Chronic lymphocytic leukemia: A prognostic model comprising only two biomarkers (IGHV mutational status and FISH cytogenetics) separates patients with different outcome and simplifies the CLL‐IPI

Rai and Binet staging systems are important to predict the outcome of patients with chronic lymphocytic leukemia (CLL) but do not reflect the biologic diversity of the disease nor predict response to therapy, which ultimately shape patients' outcome. We devised a biomarkers‐only CLL prognostic system based on the two most important prognostic parameters in CLL (i.e., IGHV mutational status and fluorescence in situ hybridization [FISH] cytogenetics), separating three different risk groups: (1) low‐risk (mutated IGHV + no adverse FISH cytogenetics [del(17p), del(11q)]); (2) intermediate‐risk (either unmutated IGHV or adverse FISH cytogenetics) and (3) high‐risk (unmutated IGHV + adverse FISH cytogenetics). In 524 unselected subjects with CLL, the 10‐year overall survival was 82% (95% CI 76%‐88%), 52% (45%‐62%), and 27% (17%‐42%) for the low‐, intermediate‐, and high‐risk groups, respectively. Patients with low‐risk comprised around 50% of the series and had a life expectancy comparable to the general population. The prognostic model was fully validated in two independent cohorts, including 417 patients representative of general CLL population and 337 patients with Binet stage A CLL. The model had a similar discriminatory value as the CLL‐IPI. Moreover, it applied to all patients with CLL independently of age, and separated patients with different risk within Rai or Binet clinical stages. The biomarkers‐only CLL prognostic system presented here simplifies the CLL‐IPI and could be useful in daily practice and to stratify patients in clinical trials.     

Monoclonal B‐cell lymphocytosis is closely related to chronic lymphocytic leukaemia and may be better classified as early‐stage CLL

The World Health Organization classification uses a cut‐off point of 5·0 × 10⁹/l cells with a chronic lymphocytic leukaemia (CLL)‐phenotype in peripheral blood to discriminate between monoclonal B‐lymphocytosis (MBL) and B‐CLL. This study analysed 298 MBL patients by multi‐parameter flow cytometry, chromosome banding analysis (CBA)/fluorescence in situ hybridization (FISH), and IGHV mutation status and compared them with 356 CLL patients. In MBL, CBA more frequently revealed a normal karyotype and FISH identified less frequently del(6q), del(13q) (as sole alterations), and del(17)(p13). Within the MBL cohort, a shorter time to treatment (TTT) was found for ZAP‐70‐positivity, 14q32/IGH‐translocations (CBA), del(11)(q22·3) (FISH) and unmutated IGHV status. Higher CD38 and ZAP‐70 expression, del(11)(q22·3) (FISH), trisomy 12 (FISH), and 14q32/IGH‐translocations (CBA) were correlated with a shorter TTT in the combined cohort (MBL + CLL); a sole del(13)(q14) (FISH) correlated with longer TTT. Regarding overall survival, unmutated IGHV status and ‘other’ alterations (CBA) had an adverse impact. There was no correlation between the concentration of CLL‐cells and TTT or overall survival. Multivariate analysis confirmed a negative impact on TTT for del(11)(q22·3)/ATM, trisomy 12 (both by FISH), and 14q32/IGH‐translocations by CBA. These data emphasize a close relationship between MBL and CLL regarding clinically relevant parameters and provide no evidence to strictly separate these entities by a distinct threshold of clonal B‐cells.     

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.     

