Flow Cytometry in the Diagnosis of Acute Promyelocytic Leukemia (APL) – What We Learn from Outliers
Weijie Li, John Pizzolo, Rasel Ahsan, Linda Cannizzaro, KH Ramesh, Howard Ratech, Christine McMahon. Albert Einstein College of Medicine Montefiore Medical Center, Bronx, NY
Background: APL is a hematologic emergency, requiring rapid diagnosis. The diagnosis rests on evidence of a retinoic acid receptor alpha (RARA) gene rearrangement, but flow cytometry (FC) serves as a rapidly accessible surrogate, in conjunction with morphology. Classic APL features include positivity for CD117 and myeloperoxidase (MPO), and negativity for CD34 and HLA-DR. Recently, absence of CD11b and CD11c was reported to be specific and sensitive for APL. We evaluated the performance of these markers and characterized exceptions to the rule.
Design: Six color FC data for all acute myeloid leukemias (AML) diagnosed at Montefiore between 5/2011 and 9/2012 and were correlated with morphology and cytogenetics, including FISH studies with a RARA break-apart probe. Individual antigens and their combinations were analyzed with respect to positive predictive value (PPV) and negative predictive value (NPV) in identifying APL.
Results: Of 36 AML cases, 12 were APL and 24 were non-APL, based on FISH results. Many APL cases were HLA-DR- (75%) and CD34- (67%). Among non-APL cases, 13% were HLA-DR-, 25% were CD34-, and 8% were negative for both. All APL cases were positive for CD117, CD33 and myeloperoxidase (MPO). Most APL cases were CD11b- (95%) and CD11c- (85%). Among non-APL cases, 79% were CD11b-, 42% were CD11c-, and 38%were negative for both. Two non-APL cases (5%) had an APL-like immunophenotype (CD117+/MPO+/CD11b/c-/CD34-/HLA-DR-); one of these had APL-like morphologic features. Clinical correlation revealed that the two CD11b/c+ APL samples came from patients who had been presumptively treated with ATRA. One patient had FC twice- before and then 16 hours after receiving ATRA. CD11b and CD11c were detected in the post-ATRA sample only. Excluding patients exposed to ATRA, 100% of APL cases were CD11b/c-/CD117+/MPO+.
Conclusions: The CD11b/c-/CD34-/HLA-DR-/CD117+/MPO+ phenotype is quite specific for APL. We identified two outlier cases that lack cytogenetic evidence of RARA disruption, one of which morphologically resembles APL. The CD11b/c- phenotype is a highly sensitive predictor of APL, but not in patients exposed to ATRA. This is likely due to induction of CD11b and CD11c (differentiation markers) by ATRA, as previously shown in vitro. To our knowledge, this is the first report of CD11b/c being induced by ATRA on APL leukemia cells in a patient.
|APL-Associated features||Sensitivity (%)||Specificity (%)||PPV (%)||NPV (%)|