Diffuse large B-cell lymphoma (DLBCL), the most common subtype of non-Hodgkin's lymphoma, is clinically heterogeneous: 40% of patients respond well to current therapy and have prolonged survival, whereas the remainder succumb to the disease. We proposed that this variability in natural history reflects unrecognized molecular heterogeneity in the tumours. Using DNA microarrays, we have conducted a systematic characterization of gene expression in B-cell malignancies. Here we show that there is diversity in gene expression among the tumours of DLBCL patients, apparently reflecting the variation in tumour proliferation rate, host response and differentiation state of the tumour. We identified two molecularly distinct forms of DLBCL which had gene expression patterns indicative of different stages of B-cell differentiation. One type expressed genes characteristic of germinal centre B cells ('germinal centre B-like DLBCL'); the second type expressed genes normally induced during in vitro activation of peripheral blood B cells ('activated B-like DLBCL'). Patients with germinal centre B-like DLBCL had a significantly better overall survival than those with activated B-like DLBCL. The molecular classification of tumours on the basis of gene expression can thus identify previously undetected and clinically significant subtypes of cancer.

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The presence or absence of somatic mutations in the expressed immunoglobulin heavy chain variable regions (IgVH) of chronic lymphocytic leukemia (CLL) cells provides prognostic information. Patients whose leukemic cells express unmutated IgVH regions (Ig-unmutated CLL) often have progressive disease, whereas patients whose leukemic cells express mutated IgVH regions (Ig-mutated CLL) more often have an indolent disease. Given the difficulty in performing IgVH sequencing in a routine diagnostic laboratory, this prognostic distinction is currently unavailable to most patients. Pilot gene expression profiling studies in patients with CLL identified genes that were differentially expressed between the Ig-unmutated and Ig-mutated CLL subtypes. Here, we have profiled an expanded cohort of 107 patients and show that ZAP-70 is the gene that best distinguishes the CLL subtypes. Ig-unmutated CLL expressed ZAP-70 5.54-fold more highly than Ig-mutated CLL (P < 10 -21 ). ZAP-70 expression correctly predicted IgVH mutation status in 93% of patients. ZAP-70 expression and IgVH mutation status were comparable in their ability to predict time to treatment requirement following diagnosis. In 7 patients, ZAP-70 expression and IgVH mutation status were discordant: 4 Ig-mutated CLLs had high ZAP-70 expression and 3 Igunmutated CLLs had low ZAP-70 expression. Among these ZAP-70 "outliers," those with Ig-mutated CLL had clinical features that are uncharacteristic of this CLL subtype: 2 required early treatment and 2 used a mutated VH3-21 gene, an IgVH gene that has been associated with progressive disease. We developed reverse transcriptase-polymerase chain reaction and immunohistochemical assays for ZAP-70 expression that can be applied clinically and would yield important prognostic information for patients with CLL. IntroductionThe clinical course of chronic lymphocytic leukemia (CLL) is heterogeneous. [1][2][3] Although some patients have an indolent disease without any need for therapeutic interventions, other patients may succumb rapidly despite intensive treatment. There has been an intense search for good prognostic markers in early-stage disease that might facilitate risk-adapted treatment strategies. 4,5 Although cytogenetic abnormalities, especially deletions of 11q and 17p, have been shown to correlate with short survival, 6,7 these changes may not be present in early disease and may only be acquired during disease progression. 8 The presence or absence of somatic mutations in the variable region of the B-cell-receptor heavy chain gene (IgVH) has been shown to distinguish between 2 disease subsets conferring important prognostic information. 9,10 Median survival in patients whose CLL cells express unmutated IgVH genes (Ig-unmutated CLL) ranges between 79 and 119 months. 7,[9][10][11] In contrast, patients whose CLL cells express mutated IgVH genes (Ig-mutated CLL) have a distinctly longer median survival, reaching 293 months in one study, and many may never require treatment. 11 Several lines of evidence indicate that th...