Jessica Blackburn from the University of Kentucky spoke at the Nanopore Community Meeting 2021 about “Detection of cell-free DNA in acute lymphoblastic leukemia patients to potentially identify CNS disease and monitor response to therapy.” Blackburn explained that acute lymphoblastic leukemia is the most common pediatric cancer, with ~3,000 children diagnosed each year. They explained that 90% of children and 40% of adults can be cured, but the prognosis of patients relapsing is grim, with <30% surviving. Minimal residual disease (MRD) is the best predictor of relapse. Patients with MRD received increased treatment. Pathology, flow cytometry, and ClonoSEQ/Illumina are three diagnostic methods, but these assays depend on the presence of lymphoblasts in the patient samples. Blackburn noted that the remaining lymphoblasts could be below the limit of detection and may not be in the CSF. CronoSEQ is not used for CSF; it is used for bone marrow and blood samples. Blackburn asked: how can we identify disease? They explained that cell-free DNA can be used as a biomarker of cancer progression. To do so, cell-free DNA is analyzed for mutations and methylation patterns. Blackburn and team are looking at VDJ recombination as an approach to identify specific cancer mutations. Leukemia clones can be tracked through barcode sequences. The workflow using Nanopore sequencing for VDJ analysis starts with cell free DNA. cDNA creation, PCR amplification of IGHV, and sequencing. Blackburn shared some data from patients, comparing a patient with MRD to one without. This approach can allow detection of MRD, for example. Cell-free DNA is now used to monitor clones. Blackburn and team are determining the false positive/negative rates and comparing the sensitivity to ClonoSEQ to improve this approach. Importantly, Blackburn is also searching for mechanisms of resistance that may be further studied by tracking clones recalcitrant to current treatments.
