A Prospective Study of Young Chronic Lymphocytic Leukemia (CLL) Patients Investigated at Diagnosis: Distribution and Clinical Significance of Prognostic Factors

In chronic lymphocytic leukemia (CLL) the distribution and prognostic impact of genetic and molecular markers has been assessed on retrospective series of patients in different phases of the disease. Our aim was to assess the distribution and clinical significance of a comprehensive panel of clinical and biologic parameters prospectively evaluated at presentation in all young patients diagnosed with CLL at our Institution, taking advantage of the fact that in Italy individuals with a lymphocytosis are referred to the hematologist for the diagnostic work-up. From November 2002 to June 2008, 105 young CLL patients (<60 years-old) were diagnosed with CLL and included in this study. There were 56 males and 49 females, with a median age of 52 years-old. 81% were in Binet stage A, 19% in stages B/C. Rai stage 0 was recorded in 63% of patients, I/II in 31%, III/IV in 6%. The median white blood cell (WBC) count was 18.8 × 109/L (range 5.8–236.6). Prognosis was evaluated as the time from diagnosis to first treatment (TFI, treatment-free interval), since only 3 patients died after a median follow-up of 32.4 months (range 1 to 88). The median TFI was 43.9 months. Clinical features included: gender, WBC count, Binet and Rai stage. Serological and biologic parameters included: beta2-microglobulin, LDH, IgG immunoglobulin levels, lymphocyte morphology, T-cell subsets, IgVH mutational status, CD38 and ZAP-70 expression, cytogenetic abnormalities evaluated by FISH, p53 protein expression and p53 gene sequencing (exon 5 to 8). The distribution of the prognostic markers is summarized in the table. Raised beta2-microglobulin and LDH were present only in 5% and 15% of cases, respectively. The CD4/CD8 ratio was normal in almost all cases. The proportion of unmutated IgVH, CD38 and ZAP-70 positive cases was about one third of the cohort of CLL patients at diagnosis. The incidence of del(17p) (cut off >20% cells) and del(11q) (cut off >10% cells) was 2% and 7%, respectively. Patients with del(17p) or del(11q) exclusively showed unmutated IgVH and ZAP-70+, and were mostly CD38+. p53 mutations were present in 4 cases, 3 with unmutated IgVH and del(17p) and 1 with mutated IgVH and no del(17p). In univariate analysis, the following variables resulted associated to a short TFI: advanced stage Binet B/C and Rai I/IV (<0.0001), WBC count (<0.0001), proportion of CD3+ cells <16% (0.0002), raised beta2-microglobulin (<0.0001) and LDH (<0.0001), unmutated IgVH (<0.0001), CD38+ (<0.0001), ZAP-70+ (<0.0018), adverse cytogenetic abnormalities (del(17p), del(11q), +12) (<0.0001). Atypical CLL morphology showed a trend for significance (0.06). Multivariate analysis on TFI - including WBC count (as continuous variable), CD3 %, LDH, IgVH mutation status, ZAP-70 and CD38 expression and corrected with interaction between WBC and IgVH status - was focused on the 84 patients with Binet stage A. High WBC count, raised LDH, unmutated IgVH resulted as unfavorable prognostic factors, whilst the proportion of CD3+ cells was associated with a better outcome. Neither ZAP-70 or CD38 showed an independent prognostic value. In CLL cases with discordant expression of ZAP-70 and IgVH mutation status (25% of cases), the latter appeared to be more relevant than ZAP-70 in determining the TFI. In conclusion, unmutated IgVH, raised LDH, WBC count and a low proportion of CD3+ cells at diagnosis are significant predictors of TFI in early stage CLL. This group represents about one third of young patients at diagnosis. Adverse FISH abnormalities are present only in a small subgroup of cases in the early phases of the disease. Young CLL patients at diagnosis N° of cases % Raised beta2-microglobulin (>3400 ng/l) 5/102 5% Raised LDH 16/105 15% Hypo IgG 21/98 21% Atypical morphology 27/104 26% CD3+ cells (<16%) 60/105 57% CD4/CD8 (<1) 3/101 3% IgVH mutated (≥98%) 67/103 65% IgVH unmutated (<98%) 36/103 35% CD38 ≥7% 25/104 24% ZAP-70 ≥10% 39/100 39% Del(17p) >20% 2/104 2% Del(11q) >10% 7/104 7% +12 >5% 7/104 7% Del(13q) >5% isolated 59/104 57% Normal (none of the above) 29/104 28% p53 protein expression 3/100 3% p53 gene mutation 4/105 4%