Suchergebnisse

[Background]: Chronic lymphocytic leukemia (CLL) is a highly genetically heterogeneous disease. Although CLL has been traditionally considered as a mature B cell leukemia, few independent studies have shown that the genetic alterations may appear in CD34+ hematopoietic progenitors. However, the presence of both chromosomal aberrations and gene mutations in CD34+ cells from the same patients has not been explored. [Methods]: Amplicon-based deep next-generation sequencing (NGS) studies were carried out in magnetically activated-cell-sorting separated CD19+ mature B lymphocytes and CD34+ hematopoietic progenitors (n = 56) to study the mutational status of TP53, NOTCH1, SF3B1, FBXW7, MYD88, and XPO1 genes. In addition, ultra-deep NGS was performed in a subset of seven patients to determine the presence of mutations in flow-sorted CD34+CD19− early hematopoietic progenitors. Fluorescence in situ hybridization (FISH) studies were performed in the CD34+ cells from nine patients of the cohort to examine the presence of cytogenetic abnormalities. [Results]: NGS studies revealed a total of 28 mutations in 24 CLL patients. Interestingly, 15 of them also showed the same mutations in their corresponding whole population of CD34+ progenitors. The majority of NOTCH1 (7/9) and XPO1 (4/4) mutations presented a similar mutational burden in both cell fractions; by contrast, mutations of TP53 (2/2), FBXW7 (2/2), and SF3B1 (3/4) showed lower mutational allele frequencies, or even none, in the CD34+ cells compared with the CD19+ population. Ultra-deep NGS confirmed the presence of FBXW7, MYD88, NOTCH1, and XPO1 mutations in the subpopulation of CD34+CD19− early hematopoietic progenitors (6/7). Furthermore, FISH studies showed the presence of 11q and 13q deletions (2/2 and 3/5, respectively) in CD34+ progenitors but the absence of IGH cytogenetic alterations (0/2) in the CD34+ cells. Combining all the results from NGS and FISH, a model of the appearance and expansion of genetic alterations in CLL was derived, suggesting that most of the genetic events appear on the hematopoietic progenitors, although these mutations could induce the beginning of tumoral cell expansion at different stage of B cell differentiation. [Conclusions]: Our study showed the presence of both gene mutations and chromosomal abnormalities in early hematopoietic progenitor cells from CLL patients. ; This work was partially supported by grants from the Spanish Fondo de Investigaciones Sanitarias PI12/00281, PI15/01471, Instituto de Salud Carlos III (ISCIII), European Regional Development Fund (ERDF) "Una manera de hacer Europa," Consejería de Educación, Junta de Castilla y León (SA085U16), Proyectos de Investigación del SACYL, Spain: GRS 1172/A/15, BIO/SA10/14, GRS 1343/A/16, and Fundación Española de Hematología y Hemoterapia (FEHH) and by a grant (RD12/0036/0069) from the Red Temática de Investigación Cooperativa en Cáncer (RTICC), Instituto de Salud Carlos III (ISCIII), Spanish Ministry of Economy and Competitiveness & European Regional Development Fund (ERDF) "Una manera de hacer Europa" (Innocampus; CEI-2010-1-0010), Fundación "Memoria Don Samuel Solórzano Barruso" 2016, and the European Union Seventh Framework Programme [FP7/2007–2013] under Grant Agreement no. 306242-NGS-PTL. MHS is fully supported by an "Ayuda predoctoral de la Junta de Castilla y León" by the Fondo Social Europeo (JCYL-EDU/346/2013 PhD scholarship). ; Peer Reviewed