Identifying patients who will likely develop prolonged concussion symptoms could lead to new clinical laboratory tests and personalized medicine treatments
Researchers are homing in on a new diagnostic assay for concussion that could potentially generate significant numbers of test referrals to the nation’s clinical laboratories. This innovative work is targeting how concussions are diagnosed and treated.
Each year, thousands of children receive sports-related injuries, including concussions. There are ways for anatomic pathologists and hospital medical laboratories to diagnose concussions; however, testing can be invasive and doesn’t always reveal a complete picture of the injury state.
Additionally, about one third of children with concussions develop prolonged symptoms. However, when prescribing treatment plans, physicians have been unable to predict which patients are likely to recover quickly versus those who will have a longer recovery.
Now, researchers at Penn State College of Medicine (Penn State) believe they have discovered five microRNAs in saliva that could be used to identify patients who will likely experience prolonged concussion symptoms even one month after the initial injury.
The study also found that certain materials in saliva can help diagnose the severity of concussions and could hold the key to more effective clinical laboratory tests and personalized medicine treatments.
The Penn State researchers published their study results in JAMA Pediatrics, a publication of the Journal of the American Medical Association (JAMA).
Concussion Leading Sports-related Brain Injury
There are approximately 3.8 million sports and recreation-related traumatic brain injuries in the United States each year and the majority of those cases are concussions, according to The Concussion Place. Most concussions treated in emergency rooms are due to falls, motor-vehicle related injuries, being struck by an object, assaults, or playing sports.
Also known as mild traumatic brain injuries (mTBI), concussions are caused by blows or jolts to the head or body that cause the brain to move with excessive force inside the skull. The sudden impact damages brain cells and causes chemical changes within the brain that alter normal functioning. Though usually not life threatening, the damage can be serious and linger for months.
Symptoms of concussion include: headaches, fatigue, nausea, vomiting, dizziness, balance problems, confusion, memory problems, sleep disturbances, and double or blurry vision. Symptoms usually occur immediately, but could take days or even weeks to appear.
Identifying Severity/Predicting Prolonged Symptoms of Traumatic Brain Injuries
After a concussion occurs, brain cells release small fragments of genetic material known as microRNAs while they attempt to repair themselves. A portion of these microRNAs appear in the injured person’s blood and saliva.
In order to determine whether these microRNAs could be used to determine the severity of a traumatic brain injury and predict whether prolonged symptoms would occur, the prospective cohort study researchers gathered saliva samples from 52 concussion patients between the ages of seven and 21:
The average age of the subjects was 14;
Twenty-two of the participants were female;
They were all athletes; and,
The majority of the samples were collected one to two weeks after the initial injury.
The researchers examined distinct microRNAs in the samples and identified some that enabled them to predict how long a patient’s concussion symptoms might last. In addition, they found one microRNA in children and young adults that accurately predicted which subjects would experience memory and problem-solving difficulties as part of their symptomatology.
The researchers also evaluated the concussion patients using the Sport Concussion Assessment Tool (SCAT-3), Third Edition. Physicians use this questionnaire to assess the symptoms and severity of concussions. The researchers also asked the parents of the concussed patients for observations about their children’s symptoms.
During follow up visits, which occurred at four- and eight-week increments following the original assessment, the Penn State researchers collected additional saliva samples and re-evaluated the patients using SCAT-3.
New Biomarkers Based on MicroRNAs Instead of Protein
“There’s been a big push recently to find more objective markers that a concussion has occurred, instead of relying simply on patient surveys,” Steven Hicks, MD, PhD, Assistant Professor of Pediatrics, Penn State College of Medicine, Hershey, Pa., one of the study researchers, told Penn State News.
“Previous research has focused on proteins, but this approach is limited because proteins have a hard time crossing the blood-brain barrier. What’s novel about this study is we looked at microRNAs instead of proteins, and we decided to look in saliva rather than blood,” he noted.
