MDS are a group of myeloid neoplasms characterized by deregulated blood cell production and a high propensity to AML. Although a number of gene alterations have been implicated in the pathogenesis of MDS, they do not fully explain the pathogenesis of MDS. So, in order to clarify a comprehensive registry of gene mutations in MDS, we performed whole-exome sequencing of 29 cases with MDS and related myeloid neoplasm. A total of 268 somatic mutations or 9.2 mutations per sample were identified. Among these 9 genes were mutated in more than 2 cases, which not only included a spectrum of known gene targets in MDS, but also affected previously unknown genes that are commonly involved in RNA splicing pathway, including U2AF35, SRSF2 and ZRSR2. Together with additional three (SF3A1, SF3B1 and PRPF40B) found in single cases, 16 (55.2%) of the 29 discovery cases carried a mutation affecting the component of the splicing machinery. To confirm the observation, we examined 9 spliceosome genes for mutations in a large set of myeloid neoplasms. In total, 219 mutations were identified in 209 out of the 582 samples of myeloid neoplasms. RNA splicing pathway mutations were highly specific to myelodysplasia, including 19 of 23 (83%) cases with RARS, 43 of 50 (86%) RCMD-RS, 68 of 155 (44%) other MDS, 48 of 88 (55%) CMML, and 16 of 62 (26%) secondary AML with MDS features with a string preference of SF3B1 mutations to RARS and RCMD-RS and of SRSF2 to CMML, while they were rare in cases with de novo AML and MPN. Significantly, these mutations occurred in an almost completely mutually exclusive manner among mutated cases, suggesting the importance of deregulated RNA splicing in the pathogenesis of MDS. RNA splicing plays critical roles in differentiation, development, and disease and is a major source for protein diversity in higher eukaryotes. Splicing pathway mutations in myelodysplasia commonly affected those components of the splicing complex that are engaged in the 3′ splice site recognition, strongly indicating production of unspliced or aberrantly spliced RNA species are incriminated for the pathogenesis of MDS. So, to clarify the effect of these splicing mutations on RNA splicing, we expressed the wild-type and the mutant U2AF35 or SRSF2 in HeLa cells and performed whole transcriptome analysis in these cells. The results of exon array showed that the wild-type U2AF35 promoted RNA splicing correctly, whereas the mutant U2AF35 inhibited this processes and rendered intronic sequences to remain unspliced. RNA sequencing additionally showed that the number of reads that encompassed the exon/intron junctions was significantly increased in mutant U2AF35-transduced cells. This result means that mutant U2AF35 actually induced impaired 3′-splice site recognition during pre-mRNA processing. In conclusion, our study demonstrated that abnormal RNA splicing caused by mutations of multiple genes on RNA splicing pathway is a common feature of myelodysplasia.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5119. doi:1538-7445.AM2012-5119