Although +12 chronic lymphocytic leukemia comprises about 20% of cases, relatively little is known about its pathophysiology. These cases often demonstrate atypical morphologic and immunophenotypic ...features, high proliferative rates, unmutated immunoglobulin heavy chain variable region genes, and a high frequency of NOTCH1 mutation. Patients with +12 chronic lymphocytic leukemia have an intermediate prognosis, and show higher incidences of thrombocytopenia, Richter's transformation, and other second cancers. Despite these important differences, relatively few transcriptional profiling studies have focused on identifying dysregulated pathways that characterize +12 chronic lymphocytic leukemia, and most have used a hierarchical cytogenetic classification in which cases with more than one recurrent abnormality are categorized according to the abnormality with the poorest prognosis. In this study, we sought to identity protein-coding genes whose expression contributes to the unique pathophysiology of +12 chronic lymphocytic leukemia. To exclude the likely confounding effects of multiple cytogenetic abnormalities on gene expression, our +12 patient cohort had +12 as the sole abnormality. We profiled samples obtained from 147 treatment-naive patients. We compared cases with +12 as the sole cytogenetic abnormality to cases with sole del(13q), del(11q), or diploid cytogenetics using independent discovery (n=97) and validation (n=50) sets. We demonstrate that chronic lymphocytic leukemia cases with +12 as the sole abnormality express a unique set of activated pathways compared to other cytogenetic subtypes. Among these pathways, we identify the NFAT signaling pathways and the immune checkpoint molecule, NT5E (CD73), which may represent new therapeutic targets.
Aberrant expression of microRNAs (miRNAs) has been associated with clinical outcome in patients with chronic lymphocytic leukemia (CLL). To identify a powerful and easily assessable miRNA bio-marker ...of prognosis and survival, we performed quantitative reverse-transcription polymerase chain reaction (qRT-PCR) profiling in 104 CLL patients with a well-defined chromosome 17p status, and we validated our findings with miRNA microarray data from an independent cohort of 80 patients. We found that miR-15a, miR-21, miR-34a, miR-155, and miR-181b were differentially expressed between CLLs with chromosome 17p deletion and CLLs with normal 17p and normal karyotype, and that miR-181b was down-regulated in therapy-refractory cases. miR-21 expression levels were significantly higher in patients with poor prognosis and predicted overall survival (OS), and miR-181b expression levels significantly predicted treatment-free survival. We developed a 21FK score (miR-21 qRT-PCR, fluorescence in situ hybridization, Karyotype) to stratify patients according to OS and found that patients with a low score had a significantly longer OS time. When we evaluated the relative power of the 21FK score with the most used prognostic factors, the score was the most significant in both CLL cohorts. We conclude that the 21FK score represents a useful tool for distinguishing between good-prognosis and poor-prognosis CLL patients.
Fludarabine, cyclophosphamide, and rituximab (FCR) has become a gold-standard chemoimmunotherapy regimen for patients with chronic lymphocytic leukaemia. However, the question remains of how to treat ...treatment-naive patients with IGHV-unmutated chronic lymphocytic leukaemia. We therefore aimed to develop and validate a gene expression signature to identify which of these patients are likely to achieve durable remissions with FCR chemoimmunotherapy.
We did a retrospective cohort study in two cohorts of treatment-naive patients (aged ≥18 years) with chronic lymphocytic leukaemia. The discovery and training cohort consisted of peripheral blood samples collected from patients treated at the University of Texas MD Anderson Cancer Center (Houston, TX, USA), who fulfilled the diagnostic criteria of the International Workshop on Chronic Lymphocytic Leukemia, had received at least three cycles of FCR chemoimmunotherapy, and had been treated between Oct 10, 2000, and Oct 26, 2006 (ie, the MDACC cohort). We did transcriptional profiling on samples obtained from the MDACC cohort to identify genes associated with time to progression. We did univariate Cox proportional hazards analyses and used significant genes to cluster IGHV-unmutated samples into two groups (intermediate prognosis and unfavourable prognosis). After using cross-validation to assess robustness, we applied the Lasso method to standardise the gene expression values to find a minimum gene signature. We validated this signature in an external cohort of treatment-naive patients with IGHV-unmutated chronic lymphocytic leukaemia enrolled on the CLL8 trial of the German Chronic Lymphocytic Leukaemia Study Group who were treated between July 21, 2003, and April 4, 2006 (ie, the CLL8 cohort).
