RARE-04. DEVELOPMENT OF A PLATFORM TO PROFILE MICRORNA IN CEREBROSPINAL FLUID FROM PATIENTS WITH INTRACRANIAL GERM CELL TUMORS

BACKGROUND: Current diagnosis of intracranial germ cell tumors (iGCTs) utilizes alpha fetoprotein (AFP) and chorionic gonadotropin (β-HCG) as biomarkers in the blood or cerebrospinal fluid (CSF). Two thirds of iGCTs are marker negative and AFP/HCG are not specific to iGCTs. Thus, there is a need to develop other biomarkers to improve the diagnosis of iGCT. Two microRNA (miRNA) clusters, miR-371-373 and miR-302/367 have previously been identified as biomarkers for extracranial GCTs and preliminary data from our lab and others confirmed their potential utility in iGCTs. However, the optimal way to profile miRNAs in CSF is not fully established. Two commonly used methods are real-time polymerase chain reaction (qPCR) or droplet digital polymerase chain reaction (ddPCR). METHODS: CSF samples were obtained from biorepositories at Connecticut Children’s and through the Children’s Brain Tumor Network. Cell culture media from an embryonal carcinoma cell line was used as positive control. MicroRNA was extracted from 50uL of CSF, reverse transcribed into cDNA and pre-amplified. Samples were then diluted 1:5 for qPCR and 1:10 or 1:50 for ddPCR. RESULTS: We profiled the CSF from 6 germinomas and 4 non-germinomatous, including two mature teratomas. Additionally, we profiled CSF from two non-iGCT brain tumors and two leukemia patients, as non-iGCT controls. For qPCR, raw CT values were evaluated with CT <35 considered detectable. Both platforms, qPCR and ddPCR were able to detect miR-371-373 and miR-302/367 in malignant iGCTs, including marker negative germinomas. miR-302/367 cluster showed varying expression in non-iGCT samples. CONCLUSIONS: We were able to use low input (50uL) of CSF to identify markers of malignant iGCT, the microRNA clusters miR-371-373 and miR-302/367. Both platforms were able to identify the microRNAs. More samples will need to be profiled to evaluate which platform ddPCR or qPCR is best to move forward towards a clinically validated platform.