Significant change in ZAP‐70 expression during the course of chronic lymphocytic leukemia

Introduction: It is widely accepted that expression of ZAP‐70 in chronic lymphocytic leukemia (CLL) remains stable in time. However, data supporting this notion are surprisingly scarce. Therefore, we assessed expression of ZAP‐70 in serial samples taken during the course of the disease. Patients and Methods: We studied 44 patients with CLL diagnosed according to NCI‐WG criteria (34 men, 10 women, median age 62, range, 36–81). A total of 104 samples were examined; all patients had at least two measurements. Median interval between the first and the second sample was 13 months (range, 2–36). ZAP‐70 expression was detected by flow cytometry using phycoerythrin‐conjugated monoclonal antibody clone 1E7.2 and negative isotype control. Twenty percent of positive cells were considered as the threshold of positivity. Results: Significant change in ZAP‐70 expression (i.e. from positivity to negativity and vice versa) was detected in 15/44 patients (34%). Interestingly, 7/8 patients whose ZAP‐70 expression converted to positivity had unmutated IgVH genes. In addition, the conversion was accompanied by clinical progression or relapse in all but one patient. On the other hand, 5/7 patients with loss of ZAP‐70 had stable clinical course. One patient became ZAP‐70‐negative during treatment with prednisone for autoimmune hemolytic anemia. Conclusions: In contrast to commonly accepted opinion, significant change in ZAP‐70 expression in time was detected in a substantial proportion of our patients with CLL. While the conversion to ZAP‐70 negativity was found predominantly in patients with stable disease, change to positivity was typical in patients with unmutated IgVH genes at the time of progression or relapse. Based on our pilot results, repeated assessment of ZAP‐70 expression might be especially useful at the time of progression or relapse in patients who were initially ZAP‐70‐negative.     

Evaluation of 230 patients with relapsed/refractory deletion 17p chronic lymphocytic leukaemia treated with ibrutinib from 3 clinical trials

Patients with chronic lymphocytic leukaemia/small lymphocytic lymphoma (CLL/SLL) with deletion 17p [del(17p)] have poor outcomes with chemoimmunotherapy. Ibrutinib is indicated for the treatment of CLL/SLL, including del(17p) CLL/SLL, and allows for treatment without chemotherapy. This integrated analysis was performed to evaluate outcomes in 230 patients with relapsed/refractory del(17p) CLL/SLL from three ibrutinib studies. With a median of 2 prior therapies (range, 1–12), 18% and 79% of evaluable patients had del(11q) or unmutated IGHV, respectively. With a median follow‐up of 28 months, overall response rate was 85% and estimated 30‐month progression‐free and overall survival rates were 57% [95% confidence interval (CI) 50–64] and 69% (95% CI 61–75), respectively. Patients with normal lactate dehydrogenase or no bulky disease had the most favourable survival outcomes. Sustained haematological improvements in haemoglobin, platelet count and absolute neutrophil count occurred in 61%, 67% and 70% of patients with baseline cytopenias, respectively. New onset severe cytopenias and infections decreased in frequency over time. Progression‐free and overall survival with ibrutinib surpass those of other therapies for patients with del(17p) CLL/SLL. These results provide further evidence of the robust clinical activity of ibrutinib in difficult‐to‐treat CLL/SLL populations.     

Comparison of flow cytometric methods for the measurement of ZAP‐70 expression in a routine diagnostic laboratory

Chronic lymphocytic leukaemia (CLL) follows a variable clinical course with patient survival ranging from only a few years despite treatment, to several decades in patients who may never require clinical intervention. Determination of the mutational status of a patient's immunoglobulin heavy chain variable region (Ig V_H) gene has been used to provide prognostic information, but this assay is not available in most laboratories. The discovery of the expression of the protein tyrosine kinase zeta‐associated protein (ZAP)‐70 in V_H‐unmutated CLL cases led to its proposal as a surrogate marker for V_H status. This study investigated the measurement of ZAP‐70 expression in CLL using different flow cytometric protocols. Two different antibodies and two different staining methods were compared. The Caltag ZAP‐70 antibody and Fix & Perm kit were the easiest to use and were the most sensitive and specific combination, with 91% concordance between ZAP‐70 expression and V_H status. Three patients (9%) were discordant (two V_H mutated/ZAP‐70 positive, and one V_H unmutated/ZAP‐70 negative). No correlation existed between CD38 and either ZAP‐70 expression or V_H status. Measurement of ZAP‐70 expression using the Caltag antibody/kit combination provides a standardized flow cytometric method that could be introduced into a routine CLL immunophenotyping panel in a clinical diagnostic laboratory.     

