updating the salivary proteome catalog.
“We developed the Human Salivary Proteome (HSP) Wiki as a public data platform for researching and retrieving custom-curated data and knowledge on the saliva proteome. … The HSP Wiki will pave the way for harnessing the full potential of the salivary proteome for diagnosis, risk prediction, therapy of oral and systemic diseases, and preparedness for emerging infectious diseases,” they concluded.
“This community-based data and knowledge base will pave the way to harness the full potential of the salivary proteome for diagnosis, risk prediction, and therapy for oral and systemic diseases, and increase preparedness for future emerging diseases and pandemics,” Stefan Ruhl, DDS, PhD (above right, with Omer Gokcumen, PhD, Associate Professor of Biological Sciences on left), Professor, Department of Oral Biology, University of Buffalo, and lead researcher of the study, told Labroots. Development of precision medicine clinical laboratory diagnostics is part of their research goals. (Photo copyright: University of Buffalo.)
Where Does Saliva Come From?
Saliva is a complex biological fluid that has long been linked to oral health and the health of the upper gastrointestinal tract. Only recently, though, have scientists begun to understand from where in the body saliva proteins originate.
The authors wrote: “Salivary proteins are essential for maintaining health in the oral cavity and proximal digestive tract, and they serve as potential diagnostic markers for monitoring human health and disease. However, their precise organ origins remain unclear.
“Through transcriptomic analysis of major adult and fetal salivary glands and integration with the saliva proteome, the blood plasma proteome, and transcriptomes of 28+ organs, we link human saliva proteins to their source, identify salivary-gland-specific genes, and uncover fetal- and adult-specific gene repertoires,” they added.
“Our results pave the way for future investigations into glandular biology and pathology, as well as saliva’s use as a diagnostic fluid,” the researchers concluded.
Saliva plays a crucial role in digestion by breaking down starches. It also provides a protective barrier in the mouth. When salivary glands malfunction, patients can face serious health consequences. Although clinicians and scientists have long understood the importance of saliva to good health, the question now is whether it contains markers of specific diseases.
“The Human Salivary Proteome Wiki contains proteomic, genomic, transcriptomic data, as well as data on the glycome, sugar molecules present on salivary glycoproteins. New data goes through an interdisciplinary team of curators, which ensures that all input data is accurate and scientifically sound,” noted Labroots.
The graphic above “shows the interconnectedness of the thousands of salivary proteins originating from blood plasma, parotid glands, and submandibular and sublingual glands. The diagram is one of many tools available to researchers and clinicians through the Human Salivary Proteome Wiki,” noted a UBNow blog post. (Graphic copyright: University of Buffalo.)
Omics and Their Role in Clinical Laboratory Diagnostics
Proteomics is just one of several hotly-researched -omics that hold the potential to develop into important personalized medicine and diagnostics tools for pathologists. Genomics is a related area of research being studied for its potential to benefit precision medicine diagnostics.
However, unlike genomes, which do not change, proteomes change constantly. That is one of the main reasons studying the human salivary proteome could lead to valuable diagnostics tools.
Combining the study of the -omes with tools like mass spectrometry, a new era of pathology may be evolving. “With the rapid decrease in the costs of omics technologies over the past few years, whole-proteome profiling from tissue slides has become more accessible to diagnostic labs as a means of characterization of global protein expression patterns to evaluate the pathophysiology of diseases,” noted Pathology News.
Saliva and the Age of Precision Medicine
The study of the -omes may be an important element in the evolution of precision medicine, because of its ability to provide information about what is happening in patients’ bodies at the point of care.
Thus, a full understanding of the proteome of saliva and what causes it to change in response to different health conditions and diseases could open the door to an entirely new branch of diagnostics and laboratory medicine. It is easy and non-invasive to gather and, given that saliva contains so much information, it offers an avenue of study that may improve patients’ lives.
It also would bring us closer to the age of precision medicine where clinical laboratory scientists and pathologists can contribute even more value to referring physicians and their patients.
Scientists
at St. Jude’s have discovered that performing different genetic tests on pediatric
cancer patients, and then combining those test results, may help guide and
improve patient care.
