Published online before print December 28, 2007

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Copyright © 2008 American Society for Investigative Pathology
Journal of Molecular Diagnostics, doi:10.2353/jmoldx.2008.070086

Accepted for publication September 20, 2007.

Article

Mutation Screening of EXT1 and EXT2 by Denaturing High-Performance Liquid Chromatography, Direct Sequencing Analysis, Fluorescence in Situ Hybridization, and a New Multiplex Ligation-Dependent Probe Amplification Probe Set in Patients with Multiple Osteochondromas

Ivy Jennes^*, Mark M. Entius, Els Van Hul^*, Alessandro Parra, Luca Sangiorgi, and Wim Wuyts^*@

From the Department of Medical Genetics,* University and University Hospital of Antwerp, Antwerp, Belgium; MRC-Holland, Amsterdam, The Netherlands; and the Medical Genetics Unit, Rizzoli Orthopaedic Institute, Bologna, Italy

^@ To whom correspondence should be addressed. E-mail: wim.wuyts{at}ua.ac.be.

	Abstract

Multiple osteochondromas (MO) is an autosomal-dominant skeletal disorder characterized by the formation of multiple cartilage-capped protuberances. MO is genetically heterogeneous and is associated with mutations in the EXT1 and EXT2 genes. In this study we describe extensive mutation screening in a set of 63 patients with clinical and radiographical diagnosis of MO. Denaturing high-performance liquid chromatography analysis revealed mutations in 43 patients. Additional deletion analysis by fluorescence in situ hybridization and a newly developed multiplex ligation-dependent probe amplification probe set identified one patient with an intragenic EXT1 translocation, three patients with a partial EXT1 deletion, and one patient with a partial EXT2 deletion. Thirty-six patients harbored an EXT1 mutation (57%), and 12 had an EXT2 mutation (19%). We show that our optimized denaturing high-performance liquid chromatography/sequencing/multiplex ligation-dependent probe amplification protocol represents a reliable and highly sensitive diagnostic strategy for mutation screening in MO patients. Clinical analysis showed no clear genotype-phenotype correlation in our cohort of MO patients.