Mutations of the SF3B1 splicing factor in chronic lymphocytic leukemia: association with progression and fludarabine-refractoriness

Abstract

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...

The SF3B1 inhibitor spliceostatin A (SSA) elicits apoptosis in chronic lymphocytic leukaemia cells through downregulation of Mcl-1

“…Due to the incidence of SF3B1 mutations in CLL, 8,10 we compared the effect of SSA on 13 patients carrying a mutation in this gene and 42 wild-type patients. The SF3B1 mutations observed reflected the spectrum reported previously (Supplementary Table 2).…”

“…Progressive disease is characterised by unmutated immunoglobulin heavy variable (IGHV) genes, ZAP70 and CD38 expression. 1 Genetic markers including deletions of 17p, 11q and 13q, trisomy 12 and TP53 mutations [2][3][4] have been known to impact prognosis for some time, however with the emergence of next generation sequencing, a plethora of new genetic alterations have been identified including NOTCH1 (10-15%) [5][6][7][8] and SF3B1 (10-17%) [8][9][10] mutations which identify patients with progressive disease. 8 SF3B1 is one of the proteins that make up the spliceosome and regulates excision of introns from premRNA 11 .…”

The genome of chemorefractory chronic lymphocytic leukemia reveals frequent mutations of NOTCH1 and SF3B1

“…SF3B1 was altered in 10/59 (17%) fludarabine-refractory CLL by missense mutations (n ¼ 9) or in-frame deletions (n ¼ 1) clustering in the HEAT3, HEAT4 and HEAT5 repeats of the SF3B1 protein. 21 Two sites that are highly conserved among species (codon 662 and codon 700) were recurrently mutated in three and five cases, respectively. SF3B1 mutations were monoallelic and were predicted to be functionally significant according to the PolyPhen-2 algorithm.…”

“…21 The study population comprised three clinical cohorts representative of the following: i) fludarabine-refractory CLL (n ¼ 59), including cases (n ¼ 11) subjected to WES; ii) newly diagnosed and previously untreated CLL (n ¼ 301); and iii) clonally related RS (n ¼ 33). Tumor samples were obtained for the following: i) fludarabine-refractory CLL, immediately before starting the treatment to which the patient eventually failed to respond, and ii) for newly diagnosed and previously untreated CLL, at disease presentation.…”

“…Mutations occurred irrespective of IGHV mutation status, CD38 expression and ZAP70 expression. 21 At the time of fludarabine refractoriness, SF3B1 mutations were enriched in cases harboring a normal fluorescence in situ hybridization karyotype (P ¼ 0.008) and distributed in a mutually exclusive manner with TP53 disruption (mutual information I ¼ 0.0609; P ¼ 0.046). By combining SF3B1 mutations with other genetic lesions enriched in chemorefractory cases (TP53 disruption, NOTCH1 mutations, ATM deletion), fludarabine-refractory CLL appeared to be characterized by multiple molecular alterations that, to some extent, are mutually exclusive.…”