The research was part of a St. Jude’s project called Genomes
for Kids (G4K), a study to determine how genetic information may be used to
diagnose and treat pediatric cancers.
Through this project, the researchers hope to learn why tumors form in
children and predict how tumors will respond to certain treatments.
‘It’s
a Whole Lot of Sequencing.’
Few tragedies are worse than cancer in children. This is where precision medicine treatments can be critical, and multiomics may play an important role in the development of new therapies.
Multiomics refers to a biological analysis approach in which
multiple “omes” are analyzed together in a collaborative way to locate relevant
biomarkers and functional relationships. These “omes” include:
To perform their research, the St. Jude scientists examined
253 pediatric cancer patients by conducting whole genome
sequencing (WGS), whole
exome sequencing (WES), and RNA
sequencing of their tumors. They also looked at the WGS and WES of
non-cancerous tissues extracted from the same cancer patients.
“It is a whole lot of sequencing. I admit that,” Scott Newman, PhD,
Group Lead, Bioinformatics Analysis at St. Jude’s, told The
Scientist.
“With results available in a clinically relevant time frame, and pricing becoming increasingly comparable to the radiology and pathology tests, WGS is becoming more accessible to pediatric oncology patients,” said Scott Newman, PhD (above), Group Lead, Bioinformatics Analysis, at St. Jude’s, in an American Society of Human Genetics (ASHG) news release. (Photo copyright: ASHG.)
As a result of their three-platform testing, the researchers
discovered there was at least one finding for each patient that could be useful
in providing a diagnosis, revealing risks for individual patients, or
pinpointing which drugs may be most beneficial for a particular patient in
nearly 200 (79%) of the cases. Such findings are at the heart of precision
medicine.
The researchers also compared their sequencing results to
cancer panels that use next-generation
sequencing (NGS) to target specific genes or mutations relevant to a
certain cancer phenotype.
During this portion of the research, they discovered that the cancer panels
missed 11% to 16% of actionable genes relating to diagnosis, prognosis, and
treatment.
“This is either good news or bad news, depending on how you
look at it,” Newman said. “Personally, I am amazed at how well these panels do
and how well they have been designed. But, if you want to know every mutation
that you would probably want to report, you have to do comprehensive
sequencing.”
First Multi-Platform Genomic Sequencing Study
“To
our knowledge, this is the first clinical study where this comprehensive three-platform
genomic sequencing approach was offered prospectively to all pediatric oncology
patients,” said Kim Nichols, MD,
Director, Division of Cancer Predisposition at St. Jude’s, in a St.
Jude’s blog post.
The testing costs $8,600 per patient, but is considered worth
it to improve patient diagnosis, prognosis, and treatment for pediatric cancer
patients.
“Compared with the cost of many
other procedures that children with cancer undergo, the cost is likely
comparable, or even less—for example, compared with complex surgical procedures
or multiple radiology tests,” Nichols said.
In addition, the test results are available in less than 30
days, which makes them more valuable, as time can be a critical asset to cancer
management.
The scientists hope this type of three-platform genetic
testing can help guide care for pediatric cancer patients.
“Because
so few of the molecular lesions in pediatric cancer are targetable by specific
drugs, currently it is the diagnostic and prognostic insights provided by the
three-platform approach that appear most clinically impactful,” said Nichols.
“From a diagnostic perspective, tumors may look the same under a microscope,
but the identification of specific genetic changes can direct you to the correct
diagnosis, and therefore, the most appropriate therapy. From a prognostic
perspective, you will have different risk stratifications depending on results.”
The results of the research were presented at the 2018
annual meeting of the American Society of Human Genetics in San Diego last
October. The St. Jude’s researchers hope that this type of research can drive
wider adoption of WGS in the assessment of pediatric tumors to improve patient
outcomes. Pathologists and medical laboratory scientists will want to watch for
additional research findings as the team at St. Jude’s uses this approach on
more pediatric cancer patients.
These “off-target” genetic alterations demonstrate that certain CRISPR base editors need further refinement in a research finding of interest to pathologists
Could CRISPR
DNA-editing technology unintentionally effect RNA as well? A new study conducted
at Massachusetts General Hospital
(MGH) suggests that it can. Clinical
laboratories doing genetic testing will want to understand why this
research implies that refinements to CRISPR may be needed for it to be accurate
in therapeutic applications.
For years, a huge value of CRISPR (Clustered Regularly
Interspaced Short Palindromic Repeats) base editors have been their ability to
edit genes or convert a specific DNA base without breaking the DNA. Now, the MGH
scientists have discovered that certain CRISPR base editors may extend beyond
the targeted DNA and perform unwanted edits to RNA, according to a news release.
“Most investigation of off-target base editing has focused
on DNA, but we have found that this technology can induce large numbers of RNA
alterations as well. This surprising finding suggests the need to look at more
than just genetic alterations when considering unintended off-target effects of
base editors in cells,” J. Keith Joung, MD,
PhD, MGH Pathologist and Professor of Pathology at Harvard Medical School, stated in the news release.
The MGH scientists published their study in Nature.
How the MGH Researchers
Found Off-Target Effects on RNA
The researchers had set their sights on developing a base
editor that targets cytosine,
according to the study.
“Previous studies of cytosine base editor specifically have
identified off-target DNA edits in human cells. Here, we show that a cytosine
base editor with rat APOBEC1
[rAPOBEC1] enzyme can cause extensive transcriptome-wide RNA
cytosine deamination in
human cells,” the scientists wrote in Nature.
According to the news
release, when the researchers put base editors into human liver and kidney cells,
they found their technology induced efficient edits at the target DNA site.
However, they also discovered tens of thousands of cytosine-to-uracil edits in the cells. They
found that deaminases, an enzyme that acts as a catalyst, which they used in
their base editor to change DNA, also altered the RNA in the cells, Science reported.
“Base editors are still incredibly powerful tools. This is just another parameter we need to understand,” J. Keith Joung, MD, PhD (above), MGH Pathologist and Professor of Pathology at Harvard Medical School, told Science. (Photo copyright: Massachusetts General Hospital.)
The researchers developed a way to reduce the unwanted RNA
edits, while maintaining the targeted DNA effects. They came up with cytosine
base editor variants, which they dubbed SElective Curbing of Unwanted RNA
Editing (SECURE).
“We engineered two cytosine base editor variants bearing
rAPOBEC1 mutations that substantially decreased the number of RNA edits in
human cells,” the researchers wrote in their study.
However, they also
called for changes to how base editors are used. “For research applications,
scientists using base editors will need to account for potential RNA off-target
effects in their experiments,” the MGH news release notes. “For therapeutic
applications, our results further argue for limiting the duration of base-editor
expression to the shortest length of time possible and the importance of
minimizing and accounting for potential impacts of these effects in safety
assessments.”
Other Studies Explore CRISPR
Other studies published earlier this year on mice and on rice also suggested
that “modified CRISPR-Cas9 technology will need to be further refined before it
can safely be used for research and therapeutic applications,” The Scientist reported.
Clinical laboratory leaders and pathologists recognize
CRISPR technology is changing the way research is done for diagnosing disease
as well as guiding treatment. Dark Daily has reported on key
CRISPR developments over many years.
And now, though the MGH study may appear to be a set-back
for CRISPR, it also may propel further research into possible therapeutic
applications of CRISPR base editing. It’s a development worth watching.
Methods that target the causes of acidity could become part of precision medicine cancer treatments and therapies
Researchers at Massachusetts
Institute of Technology (MIT) have found that acidic environments enable
tumor cells to strengthen through protein production. And that when acidic surfaces
extend beyond a tumor’s interior, and come into contact with healthy tissue,
cancer can spread.
The results of their study will interest anatomic
pathologists who review tissue biopsies to diagnose cancer and help identify
the most effective therapies for cancer patients. Currently, there are no new clinical
laboratory tests under development based on MIT’s research.
The researchers published their findings in the journal Cancer Research. Their paper also
shared how tumor acidity can be identified and reversed.
“Our findings reinforce the view that tumor acidification is an important driver of aggressive tumor phenotypes, and it indicates that methods that target this acidity could be of value therapeutically,” noted Frank Gertler, PhD (above), in a news release. Gertler is an MIT Professor of Biology, a member of MIT’s Koch Institute for Integrative Cancer Research, and a Senior Author of the study. (Photo copyright: MIT News.)
Acidity is a Tumor Cell’s
Friend
Acidity results from lack of oxygen in tumors and enables
tumor cell growth. “Acidification of the microenvironment plays established roles
in tumor progression and provides a hostile milieu that advantages tumor cell
survival and growth compared to non-cancerous cells,” the researchers wrote in Cancer Research.
In their study, the MIT scientists sought to learn:
What areas of a tumor are actually acidic?
How does acidosis propel cells to
invade surrounding healthy tissues?
They used a nanotechnology platform
called pHLIP (pH Low
Insertion Peptide) to sense pH at the surface of cancer cells and then insert a
molecular probe into the cell membranes. “This brings nanomaterial to close
proximity of cellular membrane,” noted a research study
conducted at the University of Rhode Island by scientists who developed the
pHLIP technology.
Medical News Today reported that the MIT scientists
used pHLIP to map the acidity in human breast cancer tumors implanted in mice.
When it detected a cell in an acidic environment, pHLIP sent a small protein
molecule into the cell’s membrane. The scientists found that acidosis was not
confined to the oxygen-rich tumor core. It extended to the stroma, an important boundary
between healthy tissue and malignant tumor cells.
“We characterized the spatial characteristics of acidic
tumor microenvironments using pHLIP technology, and demonstrated that
tumor-stroma interfaces are acidic, and that cells within the acidic front are
invasive and proliferative,” the scientists wrote in Cancer Research.
What Stimulates
Acidity and How to Reverse It?
The MIT researchers sought the reasons, beyond hypoxia, for
high acidity in tumor tissue.
“There was a great deal of tumor tissue that did not have
any hallmarks of hypoxia that was quite clearly exposed to acidosis. We started
looking at that, and we realized hypoxia probably wouldn’t explain the majority
of regions of the tumor that were acidic,” Gertler pointed out in the MIT news
release.
So what did explain it? The researchers pointed to aerobic
glycolysis, a “condition in which glucose is converted to lactate in the presence
of oxygen,” according to an article published by StatPearls. “Cancer
stem cells (CSC) within a tumor are notorious for aerobic glycolysis. Thus,
extensive aerobic glycolysis has been indicative of aggressive cancer,” the
paper’s authors noted.
During their study, the MIT scientists found:
Cells at the tumor surface shifted to aerobic
glycolysis, “a type of metabolism that generates lactic acid, making way
for high acidity,” and
“Tumor acidosis gives rise to the expression of molecules
involved in cell invasion and migration. This reprogramming, which is an
intracellular response to a drop in extracellular pH, gives the cancer cells
the ability to survive under low-pH conditions and proliferate,” said Nazanin Rohani, PhD, former
postdoctoral researcher in the MIT Koch Institute for Integrative Cancer
Research, and Lead Author of the study, in the news release.
Could a Reduction in Acidity Reverse Tumor Growth?
In another experiment, the researchers fed sodium
bicarbonate (baking soda) to mice with breast or lung tumors. The tumors became
less acidic and metastatic.
“It adds to the sense that this pH dynamic is not permanent.
It’s reversible. I think that’s an important addition to an ongoing discussion
about the role of pH in tumor behavior,” said Ian Robey, PhD, in an
MITblog
post. Robey is a Research Assistant Professor, Department of Medicine
at the University of Arizona, and Full Investigator at the Arizona Cancer Center. He was not
involved in the MIT research.
Spreading the Word on
How Cancer Spreads
The MIT study is important—not only to anatomic pathologists—but
also to oncologists and cancer patients worldwide. Cancer is not simple to
diagnose and treat. The MIT study may provide important insights into targeting
cancer care and precision
medicine treatments.