The MDACC cohort consisted of 101 patients and the CLL8 cohort consisted of 109 patients. Using the MDACC cohort, we identified and developed a 17-gene expression signature that distinguished IGHV-unmutated patients who were likely to achieve a long-term remission following front-line FCR chemoimmunotherapy from those who might benefit from alternative front-line regimens (hazard ratio 3·83, 95% CI 1·94–7·59; p<0·0001). We validated this gene signature in the CLL8 cohort; patients with an unfavourable prognosis versus those with an intermediate prognosis had a cause-specific hazard ratio of 1·90 (95% CI 1·18–3·06; p=0·008). Median time to progression was 39 months (IQR 22–69) for those with an unfavourable prognosis compared with 59 months (28–84) for those with an intermediate prognosis.
We have developed a robust, reproducible 17-gene signature that identifies a subset of treatment-naive patients with IGHV-unmutated chronic lymphocytic leukaemia who might substantially benefit from treatment with FCR chemoimmunotherapy. We recommend testing the value of this gene signature in a prospective study that compares FCR treatment with newer alternative therapies as part of a randomised clinical trial.
Chronic Lymphocytic Leukaemia Global Research Foundation and the National Institutes of Health/National Cancer Institute.
Clonal heterogeneity is common in many types of cancer, including chronic lymphocytic leukemia (CLL). Previous research suggests that the presence of multiple distinct cancer clones is associated ...with clinical outcome. Detection of clonal heterogeneity from high throughput data, such as sequencing or single nucleotide polymorphism (SNP) array data, is important for gaining a better understanding of cancer and may improve prediction of clinical outcome or response to treatment. Here, we present a new method, CloneSeeker, for inferring clinical heterogeneity from sequencing data, SNP array data, or both.
We generated simulated SNP array and sequencing data and applied CloneSeeker along with two other methods. We demonstrate that CloneSeeker is more accurate than existing algorithms at determining the number of clones, distribution of cancer cells among clones, and mutation and/or copy numbers belonging to each clone. Next, we applied CloneSeeker to SNP array data from samples of 258 previously untreated CLL patients to gain a better understanding of the characteristics of CLL tumors and to elucidate the relationship between clonal heterogeneity and clinical outcome. We found that a significant majority of CLL patients appear to have multiple clones distinguished by copy number alterations alone. We also found that the presence of multiple clones corresponded with significantly worse survival among CLL patients. These findings may prove useful for improving the accuracy of prognosis and design of treatment strategies.
Code available on R-Forge: https://r-forge.r-project.org/projects/CloneSeeker/.
Supplementary data are available at Bioinformatics online.
We developed and validated a two-gene signature that predicts prognosis in previously-untreated chronic lymphocytic leukemia (CLL) patients. Using a 65 sample training set, from a cohort of 131 ...patients, we identified the best clinical models to predict time-to-treatment (TTT) and overall survival (OS). To identify individual genes or combinations in the training set with expression related to prognosis, we cross-validated univariate and multivariate models to predict TTT. We identified four gene sets (5, 6, 12, or 13 genes) to construct multivariate prognostic models. By optimizing each gene set on the training set, we constructed 11 models to predict the time from diagnosis to treatment. Each model also predicted OS and added value to the best clinical models. To determine which contributed the most value when added to clinical variables, we applied the Akaike Information Criterion. Two genes were consistently retained in the models with clinical variables: SKI (v-SKI avian sarcoma viral oncogene homolog) and SLAMF1 (signaling lymphocytic activation molecule family member 1; CD150). We optimized a two-gene model and validated it on an independent test set of 66 samples. This two-gene model predicted prognosis better on the test set than any of the known predictors, including ZAP70 and serum β2-microglobulin.