According to Steven Hicks, MD, PhD (above), who worked on the Penn State College of Medicine study, microRNAs could be more accurate than the traditional questionnaire when diagnosing and forecasting the effects of concussions. “The microRNAs were able to predict whether symptoms would last beyond four weeks with about 85% accuracy,” he told Penn State News. “In comparison, using the SCAT-3 report of symptoms alone is about 64% accurate. If you just go off the parent’s report of symptoms, it goes down to the mid-50s. In this pilot study, these molecular signatures are outperforming survey tools.” (Photo copyright: MD Magazine.)
The goal of this research was to develop a way to definitively ascertain that a concussion had occurred, predict the length and type of symptoms, and then use that data to improve and personalize care for children and young adults who have had a concussion.
“With that knowledge physicians could make more informed decisions about how long to hold a child out of sports, whether starting more aggressive medication regimens might be warranted, or whether involving a concussion specialist might be appropriate,” Hicks told MD Magazine. “Anytime we can use accurate, objective measures to guide medical care, I think that represents an opportunity to improve concussion treatment.”
Further research and clinical trials will be needed to solidify the effectiveness and accuracy of these new biomarkers. However, a rapid, non-invasive saliva test that can determine the severity of a concussion, and predicted whether prolonged symptoms will likely occur, would be widely used and could be an important assay for clinical laboratories. Particularly those associated with hospital medical laboratories and emergency rooms.
Research published in JAMA Pediatrics reports that non-invasive salivary microRNA testing identifies prolonged concussion symptoms with 85% accuracy
Sports-related concussions are always tragic, but doubly so when they involve child athletes. Quick diagnoses and treatments are critical to prevent permanent brain injury. But doctors are often hampered by the pace at which traditional medical imaging modalities and clinical laboratory diagnostic technologies provide crucial feedback.
Now, researchers at Penn State Health Children’s Hospital have determined that microRNA in saliva could be used as biomarkers in point-of-care concussion testing during sports events, according to a Penn State Health news release. Such sideline saliva analyses could provide quick feedback to field doctors on whether a head injury is serious enough to put injured athletes out of play, and how long the effects of such injuries might last. But is it accurate?
Jeremiah J. Johnson, MA, BS, Department of Pediatrics, at Penn State College of Medicine in Hershey, Pa., et al, recently published a study in the Journal of the American Medical Association (JAMA) Pediatrics that evaluated the ability of salivary microRNA to identify concussion in children. The salivary test of microRNA levels, Johnson and colleagues argued, does accurately identify the “duration and character of concussion symptoms.” According to the researchers, the test demonstrated high prognostic potential as a “toolset for facilitating concussion management” and may provide an additional biomarker source for use in clinical laboratory testing.
MicroRNA Offers New Biomarkers for Concussion Diagnosis
The study tested the saliva of 52 adolescents with a clinical diagnosis of mild traumatic brain injury in the form of concussion for specific microRNA expressions. Researchers identified five microRNA molecules which “accurately identify” patients with concussion symptoms. Three of those molecules served to diagnose specific symptoms of headache, fatigue, and memory difficulties up to one month after injury with low false detection rates. Because these microRNA molecules are not specific to children, could the test maintain diagnostic accuracy for patients of all ages?
Meehan and Mannix also remarked on the speed and relative ease of obtaining saliva samples, stating that “salivary microRNAs could also offer insights into the underlying biological mechanisms of injuries, potentially identifying specific targets to modify disease.”
More Accurate than Current Concussion Diagnosis Tools
There has been a marked interest in microRNA analysis and testing in recent years. MicroRNA analysis and testing has found use in cancer prognosis and personalized medicine that help predict responses to specific treatments for individual patients with a variety of chronic diseases. The news that microRNA can be used to predict concussion and duration of symptoms further solidifies the role microRNA may play in medical laboratory testing in the near future.