Prognostic importance of soluble CD23 in B‐cell chronic lymphocytic leukemia

Soluble CD23 (sCD23) was proposed as a marker of disease activity and as an important prognostic parameter in B‐cell chronic lymphocytic leukemia (B‐CLL). In this study, prognostic significance of sCD23 in B‐CLL was examined according to its temporal relationship with the known clinical parameters of the disease, CD38 and ZAP‐70. Serum sCD23 levels of 36 B‐CLL patients, followed up in our clinic between 1999 and 2005, and 15 healthy subjects were measured with enzyme‐linked immunosorbent assay. The mean serum sCD23 level of the B‐CLL patients (210.72 ± 193.67 and 6–600 U/ml) was significantly higher than the control group (18.20 ± 14.30 and 6–50 U/ml). Seventy‐eight percent of the B‐CLL patients with lymphocyte doubling time (LDT) <12 months and 24% of patients with LDT >12 months had high sCD23 levels (P = 0.008). Meanwhile, 81% of the patients with diffuse bone marrow infiltration and 33% of patients with nondiffuse infiltration had high levels of serum sCD23 (P = 0.029). A significant difference was found between B‐CLL patients with Binet stages A and C (P = 0.009). Peripheral blood flow cytometry of the patients revealed a significant CD38 expression in patients with high serum sCD23 levels (P = 0.002). Similarly, an increased bone marrow zeta‐chain associated protein kinase‐70 (ZAP‐70) expression was seen in patients with high serum sCD23 levels (P = 0.009, correlation co‐efficient was 0.714). Cumulative and the progression free survivals of the patients with low serum sCD23 levels were 60.1 ± 5.7 months [95% confidence interval (CI); 49.0–71.2] and 51.1 ± 6.6 months (95% CI; 38.0–64.1), respectively. However, they were 43.8 ± 6.5 months (95% CI; 31.0–56.6) and 26.5 ± 6.4 months (95% CI; 14.0–39.1) in patients with high levels. Serum sCD23 is increased in B‐CLL patients and can be used in the clinical follow‐up of the disease in prediction of the tumor mass and prognosis.     

Baff serum level predicts time to first treatment in early chronic lymphocytic leukemia

We analyzed the correlation between well‐established biological parameters of prognostic relevance in B‐cell chronic lymphocytic leukemia (CLL) [i.e. mutational status of the immunoglobulin heavy chain variable region (IgVH), ZAP‐70 and CD38 expression] and serum levels of B cell–activating factor (BAFF of the TNF family) by evaluating the impact of these variables on the time to first treatment (TFT) in a series of 169 previously untreated CLL patients in Binet stage A. Higher levels of BAFF were more frequently associated with female gender (P = 0.02), younger age (P = 0.01), Rai stage 0 (P = 0.002), higher platelet count (P = 0.005), mutated IgVH disease (P = 0.002), higher occurrence of normal cytogenetic profile or presence of 13q deletion (P = 0.02), low ZAP‐70‐ (P = 0.003), and CD38‐expression (P = 0.02). Maximally selected log‐rank statistic plot identified a serum BAFF concentration of 0.313 ng/mL as the best cut‐off (P < 0.0001). This threshold recognized two subsets of patients with different TFT (P < 0.0001). Because in multivariate analysis soluble BAFF [Hazard ratio (HR), 8.23; confidence Interval (CI) 95%,3.0–22.6, P < 0.0001] and mutational status of IgVH (HR = 2.60; CI 95% 1.10–6.14, P = 0.03) maintained the discriminating power their combined effect on clinical outcome was assessed. When three groups were considered: ( 1 ) low‐risk (n = 93), patients with concordant IgVH_mut and higher soluble BAFF; ( 2 ) intermediate‐risk (n = 50), patients with IgVH_mut and low BAFF levels or IgVH_unmut and soluble higher BAFF;( 3 ) high‐risk (n = 26), patients with concordant IgVH _unmut and low soluble BAFF, the 2‐yr TFTs were, respectively, 95%, 85%, and 41% (P < 0.0001). In conclusion, our results indicate that in early B‐cell CLL, the biological profile including among other parameters soluble BAFF may provide a useful insight into the complex interrelationship of prognostic variables.     

