Research News

Cleveland Clinic research published in Neuro Oncology Advances is the first to establish microRNA (miRNA) as a factor in what makes glioblastoma that make the brain cancer more common and more deadly in males (defined here as individuals born with XY chromosomes) than in females (individuals born with XX chromosomes). The collaborative research from the laboratories of Andrew Dhawan, MD, DPhil, FRCPC, and Justin Lathia, PhD, provides a new target for glioblastoma drug discovery research. 

There are over 100 miRNAs on the X chromosome, but only two on the Y chromosome. This observation led study lead author Ellen Hong, a graduate student co-mentored by Drs. Dhawan and Lathia, to ask whether the difference could be connected to sex-based outcomes. She found her answer in the microRNA called miR-644a. 

“I figured out pretty quickly that there were no other papers in the literature that had ever looked at the role of miRNAs in sex differences in glioblastoma and the rest is history,” she says. “It was kind of crazy to me that no one had thought to look at this before, especially because miRNA has been tied to other cancers the disparity in miRNA number coded in the X and Y chromosomes has been known for a while.” 

What is miRNA? 

miRNA are small pieces of RNA in the cell nucleus that control gene regulation, or how much of a particular type of protein our cells make. In healthy cells, miR-644a ensures its target genes cannot produce too many proteins and tip the cell from healthy to cancerous.  Higher levels of miR-644a are needed to control cell development in glioblastoma tumor cells, which retain some normal cell production patterns. 

Theoretically, miRNA on the X chromosome could be more abundant in biological females (who have two) than biological males (who have one X and one Y). Hong started with a 39-person data set to identify miRNAs whose expression levels (how much miRNA there is) differed between male and female patients. She then moved through progressively larger datasets, overall analyzing data from nearly 600 tumors and more than 809 miRNAs. one consistently stuck out across her datasets: miR-644a. 

Hong, Dr. Dhawan and Dr. Lathia and her team then needed to be creative to validate their findings in larger glioblastoma databases that did not contain miRNA expression levels.  

“Glioblastoma is a relatively rare type of cancer, so we have fewer tumors available to study directly and less publicly available data to study indirectly,” explains Dr. Dhawan, who alongside Dr. Lathia holds appointments in the Department of Cardiovascular & Metabolic Sciences and the Rose Ella Burkhardt Brain Tumor & Neuro-Oncology Center (of which the latter serves as Scientific Director). “A lot of the larger publicly available data doesn’t even have information about miRNAs because certain types of RNA sequencing can’t measure miRNA expression at all.” 

How did the researchers use gene expression data to study miRNA? 

Computer analyses predicted that miR-644a negatively regulates (turns “off”) specific genes involved in cell adhesion and cell development in brain cells. Theoretically, lower levels of those genes would indicate higher levels of miR-644a, and vice versa. 

Hong analyzed the larger cancer databases for higher levels of the miR-644a's target genes. She found patients whose brains and tumors expressed higher levels of miR-644a's target gene exhibited lower rates of survival. Females also expressed lower levels of miR-644a's target genes than their male counterparts.  

Hong validated her computational findings by directly measuring expression levels for miR-644a and its target genes in dish experiments using cells derived from male and female glioblastoma patients. The models showed that cells derived from female brains expressed miR-644a much more highly than the male cell culture models. 

“It was advantageous for me to be co-mentored by two PIs for this project because we could combine expertise,” Hong says. “Once I got through most of the initial bioinformatics with Andrew and the Dhawan lab, I had a lot of help from Justin and the Lathia lab in doing the more traditional molecular biology experiments.” 

What role could miRNA gene regulation play in glioblastoma treatment? 

Because drug developers have had relatively little success in targeting single genes in glioblastoma treatments, targeting gene networks regulated by miRNAs could be one way to collapse networks and pathways integral to glioblastoma’s growth, Hong explains. 

"Since miRNA has also been tied to sex-based differences in other cancers as well, new methods to measure them and validate the results on a larger scale have potential for drug development beyond glioblastoma," Dr. Lathia says.  

“Ideally, we would like to see patient-informed therapeutics that take factors like biological sex and miRNA into account. The first step of this is understanding underlying processes that govern sex differences of the disease, which is where we still are,” Dr. Dhawan adds. “Once we understand that, we can truly start to understand and combat why glioblastoma is so much more aggressive in males.”