Digital PCR Technology Detects Brain Tumor–associated Mutation in Cerebrospinal Fluid
Researchers at Massachusetts General Hospital have used digital versions of a standard molecular biology tool to detect a common tumor-associated mutation in the cerebrospinal fluid of patients with brain tumors. In a report published in Molecular Therapy – Nucleic Acids, the investigators described using digital polymerase chain reaction (PCR) to analyze extracellular vesicles for the presence of a tumor-associated mutation in the IDH1 gene.
"Reliable detection of tumor-associated mutations in cerebrospinal fluid with digital PCR would provide a biomarker for monitoring and tracking tumors without invasive neurosurgery," said corresponding author Xandra Breakefield, PhD, of the Molecular Neurogenetics Unit at Massachusetts General Hospital. "Knowing the IDH1 mutation status of these tumors could help guide treatment decisions, since a number of companies are developing drugs that specifically target that mutant enzyme."
Both normal and tumor cells regularly release extracellular vesicles, which contain segments of RNA, DNA or proteins and can be found in blood, cerebrospinal fluid, and other bodily fluids. An earlier study identified a relatively large tumor-associated mutation in extracellular vesicles from the blood of brain tumor patients, but most current diagnostic technologies that analyze cerebrospinal fluid do not capture molecular or genetic information from central nervous system tumors.
In addition, explained co–lead author Leonora Balaj, PhD, "Tumor-specific extracellular vesicles make up only a small percentage of the total number of extracellular vesicles found in either blood or cerebrospinal fluid, so finding rare, single-nucleotide mutations in a sample of blood or cerebrospinal fluid is very challenging. These digital PCR techniques allow the amplification of such hard-to-find molecules, dramatically improving the ability to identify tumor-specific changes without the need for biopsy."
Study Details
The current study used two forms of digital PCR—BEAMing and Droplet Digital PCR—to analyze extracellular vesicles in the blood and cerebrospinal fluid of brain tumor patients and healthy controls for the presence of a single-nucleotide IDH1 mutation known to be associated with several types of cancer.
Both forms of PCR were able to detect both the presence and abundance of mutant IDH1 in the cerebrospinal fluid of five of the eight patients known to have IDH1-mutant tumors. Two of the three mutation-positive tumors that had false-negative results were low-grade and the third was quite small, suggesting a need for future studies of more samples to determine how the grade and size of the tumors affect the ability to detect mutations. The failure to detect tumor-associated mutations in blood samples with this technology may indicate that cerebrospinal fluid is a better source for extracellular vesicles from brain tumors.
Potential Effect on Patient Care
The ability to noninvasively determine the genetic makeup of brain tumors could have a significant effect on patient care, explained study coauthor Fred Hochberg, MD. "The current approach for patients who may have a brain tumor is first to have a brain scan and then a biopsy to determine whether a growth is malignant. Patients may have a second operation to remove the tumor prior to beginning radiation therapy and chemotherapy, but none of these treatments are targeted to the specific molecular nature of the tumor,” he said.
"Having this sort of molecular diagnostic assay—whether in spinal fluid or blood—would allow us to immediately initiate treatment that is personalized for that patient without the need for surgical biopsy," he added. "For some patients, the treatment could shrink a tumor before surgical removal, for others it may control tumor growth to the point that surgery is not necessary … We still have a long way to go to improve survival [in patients with] these malignancies, so every improvement we can make is valuable."
The study was funded by National Institutes of Health grants CA069246, CA141226, CA156009 and CA141150 and grants from the Brain Tumor Funders’ Collaborative and the American Brain Tumor Association.
The content in this post has not been reviewed by the American Society of Clinical Oncology, Inc. (ASCO®) and does not necessarily reflect the ideas and opinions of ASCO®.
