MANASSAS, Va. 鈥� What if you could know whether you had suffered a concussion 鈥� without any shadow of a doubt, and in just minutes 鈥� simply by spitting in a tube?
As the scope of potential short- and long-term damage from brain injuries in football becomes clearer every day, the mad scramble is on to try to address the crisis, which threatens the very existence of the sport.
And while the leading聽chronic traumatic encephalopathy, or CTE, research at Boston University has made the most headlines, two scientists at George Mason University have been working on a potentially groundbreaking diagnostic tool that could change the way we test for and treat brain injury across the sports world and beyond.
The way brain injuries are treated and managed is a nascent and evolving science, with the effectiveness of stopgap protocols being called into question on nearly a weekly basis during the football season. So, while the findings of the joint project between Dr. Shane Caswell, an athletic trainer, and Dr. Emanuel 鈥淐hip鈥� Petricoin, a proteomics expert, are still a ways from translating to a marketable product, they shed new insight into a field that many are only beginning to understand.
Coming from entirely different sides of research at George Mason, Caswell and Petricoin may never have met if not for a chance encounter at a social event put on by the university to try to draw a pharmaceutical company to Prince William County back in 2011. Each heard the other speak about their work and both realized there might be an opportunity to work together across disciplines.
The Center for Applied Proteomics and Molecular Medicine, which sits on a nondescript, suburban highway in Manassas, largely focuses on oncological issues, using nanoparticle technology for early detection tests for breast cancer and even a urine test for tuberculosis for children in Africa. Petricoin had been called in as a subject review expert on an Army grant focusing on traumatic brain injury, or TBI, and realized there were protein biomarkers being discovered in subjects.
Knowing the tools already at his disposal in the lab, he saw an opportunity.
鈥淲hat came out of it was a realization that the technologies and tools that we鈥檙e using to discover biomarkers for early detection of ovarian cancer could likely be used for TBI,鈥� Petricoin said. 鈥淚n fact, probably, we had better tools than the TBI community had been using before.鈥�
The first test Petricoin鈥檚 team developed for the sports world came on the heels of the human growth hormone doping scandals in the early 2000s. At the time, the only way to look for HGH was to test cerebral spinal fluid, which involved, obviously, a highly impractical method of collection.
Petricoin鈥檚 team developed a test that could find even trace amounts of HGH in urine, something thought to be analytically impossible.
In the past, these nanoparticles have been used primarily as a delivery device, often to help bring needed medicine to a particular part of the body. The nanoparticles being used by Petricoin鈥檚 team function the opposite way, by going in and retracting the desired protein from a fluid like urine or saliva, no matter the trace amount.
鈥淲e actually reverse-engineered it 鈥� we don鈥檛 want to deliver anything. We want it to suck up and harvest these low-abundance biomarkers that are in biofluids like urine, but very dilute.鈥�
Now, he鈥檚 applying the same science to discovering and detecting protein changes in the saliva for athletes with potential brain injuries.
Salivary bank
As part of his concussion research, Caswell has been building a salivary biobank for years. That has allowed him to acquire data at baseline, before any injury, allowing him to monitor potential changes as a result of brain injuries. This could be crucial for more longitudinal research in the future, allowing him to see changes over the course of months or years following a brain injury.
鈥淗ow does your biomarker fingerprint change as you鈥檙e developing, and are those changes typical? Are they related to potential injury that you might suffer? Those are questions that no one has the answer to,鈥� he said.
Saliva is a particularly useful fluid for research due to how easy it is to acquire and test.
鈥淪aliva鈥檚 noninvasive,鈥� said Caswell. 鈥淲e鈥檙e not doing a finger-stick; we鈥檙e not doing a lumbar puncture. We鈥檙e asking people to spit in a little tube.鈥�
It鈥檚 especially useful for Caswell, who is one of the few brain injury researchers focusing on youth sports.
鈥淚f you look at a lot of the research in sport-related concussion, a lot of the research that is driving the field is in collegiate athletes or professional athletes,” he said.
“But the largest number of participants are in youth and high school. And it鈥檚 very difficult to get access to those populations with invasive measures.鈥�
How the test works
The salivary proteum contains about 1,500 proteins. When Petricoin and his team started exploring, they found themselves looking at a Hubble Space Telescope-like universe of previously undiscovered territory. That included about 400 brand-new, never before seen proteins.