The absolute percent deviation of IGHV mutation rather than a 98% cut‐off predicts survival of chronic lymphocytic leukaemia patients treated with fludarabine,cyclophosphamide and rituximab

The degree of somatic hypermutation, determined as percent deviation of immunoglobulin heavy chain gene variable region sequence from the germline (IGHV%), is an important prognostic factor in chronic lymphocytic leukaemia (CLL). Currently, a cut‐off of 2% deviation or 98% sequence identity to germline in IGHV sequence is routinely used to dichotomize CLL patients into mutated and unmutated groups. Because dissimilar IGHV% cut‐offs of 1–5% were identified in different studies, we wondered whether no cut‐off should be applied and IGHV% treated as a continuous variable. We analysed the significance of IGHV% in 203 CLL patients enrolled on the original frontline fludarabine, cyclophosphamide and rituximab (FCR) trial with a median of 10 years follow‐up. Using the Cox Proportional Hazard model, IGHV% was identified as a continuous variable that is significantly associated with progression‐free (PFS) and overall survival (OS) (P < 0·001). Furthermore, we validated this finding in 323 patients treated with FCR off‐protocol and in the total cohort (n = 535). Multivariate analysis revealed a continuous trend. Higher IGHV% levels were incrementally associated with favorable PFS and OS in both FCR‐treated cohorts (P < 0·001, both cohorts). Taken together, our data suggest that IGHV% is a continuous variable in CLL patients treated with FCR.     

FCRL2 mRNA expression is inversely associated with clinical progression in chronic lymphocytic leukemia

Fc receptor‐like 2 (FCRL2) is highly expressed on B‐cell chronic lymphocytic leukemia (B‐CLL) cells and could possibly influence disease pathogenesis. Therefore, we investigated FCRL2 mRNA expression in a large cohort with 152 CLL patients in order to assess its role in risk prediction in B‐CLL. FCRL2 mRNA expression was found to be expressed at considerably higher levels in peripheral blood mononuclear cells (PBMC) of B‐CLL patients compared to controls (range 1.35‐ to 210‐fold upregulation; P < 0.0001) and cells of other hematological diseases. Patients with high FCRL2 expression (according to ROC‐analysis) had a significantly longer treatment‐free survival (TFS) and overall survival (OS) than patients with low FCRL2 expression (median TFS: 119 vs. 34 months, P < 0.0001; median OS: 321 months vs. not reached, P = 0.009). Univariate comparisons found that FCRL2 expression was weakly associated with IGHV mutation status (P = 0.05), CD38 status (P < 0.0001) and ZAP‐70 status (P < 0.0001). Furthermore, we show that the combination of FCRL2 with ZAP70‐, CD38‐ or IGHV‐status could further significantly refine the prognostic information provided by either of the factors alone in TFS and OS. In multivariate analysis low FCRL2 expression was a significant independent prognostic factor (HR 2.4; P = 0.005). Here we demonstrate that the level of FCRL2 expression is correlated with prognosis in B‐CLL.     