In an interview with CNN, Steve Hicks, MD, PhD, senior author of the JAMA Pediatrics research article and Assistant Professor of Pediatrics at Penn State College of Medicine, reported that the salivary microRNA test predicted concussion with 85% accuracy in comparison to current clinical survey measures, which are “approximately 65% accurate.” Hicks added that “the technology required to measure saliva RNA is already employed in medicine” as a common means of testing for upper respiratory viruses and that “modifying this approach for patients with concussions could potentially provide a rapid, objective tool for managing brain injury.”
Currently the Standard Concussion Assessment Tool, Third Edition (SCAT 3), which includes a series of cognitive and physical tests, is used on sports sidelines to detect concussion symptoms. Hicks notes that one problem with SCAT 3 is that “an athlete may have a concussion even if [his or her] score is ‘normal.’” Therefore, the microRNA saliva test could provide objective evidence of concussion in patients SCAT 3 fails to accurately diagnose.
Steven D. Hicks, MD, PhD (above), led the research team that studied the use of microRNA in saliva, rather than in blood, as a biomarker to identify concussions symptoms in children, and determine how long effects of the injury might last. (Photo copyright: Penn State Health.)
Too Early to Know How Helpful the Test May Be?
In the same CNN interview, Neurologist Jeffery Kutcher, MD, head of the Sports Neurology Clinic at The Core Institute in Brighton, Mich., stated that the Penn State study’s findings were “promising” and that “work like this is important because it does provide potential for tests that can be helpful in the clinical setting.” Kutcher cautioned however, that it was “too early to know what this type of tool can do for us.”
In an NPR article, Manish Bhomia, M.Eng., PhD, a brain injury researcher with the Uniformed Services University of the Health Sciences commented that “a saliva test could greatly improve care for young people who don’t have obvious symptoms of a concussion.” Bhomia stated that “micro-RNAs offer a promising way to assess concussions in adults as well as children,” but he is wary to laud saliva tests as the best method of measuring relevant microRNA molecules. Bhomia states that blood samples “which tend to contain greater numbers of the genetic fragments” are perhaps a better option.
Hicks disagrees. In an article from Penn State News, Hicks stated that the novel aspect of this study was that it focused on microRNA levels “in saliva rather than blood.” Thus, a test based on saliva, rather than a phlebotomy stick or more invasive blood testing, requires no need for venous blood.
“The ultimate goal is to be able to objectively identify that a concussion has happened and then predict how long the symptoms will go on for,” Hicks noted in the Penn State News article. “Then, we can use that knowledge to improve the care that we provide for children who have concussions, either by starting medicine earlier or holding them out of activities for longer.”
Quadrant Biosciences, a biotech company in Syracuse, N.Y., that helped fund the study, is hoping to “bring a saliva test for concussion to market in the next 12 to 24 months,” according to Hicks in his CNN interview. If development proceeds as planned, the saliva test could prove a “game changer” for sports medicine diagnostics and possibly open new avenues for related microRNA in clinical laboratory testing.
Researchers at UCLA have published the foundation science to use saliva as the specimen for sophisticated diagnostic testing
Someday soon, when your dentist asks you to say “Ah”, he will then collect a saliva specimen and use a chairside point-of-care test (POCT) to screen you for any number of conditions and diseases. This is the goal of a research team at the University of California, Los Angeles (UCLA), who recently developed what they call the Salivaomics Knowledge Base (SKB). It is a web-based data management system dedicated to help clinicians use saliva as a diagnostic tool.
Nearly 100,000 patients submitted saliva samples to a genetic testing laboratory, providing insights into their disease risk
Researchers at Mayo Clinic have employed next-generation sequencing technology to produce a massive collection of exome data from more than 100,000 patients, offering a detailed look at genetic variants that predispose people to certain diseases. The study, known as Tapestry, was administered by doctors and scientists from the clinic’s Center for Individualized Medicine and produced the “largest-ever collection of exome data, which include genes that code for proteins—key to understanding health and disease,” according to a Mayo Clinic news release.
