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Microarray and Next Generation Sequencing: Complementary Technologies for Diagnosing Autosomal Recessive Disorders
Abstracts 30 Precision Medicine for Autism and Related Developmental Brain Disorders Invited Speaker: David Ledbetter, PhD
31 Microarray and Next Generation Sequencing: Complementary Technologies for Diagnosing Autosomal Recessive Disorders Alka Chaubey, Michael J. Friez, Barbara R. DuPont
Molecular technologies have advanced at a staggering pace and are quickly changing the conventional paradigm of clinical diagnostics. In the past decade, microarray has become the gold standard tool in the management of constitutional genetic disorders. Microarray has the unique ability to be utilized as a high resolution molecular karyotype for identifying large genomic changes (trisomies, large duplications, and large deletions) as well as cryptic microdeletions/microduplications involving discreet genomic locations. However, the diagnostic yield of this technology is limited to ~25–35%. This limitation is addressed by clinicians ordering sequencing of single genes, Next-Generation Sequencing (NGS) panels or Whole Exome Sequencing (WES) to identify the various genetic etiologies. In turn, the unique feature of Single Nucleotide Polymorphism microarrays can be utilized as a complement to the previously mentioned sequencing technologies, especially in patients with autosomal recessive disorders. We present a variety of cases where microarray and sequencing complemented each other for a clinical diagnosis: cases where one mutation was found by sequencing and the other change was a single exon deletion (Sanfilippo B syndrome, Phenylketonuria); cases where NGS identified a single nucleotide mutation and microarray a heterozygous intragenic deletion (Brittle Cornea syndrome, Aspartylglucosaminuria, Sanfilippo A); and NGS cases which failed to amplify select exons of an associated gene and microarray confirmed a homozygous intragenic deletion (juvenile neuronal ceroid lipofuscinosis; Morquio syndrome type A). Our clinical experience with these cases demonstrates the power of these complementary molecular technologies to aid in the clinical diagnosis and management of autosomal recessive disorders.
32 Unexpected Chromosomal Abnormalities Identified by CMA Confirmation Studies Sau W. Cheung, Amber N. Pursley, Farah A. Ladha, Roger H. Song, Stephanie A. Anderson, M. Lance Cooper, Chad Shaw, Carlos A. Bacino, Ankita Patel
While Chromosomal Microarray Analysis (CMA) has become the first-tier clinical diagnostic test for individuals with developmental disabilities or congenital anomalies, practice guidelines [PMID: 20962661 still recommend cytogenetic/FISH studies when a chromosome imbalance is identified due to the limitation of CMA in detecting chromosome structural abnormalities associated with copy number changes. In the course of performing CMA confirmation studies for samples submitted for routine diagnostic testing, we encountered unexpected structural changes not predicted by the CMA results. Here we present case examples of each catagories of the following atypical structural abnormalities identified with CMA results indicating apparent simple copy number gains/losses: [1] marker chromosomes stabilized by attachment
239 to other chromosomes, [2] insertion translocations in the patient or unaffected parent, [3] fusion of two chromosomes and [4] parental inversion of an acrocentric chromosome. In addition, the combination of confirmation studies required to detect these changes will be reviewed with a special emphasis on the importance of examining metaphase cells by G-banded chromosome analysis and/or FISH to ensure detection of these types of abnormalities. Accurate elucidation of the underlying structural changes has a direct impact on recurrence risk calculation, differential diagnosis and clinical care of the patient and their family.
33 Gamification of Chromosome Identification: Creation of a Software Tool to Introduce Cytogenetic Analysis Cate Randall Paschal *, Kimberly Anderson, Angela M. Lager, Kate Thompson, Jennifer Laffin
Despite the expanding role of genomic sequencing and other molecular techniques in today’s healthcare system, conventional cytogenetic analysis remains an important tool in prenatal, constitutional, and oncology studies. The challenge remains in how to engage new learners in chromosome identification and analysis. Our laboratory has addressed this issue through the creation of an interactive software tool called Karyotyper. This program, designed by an experienced cytotechnologist, utilizes a gamecentric approach to train users in cytogenetic analysis. Through a series of modules, the learner is taught how to recognize and identify individual chromosomes. As the learner progresses through the game modules, the feedback provided becomes less immediate. Separate units have been created for different band levels of resolution, and a variety of normal, aneuploid, and structurally abnormal cells have been included. In addition, modules have been developed to assess competency of experienced technicians. The gamification of this process, along with the range of units created, has allowed us to actively engage learners at every level, from grade school children at outreach events to residents and fellows doing rotations through the cytogenetics laboratory. The immediate and incremental feedback built into the software not only reinforces the learner’s growing confidence and skill, but also allows users to progress at their own pace and without the need for constant observation. From marketing to education, gamebased applications are a constant in today’s society. By applying game design elements to the identification of chromosomes, we are able to sustain high learner involvement with minimal supervision.
34 Exon-Targeted Array CGH for Identification of Clinically Relevant Small-Sized, Intragenic CNVs Ankita Patel, J. Rosenfeld-Mokry, T. Gambin, P. Liu, W. Bi, A. Breman, J. Smith, S. Lalani, C. Bacino, A.L. Beaudet, J.R. Lupski, C.A. Shaw, S.W. Cheung, P. Stankiewicz
Advances in genetic diagnostic technologies has allowed for monumental improvements in the diagnosis of genetic diseases. With newer technologies, offering improved diagnostic results, unexpected findings have also been delineated in some cases. Many groups have addressed this issue by providing * Corresponding author.
