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Analysis of C9orf72 hexanucleotide repeat expansions in SOD1 negative patients with amyotrophic lateral sclerosis
Pathology (2015) 47(S1), pp. S27–S30
Genetics
THE CHANGING LANDSCAPE OF CHROMOSOME MARKERS Dimitar N. Azmanov1, Con Ngo1, Soruba Sivamoorty1, Vikki Britton1, Joanne Peverall1, Karen Woodward1, Ashleigh Murch1, David Ravine1 and Kym Mina1,2 1Department of Diagnostic Genomics, PathWest Laboratory Medicine, and 2School of Pathology and Laboratory Medicine, University of Western Australia, WA, Australia The phenomenon of multiple supernumerary marker chromosomes (SMCs) is a rare event (http://ssmc-tl.com/sSMC.html). It is typically accompanied by somatic mosaic variation in the number of intracellular marker chromosomes and their morphology. This dynamic variation is associated with mitosis and complicates diagnosis, as well as predictions of the clinical severity. Until now, the mechanism of SMC formation remains poorly understood, with associated uncertainties about the familial recurrence risk. We describe a paediatric patient with multiple congenital abnormalities and developmental delay associated with dynamic mosaicism for up to three SMCs. Using a combination of conventional and molecular cytogenetic techniques, we clarified the origins of SMCs from five different genomic locations (1q21.1q21.3, 1q21.3q22, 8q11.1q11.23, 9p13.1p12 and 14q11.2). SNP microarray data and family-trio analysis excluded the possibility of secondary uniparental disomy, which may sometimes occur as a result of an accompanying chromosomal rescue event. It also demonstrated that the markers were of maternal origin and consistent with a meiosis II error. Our observations, in concert with published findings from in vitro fertilisation experiments, allow us to propose a novel mechanism to explain the origin of multiple SMCs.
Cheng Yee Chan, Howard Potter and Peter George Department of Molecular Pathology, Canterbury Health Laboratories, Christchurch, New Zealand Amyotrophic lateral sclerosis (ALS) is a neurodegenerative condition that forms part of the spectrum of motor neurone disease, which results in progressive weakness and has a poor prognosis. Over the last twenty years, mutations in the SOD1 gene were thought to be the most common cause of the familial form of ALS, accounting for 20% of cases. However, in the last couple of years, hexanucleotide repeat expansions within the C9orf72 gene have been discovered to account for up to 40% of familial ALS, and are now recognised to be the most common known cause. We have recently developed a new assay to analyse C9orf72 expansions, and sought to investigate this in samples that had previously been investigated for SOD1 mutations and found to be negative, over the period of 2003 to 2014. A total of 32.9% (23/70) samples were found to carry pathogenic C9orf72 expansions (>30 repeats), while 2.9% (2/70) were borderline (20–30 repeats) and 50.0% (35/70) were normal (<20 repeats). This analysis has enabled the genetic diagnosis of ALS in a large proportion of ALS patients in whom a genetic cause has not previously been identified, and opens up the possibility of screening for these expansions in appropriately-counselled family members.
PROTEOMICS AND MITOCHONDRIAL DISEASE Michael T. Ryan and David A. Stroud Department of Biochemistry and Molecular Biology, Monash University, Vic, Australia
ARRAYS: HAS THE TIME COME FOR HAEMATOLOGICAL MALIGNANCY? Peter A. Kaub, Janice M. Fletcher and Sarah Moore Genetics and Molecular Pathology, SA Pathology, Adelaide, SA, Australia Single nucleotide polymorphism (SNP) microarray technology allows elucidation of a virtual karyotype at much greater resolution than traditional cytogenetic techniques. The size of deletions of a bi- or mono-allelic nature, clone size and concomitant abnormalities can be assessed in a single test. Is array therefore inherently better than traditional karyotyping? At what stage should it become the new standard of care? A case presentation for a patient with chronic lymphocytic leukaemia (CLL) will be presented to provide the evidence for advantages of SNP array over classical karyotyping. A summary of recent CLL cases analysed by SNP array from our centre will be discussed in terms of diagnostic and prognostic value, supporting the case that, at least for CLL, SNP arrays are ready to supplant classical karyotyping for diagnosis. Print ISSN 0031-3025/Online ISSN 1465-3931
ANALYSIS OF C9orf72 HEXANUCLEOTIDE REPEAT EXPANSIONS IN SOD1 NEGATIVE PATIENTS WITH AMYOTROPHIC LATERAL SCLEROSIS
#
Disorders of mitochondrial oxidative phosphorylation (OXPHOS) present with almost any symptom, at any age with any mode of inheritance. OXPHOS disorders are the most common group of inherited metabolic diseases and occur in 1/5000 births. There is no ‘gold standard’ diagnostic test and most paediatric diagnoses rely on finding an enzyme defect while molecular investigations are typically restricted to testing for common mutations or sequencing of specific candidate genes. While next generation sequencing (NGS) approaches have yielded great insights into the identification of pathogenic mutations, these generally focus on sequencing of known mitochondrial and disease related genes, yielding a molecular diagnosis in 60% of cases. Recent advances in proteomics have improved such that much of the human cellular proteome can now be determined in a matter of hours. In addition, with the use of quantitaive approaches including stable isotope labeling of cells in culture (SILAC), differences between the proteomes of diseased and controls cells can be measured. We are now investigating the use of quantitative proteomics as a complementery tool to NGS in the molecular diagnosis of
2015 Royal College of Pathologists of Australasia
Copyright © Royal College of pathologists of Australasia. Unauthorized reproduction of this article is prohibited.
