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Exosome-Based Screening Analysis of Cervical Mucus May Enable Early Diagnosis of Ovarian Cancer, Leading to Earlier Intervention & Increased Survival; Clemson-Led Collaboration Employs Special Chromatography Method for Rapid Isolation of Exosomes

A team of researchers from Clemson University and Prisma Health–Upstate, both in South Carolina, are working to create a screening process to detect ovarian cancer in the early or pre-cancerous stages, according to a June 4, 2019 release from Clemson University. Their goal of the researchers is to make this screening as simple and easy for women as getting a pap smear. The idea is to identify the pre-cancerous changes by analyzing the makeup of the cervical mucus. Dr. Larry Puls (right in photo), the Director of Gynecologic Oncology at Prisma Health Cancer Institute; Terri Bruce (center in photo), Director of the Clemson University Light Imaging Facility and Research Assistant Professor of Bioengineering, and Ken Marcus (left in photo), PhD, Professor of Chemistry at Clemson and 2019 University Researcher of the Year, have been working together for more than a year to create this process. Through their research of cervical mucus, exosomes, and chromatography, the scientists are working to find a way to detect and identify pre-cancerous changes in a diagnostic setting. And now they are putting their screening tool to the test through trials. According to the Centers for Disease Control and Prevention (CDC), there is no simple, reliable way to screen for ovarian cancer, especially in women who do not show symptoms. And none of the usual cancer exams work as a screening tool to catch the disease in the pre-cancerous stage, Dr. Puls said. “People have looked at pelvic exams, blood tests, and transvaginal ultrasonography as screening tools and none of it has worked as we would like,” Dr. Puls said. “To this day, there is no adequate screening protocol for ovarian cancer and that’s part of the reason why it’s such a lethal disease. We can’t find it early enough.”

Ovarian cancer is the fifth deadliest cancer for women, just behind pancreas, colon, breast, and lung cancers according to the CDC. In 2015, the latest year for which data is available, nearly 14,000 women died of ovarian cancer in the United States, according to the CDC.

While there are ways to diagnose ovarian cancer in the early stages, it can be difficult, because the term ovarian cancer can be used to describe cancer of the ovary, fallopian tube, or peritoneum, Dr. Puls said. However, Dr. Puls said he believes the majority of cases of this type of cancer most likely originate in the fallopian tubes. One of the best ways to potentially impact survival with ovarian cancer is to screen for it before symptoms occur and treat it before it is advanced, Dr. Puls said.

“Of all the advanced stage ovarian cancer patients, which makes up about 75 percent of the cases, only 10-15 percent will survive,” Dr. Puls said. “But if you can find it when it’s confined to just the ovary alone, 90 percent of patients beat their cancer. If we could shift women out of stage 3 and into stage 1, we can make a huge impact on this disease. We can even make a bigger impact if we could find a pre-cancerous change.”

Citing previous research, Dr. Puls said that researchers have begun to identify this change because of a genetic mutation that predisposes a woman to ovarian cancer.

If most ovarian cancer cases are actually fallopian tube cancer, Dr. Bruce, Dr. Marcus, and Dr. Puls said precancerous changes could be identified using the cervical mucus because there’s a direct conduit from the fallopian tube to the mucus.

To do this, they used Dr. Bruce’s already established work with exosomes, which are tiny, subcellular, membrane-bounded vesicles derived from a cell.

“Exosomes were once simply dismissed as a way of eliminating ‘trash’ from cells, but researchers now realize that they may be the key to the development of liquid biopsies,” Dr. Bruce said.

This is because exosomes are derived from virtually all cells, giving a snapshot of what is going on within the cell, and can be found in any liquid within the human body, such as cervical mucus.

“You can tell a lot about somebody by looking through their trash, so it turns out that you learn a lot about a cell from its exosomes,” Dr. Bruce said. “This can include an overview of the types of RNA and proteins that are being made within the cells. Changes in these profiles could hold the key to early diagnostics.”

To make this idea a reality, the exosomes first must be captured. Because they are so small, exosomes are routinely separated from body fluids using a two- to three-hour process of ultracentrifugation steps or kits that can leave behind contaminants. These methods are costly and time-consuming, making a simple diagnostic test based on these protocols unrealistic, Dr. Bruce said.

To separate the exosomes from the cervical mucus, the researchers used Dr. Marcus’ method that involves using capillary-channeled polyester fibers and hydrophobic interaction chromatography in a process that takes 8 to 10 minutes and leaves behind no appreciable contaminants. This method, which was studied by Dr. Bruce and Dr. Marcus and was recently described in the journal Electrophoresis (https://onlinelibrary.wiley.com/doi/10.1002/elps.201800417), could evolve in the future to be used for a “dipstick” test, Dr. Bruce said.

“We’ve been getting patient samples so we can really start looking at possible ways of isolating very specific exosomes to identify common marker proteins, as well as mRNA messages, to help identify early-stage ovarian cancer,” Dr. Bruce said.

Through private funding, Drs. Puls, Bruce, and Marcus have started trials and will continue for several more months. They plan to use the data this preliminary study provides to apply for larger, federal grants from agencies such as the National Institutes of Health, the National Cancer Institute, and the Department of Defense. With the data they’ve collected thus far, they are hopeful this could be a way to help identify ovarian cancer earlier.

“The hope is to be able to come up with an early diagnostic test so that we can catch this deadly cancer much earlier and give women a fighting chance,” Dr. Bruce said.

Their research is a product of the mission of the Clemson University (CU) School of Health Research (https://www.clemson.edu/health-research/), a virtual school that works to bring researchers together for collaboration and advance the research initiative at Clemson.

Windsor Sherrill, PhD, Associate Vice President of Health Research at Clemson, brought Dr. Puls and Dr. Bruce together for this type of research because Dr. Puls had been looking to create a screening tool for ovarian cancer, and Dr. Bruce had been looking at cellular communication and exosomes in previous cancer and disease progression research. Because of previous collaboration, Dr. Bruce knew that Dr. Marcus’ work with chromatography would be an asset in their project.

“This initiative illustrates the power of collaboration between Clemson faculty and clinical investigators at Prisma Health–Upstate and is the type of innovation we envisioned with the CU School of Health Research,” Dr. Sherrill said.

“Professors Bruce and Marcus bring unique expertise to the challenges of cancer care and working with Dr. Puls and the oncology team at Prisma Health, they are making great strides toward their goal,” she added.

[Clemson news release] [Electrophoresis article]

IMAGES

In photo at left below, Clemson University’s Dr. Ken Marcus (left) and Dr. Terri Bruce (center) are working with Prisma Health’s Dr. Larry Puls (right) to develop a screening process for ovarian cancer. (Image Credit: Clemson University School of Health Research).

In photo at right below, Tyler Slonecki (left), a bioengineering graduate student, looks at exosomes using the Leica SP8 spectral confocal microscope with hyvolution super resolution and discusses the findings with Dr. Terri Bruce. (Image Credit: Clemson University School of Health Research).