Plasma RNA Is More Reliable Than Peripheral Blood Cells for Monitoring Molecular Response to Imatinib in Patients with Chronic Myeloid Leukemia.
Authors: Ma W. , Tseng R. , Gorre M. , Jilani I. , Keating M. , Kantarjian H. , Cortes J. , O’Brien S. , Giles F. , Albitar M. 
Published: October 2019

Abstract
With the use of imatinib for treatment of chronic myeloid leukemia (CML), monitoring treatment response and quantifying leukemic cells in the body by specific detection of the bcr-abl fusion gene or its mRNA is becoming the standard of care. The high frequency of complete cytogenetic response in CML patients has led to reliance on quantitative reverse-transcription-polymerase chain reaction (qRT-PCR) for detection of the bcr-abl mRNA when monitoring therapy. Peripheral blood (PB) and bone marrow (BM) specimens are usually used for qRT-PCR assays. However, standardization of the qRT-PCR assay and issues regarding sampling and the number of cells that need to be analyzed make reliance on such assays problematic. We previously showed that leukemic cells pour their RNA, DNA, and protein into circulation and, because of their high turnover rate, enrich the plasma with these components. We also hypothesize that plasma mRNA reflects disease activity in the entire body rather than the few cells in the analyzed sample, making plasma a more reliable source than PB cells. In this study we compared qRT-PCR results obtained from PB cells with those obtained from plasma in CML patients being treated with imatinib. PB cell and plasma samples obtained at baseline (n=67) and at 3 (n=43), 6 (n=22), 9 (n=19), and 12 (n=9) months of therapy were compared. The same qRT-PCR assay was used for cell- and plasma-based testing. However, the RNA extraction from plasma was standardized by using equal amounts from all samples and the RNA from 50μL plasma for each qRT-PCR assay. All plasma samples from CML patients were positive at baseline, whereas testing of more than 180 plasma samples from normal individuals or patients with leukemia other than CML showed no detectable bcr-abl transcript. In CML patients, the pattern of changes with therapy in the qRT-PCR in the plasma paralleled that obtained from cell-based testing. At 3 months, all patients who were negative by plasma-based testing were also negative by cell-based testing, whereas 6 of the 14 negative patients by cell-based testing were positive by plasma-based testing. At 6 months of therapy, 8 patients were negative by cell-based testing and all but 1 of these were negative by plasma; this patient was positive by plasma-based testing and became positive by cell-based testing at 9 and 12 months. 10 patients were negative by plasma-based testing at 6 months, 3 of whom were positive by cell-based testing. However, 2 of these 3 became negative by cell-based testing at 9 months and remained negative at 12 months. At 9 months of therapy, 9 patients were negative by cell-based testing, of whom 3 were positive by plasma-based testing; these 3 became positive by cell-based testing at 12 months. 8 patients were negative by plasma-based testing at 9 months, 1 of whom was positive by cell-based qRT-PCR. However, this patient became negative by cell-based testing at 12 months. This data show not only that plasma is a reliable source for testing and monitoring patients with CML, but that it is more reliable than PB cells for monitoring molecular response in CML. Furthermore, because plasma-based test results can be reported as mRNA copies/μL plasma, this platform has the potential to allow better standardization of testing among laboratories.