鈥淲e鈥檙e plumbing new analytical depths of saliva,鈥� Petricoin said. 鈥淪ome of those proteins are the ones that we found, and they shouldn鈥檛 be there.鈥�
So, how does the test work?
鈥淚magine if you had a swimming pool that was filled with baseballs and iron filings,鈥� Petricoin said.
And water?
鈥淎nd water. A swimming pool — you鈥檝e dumped in iron filings, you鈥檝e dumped in baseballs. And you鈥檙e like, 鈥楲ook, I want you, quickly, to get all the iron filings out of this and leave all the baseballs in.鈥欌��
In this analogy, the iron filings are the proteins that would indicate a positive test. The nanoparticles are like mesh balls 鈥� magnetized to attract the proteins the researchers are looking for 鈥� dropped into the pool. The openings on the mesh ball are wide enough to allow the iron filings to slip through, but not enough to allow the baseballs in.
After dropping the nanoparticles into the metaphorical pool, you retrieve them, reverse the polarity on the magnet, and out spill the iron filings, or the proteins.
Findings
鈥淲e have protein biomarkers that appear to be differentially expressed in the saliva of individuals,鈥� Petricoin said.
In layman鈥檚 terms: They have found proteins, about 10 different ones, that are appearing in saliva of kids with concussions that aren鈥檛 appearing in healthy brains.
But, perhaps more significantly, the researchers separated football players who were not actually diagnosed with any concussions into different groups, based on the frequency and force of impacts they encountered over the football season, measured by sensors in helmets.
鈥淎nother aspect of what we鈥檝e been looking at is not just when someone has suffered an overt concussion, but when someone has no clinical symptoms of concussion that are being manifested, nothing has been reported,” Caswell said.
“Is there something that we can identify in their saliva that would be indicative of changes in their biology that would be as a result of repetitive head trauma?鈥�
Sure enough, they found similar proteins in the saliva of the high-quantity, high-velocity impact group that did not present any official concussion symptoms as those who actually suffered diagnosed concussions.
鈥淜ids who had very few impacts looked different from kids who had a large volume of impacts over the course of the season,鈥� Caswell said.
That certainly seems to support the idea that an eye-test sideline diagnosis is not adequate to determine whether or not a traumatic brain injury has occurred.
So, while football organizations from youth leagues to the NFL may employ a return-to-play protocol, the damage inflicted from contact may well not be able to be detected by the person administering an observational test.
Also disconcerting are the types of proteins Petricoin鈥檚 team has found. One, in particular, is a cell receptor only ever before found in brain tissue.
鈥淚 don鈥檛 want to say you have a piece of your brain in your saliva, but, biochemically, it鈥檚 like, your brain got knocked around, it released 鈥� cellularly 鈥� things that only came from there, and it鈥檚 in your saliva,鈥� he said. 鈥淧retty good indication things are going on.鈥�
These results need to be independently validated, but both feel strongly that there鈥檚 a 鈥渢here鈥� there.
鈥淭hey鈥檙e very intriguing, and we have an engine to get more samples,鈥� Petricoin said, referring to the salivary bank. 鈥淲e have a technology that鈥檚 finding some signal that seems to indicate maybe something鈥檚 going on here.鈥�
What鈥檚 next
The research is currently undergoing peer review by a group at Lund University in Sweden, headed up by a team that has done research into the autoimmune response that the body creates upon suffering a brain injury. They hope to have two academic papers published this spring 鈥� one on protein identification and one on the autoimmune side.
The hope is to secure funding to help develop and implement a test that they could deploy in the field as a trial to test utility with a local club or school system. That could take the form of a tube, or even a mouth guard that players wear that might turn a different color if certain proteins presented themselves.
Even if such a product makes it to market, both Caswell and Petricoin are cautious about anyone latching onto it as 鈥渢he answer鈥� to football鈥檚 concussion crisis.
鈥淭here鈥檚 never been a biomarker, not one biomarker ever 鈥� except death 鈥� that鈥檚 100 percent sensitive and specific,鈥� Petricoin said. 鈥淎t the end of the day, we want to make sure the markers are rigorously validated, and we鈥檙e hesitant to be over-exuberant until we get that.鈥�
But if the findings lead to a potential sideline test, it could revolutionize the way people manage brain injury, removing much of the uncertainty and fear that surrounds the issue today.
鈥淚 don鈥檛 think this is going to be a silver bullet that solves the concussion issue,鈥� Caswell said. 鈥淏ut I think that a measure like this that could be ‘deployable,’ particularly on the field, I think could be very beneficial.鈥�
