Nodal mantle cell lymphoma (MCL) is a well-defined entity, but non-nodal leukemic cyclin D1 positive lymphoproliferative disorders have been reported and their relationship with MCL remains controversial and their prognosis heterogeneous. We prospectively studied the expression of cyclin D1 in CD5 positive leukemic B lymphoproliferative disorders at diagnosis and identified 65 cases overexpressing cyclin D1. We did not distinguish any clinical or biological criteria allowing one to identify a non-MCL group. Multivariate analysis identified age, anemia and p27kip1 expression as independent prognostic factors of survival. By univariate analysis, p27kip1 high expression proved to be the strongest predictor of prolonged survival. The median survival of p27 low expressors was 30 months, while it was not reached for p27 high expressors. A high level of p27 expression was often found associated with the absence of nodal involvement and the presence of somatic mutations, but neither of them was restricted to the p27 high expression group. In conclusion, we hypothesize that MCL and these cyclin D1 positive leukemic lymphoproliferative disorders represent a continuous spectrum of diseases. Determination of p27 expression level appears as a routine applicable test allowing identification of a subset of patients who could be considered for different therapeutic approaches.

Recent gene profiling studies demonstrated that signalisation through the BCR plays a central role during CLL disease progression. However, incidence of BCR engagement on cell survival remains unclear as both increased and decreased apoptosis have been reported after BCR ligation by anti-mu antibodies. We investigated the effect of anti-mu exposure on fresh cells from 32 untreated CLL cases. Cell viability was assessed both by MTT assay and Annexin V/ IP labelling. Culture conditions were set after time and dose experiments. A 25% increase of viability, obtained by MTT assay after 72h of culture, was considered as significant. When a coated rabbit anti-human IgH mu specific antibody was used, it significantly prevented apoptosis in 20/32 cases (named responders) and never increased it. On the same CLL cases, soluble goat F(ab')2 anti-human IgH mu specific antibody increased apoptosis in all but one case. However, coating of this latter antibody to the culture plate cancelled the pro-apoptotic effect and restored cell survival promoting effect in 13/14 cases tested. Unspecific effects were ruled out using control antibodies. Cytometry experiments and confocal microscopy demonstrated that uncoated antibody was rapidly internalized and degraded. In responders cases, enhanced survival was supported by increased expression of early cell cycle protein cyclin D2 and cdk4. There was no subsequent cell cycle progression as shown by IP staining and absence of pRB phosphorylation most likely related to the lack of p27 down regulation. There was a strong correlation between the extent of apoptosis prevention after anti-mu exposure and disease progression. Indeed, the 12 over 32 cases who were found to have no IgVH somatic hypermutations, happened to be all responders. Conversely among the 20 mutated cases, 12/20 were non responders and had a clinically stable disease. As for the 8 responders with somatic mutations, four had detectable levels of ZAP 70 protein and 2 others had a clinically progressive disease. In conclusion, coated anti-mu antibody is likely to mimic in vivo conditions of stimulation, suggesting that BCR ligation could play a major role in the pathophysiology of the progressive disease, giving a survival advantage to these CLL cells

In B-CLL IgV(H) genes mutational status is a major prognostic factor. Since sequencing of IgV(H) genes is not available in most laboratories, an easily performed surrogate assay is desirable. To identify the best surrogate assay, and to better discriminate prognostic subgroups we analyzed clinical and biological data from 58 typical CLL cases. A higher serum thymidine kinase level (>15 U/l) proved to be a strong predictor of mutational status, and the only independent one among the studied parameters. To further identify prognostic subgroups, cluster analysis was employed on 38 cases on which all data were available, which segregated two groups including 25 and 13 patients, respectively. These two clusters differed by their proliferative potential and appeared to discriminate patients with very different clinical course and outcome. s-TK was strikingly different among these two clusters, suggesting that s-TK level could be used routinely to identify patients at risk of progression.

BACKGROUNDWithin a period of three years, we identified 13 patients in whom pure red-cell aplasia developed during treatment with recombinant human erythropoietin (epoetin). We investigated whether there was an immunologic basis for the anemia in these patients.METHODSSerum samples from the 13 patients with pure red-cell aplasia were tested for neutralizing antibodies that could inhibit erythroid-colony formation by normal bone marrow cells in vitro. The presence of antierythropoietin antibodies was identified by means of binding assays with the use of radiolabeled intact, deglycosylated, or denatured epoetin.RESULTSSerum from all 13 patients blocked the formation of erythroid colonies by normal bone marrow cells. The inhibition was reversed by epoetin. Antibodies from 12 of the 13 patients bound only conformational epitopes in the protein moiety of epoetin; serum from the remaining patient bound to both conformational and linear epitopes in erythropoietin. In all the patients, the antibody titer slowly decreased after the discontinuation of treatment with epoetin.CONCLUSIONSNeutralizing antierythropoietin antibodies and pure red-cell aplasia can develop in patients with the anemia of chronic renal failure during treatment with epoetin.