Abstract 2428: Genetic aberrations in the DNA repair pathway among children with Philadelphia chromosome positive leukemias
Authors: Andriano Nellina , Bonaccorso Paola , Iachelli Valeria , La Rosa Manuela , Cannata Emanuela , Lo Nigro Luca 
Published: July 2016

Abstract
Background. Aberrations in the DNA repair pathway among children with leukemia are largely unknown. Knowledge of such mutations may improve the understanding of tumorigenesis, direct patient care, and enable genetic counseling of patients and families. Germline aberrations of the DNA-repair machinery are crucial for generating chromosomal instability and occurrence of acute leukemia in children. In order to better understand this mechanism, we studied a specific subgroup of childhood leukemia, e.g. t(9;22) or chromosome Philadelphia positive (Ph+) acute lymphoblastic and chronic myeloid leukemias. We addressed our research to specific genes (Nijmegen Breakage Syndrome (NBS1) and Fanconi Anemia) involved in the DNA repair pathway, in the attempt to identify mutations and aberrant expressions, predisposing and/or cooperating with the leukemogenic process.
Materials and Methods. We analyzed diagnostic and remission samples of 12 children: 8 with a Ph+ acute lymphoblastic leukemia (ALL) and 4 with a chronic myeloid leukemia (CML) diagnosed and treated at our institution from 1999 to 2013. We performed RT-PCR analyses to detect mutations in exons 3-6 of the NBS1 gene, followed by a direct sequencing analysis (BMR - Padova - Italy). The status of FANCD2 and PALB2 genes was studied by a multiplex ligation-dependent probe amplification (MLPA) using the Salsa MLPA P057-025R FANCD2-PALB2 Probemix and SALSA MLPA EK1 kit (MRC-Holland, Amsterdam, the Netherlands). Samples from five healthy donors (HDs) were used as wild-type controls. For data elaboration, we used Coffalyser.Net software for MLPA. Moreover we performed a real-time PCR amplification of BRCA1 (exons 14-15) and BRCA2 (exons 15-16) genes, respectively, calculating the median fold-changes (MFC) in patients comparing with HDs.
Results. We detected the NBS1 E185Q polymorphism (c533G&gt;C, rs1805794) in both diagnostic and remission samples in 3 out of 12 children with a Ph positive acute leukemia (2 ALL, 1 CML). We also found two pairs of deletions in FANCD2 (Δ32-35; Δ35-38) and PALB2 (Δ2; Δ2-6) in 4 out of 12 Ph positive cases (2 ALL and 2 CML). These aberrations were mutually exclusive. Thus a total of 7 cases (58%) showed an aberrant pathway. Moreover we determined that the MFC of BRCA1 expression among children with Ph positive ALL and CML was slightly higher than HDs (1,388 vs 0,433 and 1,625 vs 0,451, respectively). Surprisingly the MFC of BRCA2 expression was strongly higher than HDs in both children with Ph positive ALL (5,924 vs 0,002) and CML (54,504 vs 0,256).
Conclusion. Our preliminary findings strongly suggest that germline mutations or deletions in association with aberrant expression of specific genes that regulate the DNA repair machinery may contribute to the chromosomal instability in children with Ph positive leukemias. These data will be confirmed in a larger population of different subtypes of pediatric leukemia.
Citation Format: Nellina Andriano, Paola Bonaccorso, Valeria Iachelli, Manuela La Rosa, Emanuela Cannata, Luca Lo Nigro. Genetic aberrations in the DNA repair pathway among children with Philadelphia chromosome positive leukemias. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2428.