Introduction: Through previous studies using Sanger sequencing and targeted gene sequencing, we found that the frequencies of certain genes in Chinese patients with chronic lymphocytic leukemia (CLL) were different from those in CLL patients from the western countries. However, until now, there are no published whole-exome sequencing (WES) studies in Chinese CLL patients. To better define the genomic landscape of CLL cases in China, we conducted WES studies of CLL cases in our center.
Methods: This study include 43 CLL patients, of whom 40 were untreated at the time of sampling. The median age of the 43 patients were 61 years, the male-to-female ratio was 2:1. We isolated peripheral blood mononuclear cells, then CD19 positive cells were purified by using magnetic microbeads in some cases. The buccal cells or the leukocytes that were depleted of CD19 positive cells were used as germline controls. We purified genomic DNA and used the purified DNA for library construction, after which we sequenced the library using the Illumina HiSeq2500 platform. After sequencing, software including Burrows-Wheeler Aligner (BWA) and GATK were used for the bioinformatic analysis.
Results: In the 43 Chinese patients with CLL, the median number of somatic mutations was 67 (range:10-128) and the median number of nonsynonymous mutations was 26 (range: 5-55). We found that the number of mutations in patients older than 60 was significantly higher than that in the younger patients. Somatic mutations were detected across 991 different genes, of which 24 genes were identified as drivers in the study from the Dana-Faber Cancer Institute (DFCI). The 24 genes included NOTCH1, SF3B1, MYD88, POT1, TP53 and so on. Seventy-nine genes were recurrently mutated and 13 genes were found to be mutated in 3 or more cases. The most frequently mutated gene was NOCTH1 (6 cases, 14.0%). FBXW7, another gene that is involved in the regulation of the NOTCH1 pathway, was also mutated in 3 cases. IGLL5, MUC4 and MYD88 were also among the most frequently mutated genes, with each of them being mutated in 5 cases (11.6%). We found that the frequencies of mutations in IGLL5 (11.6% vs. 2.2%, P=0.0056), MYD88 (11.6% vs. 3.0%, P=0.0147), BCOR (9.3% vs. 2.2%, P=0.0246), and SF3B1 (4.7% vs. 21.0%, P=0.0084) were significantly different between Chinese CLL patients and patients from western countries. BCOR-mutated patients had more frequent aberrations involving NOTCH1 pathway (NOTCH1 and/or FBXW7 mutation) than BCOR-unmutated patients (3/4 [75.0%] vs. 6/39 [15.4%], P=0.0242) (Figure 1A). Analysis of 10967 tumor samples from the Cancer Genome Atlas (TCGA) program revealed that tumors with BCOR mutations had a higher frequency of NOTCH1 aberrations (NOTCH1 and/or FBXW7 mutation) than those without BCOR mutations (116/349 [33.2%] vs. 722/10618 [6.8%]; odds ratio: 6.824, 95%CI: 5.393-8.634, P<0.0001) (Figure 1B). These data suggest BCOR mutations and NOTCH1 aberration may cooperate in the pathogenesis of CLL and other types of tumors. NFKBIA mutations were identified in 2 cases (Figure 1C) and we found that the mRNA level of NFKBIA was significantly lower in the NFKBIA mutated CLL cells than in the normal B cells (Figure 1D-E); in the MEC-1 cell line, significant activation of NF-кB pathway was observed after knockdown of NFKBIA using siRNA (Figure 1F).
Conclusion: Patients with CLL in China and patients with CLL in the western countries have different mutational landscapes. BCOR gene mutations may cooperate with aberrations involving the NOTCH1 pathway in the pathogenesis of CLL. NFKBIA could be a potential tumor suppressor in the pathogenesis of CLL.
No relevant conflicts of interest to declare.
