Clinical significance of DNA methylation in chronic lymphocytic leukemia patients: results from 3 UK clinical trials.

Chronic lymphocytic leukemia patients with mutated immunoglobulin heavy-chain genes (IGHV-M), particularly those lacking poor-risk genomic lesions, often respond well to chemoimmunotherapy (CIT). DNA methylation profiling can subdivide early-stage patients into naive B-cell-like CLL (n-CLL), memory B-cell-like CLL (m-CLL), and intermediate CLL (i-CLL), with differing times to first treatment and overall survival. However, whether DNA methylation can identify patients destined to respond favorably to CIT has not been ascertained. We classified treatment-naive patients (n = 605) from 3 UK chemo and CIT clinical trials into the 3 epigenetic subgroups, using pyrosequencing and microarray analysis, and performed expansive survival analysis. The n-CLL, i-CLL, and m-CLL signatures were found in 80% (n = 245/305), 17% (53/305), and 2% (7/305) of IGHV-unmutated (IGHV-U) cases, respectively, and in 9%, (19/216), 50% (108/216), and 41% (89/216) of IGHV-M cases, respectively. Multivariate Cox proportional analysis identified m-CLL as an independent prognostic factor for overall survival (hazard ratio [HR], 0.46; 95% confidence interval [CI], 0.24-0.87; P = .018) in CLL4, and for progression-free survival (HR, 0.25; 95% CI, 0.10-0.57; P = .002) in ARCTIC and ADMIRE patients. The analysis of epigenetic subgroups in patients entered into 3 first-line UK CLL trials identifies m-CLL as an independent marker of prolonged survival and may aid in the identification of patients destined to demonstrate prolonged survival after CIT.

Cell surface phenotype profiles distinguish stable and progressive chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL) is clinically heterogeneous. While some patients have indolent disease for many years, 20-30% will progress and ultimately die of their disease. CLL may be classified by the Rai or Binet staging system, mutational status of the immunoglobulin variable heavy-chain gene (IGVH), ZAP-70 overexpression, cytogenetic abnormalities (13q-, + 12, 11q-, 17p-) and expression of several cell surface antigens (CD38, CD49d) that correlate with risk of disease progression. However, none of these markers identify all cases of CLL at risk. In a recent review, we summarized those CD antigens known to correlate with the prognosis of CLL. The present study has identified surface profiles of CD antigens that distinguish clinically progressive CLL from slow-progressive and stable CLL. Using an extended DotScan(™) CLL antibody microarray (Version 3; 182 CD antibodies), and with refined analysis of purified CD19 + B-cells, the following 27 CD antigens were differentially abundant for progressive CLL: CD11a, CD11b, CD11c, CD18, CD19, CD20 (two epitopes), CD21, CD22, CD23, CD24, CD25, CD38, CD40, CD43, CD45, CD45RA, CD52, CD69, CD81, CD84, CD98, CD102, CD148, CD180, CD196 and CD270. The extensive surface profiles obtained provide disease signatures with an accuracy of 79.2%, a sensitivity of 83.9% and a specificity of 72.5% that could provide the basis for a rapid test to triage patients with CLL according to probability of clinical progression and potential earlier requirement for treatment.

Defining the prognosis of early stage chronic lymphocytic leukaemia patients.

Approximately 70% of chronic lymphocytic leukaemia (CLL) patients present with early stage disease, therefore defining which patients will progress and require treatment is a major clinical challenge. Here, we present the largest study of prognostic markers ever carried out in Binet stage A patients (n = 1154) with a median follow-up of 8 years. We assessed the prognostic impact of lymphocyte doubling time (LDT), immunoglobulin gene (IGHV) mutation status, CD38 expression, ZAP-70 expression and fluorescence in situ hybridization (FISH) cytogenetics with regards to time to first treatment (TTFT) and overall survival (OS). Univariate analysis revealed LDT as the most prognostic parameter for TTFT, with IGHV mutation status most prognostic for OS. CD38 expression, ZAP-70 expression and FISH were also prognostic variables; combinations of these markers increased prognostic power in concordant cases. Multivariate analysis revealed that only LDT, IGHV mutation status, CD38 and age at diagnosis were independent prognostic variables for TTFT and OS. Therefore, IGHV mutation status and CD38 expression have independent prognostic value in early stage CLL and should be performed as part of the routine diagnostic workup. ZAP-70 expression and FISH were not independent prognostic markers in early stage disease and can be omitted at diagnosis but FISH analysis should be undertaken at disease progression to direct treatment strategy.

Determination of how many immunoglobulin variable region heavy chain mutations are allowable in unmutated chronic lymphocytic leukaemia - long-term follow up of patients with different percentages of mutations.

The choice of 98% sequence homology for immunoglobulin heavy chains to distinguish between mutated and unmutated versions of chronic lymphocytic leukaemia (CLL) was arbitrary and was chosen to account for supposed polymorphisms. Some authors chose 97% or even 95%. This study examined survival curves for cohorts of patients with varying degrees of sequence homology. All patients with <97% homology behaved as if mutated. Those with 97-98% homology were more aggressive than the mutated cases, but less aggressive than those with >98% homology.

Multivariate analysis of prognostic factors in CLL: clinical stage, IGVH gene mutational status, and loss or mutation of the p53 gene are independent prognostic factors.

This study evaluates the prognostic significance of genetic abnormalities (detected at or shortly after presentation), clinical stage, lymphocyte morphology, CD38 expression, and IGVH gene status in 205 patients with chronic lymphocytic leukemia (B-CLL). Deletion of chromosome 11q23, absence of a deletion of chromosome 13q14, atypical lymphocyte morphology, and more than 30% CD38 expression are significantly associated with the presence of unmutated IGVH genes. Advanced stage, male sex, atypical morphology, more than 30% CD38 expression, trisomy 12, deletion of chromosome 11q23, loss or mutation of the p53 gene, and unmutated IGVH genes are all poor prognostic factors in a univariate analysis. However, only 98% or more homology of IGVH genes to the germline sequence, loss or mutation of the p53 gene, and clinical stage retain prognostic significance in a multivariate analysis. The median survival of patients with mutated IGVH genes, unmutated IGVH genes, and loss or mutation of the p53 gene regardless of IGVH gene status is 310, 119, and 47 months, respectively. These data should facilitate the design of new trials for the management of patients presenting with advanced disease or poor prognosis early stage disease.