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Mutational analysis of EXT1 and EXT2 genes and a newsystem for “grading”
clinical expression of disease in patients with Hereditary Multiple Exostoses.
Abstract 2005 MHE Conference
1Luca Sangiorgi, 2Nicola Fabbri, 2Laura Campanacci,1Elena Pedrini,
1Veronica Maini, 1Silvia Capponcelli, 1Marina Mordenti and
2Mario Mercuri.1Genetics Unit, 2V Surgery Division,
Rizzoli Orthopedic Institute, Bologna, Italy.
Introduction: Several mutations dispersed throughout the EXT1 and EXT2 genes have been
identified in HME patients. Most of these are responsible of the truncation of EXT1 or EXT2
proteins which are likely to become inactive and degrade rapidly, resulting in a nearly complete loss-
of-function. A prospective longitudinal study was undertaken to correlate genotype and
phenotype of the disease in families and sporadic cases, with the purpose of defining the disease
spectrum of expression.
Method: A multidisciplinary clinic, involving geneticists and orthopaedic surgeons, is weekly
carried out at the Rizzoli Institute. In order to evaluate the spectrum and distribution of mutations
leading to HME in the Rizzoli cohort of patients, a new multi-step DHPLC-based mutation screening
method was optimised. For informative families, we adopted a pre-screening linkage analysis to
selectively focus the DHPLC testing on either EXT1 or EXT2. Patient clinical assessment is carried
out using a new classification system substantially based on deformity and functional limitation and
the genotype-phenotype study is performed using this system.
Results: In a small pilot we enrolled 36 unrelated probands, representing 20 families and 14
sporadic cases for the presence of mutations in either EXT1 or EXT2 genes. Thirty-one out of 36
probands (86%) had mutations either in EXT1 (24/31; 77%) or EXT2 (7/31; 23%), mainly
distributed in the amino-terminal region of the proteins and the vast majority of them are
responsible of a premature protein truncation.
Most mutations were distributed towards the 5’ end of EXT1 and EXT2 genes.In accordance with
the most recent data, 23 of 31 mutations (74%) were novel. No novel missense or splice site
mutations were detected in 200 control chromosomes. All splice site mutations were present in the
highly conserved AG and GT positions of the splice acceptor and donor junctions, and only one of
these (c.IVS6+1G>T in the EXT1 gene) had been previously reported in the HGMD database. The
3 missense mutations clustered within exon 2 of the EXT1 gene and caused amino acid
substitutions in residues 339 and 340. Two of these (c.1018C>T (p.R340C), c.1019G>A (p.
R340H)) had been already described whereas the third one(c.1016G>T (p.G339V)) was a novel
mutation responsible for substitution of a glycine codon with a valine within an important key
element for EXT1 function. The overall mutation frequency in 36 probands was 86% (67% in EXT1
and 19% in EXT2). Mutations were found in 10 of 14 sporadic cases (71%) and in 20 of 21 familial
cases (95%). No disease-causing mutation has been detected in 5 of 36 patients.
A new clinical classification based on clinical evaluation and orthopaedic problems has been defined.
A preliminary study performed on 153 pts have shown that most (80%) are not substantially
limited by the disease. Using the DHPLC based screening technique and the clinical classification
system, we are currently running a genotype/phenotype study on 170 patients. Preliminary results
indicate that mutations on specific exons of the EXT1 gene correlate with more relevant clinical
limitations.
Conclusions: Our results identified several novel and many private mutations in a large cohort of
patients, confirming the strong allelic heterogeneity of EXT1 and EXT2 genes. Our optimised
DHPLC-based approach represents a reliable, efficient and highly sensitive diagnostic strategy for
rapid detection of germline mutations in HME patients. The genotype-phenotype study is giving
indication regarding association of alterations and clinical presentation of the disease.
clinical expression of disease in patients with Hereditary Multiple Exostoses.
Abstract 2005 MHE Conference
1Luca Sangiorgi, 2Nicola Fabbri, 2Laura Campanacci,1Elena Pedrini,
1Veronica Maini, 1Silvia Capponcelli, 1Marina Mordenti and
2Mario Mercuri.1Genetics Unit, 2V Surgery Division,
Rizzoli Orthopedic Institute, Bologna, Italy.
Introduction: Several mutations dispersed throughout the EXT1 and EXT2 genes have been
identified in HME patients. Most of these are responsible of the truncation of EXT1 or EXT2
proteins which are likely to become inactive and degrade rapidly, resulting in a nearly complete loss-
of-function. A prospective longitudinal study was undertaken to correlate genotype and
phenotype of the disease in families and sporadic cases, with the purpose of defining the disease
spectrum of expression.