Comparison of familial and sporadic chronic lymphocytic leukaemia using high resolution array comparative genomic hybridization

Approximately 10% of patients with chronic lymphocytic leukaemia (CLL) have a family history of the disease or a related lymphoproliferative disorder, yet the relationship of familial CLL to genomic abnormalities has not been characterized in detail. We therefore studied 75 CLL patients, half familial and half sporadic, using high‐resolution array comparative genomic hybridization (CGH), in order to better define the relationship of genomic abnormalities to familial disease and other biological prognostic factors. Our results showed that the most common high‐risk deletion in CLL, deletion 11q, was significantly associated with sporadic disease. Comparison of familial to sporadic disease additionally identified a copy number variant region near the centromere on 14q, proximal to IGH@, in which gains were associated both with familial CLL, and with mutated IGHV and homozygous deletion of 13q. Homozygous deletion of 13q was also found to be associated with mutated IGHV and low expression of ZAP‐70, and a significantly longer time to first treatment compared to heterozygous deletion or lack of alteration. This study is the first high resolution effort to investigate and report somatic genetic differences between familial and sporadic CLL.     

IGHV-associated methylation signatures more accurately predict clinical outcomes of chronic lymphocytic leukemia patients than IGHV mutation load

Currently, no molecular biomarker indices are used in standard care to make treatment decisions at diagnosis of chronic lymphocytic leukemia (CLL). We used Infinium MethylationEPIC array data from diagnostic blood samples of 114 CLL patients and developed a procedure to stratify patients based on methylation signatures associated with mutation load of the IGHV gene. This procedure allowed us to predict the time to treatment with a hazard ratio (HR) of 8.34 (95% confidence interval [CI]: 4.54-15.30), as opposed to a HR of 4.35 (95% CI: 2.60-7.28) using IGHV mutation status. Detailed evaluation of 17 cases for which the two classification procedures gave discrepant results showed that these cases were incorrectly classified using IGHV status. Moreover, methylation-based classification stratified patients with different overall survival (HR=1.82; 95% CI: 1.07-3.09), which was not possible using IGHV status. Furthermore, we assessed the performance of the developed classification procedure using published HumanMethylation450 array data for 159 patients for whom information on time to treatment, overall survival and relapse was available. Despite 450K array methylation data not containing all the biomarkers used in our classification procedure, methylation signatures again stratified patients with significantly better accuracy than did IGHV mutation load regarding all available clinical outcomes. Thus, stratification using IGHV-associated methylation signatures may provide better prognostic power than IGHV mutation status.     

CD49d (ITGA4) expression is a predictor of time to first treatment in patients with chronic lymphocytic leukaemia and mutated IGHV status

We investigated CD49d (also termed ITGA4) expression and its biological and clinical correlations in 415 patients with chronic lymphocytic leukaemia. CD49d expression was stable over the course of the disease. A high expression of CD49d (>30%) was found in 142/415 (34%) patients and was associated with progressive disease (advanced clinical stage, high serum lactate dehydrogenase or β₂‐microglobulin levels; all p < 0·05) and aggressive disease biology (increased ZAP70 or CD38, unmutated IGHV, trisomy 12, mutations of NOTCH1 and SF3B1; all P < 0·05). A higher CD49d expression was also associated with a lower blood lymphocyte count and a higher number of lymphoid areas involved by the disease. Patients with high CD49d expression were treated more frequently (55% vs. 27%; P < 0·001) and earlier (median time to treatment [TTT] 65·4 months vs. not reached; P < 0·001) than those with low CD49d expression. However, no significant differences in response rates were observed. In the subgroup of patients with mutated IGHV, high CD49d expression was predictive of a shorter TTT while other markers, such as ZAP70 and CD38, were not. In conclusion, in this study CD49d expression correlated with high‐risk CLL biomarkers and proved to be useful for separating patients with mutated IGHV into two different prognostic groups.     

