Through the European Research Initiative on chronic lymphocytic leukemia (CLL) (ERIC), we screened 3490 patients with CLL for mutations within the NOTCH1 (n=3334), SF3B1 (n=2322), TP53 (n=2309), MYD88 (n=1080) and BIRC3 (n=919) genes, mainly at diagnosis (75%) and before treatment (>90%). BIRC3 mutations (2.5%) were associated with unmutated IGHV genes (U-CLL), del(11q) and trisomy 12, whereas MYD88 mutations (2.2%) were exclusively found among M-CLL. NOTCH1, SF3B1 and TP53 exhibited variable frequencies and were mostly enriched within clinically aggressive cases. Interestingly, as the timespan between diagnosis and mutational screening increased, so too did the incidence of SF3B1 mutations; no such increase was observed for NOTCH1 mutations. Regarding the clinical impact, NOTCH1 mutations, SF3B1 mutations and TP53 aberrations (deletion/mutation, TP53ab) correlated with shorter time-to-first-treatment (P<0.0001) in 889 treatment-naive Binet stage A cases. In multivariate analysis (n=774), SF3B1 mutations and TP53ab along with del(11q) and U-CLL, but not NOTCH1 mutations, retained independent significance. Importantly, TP53ab and SF3B1 mutations had an adverse impact even in U-CLL. In conclusion, we support the clinical relevance of novel recurrent mutations in CLL, highlighting the adverse impact of SF3B1 and TP53 mutations, even independent of IGHV mutational status, thus underscoring the need for urgent standardization/harmonization of the detection methods.

) and 8q22.3 (rs2511714, P=2.90x10 -9 ). These findings provide further insights into the genetic and biological basis of inherited genetic susceptibility to CLL. Speedy et al 3Chronic lymphocytic leukemia (CLL) is the most common hematological malignancy in Western countries 1 and is characterized by a 8-fold increased risk in first-degree relatives 2 . Genome-wide association studies (GWASs) have so far identified common variants at 24 loci that contribute to the heritable risk of CLL [3][4][5][6] . Current projections for the number of independent regions harboring common variants associated with CLL suggest that additional risk loci conferring modest effects should be identified by expansion of discovery GWAS datasets.To identify additional novel susceptibility loci for CLL, we conducted an independent primary scan of CLL and performed a genome-wide meta-analysis with a previously published GWAS followed by analysis of the top single nucleotide polymorphisms (SNPs) in two separate case-control series.In the primary scan (UK-CLL-2), 1,271 CLL cases were genotyped using the Illumina Omni Express Figure 1). To harmonize the two GWAS datasets, we imputed UK-CLL-1 to recover untyped SNPs directly genotyped in UK-CLL-2, using data from the 1000 Genomes Project as reference. Using data on all cases and controls from each GWAS, we derived joint odds ratios (ORs) and confidence intervals (CIs) under a fixed effects model for each SNP and associated P-values, restricting analysis to SNPs with MAF >1%. After filtering on the basis of pre-specified quality-control measures Table 2). We also identified promising association signals (i.e. P<1.0×10−5 ) at 11 additional loci (Supplementary Table 2). We applied 1000 Genomes imputation to UK-CLL-1 and UK-CLL-2 at these loci to investigate if a statistically significant stronger SNP association could be identified, recovering an additional SNP which was significant at the genome-wide threshold (rs6858698; Supplementary Table 2). We performed replication genotyping of six SNPs selected on the basis of statistical significance (rs2236256, rs6062501, rs6858698) and gene centricity coupled with Table 4). While we found no evidence for a relationship between rs10936599, and telomere length in 246 CLL patients (Supplementary Table 5), carrier status for the rs10936599-C risk allele is previously been associated with significantly longer telomeres in leukocytes 10,11 .The third significant association was at rs6858698 on 4q26 (OR=1.31, 95% C.I. 1.20-1.44; P=3.07x10 linked to cis-platinum resistance by enhancing apoptosis. A recent GWAS of CLL has reported promising associations at 5p15.33 defined by rs10069690 and at 8q22.33 defined by rs2511714 6 . Combining the Pvalues for rs10069690 and rs2511714 obtained in our meta-analysis (P=1.0x10 -4 and 1.0x10respectively) with published data 6 provides robust evidence for both associations (combined P-values 1.10x10 -10 and 2.90x10 -9 respectively; Supplementary Figure 4). rs10069690 maps to intron 4 of TERT (telomerase reverse trans...

Key Points• Whole-exome sequencing of CLL patients who relapsed after FCR treatment revealed frequent mutations in RPS15.• RPS15 mutations are likely to be early clonal events and confer poor prognosis.Fludarabine, cyclophosphamide, and rituximab (FCR) is first-line treatment of medically fit chronic lymphocytic leukemia (CLL) patients; however, despite good response rates, many patients eventually relapse. Although recent high-throughput studies have identified novel recurrent genetic lesions in adverse prognostic CLL, the mechanisms leading to relapse after FCR therapy are not completely understood. To gain insight into this issue, we performed whole-exome sequencing of sequential samples from 41 CLL patients who were uniformly treated with FCR but relapsed after a median of 2 years. In addition to mutations with known adverse-prognostic impact (TP53, NOTCH1, ATM, SF3B1, NFKBIE, and BIRC3), a large proportion of cases (19.5%) harbored mutations in RPS15, a gene encoding a component of the 40S ribosomal subunit. Extended screening, totaling 1119 patients, supported a role for RPS15 mutations in aggressive CLL, with one-third of RPS15-mutant cases also carrying TP53 aberrations. In most cases, selection of dominant, relapse-specific subclones was observed over time. However, RPS15 mutations were clonal before treatment and remained stable at relapse. Notably, all RPS15 mutations represented somatic missense variants and resided within a 7 amino-acid, evolutionarily conserved region. We confirmed the recently postulated direct interaction between RPS15 and MDM2/MDMX and transient expression of mutant RPS15 revealed defective regulation of endogenous p53 compared with wild-type RPS15. In summary, we provide novel insights into the heterogeneous genetic landscape of CLL relapsing after FCR treatment and highlight a novel mechanism underlying clinical aggressiveness involving a mutated ribosomal protein, potentially representing an early