Rapid assessment of <i>MGMT</i> promoter methylation status by droplet digital polymerase chain reaction on intraoperative smear preparation: A proof-of-concept study

MGMT promoter methylation is an important predictor of response to chemotherapy in adult-type diffuse gliomas. Currently, MGMT promoter methylation assessment requires formalin-fixed, paraffin-embedded (FFPE) tissue. We developed a droplet digital PCR (ddPCR) MGMT assay performed on bisulfite converted DNA utilizing primers targeting CpG positions with known clinical utility, which we validated against DNA methylation array (98.7% concordance). Smear preparations are routine for intraoperative neurosurgical consultation; thus, we sought to validate ddPCR using smear preparations. Thirty-six smears were collected during intraoperative consultation. For the first 28 samples, DNA from smears was extracted and bisulfite converted, and MGMT promoter methylation was quantified by ddPCR. These results were compared to DNA methylation array. For the final 8 samples, slides were H&E-stained and cover slipped for cellularity assessment prior to DNA extraction and directly compared with corresponding FFPE specimens. The smear ddPCR method showed concordance in 81.6% of cases, with a sensitivity of 85%, specificity of 100%, positive predictive value of 100% and negative predictive value of 81.3%. This novel approach will reduce turnaround time (approximately 2 days) over conventional MGMT analysis with FFPE and could provide clinically pertinent information for treatment planning and clinical trial enrollment options prior to discharge of the patient.