GlycoNet researchers discover enzymes that can aid pulmonary infections and chronic wounds

Study published in Science Advances discovered way to break down biofilms

By Rebecca Medel

GlycoNet Network Investigator Dr. Lynne Howell and her team at The Hospital for Sick Children are developing new methods to break down the wall that protects bacteria against current therapeutic interventions. In a study published on May 20 in Science Advances, the lab identified and produced two enzymes that are capable of both degrading the sugar component of the biofilm wall from Pseudomonas aeruginosa, and removing the protective structure employed by this bacterium.

Microbes, including bacteria and fungi, protect themselves by living in “walled cities” called biofilm, which is composed of DNA, proteins, and chains of sugars known as exopolysaccharides. Howell says the microbes within these biofilms are a significant problem both medically and environmentally as they are up to 1000 times more tolerant to antibiotics and disinfectants than other microbes. The enzymes are designed to work as adjuvants to make current therapies more effective. Using the analogy of an ancient walled city, the enzymes function as battering rams to break down the city’s wall, in this case the biofilm, which then allows an army—the antibiotics—to enter and target the bacterial cells.

The work has been a team effort and includes GlycoNet investigator Dr. Don Sheppard. Howell, who is also Associate Chief, Research Integration and Communications at SickKids, says the research started about 15 years ago when the lab was looking at different metabolic pathways in bacteria, which produces a molecule that is involved in quorum sensing.

“I was looking for some new projects and we started to look at what types of processes quorum sensing controlled,” Howell recalls. “It is involved in the biofilm life-cycle process. As a consequence of reading and talking to colleagues, we started two new projects.”

One of the projects started to examine the production of chains of sugars called exopolysaccharides, a critical component of many bacterial biofilms.

“And so as a fundamental, basic research scientist, I wanted to know how were those polymers produced? How were they synthesized? How did the building blocks get put together? … That’s how it started,” Howell says. “We started on one polymer and we now have five different polymers and biosynthetic systems that we are interested in studying.”

It was found that within each of the systems, and within each of the genetic components involved in the making of a polymer, there is a protein or part of a protein that is capable of degrading the polymer. An initial inhibition experiment to determine what would happen if the enzyme was added as the bacteria started to grow found that no biofilm grew. After a series of experiments looking at other systems in the Pseudomonas field, Banting Post Doctoral Fellow at SickKids, Dr. Perrin Baker, found that the two enzymes were also capable of disrupting preformed biofilms.

“It’s this disruption, which is sort of the unique aspect,” Baker explains. “There are a number of molecules that have been demonstrated to prevent biofilms from forming through various mechanisms, many of which are not fully understood, but the enzymes being able to break down and rapidly destroy bacterial walls is what makes this sort of unique.”

The rapidity of how the enzymes work has led to a number of applications being considered including their use to treat pulmonary infections and chronic wounds and burns. “The emergence of multi-drug resistant and completely drug resistant Pseudomonas aeruginosa in Canada is a major health concern,” Sheppard says. “The ability of these enzymes to disrupt biofilms and enhance antimicrobial agents suggests that theses molecules may provide us with a new strategy to combat with these superbugs.”

The team is currently working on an application to coat the enzymes on surfaces to prevent biofilms from forming. Howell says if they are successful with the coating application, they should be able to prevent a lot of hospital-acquired infections. Baker adds that 65 percent of hospital-acquired infections are due to biofilm formation on indwelling medical devices. They have also formulated a wound gel that is liquid at four degrees, but is a polymer at room temperature, and were able to demonstrate that even mixed within the gel, the enzyme was still active and able to degrade the biofilm.

“The downstream [benefit] for Canadians, if we’re successful in the animal and clinical trials, is that we can more rapidly treat wounds and chronic wounds as well as individuals who have various pulmonary diseases that are affected by these organisms,” Baker says.

The next steps for Howell and Sheppard, which have been funded in part through GlycoNet’s translational grant program, involve taking the enzymes and showing their efficacy in an animal model infection.

“If we can show efficacy [in these models], then we can move to clinical trials,” Howell explains. “The efficacy and animal models are going to target—in the case of the funding from GlycoNet—[Pseudomonas and Aspergillus] pulmonary infections … while the funding we have from [National Institutes of Health] is looking at testing the efficacy of these enzymes for burn and chronic wounds.”

The potential use of these two enzymes could be life changing for those with lung infections and chronic wounds.

“I am cautiously optimistic that we that have something that will be impactful and which holds promise to improve patient outcomes,” says Howell. “This project epitomized the type of research SickKids encourages— the translation of basic research into the clinical realm.”

The Canadian Glycomics Network (GlycoNet) is a pan-Canadian, multidisciplinary research network aiming to deliver solutions to important health issues and improve the quality of life of Canadians through the study of glycomics. GlycoNet is a member of the Networks of Centres of Excellence, a Government of Canada program that funds large scale, academic-led research networks to build research capacity and accelerate the creation of new knowledge in a specific research area.

Citation: P. Baker, P. J. Hill, B. D. Snarr, N. Alnabelseya, M. J. Pestrak, M. J. Lee, L. K. Jennings, J. Tam, R. A. Melnyk, M. R. Parsek, D. C. Sheppard, D. J. Wozniak, P. L. Howell, Exopolysaccharide biosynthetic glycoside hydrolases can be utilized to disrupt and prevent Pseudomonas aeruginosa biofilms. Sci. Adv. 2, e1501632 (2016).