For our clinical laboratory professionals, this shows the keen interest that a substantial portion of the population has in using their personal genetic data to help physicians identify their risk for many diseases and types of cancer. This support by healthcare consumers is a sign that labs should be devoting attention and resources to providing these types of gene sequencing services.
As Mayo explained in the news release, the exome includes nearly 20,000 genes that code for proteins. The researchers used the dataset to analyze genes associated with higher risk of heart disease and stroke along with several types of cancer. They noted that the data, which is now available to other researchers, will likely provide insights into other diseases as well, the news release notes.
“What we’ve accomplished with the Tapestry study is a blueprint for future endeavors in medical science,” said gastroenterologist and lead researcher Konstantinos Lazaridis, MD (above), in the news story. “It demonstrates that through innovation, determination and collaboration, we can deeply advance our understanding of DNA function and eventually other bio-molecules like RNA, proteins and metabolites, turning them into novel diagnostic tools to improve health, prevent illness, and even treat disease.” Some of these newly identified genetic markers may be incorporated into new clinical laboratory assays. (Photo copyright: Mayo Clinic.)
How Mayo Conducted the Tapestry Study
One notable aspect of the study was its methodology. The study launched in July 2020 during the COVID-19 pandemic. Since many patients were quarantined, researchers conducted the study remotely, without the need for the patients to visit a Mayo facility. It ran for five years through May 31, 2024. The news release notes that it’s the largest decentralized clinical trial ever conducted by the Mayo Clinic.
The researchers identified 1.3 million patients from the main Mayo Clinic campuses in Minnesota, Arizona, and Florida who met the following eligibility criteria:
Participants had to be 18 or older,
they had to have internet and email access, and
be sufficiently proficient in speaking and reading English.
More than 114,000 patients consented to participate, but some later withdrew, resulting in a final sample of 98,222 individuals. Approximately two-thirds were women. Mean age was 57 (61.9 for men and 54.3 for women).
“It was a tremendous effort,” said Mayo Clinic gastroenterologist and lead researcher Konstantinos Lazaridis, MD, in the news release. “The engagement of such a number of participants in a relatively short time and during a pandemic showcased the trust and the dedication not only of our team but also of our patients.”
He added that the researchers “learned valuable lessons about some patients’ decisions not to participate in Tapestry, which will be the focus of future publications.”
Three Specific Genes
Enrolled patients were invited to visit a website, where they could view a video and submit an eligibility form. Once approved, they completed a digital consent agreement and received a saliva collection kit. Participants were also invited to provide information about their family history.
Helix, a clinical laboratory company headquartered in San Mateo, Calif., performed the exome sequencing.
Though Helix performed whole exome sequencing, the researchers were most interested in three specific sets of genes:
Patients received clinical results directly from Helix along with information about their ancestry. Clinical results were also transmitted to Mayo Clinic for inclusion in patients’ electronic health records (EHRs).
Among the participants, approximately 1,800 (1.9%) had what the researchers described as “actionable pathogenic or likely pathogenic variants.” About half of these were BRCA1/2.
These patients were invited to speak with a genetic counselor and encouraged to undergo additional testing to confirm the variants.
Tapestry Genomic Registry
In addition to the impact on the participants, Mayo Clinic’s now has an enormous amount of raw sequencing data stored in the Tapestry Genomic Registry, where it will be available for future research.
The database “has become a valuable resource for Mayo’s scientific community, with 118 research requests submitted,” the researchers wrote in the news release. Mayo has distribution more than a million exome datasets to other genetic researchers.
“What we’ve accomplished with the Tapestry study is a blueprint for future endeavors in medical science,” Lazaridis noted. “It demonstrates that through innovation, determination, and collaboration, we can deeply advance our understanding of DNA function and eventually other bio-molecules like RNA, proteins and metabolites, turning them into novel diagnostic tools to improve health, prevent illness, and even treat disease.”
