ABSENT CD33 IN AML PATIENT MIMICS MRD POSITIVE: A SINGLE CASE REPORT

Abstract

Diagnosis and treatment of acute myeloid leukemia (AML) remains difficult to this day, cure rates remain low and measurable residual disease (MRD) assessment is still challenging. Early detection of residual leukemic cells is essential to predict therapeutic success and Multiparametric Flow Cytometry (MFC) has been an effective tool for it. Here, we focus on the importance in caution when interpreting MDR in AML. Bone marrow (BM) sample was assessed to FLT3 and NPM1 mutation, RUNX1/RUNX1T1 and CBFB-MYH11 rearrangements, karyotype (KT) and immunophenotyping. Using an 8-colour antibody panel (CD45, CD34, CD117, CD 33, HLA-DR, CD38, CD123, CD13, CD19, CD7, CD11b, CD4, CD56, CD64, CD11c, CD15, CD133, CD41a and NG2), we applied a gating strategy with fixed gates comparing normal to AML BM, settled to catch LAIPs (Leukemia Associated-Immunophenotypes), abnormal cell maturation pattern and Leukemic Stem Cell, that were detected on diagnosis and evaluated for MRD. Assays were performed at the Hematology Laboratory of the Medical School of Ribeirão Preto, University of São Paulo, in accordance to Local Ethical Boards. A 21-year-old male patient with AML without maturation, diagnosed after 60 days of symptoms, presented anemia (7.9 g/dL), thrombocytopenia (26 000/μL) and white blood cell count of 5710/μL with 73% blasts in peripheral blood and 84% in BM. MFC revealed CD45dim and low side scatter; positive staining for CD117, CD34, HLA-DR, CD38, CD123, CD11c, CD64, CD15 and CD133; negative staining for CD33, CD14, CD7, CD11b, CD4, CD56, CD19, CD41a and NG2. We found no metaphases (KT), no molecular abnormality and he failed risk stratification. Treatment consisted of 2 induction cycles (cycle 1: 3 days of Daunorubicin 60 mg/m2 and 7 days of cytarabine 200 mg/m2; cycle 2: 6 days of cytarabine 1 g/m2 twice a day) and 1 or 2 consolidation cycles (6 days of cytarabine 1 g/m2 twice a day). BM was obtained at diagnosis, 30 and 33 days after 1 st and 2 nd induction, and 3, 9, 12 and 15 months after consolidation of chemotherapy. The positivity of a specific phenotype (CD117+/CD34+/CD33wk/CD13+ ≥ 0.1%) was preserved at diagnosis and follow-ups: 0,506%, 0,681%, 0,402%, 0,101%, 0,158% and 0,122%. Complete remission was achieved by day 30 after 1st induction and no change in the outcome was reported. Absent CD33 was observed in diagnosis and follow-ups, during and after treatment. Post-analytical phase of MFC is the most vulnerable to wrong interpretations. CD33 is a common myeloid antigen expressed on malignant blasts in AML and had been reported as a potential target therapy. CD33 low blasts are associated to a more mature AML and its high expression is related to adverse landscapes, highlighting the importance of CD33 evaluation. Standard care for AML enrolls MRD monitoring and failure to achieve an MRD-negative, CR or detected MRD during or after therapy is associated with relapse or poor outcomes. Still, MRD analysis must embrace a set of standards in order to prevent post-analytic errors. Here we present a case report of AML with absent CD33 maintained from diagnosis to MRD follow-ups, mimicking MRD positivity. This data emphasizes the importance of caution in MFC analysis and correlation of diagnostic and follow-ups results interpretation, as well as constant training of the team.