Abstract
Central nervous system cancers are the tenth leading cause of death for adults and the leading cause of cancer mortality in children. Non-invasive detection methods are desperately needed for earlier diagnosis and recurrence monitoring. Aneuploidy is common to nearly all cancers and can be detected in circulating tumor DNA (ctDNA) isolated from a variety of biofluids, including plasma. We utilized a sensitive PCR-based assay called Repetitive Element AneupLoidy Sequencing System (RealSeqS) on plasma samples from 90 patients diagnosed with the most common primary brain tumors including pilocytic astrocytoma, Grade II and III astrocytoma, glioblastoma (GBM), oligodendroglioma, ependymoma, and medulloblastoma to detect aneuploidy in ctDNA. RealSeqS employs PCR amplification and sequencing of ~350,000 genome-wide loci to identify chromosomal abnormalities and then aggregate them into a single Genome Aneuploid Score via a supervised machine learning algorithm. RealSeqS identified aneuploidy in 16 of 90 (17.8%) patients, including 20.9% (9/43) of anaplastic astrocytomas and GBMs, 57.1% (4/7) ependymomas, and 18.8% (3/16) pilocytic astrocytomas. Notably, ependymomas have significant chromothripsis and aneuploidy, which was reflected in our plasma analyses. Detection of aneuploidy also correlated with higher mortality rates at the time of last follow-up (p < 0.001). RealSeqS did not detect aneuploidy in the plasma of patients with Grade II astrocytoma, medulloblastoma, or oligodendroglioma. RealSeqS detected focal amplifications of 11q in 16.7% (2/12) and a focal deletion of TERT in 8.3% (1/12) of Grade III astrocytomas, focal deletions of EXT1 (14.3%; 1/7) and focal amplifications of EGFR (14.3%; 1/7) in the plasma of patients with ependymomas, and focal deletions of EXT1 (3.2%, 1/31) and focal amplifications of CDKN2A and CDKN2B in (3.2%; 1/31) GBM. These findings suggest detection of aneuploidy by RealSeqS may be a rapid, cheaper, and more sensitive alternative to low pass whole genome sequencing for the earlier detection of certain CNS neoplasms.
Restricted PCR: amplification of an individual sequence flanked by a highly repetitive element from total human DNA