Prognostic Value of ZAP-70 Expression Detected by Immunohistochemistry on Bone Marrow Biopsies in Early Phase Chronic Lymphocytic Leukaemia.

New biological prognostic factors have been developed over the last years with the aim of predicting at presentation the heterogeneous clinical course of B chronic lymphocytic leukaemia (B-CLL) and of planning a risk-adapted treatment strategy. Among them ZAP-70 expression on leukemic cells as evaluated by molecular analysis or flow cytometry, initially proposed as surrogate of IgVH mutational status, appears a strong prognostic marker in B-CLL. However the optimal methodological approach to immunophenotipical demonstration of ZAP-70 expression as still matter of debate. We evaluated the cytoplasmic expression of ZAP-70 protein in leukemic cells by immunohistochemical method on bone marrow trephine biopsies taken at diagnosis of 84 patients with B-CLL. We used a mouse anti-human monoclonal antibody to ZAP-70 (clone 2F3.2, 1/200, Upstate, Lake Placid NY) with a polymeric labelling two-step method (Dako cytomation EnVision+, HRP). The results were correlated with age, sex, Binet stage, pattern of bone marrow infiltration, survival and clinical outcome. They were 54 males (64%), aged 34 to 80 years (median 62.5). At presentation 69 (82%) were Binet stage A, 9 (11%) stage B and 6 (7%) stage C. Among the 60 survivors, the median follow-up period from diagnosis was 78.5 months (range 22 – 236 months) Thirty-nine cases (46%) of B-CLL patients had cytoplasmic expression of ZAP-70. This group of patients presented higher percentage of advanced Binet stage (B–C) (p= 0.001). The ZAP-70 positivity was significantly related to inferior OS (55% vs 90% at 7 years)(HR 4.67 CI 1.95–11.14) and PFS (15% vs 57% at 7 years) (HR 5.52 CI 2.57–11.86), confirmed in multivariate analysis. ZAP-70 expression was correlated to poorer outcome also when we evaluated only the 69 stage A patients and the 56 cases with non-diffuse bone marrow infiltration, whereas in patients with diffuse pattern ZAP-70 expression had no prognostic significance. In conclusion, immunohistological detection of ZAP-70 on bone marrow samples at diagnosis appears a useful methodological approach to identify patients with different prognosis in B-CLL.