The genetic lesions identified in chronic lymphocytic leukemia (CLL) do not entirely recapitulate the disease pathogenesis and the development of serious complications, such as chemorefractoriness. While investigating the coding genome of fludarabine-refractory CLL, we observed that mutations of SF3B1, encoding a splicing factor and representing a critical component of the cell spliceosome, were recurrent in 10 of 59 (17%) fludarabinerefractory cases, with a frequency significantly greater than that observed in a consecutive CLL cohort sampled at diagnosis (17/301, 5%; P ‫؍‬ .002). Mutations were somatically acquired, were generally represented by missense nucleotide changes, clustered in selected HEAT repeats of the SF3B1 protein, recurrently targeted 3 hotspots (codons 662, 666, and 700), and were predictive of a poor prognosis. In fludarabine-refractory CLL, SF3B1 mutations and TP53 disruption distributed in a mutually exclusive fashion (P ‫؍‬ . IntroductionThe clinical course of chronic lymphocytic leukemia (CLL) ranges from a very indolent disorder with a normal lifespan for the patient to a rapidly progressive disease that leads to death. Occasionally, CLL undergoes a transformation to Richter syndrome (RS). [1][2][3] The variable clinical course of CLL is driven, at least in part, by the disease's immunogenetic and molecular heterogeneity. 4 Despite recent advances, the genetic lesions identified to date do not fully recapitulate the molecular pathogenesis of CLL and do not entirely explain the development of severe complications, such as chemorefractoriness, which still represent unmet clinical needs. 5 In approximately 40% of cases, refractoriness to fludarabine is attributable to the disruption of TP53, but in a sizeable fraction of patients, the molecular basis of this aggressive phenotype remains unclear. 6 Recently, 2 independent studies of the CLL coding genome investigated at disease presentation have revealed a restricted number of mutated genes, including NOTCH1. 7,8 These studies have provided a proof of concept that, similar to other malignancies, genome-wide mutational analysis might identify novel lesions of biologic and clinical relevance in CLL. On these grounds, we have embarked on the investigation of the coding genome of fludarabine-refractory CLL to identify genetic lesions associated with chemorefractoriness. The initial phases of this analysis have revealed recurrent mutations of SF3B1, a critical component of the cell spliceosome, pointing to the potential involvement of splicing regulation in CLL pathogenesis and chemorefractoriness. Methods PatientsThe study population comprised 3 cohorts representative of different disease phases: (1) fludarabine-refractory CLL (n ϭ 59), including cases (n ϭ 11) subjected to whole-exome sequencing (supplemental Table 1, available on the Blood Web site; see the Supplemental Materials link at the top of the online article); (2) a consecutive series of newly diagnosed and previously untreated patients with CLL (n ϭ 301; supplemental Table 2 For pers...

Analysis of the chronic lymphocytic leukemia (CLL) coding genome has recently disclosed that the NOTCH1 proto-oncogene is recurrently mutated at CLL presentation. Here, we assessed the prognostic role of NOTCH1 mutations in CLL. Two series of newly diagnosed CLL were used as training (n ‫؍‬ 309) and validation (n ‫؍‬ 230) cohorts. NOTCH1 mutations occurred in 11.0% and 11.3% CLL of the training and validation series, respectively. In the training series, NOTCH1 mutations led to a 3.77-fold increase in the hazard of death and to shorter overall survival (OS; P < .001). Multivariate analysis selected NOTCH1 mutations as an independent predictor of OS after controlling for confounding clinical and biologic variables. The independent prognostic value of NOTCH1 mutations was externally confirmed in the validation series. The poor prognosis conferred by NOTCH1 mutations was attributable, at least in part, to shorter treatment-free survival and higher risk of Richter transformation. Although NOTCH1 mutated patients were devoid of TP53 disruption in more than 90% cases in both training and validation series, the OS predicted by NOTCH1 mutations was similar to that of TP53 mutated/deleted CLL. NOTCH1 mutations are an independent predictor of CLL OS, tend to be mutually exclusive with TP53 abnormalities, and identify cases with a dismal prognosis. (Blood. 2012;119(2): 521-529) IntroductionChronic lymphocytic leukemia (CLL) is the most common leukemia in adults. [1][2][3][4] The clinical course of CLL ranges from very indolent, with a nearly normal life expectancy, [5][6][7][8][9] to rapidly progressive leading to death and occasionally undergoing transformation to aggressive lymphoma, known as Richter syndrome (RS). [10][11][12][13][14][15][16][17][18] At presentation, several clinical and biologic features may help to predict, at least in part, the clinical course of CLL. [19][20][21] Of the biologic prognosticators that have been developed, current guidelines for clinical practice recommend screening only for TP53 disruption by mutation, deletion, or both of the locus, that identifies a fraction of high-risk CLL destined to experience a very short survival. 2,[21][22][23][24][25][26][27][28] High-risk CLL, however, cannot be fully recapitulated by TP53 disruption, and other lesions of cancer genes may be implicated in this aggressive phenotype. 29 Recently, two independent investigations of the CLL coding genome have revealed that activating mutations of the NOTCH1 proto-oncogene are recurrently associated with CLL. 30,31 Based on current knowledge, NOTCH1 mutations occur in ϳ 10% CLL at diagnosis and their frequency increases in advanced disease phases, as exemplified by the case of RS. 30,31 The relevance of NOTCH1 mutations in CLL is reinforced by knowledge of activation of the NOTCH1 pathway in this leukemia, 32 and by the possibility of targeting NOTCH1 with drugs currently under development in other clinical contexts. 33 Although not designed to fully assess clinical correlates, the pivotal studies that have ident...