The morphology of CLL revisited: the clinical significance of prolymphocytes and correlations with prognostic/molecular markers in the LRF CLL4 trial

Historically, an increase in the percentage and number of circulating prolymphocytes in chronic lymphocytic leukaemia (CLL) has been associated with strong expression of surface immunoglobulin, trisomy 12 and a poor outcome. This study re‐examines the biological and clinical significance of increased peripheral blood prolymphocytes in 508 patients at entry into the randomized UK Leukaemia Research Fund CLL4 trial. It also investigates the associations between increased prolymphocytes and a comprehensive array of biomarkers. 270 patients (53%) had <5% prolymphocytes, 167 (33%) had 5–9%, 60 (12%) had 10–14% and 11 (2%) had ≥15% prolymphocytes. We show that a higher proportion of prolymphocytes (≥10%) was independently associated with NOTCH1 mutations (P = 0·006), absence of 13q deletion (P = 0·001), high CD38 expression (P = 0·02) and unmutated IGHV genes (P = 0·01). Deaths due to Richter syndrome were significantly more common amongst patients who had ≥10% vs <10% prolymphocytes (13% vs 2%) respectively (P < 0·0001). ≥10% prolymphocytes was also associated with a shorter progression‐free survival (Hazard ratio [HR] 1·50 [95% confidence interval [CI]: 1·16–1·93], P = 0·002) and overall survival (HR 1·99 [95% CI: 1·53–2·59], P < 0·0001). Our data support the routine examination of blood films in CLL and suggest that a finding of an increased proportion of prolymphocytes may be a trigger for further evaluation of clinical and laboratory features of progressive disease.     

Novel X-linked inhibitor of apoptosis inhibiting compound as sensitizer for TRAIL-mediated apoptosis in chronic lymphocytic leukaemia with poor prognosis

Given that aggressive DNA damaging chemotherapy shows suboptimal efficacy in chronic lymphocytic leukaemia (CLL), alternative therapeutic approaches are needed. Tumour necrosis factor‐related apoptosis‐inducing ligand (TRAIL) is able to induce tumour‐specific apoptosis. However, apoptosis might be inhibited by elevated levels of X‐linked inhibitor of apoptosis (XIAP). Use of XIAP‐inhibiting compounds might sensitize primary CLL cells towards TRAIL‐mediated apoptosis. A novel small molecule, compound A (CA), an inhibitor of XIAP, was used in combination with TRAIL to induce apoptosis in primary CLL cells (n = 48). XIAP was significantly more highly expressed in primary CLL cells (n = 28) compared to healthy B cells (n = 16) (P = 0·02). Our data obtained by specific knock‐down of XIAP by siRNA identified XIAP as the key factor conferring resistance to TRAIL in CLL. Combined treatment with CA/TRAIL significantly increased apoptosis compared to untreated (P = 8·5 × 10^?10), solely CA (P = 4·1 × 10^?12) or TRAIL treated (P = 4·8 × 10^?10) CLL cells. CA rendered 40 of 48 (83·3%) primary CLL samples susceptible to TRAIL‐mediated apoptosis. In particular, cells derived from patients with poor prognosis CLL (ZAP‐70⁺, IGHV unmutated, 17p‐) were highly responsive to this drug combination. Our highly‐effective XIAP inhibitor CA, in concert with TRAIL, shows potential for the treatment of CLL cases with poor prognosis and therefore warrants further clinical investigation.     

MFI ratio estimation of ZAP-70 in B-CLL by flow cytometry can be improved by considering the isotype-matched antibody signal

Introduction: The IgV_H mutational status of B‐cell chronic lymphocytic leukemia (B‐CLL) is of prognostic value. Expression of ZAP‐70 in B‐CLL is a surrogate marker for IgV_H unmutated (UM). As determination of IgV_H mutational status involves a methodology currently unavailable for most clinical laboratories, it is important to have available a reliable technique for ZAP‐70 estimation in B‐CLL. Flow cytometry (FC) is a convenient technique for this purpose. However, there is still no adequate way for data analysis, which would prevent the assignment of false positive or negative expression. Methods: We have modified the currently most accepted technique, which uses the ratio of the mean fluorescent index (MFI) of B‐CLL to T cells. The MFI for parallel antibody isotype staining is subtracted from the ZAP‐70 MFI of both B‐CLL and T cells. We validated this technique comparing the results obtained for ZAP‐70 expression by FC with those obtained with quantitative PCR for the same patients. Results: We applied the technique in a series of 53 patients. With this modification, a better correlation between ZAP‐70 expression and IgV_H UM was obtained. Conclusions: Thus, the MFI ratio B‐CLL/T cell corrected by isotype is a reliable analysis technique to estimate ZAP‐70 expression in B‐CLL.     

