Biomarker Discovery Aids in Tooth Decay Risk Assessment

The internal process of a mass spectrometer. It looks like a small needle amongst various metal objects.

By Vivian Mason

“It’s difficult to believe that in this world of state-of-the-art technology that there’s still no ‘cure’ for dental caries or any definitive way to predict it,” says Air Force Capt. Jun Kyung Ko, a comprehensive dentist with the 374th Dental Squadron in Yokota, Japan.  Ko recently graduated from the Uniformed Services University (USU) Postgraduate Dental College Master of Oral Biology degree program, and the two-year Advanced Education in General Dentistry residency at the Air Force Postgraduate Dental School in Texas.  

Dental caries (or cavities), more commonly known as tooth decay, is one of the most prevalent chronic diseases of people worldwide. Caries can occur throughout life. More than 91% of adults have a history of dental caries. 

“Oral health is essential to soldiers on the battlefield, but dental caries continues to be a major threat to warfighter health and readiness,” Ko continues. “Dental caries can lead to severe oral pain and dysfunction, and can interfere with a service member’s ability to eat, communicate, sleep, and perform assigned duties. The disease remains highly prevalent among military recruits, and it’s crucial to identify and treat it early.” 

Thankfully, research is being conducted to determine new ways to do exactly that―identify and treat tooth decay early.

Sample isolation of DNA and RNA leads to Library Prep, and finally Data Analysis when preparing a plaque sample.

Sample plaque preparation. (Photo credit: Capt. Jun Kyung Ko, Air Force Postgraduate Dental School, USU)

Through a two-year research project collaboration with Dr. Wen Chen―a Leidos research scientist III at the Navy Medical Research Unit-San Antonio (NAMRU-SA), and an adjunct associate professor of USU’s Postgraduate Dental College―six peptides were defined as novel microbial peptides and caries-associated saliva native biomarkers using peptidomics and mass spectrometric techniques.

So what exactly is a biomarker?  Biomarker is an abbreviated term for “biological marker,” and it’s also known as a molecular marker or a signature molecule. A biomarker is a biological molecule found in blood and other body fluids or tissues that is a sign of a normal or abnormal process, or a sign of a condition or disease. A biomarker can indicate that a biological process in the body has happened or is ongoing, or it may be used to identify how well the body responds to a treatment, disease, or condition. 

Initially, Capt. Ko says, they had no idea if they would find any of these biomarkers. The team was curious as to whether there was anything out there that could be used as a biomarker to distinguish a person at high risk for tooth decay from a person at low risk.

“We already know how dental caries start and progress,” Ko continues, “but we can’t really predictably and definitively measure who is more susceptible to getting them.”

To identify these biomarkers, Ko collected samples of saliva and plaque from more than sixty caries-free and caries-active patients. The samples were treated for peptidome extraction and analyzed by Chen using state-of-the-art liquid chromatography-mass spectrometry (LC-MS) instruments at NAMRU-SA. 

LC-MS technology with peptidomics has been used in the medical community, but not in the dental community. In the study, they first found a total of eight and then twelve biomarkers. Eight of them were in the higher expression level in caries-free saliva. The remaining four were lower expression caries-free saliva. 

State-of-the-art liquid chromatography-mass spectrometry (LC-MS) instruments housed at the Navy Medical Research Unit-San Antonio (NAMRU-SA). (Photo credit: Capt. Jun Kyung Ko, Air Force Postgraduate Dental School, USU)
State-of-the-art liquid chromatography-mass spectrometry (LC-MS) instruments housed at the Navy Medical Research Unit-San Antonio (NAMRU-SA).
(Photo credit: Capt. Jun Kyung Ko, Air Force Postgraduate Dental School, USU)

Of the 12 biomarkers found, Chen was able to sequence six of them with de novo peptide sequencing technology. 

“So now we know the exact sequences of these peptides,” Ko says. “Finding this sequence of unknown peptides takes a lot of manual labor, as well as Dr. Chen’s brain. He spent many, many hours of hard work to sequence those six peptides and that’s an incredible discovery!” 

Their research identified that these biomarkers were actually novel, proline-rich antimicrobial peptides that had never been identified before. These peptides are an innate part of the immune response system that can be found in all classes of life. 

“They are really special because they can be possibly considered as broad-spectrum antibiotics,” Ko explains. “We’re going to have to do a lot more research and study, but these peptides have the potential to be therapeutic agents for inflammation, wound repair, or injury requiring angiogenesis. It’s exciting stuff.”

The novel and native proline-rich antimicrobial peptides have variant and complementary properties, such as being multicharged, highly basic or acidic, and hydrophobic or hydrophilic, and could also be potential therapeutic agents against multidrug-resistant pathogenic bacteria, as well as dental caries biomarkers. 

“These discovered native peptide biomarkers for dental caries have been predicted to have antibacterial properties with bioinformatic tools,” Chen explains. “We’re excited with this discovery because it has key implications in preventive, diagnostic, and therapeutic application in dental caries diseases.

Dr. Chen remarks that the team’s achievements have the incredible potential to develop brand new therapies against tooth decay and bacteria that may be resistant to other drugs; however, the peptide biomarkers still have plenty of testing in store for them before they’re able to be fully utilized.

An illustration of the mass spectrometry process.

Mass spectrometry is used to calculate the exact molecular weight of a component.

(Photo credit: Capt. Jun Kyung Ko, Air Force Postgraduate Dental School, USU)

So far, none of this research has appeared in scientific literature. In fact, they’ll need more data in order to publish their findings. Dr. Chen and Capt. Ko hope to discover more dental caries biomarkers from ongoing research. Therefore, this study could improve caries risk assessment, preventive management, and effective/sustainable treatment for dental caries, as well as upgrade the oral health and readiness of active duty service members.

“Dr. Chen has a PhD in mass spectrometry and is a subject matter expert,” offers Ko. “He takes care of the technology side of things, whereas I look at the clinical aspects of dental caries. It’s a good combination of our skills, as well as a good collaboration between the Navy and the Air Force.”

Ko concludes, “I wanted to participate in this study to have some sort of impact. I wanted my efforts to count for something, and this discovery has more than fulfilled that.”