Purpose: Del17p13 predicts poor outcome and chemorefractoriness in chronic lymphocytic leukemia (CLL). Conversely, it is unknown whether TP53 mutations carry any prognostic value independent of del17p13.We tested the independent prognostic value of TP53 mutations in CLL. Experimental Design: The study was based on a consecutive series of 308 CLL. DNA sequencing of TP53 exons 2 to 10 and del17p13 interphase fluorescence in situ hybridization were done at CLL diagnosis. Study end points were survival and chemorefractoriness. Results: At diagnosis, TP53 mutations (n = 32) occurred in 31 of 308 (10.0%) patients. Of all CLL showing TP53 disruption by either mutation and/or deletion (n = 44), 10 cases (22.7%) showed TP53 mutations in the absence of del17p13. Multivariate analysis selected TP53 mutations (hazard ratio, 3.20; P = 0.002) as an independent predictor of overall survival after adjustment for del17p13. Also, multivariate analysis selected TP53 mutations (hazard ratio, 3.97; P < 0.001) as an independent predictor of chemorefractoriness after adjustment for del17p13.Compared with cases without TP53 alterations, CLL harboring any type of TP53 disruption (mutation only, del17p13 only, or both mutation and del17p13) uniformly displayed a high prevalence of unfavorable prognosticators and poor outcome. Analysis of sequential CLL samples showed the acquisition of new or additional TP53 alterations at the time of chemorefractoriness. Conclusions: These data show that (a) TP53 mutations are an independent predictor of short survival and chemorefractoriness, and (b) that CLL presenting withTP53 mutations without del17p13 fare as poorly as CLL carrying del17p13. Because CLL harboringTP53 mutations without del17p13 are currently not recognized by conventional diagnostic strategies, these results may be relevant for a comprehensive prognostic characterization of CLL.

Summary Predictors of chronic lymphocytic leukaemia (CLL) transformation to Richter syndrome (RS) are not established and were investigated in 185 consecutive CLL cases. Actuarial incidence of RS (n = 17; all diffuse large B‐cell lymphomas) at 10 years was 16·2% (95% confidence interval: 8·0–24·4%). At CLL diagnosis, prognosticators of RS by univariate analysis were IGHV homology ≥98% (P = 0·006), IGHV4‐39 usage (P < 0·001), del13q14 absence (P = 0·004), expression of CD38 (P < 0·001) and ZAP70 (P = 0·004), size (P < 0·001) and number (P < 0·001) of lymph nodes, advanced Binet stage (P = 0·002), and lactate dehydrogenase (P < 0·001). Multivariate analysis, performed separately for biological and clinical variables, identified CD38 expression [Hazard ratio (HR) = 4·26; P = 0·018], IGHV4‐39 usage (HR = 4·29; P = 0·018), and lymph node size ≥3 cm (HR = 9·07; P < 0·001) as independent RS prognosticators. A multivariate model simultaneously analysing biological and clinical variables identified lymph node size ≥3 cm (HR = 6·51; P = 0·001) and del13q14 absence (HR = 4·08; P = 0·031) as independent RS prognosticators. Risk factors of CLL transformation differed from risk factors of CLL progression. These results suggest that CD38 and del13q14 may identify biological subsets of CLL with different RS predisposition. Predominant nodal disease, CD38 expression, IGHV4‐39 usage, and absence of del13q14 may help in predicting RS at CLL diagnosis. Close monitoring and a careful biopsy policy are needed in patients carrying transformation risk factors.