Everything about this project is consistent with precision medicine, and the number of individuals discovered to have risk of cancers is relevant. Clinical laboratory professionals understand these ratios and the importance of early detection and early intervention.
Is it possible that there is a connection between an individual’s gut microbiota and the ability to fight off gastrointestinal (GI) cancer? Findings from a preliminary research study performed by researchers in South Korea suggest that a link between the two may exist and that fecal microbiota transplants (FMTs) may enhance the efficacy of immunotherapies for GI cancer patients.
The proof-of-concept clinical trial, conducted at the Gwangju Institute of Science and Technology (GIST), Gwangju 61005, Republic of Korea, analyzed how an FMT could help 13 patients with metastatic solid tumors that were resistant to the anti-PD-1 antibody drug known as nivolumab (Opdivo). Anti-PD-1 drugs are immunotherapies that help treat cancer by improving an individual’s immune response against cancer cells.
Four of the trial participants had gastric cancer, five had esophageal cancer, and the remaining four had hepatocellular carcinoma. The patients were given a colonoscopy to implant the FMTs. The recipients also received antibiotics to reduce the response of their existing microbiotas.
The FMT donors also had gastric cancer, esophageal cancer, or hepatocellular carcinoma. Prior to donating their fecal matter, the donors experienced complete or partial response to the anti-PD-1 drugs nivolumab or pembrolizumab (Keytruda) for at least six months after receiving initial treatments.
“This research highlights the complex interplay between beneficial and detrimental bacteria within the gut microbiota in determining treatment outcomes,” co-senior study author Hansoo Park, MD, PhD, Assistant Professor, Biomedical Science and Engineering, Gwangju Institute of Science and Technology, told The ASCO Post. “While the connection between gut microbiota and immune response to cancer therapy has been a growing area of interest, our study provides concrete evidence and new avenues for improving treatment outcomes in a broader range of cancers,” he added. Further studies may confirm the need for microbiome testing by clinical laboratories to guide clinicians treating patients with colon cancers. (Photo copyright: Gwangju Institute of Science and Technology.)
Surprising Results
Fecal material for an FMT procedure combines donated fecal matter with a sterile saline solution which is then filtered to produce a liquid solution. That solution is then administered to the recipient via colonoscopy, upper GI endoscopy, enema, or an oral capsule. The solution may also be frozen for later use.
Upon analyzing the recipients, the scientists found that six of the patients (46.2%) who had experienced resistance to immunotherapies for their cancers, benefitted from the FMTs.
“Both donors were long-lasting, good responders to anti-PD-1 inhibitors, but because we did not yet know the causative bacteria responsible for the [FMT] response, we could not predict whether the treatment would be effective,” she added.
The researchers also determined that the presence of a bacterial strain known as Prevotella merdae helped to improve the effectiveness of the FMTs, while two strains of bacteria—Lactobacillus salivarius and Bacteroides plebeius (aka, Phocaeicola plebeius)—had a detrimental impact on the transplants.
Challenges to Widespread Adoption of FMTs
The researchers acknowledge there are challenges in widespread acceptance and use of FMTs in treating cancers but remain optimistic about the possibilities.
“Developing efficient and cost-effective methods for production and distribution is necessary for widespread adoption,” Sook Ryun Park told The ASCO Post. “Addressing these challenges through comprehensive research and careful planning will be essential for integrating FMT into the standard of care for cancer treatment.”
More research and clinical trials are needed before this use of FMTs can be utilized in clinical settings. However, the study does demonstrate that the potential benefits of FMTs may improve outcomes in patients with certain cancers. As this happens, microbiologists may gain a new role in analyzing the microbiomes of patients with gastrointestinal cancers.
“By examining the complex interactions within the microbiome, we hope to identify optimal microbial communities that can be used to enhance cancer treatment outcomes,” Hansoo Park told The ASCO Post. “This comprehensive approach will help us understand how the microbial ecosystem as a whole contributes to therapeutic success.”