Long-Term Immune Reconstitution Following Stem Cell Transplantation in Chronic Lymphocytic Leukemia.

Abstract 4649

Chronic lymphocytic leukemia (CLL) is characterized by the accumulation of monoclonal CD5+ B lymphocytes. In addition, immune disturbances are commonly present. Although CLL is still considered incurable, long term remissions can be observed after allogeneic stem cell transplantation (SCT). Whether immune function is restored in patients responding to treatment is largely unknown. We report on the immune status in 19 patients with CLL in long-lasting complete remission (CR) after SCT (13 allogeneic, 6 autologous). Median age was 51 (range, 33- 64) and median follow-up since transplantation was 6 years (range, 2-17). Three patients had chronic graft-versus host disease at sample collection, two of whom were receiving immunosuppression. Lymphocyte subsets were studied using multiparameter flow cytometry and compared to healthy controls. We quantified immunoglobulin subtypes, complement proteins, and β₂-microglobulin (B2M) by standard techniques and IL-10 and VEGF by using flow-based cytometric bead array technology. CD8+ T cell response to CMV was assessed using a pentameric HLA-A2 binding CMV pp65-derived peptide. Three patients (all following allogeneic SCT) had detectable residual CLL cells (> 10^-4) in peripheral blood at the time of the analysis. In the remaining 16 patients with no detectable minimal residual disease (MRD) normal CD19+CD5- and CD19+CD5+ B cell populations were lower than in healthy individuals (10.5% vs. 14.5%; p=0.02 and 1.6% vs. 3.9 %; p=0.002 respectively). A significant increase in the proportion of CD8+ T cells (median 28.1% vs. 18.7%, p=0.03), particularly those with a chronically activated phenotype CD3+CD8+DR+ (10.2% of CD3+ cells vs. 4.9% in controls), was observed (p=0.013). The nine CMV+/HLA-A2 patients all showed specific cytotoxic CD8+ T cells which exhibit predominantly a CD45RA+CD27- phenotype, being better preserved in autologous SCT than in allogeneic SCT patients. Also, higher numbers of CD8+CD45RA+CD27- T cells were observed in patients with a longer follow-up. CD4+ T cell count was < 400/mm³ in 4 patients. An abnormal CD4:CD8 ratio was seen in 7 out of 19 patients. Interestingly, a significant increase of double positive CD4 and CD8 T cells was detected in most patients comprising 2.6% vs.1.6% of lymphocytes in normal subjects (p=0.02). There were no quantitative abnormalities in CD3-CD56+ cells. Not surprisingly, hypogammaglobulinemia was present in all but four patients immediately prior to transplant. Whereas IgM levels normalized in all patients, 4 and 6 patients respectively still had low IgG and IgA levels more than two years after transplantation. Of note, low serum immunoglobulin levels were seen in 6 out of 14 MRD negative-CR patients and all three MRD positive-CR patients, including one who had normal immunoglobulin levels at the time of transplant. Complement proteins C3 and C4 were within the normal range in all cases. The direct Coombs test (DCT) was also negative in all patients although one patient had indirect signs of hemolysis. Regarding serum markers, B2M was increased (>2.5mg/dl) in 5 out of 19 patients. No significant differences were found in IL-10 and VEGF levels between patients and normal controls (median levels, 2.14 vs. 1.06 pg/ml and 156.88 vs. 104.14 pg/ml), but there were 2 patients with markedly elevated IL-10. In summary, these data demonstrate that immune defects persist over time in CLL patients with a long lasting CR (including MRD-negative CRs) after SCT.