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BioMed Research International
Volume 2016, Article ID 4247908, 5 pages
http://dx.doi.org/10.1155/2016/4247908
Research Article

A Novel Assay for the Identification of NOTCH1 PEST Domain Mutations in Chronic Lymphocytic Leukemia

1Departments of Hematology and Clinical Pathology, and Research Institute, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil
2Department of Hematology, Universidade Estadual de Campinas (Hemocentro-Unicamp), Campinas, SP, Brazil
3Department of Clinical Pathology, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil
4Hematology Service, Hospital das Clínicas, Faculdade de Medicina da Universidade de São Paulo São Paulo, SP, Brazil
5Cytogenetics Laboratories, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil
6Hemocentro, Faculdade de Medicina da Universidade Estadual de Campinas (Hemocentro-Unicamp), Campinas, SP, Brazil
7Bone Marrow Transplantation Center, Instituto Nacional de Câncer (INCA), Rio de Janeiro, RJ, Brazil
8Paediatric Haematology-Oncology Program, Instituto Nacional de Câncer (INCA), Rio de Janeiro, RJ, Brazil
9Department of Hematology, Faculdade de Medicina da Santa Casa de Misericórdia de São Paulo, São Paulo, SP, Brazil
10Department of Hematology, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil

Received 9 June 2016; Revised 5 October 2016; Accepted 18 October 2016

Academic Editor: Carlo Visco

Copyright © 2016 Paulo Vidal Campregher et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Aims. To develop a fast and robust DNA-based assay to detect insertions and deletions mutations in exon 34 that encodes the PEST domain of NOTCH1 in order to evaluate patients with chronic lymphocytic leukemia (CLL). Methods. We designed a multiplexed allele-specific polymerase chain reaction (PCR) combined with a fragment analysis assay to detect specifically the mutation c.7544_7545delCT and possibly other insertions and deletions in exon 34 of NOTCH1. Results. We evaluated our assay in peripheral blood samples from two cohorts of patients with CLL. The frequency of NOTCH1 mutations was 8.4% in the first cohort of 71 unselected CLL patients. We then evaluated a second cohort of 26 CLL patients with known cytogenetic abnormalities that were enriched for patients with trisomy 12. NOTCH1 mutations were detected in 43.7% of the patients with trisomy 12. Conclusions. We have developed a fast and robust assay combining allele-specific PCR and fragment analysis able to detect NOTCH1 PEST domain insertions and deletions.