Peripheral Blood Monoclonal B-Cell and/or Plasma Cell Detection By Flow Cytometry in Screening and Monitoring EBV-Positive Post-Transplant Lymphoproliferative Disorders

Background Post-transplant lymphoproliferative disorder (PTLD) is a serious complication that can occur following an allogenic hematopoietic stem cell transplant (allo-HSCT). PTLD occurs in approximately 0.8% to 20% of patients following an allo-HSCT and is associated with Epstein-Barr virus (EBV) infection in about 60% to 80% of patients. EBV positive (EBV+) PTLD generally arises early, several months following transplantation. The routine methods to diagnose EBV+ PTLD are clinical symptoms, EBV copy number, imageological examination, and pathological diagnosis, of which pathological diagnosis is the gold standard. Yet obtaining a biopsy is not possible with some patients and is not applicable for post-therapy monitoring. Therefore, there is an urgent need to find a simple, highly efficient, feasible, sensitive, and specific diagnostic and monitoring tool. Flow cytometry (FCM) has been established as a highly cost-effective method to diagnose lymphoma over the past several decades, and particularly for screening monoclonal B and/or plasma cells (MC B/P). Several recent publications as well as our own published and unpublished data have found that MC B/P are present in the peripheral blood of most PTLD patients. To this end, FCM detection of MC B/P in PB is a promising screening and monitoring method for EBV(+) PTLD. Objective To investigate the effectiveness of detecting MC B/P in PB by FCM for EBV+ PTLD screening and monitoring. Methods A total of 1470 patients received allo-HSCT at the Hebei Yanda Ludaopei Hospital, China from January 2018 through December 2019. We conducted a retrospective study of 481 patients with fever and lymphadenopathy following allo-HSCT. Patient PB was extracted for FCM MC B/P analysis. Plasma EBV viral loads were detected by PCR. The median fellow-up time was 6 months (range: 2 days to 21 months). The relationships of PB MC B/P, EBV load and clinical EBV-associated PTLD symptoms were investigated. Results: MC B/Ps were detected in the PB of 47 patients, of which 29 cases had monoclonal B cells, 14 cases had detectable co-existence of monoclonal B cells and monoclonal plasma cells, and 4 cases had detectable monoclonal plasma cells. The median time to PTLD onset following allo-HSCT was 70 days (range: 33 days to 491 days). The median percentage of monoclonal B cells was 0.43% (range: 0.1% to 23.41%. The median percentage of monoclonal plasma cells was 0.25%). Forty of 47 patients (85.1%) were finally diagnosed with PTLD using clinical comprehensive examination of symptoms. The incidence of clinical EBV+ PTLD was 2.9% (44 of 1470 cases). By FCM, MC B/Ps were observed in the PB of 91% of the patients (40 of 44 cases). No MC B/Ps were found in the PB of 4 patients. MC B/P detection in PB by FCM screening could effectively predict the incidence of EBV+ PTLD with a sensitivity of 90.91% and specificity of 98.40%. The positive predictive value was 85.17% and the negative predictive value was 99.08%. There was no significant correlation between EBV viral copy number and percentage of MC B/P. The median fellow-up time was 6 months (range: 2 days to 21 months) for all patients. The therapeutic response rate was 87.5% (35 of 40 patients) for MC B/P positive EBV+ PTLD patients. In those patients with a therapeutic response, clinical symptoms improved, lymph nodes significantly shrank, EBV viral loads decreased, MC B/Ps decreased or vanished, and CD20+ cell proportion decreased or vanished. Among the 7 patients who had MC B/Ps in PB but were not diagnosed with PTLD , their clinical symptoms recovered after treatment with immunosuppressive therapy. Conclusion By detecting PB MC B/Ps using FCM, we can successfully screen for EBV+ PTLD and monitor patient therapeutic responses in most cases. FCM to detect MC B/Ps in PB is a new, promising method for the diagnosis and monitoring of EBV+ PTLD. Compared to a biopsy, this appears to be a more simple, easier and more applicable tool to diagnose and monitor EBV+ PTLD.