Whole-Exome Sequencing in Myelodysplastic Syndromes with 5q Deletion
Authors: Adema Vera , Palomo Laura , Díez-Campelo María , Mallo Mar , Arenillas Leonor , Luño Elisa , Perez-Ladaga Albert , Yost Shawn , Zamora Lurdes , Gomez-Marzo Paula , Xicoy Blanca , Pedro Carmen , Cañizo María Consuelo , Sanzo Carmen , Lopez Rosario , Lemes Angelina , Raya Jose Maria , Rios Eduardo , Arnan Montserrat , Paz Raquel , sanchez-Garcia Joaquin , Florensa Lourdes , Gut Ivo G. , Bayes Monica , Bejar Rafael , Sole Francesc 
Published: October 2019

Abstract
INTRODUCTION
Myelodysplastic syndromes (MDS) are a heterogeneous group of clonal myeloid stem cell disorders that are highly prevalent in elderly populations. MDS are characterized by inefficient hematopoiesis, peripheral blood (PB) cytopenias, and increased risk of transformation to acute myeloid leukemia (AML; 20–30% of patients with MDS). Around 50% of MDS patients carry at least one karyotypic aberration. The interstitial deletion of the long arm of chromosome 5 ([del(5q)] is the most common aberration, accounting for almost 30% of abnormal MDS karyotype. Various studies supports a favorable prognosis of MDS with isolated del(5q) with an excellent response to lenalidomide treatment. In order to describe the molecular events associated with MDS and del(5q) we performed whole-exome sequencing (WES)(assessing 334,378 exons) of tumor-normal paired samples from 20 MDS patients to unravel the genetic basis of MDS with del(5q). The analysis is ongoing and the complete results will be presented in the meeting.
METHODS
A total of 50 samples from 20 patients with MDS, with del(5q) were collected. For each diagnostic sample, we performed Conventional G-banding cytogenetics and fluorescence in situ hybridization (FISH, to confirm or dismiss del(5q)) and SNP arrays with Cytoscan HD (Affymetrix). These samples included: 20 tumor samples at diagnosis, 20 control samples and 10 samples after diagnosis, during lenalidomide treatment (5) or at the moment of relapse (5) in order to compare the genetic status before and during the treatment. Genomic DNA from tumor cells was obtained from bone marrow (BM) samples or from PB granulocytes. As a source of constitutional DNA we used CD3+T cells from each patient by isolating by magnetic-activated cell sorting.
WES targeted capture was carried out on 7μg of genomic DNA, using the SureSelect Human Exome Kit 51Mb version 4.Libraries were sequenced on an Illumina HiSeq2000. Sequencing data will be analyzed using an in-house bioinformatics pipeline as previously reported.
RESULTS
Our preliminary analysis of these 20 new patients by WES confirmed our previous analyses with mutations in well described genes as ASXL1, JAK2 and TET2, but not in genes RUNX1, SF3B1 and SRSF2. In those patients we found two patients with missense mutation in TP53, one of the patients had an isolated del(5q) and is receiving lenalidomide treatment, and the other one had a complex karyotype. According to our prior analyses, in which 249 non-silent somatic variants were detected, we look forward to validate these mutations in this new series of patients.
CONCLUSIONS
We envision to validate these previous results with the new sequencing data of more patients with MDS and del(5q). We expect to measure somatic mutations that vary in abundance after lenalidomide treatment, potentially identifying mutations associated with resistance or relapse.
ACKNOWLEDGEMENTS: This work has been supported (in part) by a grants from Instituto de Salud Carlos III, Ministerio de Sanidad y Consumo, Spain (PI 11/02010); by Red Temática de Investigación Cooperativa en Cáncer (RTICC, FEDER) (RD07/0020/2004; RD12/0036/0044); 2014 SGR225 (GRE) Generalitat de Catalunya; Fundació Internacional Josep Carreras; Obra Social “la Caixa”; Sociedad Española de Hematología y Hemoterapia (SEHH)and Celgene Spain.
FOOTNOTES
Rafael Bejar and Francesc Sole contributed equally.

Disclosures
Díez-Campelo: Novartis, Celgene: Honoraria, Research Funding. Xicoy:Celgene: Honoraria. Cañizo:Celgene, Jansen-Cilag, Arry, Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. sanchez-Garcia:Celgene: Honoraria, Research Funding. Bejar:Celgene: Membership on an entity's Board of Directors or advisory committees; Genoptix Medical Laboratory: Consultancy, Honoraria, Licensed IP, no royalties Patents & Royalties, Membership on an entity's Board of Directors or advisory committees. Sole:Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding.