Immunophenotypic Investigation of Minimal Disease Levels in Bclpd Other Than CLL

Assessment of minimal disease in already diagnosed B-CLPD is of utility not only for the evaluation of response to treatment but also, in primary lymphomas, to identify small numbers of neoplastic B-cells in PB, BM, and other body fluids for either diagnostic or staging purposes (25-27,90-92). Until recently, assessment of the quality of remission in B-CLPD using immunophenotyping has been almost restricted to CLL patients and mainly based on monitoring either the K+/X+ B-cell ratio or the number of cells showing CD5 overexpression (91,92). As discussed above in Subheading 2., the balance between k+ and X+ B-lymphocytes is not a specific marker for leukemic B-cells, and its sensitivity is relatively low (10-2-10-3), being limited by the presence of normal B-cells in the sample (3,94). In turn, CD5 overexpression, although a useful marker for the detection of minimal disease levels in CLL (91), can only be applied to a minor subset of other leukemic B-CLPDs (<15%) (25); the same applies for CD79b (93).

Information currently available on the incidence and type of aberrant phenotypes displayed by neoplastic B-cells in CLPD other than CLL is limited to a few reports. In a recent study (25) based on the analysis of 85 B-CLPD other than CLL, we have clearly shown that PB and/or BM B-cells from most patients (99%) display aberrant phenotypes. Serial dilutional experiments demonstrated that, based on these aberrant phenotypes, unequivocal identification of neoplastic B-cells can be achieved even once they are present at frequencies as low as one neoplastic B-cell among 104 to 105 normal PB or BM cells (10-4-10-5) (25) (Fig. 2). Interestingly, distinct patterns of phenotypic aberrations were found in the different diagnostic subgroups. Accordingly, antigen overexpression was the most common aberration in PLL (80%), HCL (100%), and SMZL (79%),

Fig. 2. Representative dilutional experiment of neoplastic B-cells in normal peripheral blood cells. This case corresponds to a hairy cell leukemia in which neoplastic B-cells overexpress CD11c. Aberrant B-cells could be detected at a level of 10-4 (one aberrant B-cell among 10,000 normal PB cells).

whereas in LPL, FL, and MCL the most frequent abnormality consisted of the presence of asynchronous antigen expression: 83, 95, and 100% of the cases, respectively (25). The most frequently overexpressed markers (25) included sIg (60%) and FMC7 (40%) in the context of abnormally high FSC/SSC values (85%) in HCL and FMC7 (57%) in SMZL (25). In turn, the most frequent patterns of asynchronous antigen expression involved CD22-(weak)/CD10-/ CD19+ B-cells found in 67 and 100%, respectively, of the LPL and MCL analyzed; in addition, in MCL, neoplastic B-cells frequently showed coexpression of CD5 on B-cells showing abnormally low levels of CD22 (25). Additional aberrations that have been reported as rather frequent in FL (>80% of cases) include CD10 and bcl2 overexpression on CD20 strongly positive B-cells (4,50,52) and abnormally low CD44 expression on CD38+ clonal B-cells (50).

Despite the apparently high applicability of immunophenotyping for the detection of minimal disease in leukemic B-CLPD other than CLL, few studies have explored its potential clinical utility. Wells et al. (26) have clearly shown that occult B-cell malignancies can be successfully detected by multiparameter flow cytometry in patients with cytopenias, mainly including the diagnosis of HCL and, to a lesser extent, other primary B-cell lymphomas. In a similar way, Subira et al. (90) have demonstrated that multiparameter flow cytometry immunophenotyping is a rather sensitive and specific approach for the detection of central nervous system involvement by B-cell neoplasias. Further studies are still necessary to confirm and potentially extend these observations.

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