Flow cytometric detection of peripheral blood involvement by mycosis fungoides and Sezary syndrome using T-cell receptor Vβ chain antibodies and its application in blood staging

Peripheral blood involvement has been recognized as an adverse prognostic factor in patients with mycosis fungoides and Sezary syndrome. However, accurate identification and enumeration of the neoplastic cells in these diseases can be challenging. We assessed the clinical utility of flow cytometric immunophenotypic analysis of T-cell receptor V beta expression in 82 mycosis fungoides and 6 Sezary syndrome patients, with an atypical T-cell immunophenotype, or abnormal CD4:CD8 ratio, identified from peripheral blood specimens of 723 patients submitted for routine mycosis fungoldes/Sezary syndrome blood staging. To improve detection sensitivity, V beta expression was analyzed on gated CD3 + CD4 + T cells or T cells with an aberrant immunophenotype, if present. The flow cytometric results were compared with traditional morphologic assessment (n=88) and molecular methods to assess the T-cell receptor gamma or beta genes (n=41 tested in parallel). Flow cytometric immunophenotyping yielded a clonal V beta pattern in 60/82 mycosis fungoides and 6/6 Sezary syndrome patients. By contrast, flow cytometric V beta was negative in all 10 healthy donors and 18 control patients, showing a specificity of 100% and concordance with molecular testing of 86%. Using flow cytometric V beta results instead of morphologic assessment, 12 patients were upstaged from B1 to B2, and 20 patients from B0 to B1 (P<0.0001). The 12 upstaged B2 patients had no morphologic evidence of involvement, but had an aggressive clinical course similar to those staged by traditional morphologic assessment (median survival 27 vs 41 months, log-rank P=0.701). In 30/44 patients with a tumor-associated V beta expression, a single V beta tube was used to monitor treatment response. In conclusion, flow cytometric V beta analysis is rapid and convenient, can assess T-cell clonality and tumor quantity simultaneously, and is useful both in initial blood staging and monitoring tumor burden during therapy in patients with mycosis fungoides or Sezary syndrome. Modern Pathology (2010) 23, 284-295; doi: 10.1038/modpathol.2009.175; published online 20 November 2009