Diffuse large B‐cell lymphoma (Richter syndrome) in patients with chronic lymphocytic leukaemia (CLL): a cohort study of newly diagnosed patients

Nearly all information about patients with chronic lymphocytic leukaemia (CLL) who develop diffuse large B‐cell lymphoma [Richter syndrome (RS)] is derived from retrospective case series or patients treated on clinical trials. We used the Mayo Clinic CLL Database to identify patients with newly diagnosed CLL between January 2000 and July 2011. Individuals who developed biopsy‐proven RS during follow‐up were identified. After a median follow‐up of 4 years, 37/1641 (2·3%) CLL patients developed RS. The rate of RS was approximately 0·5%/year. Risk of RS was associated with advanced Rai stage at diagnosis (P < 0·001), high‐risk genetic abnormalitites on fluorescence in situ hybridization (P < 0·0001), unmutated IGHV (P = 0·003), and expression of ZAP70 (P = 0·02) and CD38 (P = 0·001). The rate of RS doubled in patients after treatment for CLL (1%/year). Stereotyped B‐cell receptors (odds‐ratio = 4·2; P = 0·01) but not IGHV4‐39 family usage was associated with increased risk of RS. Treatment with combination of purine analogues and alkylating agents increased the risk of RS three‐fold (odds‐ratio = 3·26, P = 0·0003). Median survival after RS diagnosis was 2·1 years. The RS prognosis score stratified patients into three risk groups with median survivals of 0·5 years, 2·1 years and not reached. Both underlying characteristics of the CLL clone and subsequent CLL therapy influence the risk of RS. Survival after RS remains poor and new therapies are needed.     

Final analysis from RESONATE: Up to six years of follow-up on ibrutinib in patients with previously treated chronic lymphocytic leukemia or small lymphocytic lymphoma

Ibrutinib, a once-daily oral inhibitor of Bruton's tyrosine kinase, is approved in the United States and Europe for treatment of patients with chronic lymphocytic leukemia (CLL) or small lymphocytic lymphoma (SLL). The phase 3 RESONATE study showed improved efficacy of single-agent ibrutinib over ofatumumab in patients with relapsed/refractory CLL/SLL, including those with high-risk features. Here we report the final analysis from RESONATE with median follow-up on study of 65.3 months (range, 0.3-71.6) in the ibrutinib arm. Median progression-free survival (PFS) remained significantly longer for patients randomized to ibrutinib vs ofatumumab (44.1 vs 8.1 months; hazard ratio [HR]: 0.148; 95% confidence interval [CI]: 0.113-0.196; P˂.001). The PFS benefit with ibrutinib vs ofatumumab was preserved in the genomic high-risk population with del(17p), TP53 mutation, del(11q), and/or unmutated IGHV status (median PFS 44.1 vs 8.0 months; HR: 0.110; 95% CI: 0.080-0.152), which represented 82% of patients. Overall response rate with ibrutinib was 91% (complete response/complete response with incomplete bone marrow recovery, 11%). Overall survival, censored for crossover, was better with ibrutinib than ofatumumab (HR: 0.639; 95% CI: 0.418-0.975). With up to 71 months (median 41 months) of ibrutinib therapy, the safety profile remained consistent with prior reports; cumulatively, all-grade (grade ≥3) hypertension and atrial fibrillation occurred in 21% (9%) and 12% (6%) of patients, respectively. Only 16% discontinued ibrutinib because of adverse events (AEs). These long-term results confirm the robust efficacy of ibrutinib in relapsed/refractory CLL/SLL irrespective of high-risk clinical or genomic features, with no unexpected AEs. This trial is registered at www.clinicaltrials.gov (NCT01578707).