Although immunoglobulin VH mutation status (IgVH MS) is prognostic in patients with chronic lymphocytic leukemia (CLL) who are treated with alkylating agents or single-agent fludarabine, its ...significance in the era of chemoimmunotherapy is not known. We determined the IgVH somatic mutation status (MS) in 177 patients enrolled in a phase 2 study of fludarabine, cyclophosphamide, and rituximab (FCR) and in 127 patients treated with subsequent chemoimmunotherapy protocols. IgVH MS did not impact significantly on the complete remission (CR) rate of patients receiving FCR or related regimens. However, CR duration was significantly shorter in patients with CLL that used unmutated IgVH than those whose CLL used mutated IgVH (TTP 47% vs 82% at 6 years, P < .001). In a multivariate model considering all baseline characteristics, IgVH MS emerged as the only determinant of remission duration (hazard ratio 3.8, P < .001). Our results suggest that postremission interventions should be targeted toward patients with unmutated IgVH status.
In chronic lymphocytic leukemia (CLL), analysis of immunoglobulin heavy chain variable regions for somatic hypermutation identifies 2 prognostic subsets, mutated and unmutated. Investigators have ...postulated that unmutated and mutated CLL arises from malignant transformation of pre– and post–germinal center (GC) B cells, respectively. Alternatively, unmutated cases may arise from B cells stimulated by T-cell–independent antigens or from GC B cells with inactive somatic hypermutation. Activation-induced cytidine deaminase (AID), a protein essential for somatic hypermutation, is expressed by GC B cells in which this process occurs. We investigated AID mRNA expression in 20 CLL cases. In 8 cases we detected high expression of wild-type AID mRNA and 2 splice variants; in 12 cases and 5 normal peripheral blood B-cell samples we detected no expression using standard conditions. Of 8 CLL cases that highly expressed AID, 7 were unmutated, suggesting that this subset may arise from GC-experienced B cells with inactive somatic hypermutation, and may predict prognosis.
B-lymphoblastic leukemia (B-LBL) arising in patients with chronic lymphocytic leukemia (CLL) is exceedingly rare and poorly characterized.
We describe four patients with CLL and concurrent or ...subsequent B-LBL diagnosed by morphologic, immunophenotypic, cytogenetic, and molecular analysis and reviewed the literature.
In three patients, B-LBL followed CLL by 5 to 15 years, and in one patient, B-LBL was diagnosed simultaneously with CLL. In all cases, the CLL had a typical immunophenotype, and the B-LBL blasts showed an immature B-cell immunophenotype with expression of CD10, CD19, and TdT and absence of surface immunoglobulin. In two patients, B-LBL blasts harbored t(9;22)(q34;q11.2)/BCR-ABL1. We sequenced the IGHV genes in both CLL and B-LBL in two patients and showed that IGHV usage differed.
Our data suggest that at least some cases of B-LBL arising in patients with CLL are independent, secondary neoplasms rather than a manifestation of histologic transformation.
Objectives:
B-lymphoblastic leukemia (B-LBL) arising in patients with chronic lymphocytic leukemia (CLL) is exceedingly rare and poorly characterized.
Methods:
We describe four patients with CLL and ...concurrent or subsequent B-LBL diagnosed by morphologic, immunophenotypic, cytogenetic, and molecular analysis and reviewed the literature.
Results:
In three patients, B-LBL followed CLL by 5 to 15 years, and in one patient, B-LBL was diagnosed simultaneously with CLL. In all cases, the CLL had a typical immunophenotype, and the B-LBL blasts showed an immature B-cell immunophenotype with expression of CD10, CD19, and TdT and absence of surface immunoglobulin. In two patients, B-LBL blasts harbored t(9;22)(q34;q11.2)/BCR-ABL1. We sequenced the IGHV genes in both CLL and B-LBL in two patients and showed that IGHV usage differed.
Conclusions:
Our data suggest that at least some cases of B-LBL arising in patients with CLL are independent, secondary neoplasms rather than a manifestation of histologic transformation.
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