Method: A multidisciplinary clinic, involving geneticists and orthopaedic surgeons, is weekly
carried out at the Rizzoli Institute. In order to evaluate the spectrum and distribution of mutations
leading to HME in the Rizzoli cohort of patients, a new multi-step DHPLC-based mutation screening
method was optimised. For informative families, we adopted a pre-screening linkage analysis to
selectively focus the DHPLC testing on either EXT1 or EXT2. Patient clinical assessment is carried
out using a new classification system substantially based on deformity and functional limitation and
the genotype-phenotype study is performed using this system.
Results: In a small pilot we enrolled 36 unrelated probands, representing 20 families and 14
sporadic cases for the presence of mutations in either EXT1 or EXT2 genes. Thirty-one out of 36
probands (86%) had mutations either in EXT1 (24/31; 77%) or EXT2 (7/31; 23%), mainly
distributed in the amino-terminal region of the proteins and the vast majority of them are
responsible of a premature protein truncation.
Most mutations were distributed towards the 5’ end of EXT1 and EXT2 genes.In accordance with
the most recent data, 23 of 31 mutations (74%) were novel. No novel missense or splice site
mutations were detected in 200 control chromosomes. All splice site mutations were present in the
highly conserved AG and GT positions of the splice acceptor and donor junctions, and only one of
these (c.IVS6+1G>T in the EXT1 gene) had been previously reported in the HGMD database. The
3 missense mutations clustered within exon 2 of the EXT1 gene and caused amino acid
substitutions in residues 339 and 340. Two of these (c.1018C>T (p.R340C), c.1019G>A (p.
R340H)) had been already described whereas the third one(c.1016G>T (p.G339V)) was a novel
mutation responsible for substitution of a glycine codon with a valine within an important key
element for EXT1 function. The overall mutation frequency in 36 probands was 86% (67% in EXT1
and 19% in EXT2). Mutations were found in 10 of 14 sporadic cases (71%) and in 20 of 21 familial
cases (95%). No disease-causing mutation has been detected in 5 of 36 patients.
A new clinical classification based on clinical evaluation and orthopaedic problems has been defined.
A preliminary study performed on 153 pts have shown that most (80%) are not substantially
limited by the disease. Using the DHPLC based screening technique and the clinical classification
system, we are currently running a genotype/phenotype study on 170 patients. Preliminary results
indicate that mutations on specific exons of the EXT1 gene correlate with more relevant clinical
limitations.
Conclusions: Our results identified several novel and many private mutations in a large cohort of
patients, confirming the strong allelic heterogeneity of EXT1 and EXT2 genes. Our optimised
DHPLC-based approach represents a reliable, efficient and highly sensitive diagnostic strategy for
rapid detection of germline mutations in HME patients. The genotype-phenotype study is giving
indication regarding association of alterations and clinical presentation of the disease.
Dr. Sangiorgi serves on the Scientific and Medical Advisory Board of the MHE Research Foundation
Research authored by Dr. Sangiorgi
Click the tab and a window will appear.
List of Publications via PubMed
(NIH National Library of Medicine)
Research authored by Dr. Sangiorgi
Click the tab and a window will appear.
List of Publications via PubMed
(NIH National Library of Medicine)
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Poster of MHE Research at the CTOS conference, Genotype-Phenotype Study in 240 MO Patients
PDF file link
PDF file link
Poster of MHE Research at the CTOS conference EXT - Mutation Analysis in Sporadic and
Hereditary Osteochondroma Secondary Chondrocarcoma Tissues and Peripheral Linphocytes PDF file link
Hereditary Osteochondroma Secondary Chondrocarcoma Tissues and Peripheral Linphocytes PDF file link
| Dr. Sangiorgi's research |
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Press Release 1 / 19 / 08
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{dagger}, Els Van Hul*, Alessandro Parra{ddagger}, Luca Sangiorgi
{ddagger} and Wim Wuyts*
To Read this publication Click Here
Both Luca Sangiorgi, M.D., Ph.D. and Wim Wuyts, Ph.D. are members of our Scientific & Medical
Advisory Board and our foundation helped support this research
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{dagger}, Els Van Hul*, Alessandro Parra{ddagger}, Luca Sangiorgi
{ddagger} and Wim Wuyts*
To Read this publication Click Here
Both Luca Sangiorgi, M.D., Ph.D. and Wim Wuyts, Ph.D. are members of our Scientific & Medical
Advisory Board and our foundation helped support this research
|
Questions to the Scientific & Medical Advisory Board,
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AdvisoryBoard@mheresearchfoundation.org
Please use the contact Scientific & Medical Advisory Board Tab or you may email directly
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Have a Question or Comments for the Board of Directors,
Please use the contact Board of Directors Tab or General enquiries can be emailed to
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By the Health On the Net Foundation. Click here to verify.# HON Conduct 282463 and is linked on the NIH
National Library of Medicine, Directory of Health Organizations (SIS) website, as well as the link for Patient
Information on The Diseases Database a cross-referenced index of human disease, and the Intute: health & life
sciences a free online service providing access to the very best Web resources for education and research
located in the UK
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