Identification of Males with Cryptic Fragile X Alleles by Methylation-Specific Quantitative Melt Analysis

Clinical Chemistry

Lorena Santa Maria⁴  Xin Li⁵  Paulina Morales⁴  David Francis⁵  David J. Amor³  Desiree Du Sart⁵  Solange M. Aliaga¹  Bianca Curotto⁴  Cesar Trigo²  Víctor Faundes⁴  Angelica M. Alliende²  David E. Godler⁵  Howard R. Slater³  Isabel Salas² 

[1] Cyto-molecular Diagnostic Research Laboratory, Victorian Clinical Genetics Services and Murdoch Children`s Research Institute, Royal Children`s Hospital, Melbourne, Victoria, Australia;Department of Paediatrics, University of Melbourne, Melbourne, Victoria, Australia;Cytogenetics and Molecular Laboratory, Institute of Nutrition and Food Technology, University of Chile, Santiago, Chile;;Centre for Diagnosis and Treatment of Fragile X Syndrome, INTA University of Chile, Santiago, Chile.Cyto-molecular Diagnostic Research Laboratory, Victorian Clinical Genetics Services and Murdoch Children`s Research Institute, Royal Children`s Hospital, Melbourne, Victoria, Australia;Department of Paediatrics, University of Melbourne, Melbourne, Victoria, Australia;;Cytogenetics and Molecular Laboratory, Institute of Nutrition and Food Technology, University of Chile, Santiago, Chile;Centre for Diagnosis and Treatment of Fragile X Syndrome, INTA University of Chile, Santiago, Chile.Cyto-molecular Diagnostic Research Laboratory, Victorian Clinical Genetics Services and Murdoch Children`s Research Institute, Royal Children`s Hospital, Melbourne, Victoria, Australia;

DOI  :  10.1373/clinchem.2015.244681

学科分类：过敏症与临床免疫学

来源: American Association for Clinical Chemistry

【 摘 要 】

BACKGROUND: FMR1 full mutations (FMs) (CGG expansion >200) in males mosaic for a normal (<45 CGG) or gray-zone (GZ) (45–54 CGG) allele can be missed with the standard 2-step fragile X syndrome (FXS) testing protocols, largely because the first-line PCR tests showing a normal or GZ allele are not reflexed to the second-line test that can detect FM.METHODS: We used methylation-specific quantitative melt analysis (MS-QMA) to determine the prevalence of cryptic FM alleles in 2 independent cohorts of male patients (994 from Chile and 2392 from Australia) referred for FXS testing from 2006 to 2013. All MS-QMA–positive cases were retested with commercial triplet primed PCR, methylation-sensitive Southern blot, and a methylation-specific EpiTYPER-based test.RESULTS: All 38 FMs detected with the standard 2-step protocol were detected with MS-QMA. However, MS-QMA identified methylation mosaicism in an additional 15% and 11% of patients in the Chilean and Australian cohorts, respectively, suggesting the presence of a cryptic FM. Of these additional patients, 57% were confirmed to carry cryptic expanded alleles in blood, buccal mucosa, or saliva samples. Further confirmation was provided by identifying premutation (CGG 55–199) alleles in mothers of probands with methylation-sensitive Southern blot. Neurocognitive assessments showed that low-level mosaicism for cryptic FM alleles was associated with cognitive impairment or autism.CONCLUSIONS: A substantial number of mosaic FM males who have cognitive impairment or autism are not diagnosed with the currently recommended 2-step testing protocol and can be identified with MS-QMA as a first-line test.

【 授权许可】

Unknown