ALBERT

Description: We studied the expression of CD34 in 169 bone marrow trephines of 53 patients with CML in chronic, accelerated and blastic phases. The bone marrow blast count ranged from 0–82% (median: 4.5%). All were receiving imatinib treatment. We aimed to establish which counting method in bone marrow trephines provided the best correlation with the blast count of the corresponding bone marrow aspirate in all phases of the disease. Paraffin embedded sections were stained with the monoclonal antibody QBEND10. CD34 expression in each marrow trephine was evaluated by three counting methods. The first method, established in our department for several years, expressed the percentage of CD34 positive cells vs. all nucleated cells in a minimum 500-cell count (% CD34+/500 cells). The second recorded the total number of CD34 positive cells in ten high power fields (CD34+/10hpf) using x 40 magnification /numerical aperture 0.65. The third method evaluated the highest number of CD34 positive cells present in a single high power field (CD34+/hpf), this value being taken from the previous 10 hpf count. The two latter methods were employed because they may be more applicable for general use by practicing histopathologists. In addition, counting over 10 hpf provides a much larger sample size than a 500-cell count. CD34 expression in endothelial cells served as a positive internal control. All three methods showed significant positive correlation with blast count (% CD34+/500 cells: Pearson’s Correlation - 0.723, sig -

Description: CD34 is a surface glycophosphoprotein expressed on developmentally early lymphohematopoietic stem and progenitor cells, small-vessel endothelial cells, and embryonic fibroblasts. We studied the prognostic value of presence of CD34 expressing cells in the marrow trephine biopsies of 58 patients with chronic myeloid leukemia in chronic phase treated with imatinib (21 newly diagnosed and 37 IFN failures). Briefly, patients received 400 mg daily and the dose was then adjusted according to tolerance and response. Bone marrow examinations were performed before starting therapy with imatinib and then every 3–6 months. All patients achieved complete haematological remission and 48% achieved complete cytogenetic remission. Chronic, accelerated and blastic phases were defined as described by Kantarjian 1988. Paraffin embedded sections were stained with the monoclonal antibody QBEND10. A minimum of 500 nucleated cells were counted per trephine section. Results were expressed as percentage of CD34+ vs all nucleated marrow cells. The expression of CD34 by the endothelial cells was used as an internal control. The median CD34 percentage before the onset of the imatinib therapy was 0.8% (25 and 75 percentiles 0.8 and 2%). We performed univariate and multivariate analysis for PFS with variables defined at the time of starting imatinib. Sokal risk group (defined at diagnosis), presence or absence of additional cytogenetic abnormalities, percentage of blasts in the bone marrow aspirate and the percentage of CD34+ cells in the trephine (considered as a continuos variable) were significant in the univariate analysis. Only CD34 expression and Sokal risk group were significant in the multivariate model. Sokal risk group lost significance when the percentage of CD34 cells was categorized as 2% (RR for PFS=11.7, 95 CI=1.9–70.4, p=0.007; RR for survival = 10.5, 95CI= 1.01–80.3, p=0.05). The 3 years PFS for patients with a CD34 percentage 2% (n=14) it was 36% (p=0.006), similarly the three year overall-survival was 80 and 45% respectively (p=0.009). The median value of CD34 in the first bone marrow examination performed after the start of imatinib (typically performed at 3–6 months) was 0.8 (25 and 75 percentiles 0.8 and 1.5%). The degree of reduction from the baseline level achieved in the first bone marrow examination had favourable prognostic significance (as a continuos variable) with a RR for PFS of 0.88, 95CI 0.53–0.93 (p= 0.006) and RR for overall survival of 0.93, 95CI 0.19–0.998 (p=0.05). During follow up, an increment of 2 percentage points over the pre therapy CD34 value in patients in otherwise complete haematological remission had an adverse prognostic implication with a RR for PFS of 10.2 95 CI 2.1–17, (p=0.006) and a RR for survival of 6.8 95 CI 1.01–101-(p=0.05). We conclude that the use of CD34 staining in bone marrow trephines is a valuable tool for both diagnosis and monitoring in patients with CML treated with imatinib.