Genetics
THE CHANGING LANDSCAPE OF CHROMOSOME MARKERS Dimitar N. Azmanov1, Con Ngo1, Soruba Sivamoorty1, Vikki Britton1, Joanne Peverall1, Karen Woodward1, Ashleigh Murch1, David Ravine1 and Kym Mina1,2 1Department of Diagnostic Genomics, PathWest Laboratory Medicine, and 2School of Pathology and Laboratory Medicine, University of Western Australia, WA, Australia The phenomenon of multiple supernumerary marker chromosomes (SMCs) is a rare event (http://ssmc-tl.com/sSMC.html). It is typically accompanied by somatic mosaic variation in the number of intracellular marker chromosomes and their morphology. This dynamic variation is associated with mitosis and complicates diagnosis, as well as predictions of the clinical severity. Until now, the mechanism of SMC formation remains poorly understood, with associated uncertainties about the familial recurrence risk. We describe a paediatric patient with multiple congenital abnormalities and developmental delay associated with dynamic mosaicism for up to three SMCs. Using a combination of conventional and molecular cytogenetic techniques, we clarified the origins of SMCs from five different genomic locations (1q21.1q21.3, 1q21.3q22, 8q11.1q11.23, 9p13.1p12 and 14q11.2). SNP microarray data and family-trio analysis excluded the possibility of secondary uniparental disomy, which may sometimes occur as a result of an accompanying chromosomal rescue event. It also demonstrated that the markers were of maternal origin and consistent with a meiosis II error. Our observations, in concert with published findings from in vitro fertilisation experiments, allow us to propose a novel mechanism to explain the origin of multiple SMCs.
Cheng Yee Chan, Howard Potter and Peter George Department of Molecular Pathology, Canterbury Health Laboratories, Christchurch, New Zealand Amyotrophic lateral sclerosis (ALS) is a neurodegenerative condition that forms part of the spectrum of motor neurone disease, which results in progressive weakness and has a poor prognosis. Over the last twenty years, mutations in the SOD1 gene were thought to be the most common cause of the familial form of ALS, accounting for 20% of cases. However, in the last couple of years, hexanucleotide repeat expansions within the C9orf72 gene have been discovered to account for up to 40% of familial ALS, and are now recognised to be the most common known cause. We have recently developed a new assay to analyse C9orf72 expansions, and sought to investigate this in samples that had previously been investigated for SOD1 mutations and found to be negative, over the period of 2003 to 2014. A total of 32.9% (23/70) samples were found to carry pathogenic C9orf72 expansions (>30 repeats), while 2.9% (2/70) were borderline (20–30 repeats) and 50.0% (35/70) were normal (<20 repeats). This analysis has enabled the genetic diagnosis of ALS in a large proportion of ALS patients in whom a genetic cause has not previously been identified, and opens up the possibility of screening for these expansions in appropriately-counselled family members.
PROTEOMICS AND MITOCHONDRIAL DISEASE Michael T. Ryan and David A. Stroud Department of Biochemistry and Molecular Biology, Monash University, Vic, Australia
ARRAYS: HAS THE TIME COME FOR HAEMATOLOGICAL MALIGNANCY? Peter A. Kaub, Janice M. Fletcher and Sarah Moore Genetics and Molecular Pathology, SA Pathology, Adelaide, SA, Australia Single nucleotide polymorphism (SNP) microarray technology allows elucidation of a virtual karyotype at much greater resolution than traditional cytogenetic techniques. The size of deletions of a bi- or mono-allelic nature, clone size and concomitant abnormalities can be assessed in a single test. Is array therefore inherently better than traditional karyotyping? At what stage should it become the new standard of care? A case presentation for a patient with chronic lymphocytic leukaemia (CLL) will be presented to provide the evidence for advantages of SNP array over classical karyotyping. A summary of recent CLL cases analysed by SNP array from our centre will be discussed in terms of diagnostic and prognostic value, supporting the case that, at least for CLL, SNP arrays are ready to supplant classical karyotyping for diagnosis. Print ISSN 0031-3025/Online ISSN 1465-3931
ANALYSIS OF C9orf72 HEXANUCLEOTIDE REPEAT EXPANSIONS IN SOD1 NEGATIVE PATIENTS WITH AMYOTROPHIC LATERAL SCLEROSIS
#
Disorders of mitochondrial oxidative phosphorylation (OXPHOS) present with almost any symptom, at any age with any mode of inheritance. OXPHOS disorders are the most common group of inherited metabolic diseases and occur in 1/5000 births. There is no ‘gold standard’ diagnostic test and most paediatric diagnoses rely on finding an enzyme defect while molecular investigations are typically restricted to testing for common mutations or sequencing of specific candidate genes. While next generation sequencing (NGS) approaches have yielded great insights into the identification of pathogenic mutations, these generally focus on sequencing of known mitochondrial and disease related genes, yielding a molecular diagnosis in 60% of cases. Recent advances in proteomics have improved such that much of the human cellular proteome can now be determined in a matter of hours. In addition, with the use of quantitaive approaches including stable isotope labeling of cells in culture (SILAC), differences between the proteomes of diseased and controls cells can be measured. We are now investigating the use of quantitative proteomics as a complementery tool to NGS in the molecular diagnosis of
2015 Royal College of Pathologists of Australasia
Copyright © Royal College of pathologists of Australasia. Unauthorized reproduction of this article is prohibited.