Combined use of reverse transcriptase polymerase chain reaction and flow cytometry to study minimal residual disease in Philadelphia positive acute lymphoblastic leukemia

Background and Objectives. The Philadelphia chromosome in acute lymphoblast ic leukemia (Ph+ ALL) is associated with a poor prognosis given the high fr equency of chemoresistance and leukemia relapse. Minimal residual disease ( MRD) detection before cytogenetic and hematologic relapse could be useful i n early therapy. The most suitable methods for detecting MRD in Ph+ ALL are now cytometry (FC) and reverse transcriptase polymerase chain reaction (RT -PCR). However, since both techniques carry the risk of false-negative resu lts the combined use of these two techniques could overcome this problem. Design and Methods. We report our experience using this approach in 47 bone marrow samples obtained from 10 Ph+ ALL patients. Twenty-seven marrow aspi rates were taken from patients in clinical remission (CR). The samples were considered positive for MRD by FC when two conditions were met: 1) detecti on of an abnormal B-cell differentiation pattern and 2) presence of more th an 1x10(-3) cells coexpressing CD22/CD34/CD45 or CD66/CD34/CD10. After FC a nalysis, RNA was purified using standard methods. Results. FC was positive in 23/27 samples in CR (sensitivity 85%). RT-PCR w as successfully performed in 23 samples in CR. RT-PCR was positive in 18/23 samples (sensitivity 78%). There were 5 samples with discordant results. F C was positive in 3 samples with a negative RT-PCR and FC was negative in 2 samples with a positive RT. All the 10 patients relapsed and only 1 is cur rently alive after an allogeneic stem cell transplantation. The median (ran ge) time from MRD detection to relapse in patients treated with chemotherap y was 42 (39-71) days. Interpretation and Conclusions. These data suggest that RT-PCR may be negat ive despite the presence of neoplastic cells identified by their immunophen otypic traits. We conclude that immunologic and molecular techniques can be used in tandem for monitoring MRD in Ph+ ALL. (C) 2000, Ferrata Storti Fou ndation.