31 Microarray and Next Generation Sequencing: Complementary Technologies for Diagnosing Autosomal Recessive Disorders Alka Chaubey, Michael J. Friez, Barbara R. DuPont
Molecular technologies have advanced at a staggering pace and are quickly changing the conventional paradigm of clinical diagnostics. In the past decade, microarray has become the gold standard tool in the management of constitutional genetic disorders. Microarray has the unique ability to be utilized as a high resolution molecular karyotype for identifying large genomic changes (trisomies, large duplications, and large deletions) as well as cryptic microdeletions/microduplications involving discreet genomic locations. However, the diagnostic yield of this technology is limited to ~25–35%. This limitation is addressed by clinicians ordering sequencing of single genes, Next-Generation Sequencing (NGS) panels or Whole Exome Sequencing (WES) to identify the various genetic etiologies. In turn, the unique feature of Single Nucleotide Polymorphism microarrays can be utilized as a complement to the previously mentioned sequencing technologies, especially in patients with autosomal recessive disorders. We present a variety of cases where microarray and sequencing complemented each other for a clinical diagnosis: cases where one mutation was found by sequencing and the other change was a single exon deletion (Sanfilippo B syndrome, Phenylketonuria); cases where NGS identified a single nucleotide mutation and microarray a heterozygous intragenic deletion (Brittle Cornea syndrome, Aspartylglucosaminuria, Sanfilippo A); and NGS cases which failed to amplify select exons of an associated gene and microarray confirmed a homozygous intragenic deletion (juvenile neuronal ceroid lipofuscinosis; Morquio syndrome type A). Our clinical experience with these cases demonstrates the power of these complementary molecular technologies to aid in the clinical diagnosis and management of autosomal recessive disorders.
32 Unexpected Chromosomal Abnormalities Identified by CMA Confirmation Studies Sau W. Cheung, Amber N. Pursley, Farah A. Ladha, Roger H. Song, Stephanie A. Anderson, M. Lance Cooper, Chad Shaw, Carlos A. Bacino, Ankita Patel
While Chromosomal Microarray Analysis (CMA) has become the first-tier clinical diagnostic test for individuals with developmental disabilities or congenital anomalies, practice guidelines [PMID: 20962661 still recommend cytogenetic/FISH studies when a chromosome imbalance is identified due to the limitation of CMA in detecting chromosome structural abnormalities associated with copy number changes. In the course of performing CMA confirmation studies for samples submitted for routine diagnostic testing, we encountered unexpected structural changes not predicted by the CMA results. Here we present case examples of each catagories of the following atypical structural abnormalities identified with CMA results indicating apparent simple copy number gains/losses: [1] marker chromosomes stabilized by attachment
239 to other chromosomes, [2] insertion translocations in the patient or unaffected parent, [3] fusion of two chromosomes and [4] parental inversion of an acrocentric chromosome. In addition, the combination of confirmation studies required to detect these changes will be reviewed with a special emphasis on the importance of examining metaphase cells by G-banded chromosome analysis and/or FISH to ensure detection of these types of abnormalities. Accurate elucidation of the underlying structural changes has a direct impact on recurrence risk calculation, differential diagnosis and clinical care of the patient and their family.
33 Gamification of Chromosome Identification: Creation of a Software Tool to Introduce Cytogenetic Analysis Cate Randall Paschal *, Kimberly Anderson, Angela M. Lager, Kate Thompson, Jennifer Laffin
Despite the expanding role of genomic sequencing and other molecular techniques in today’s healthcare system, conventional cytogenetic analysis remains an important tool in prenatal, constitutional, and oncology studies. The challenge remains in how to engage new learners in chromosome identification and analysis. Our laboratory has addressed this issue through the creation of an interactive software tool called Karyotyper. This program, designed by an experienced cytotechnologist, utilizes a gamecentric approach to train users in cytogenetic analysis. Through a series of modules, the learner is taught how to recognize and identify individual chromosomes. As the learner progresses through the game modules, the feedback provided becomes less immediate. Separate units have been created for different band levels of resolution, and a variety of normal, aneuploid, and structurally abnormal cells have been included. In addition, modules have been developed to assess competency of experienced technicians. The gamification of this process, along with the range of units created, has allowed us to actively engage learners at every level, from grade school children at outreach events to residents and fellows doing rotations through the cytogenetics laboratory. The immediate and incremental feedback built into the software not only reinforces the learner’s growing confidence and skill, but also allows users to progress at their own pace and without the need for constant observation. From marketing to education, gamebased applications are a constant in today’s society. By applying game design elements to the identification of chromosomes, we are able to sustain high learner involvement with minimal supervision.
34 Exon-Targeted Array CGH for Identification of Clinically Relevant Small-Sized, Intragenic CNVs Ankita Patel, J. Rosenfeld-Mokry, T. Gambin, P. Liu, W. Bi, A. Breman, J. Smith, S. Lalani, C. Bacino, A.L. Beaudet, J.R. Lupski, C.A. Shaw, S.W. Cheung, P. Stankiewicz
Advances in genetic diagnostic technologies has allowed for monumental improvements in the diagnosis of genetic diseases. With newer technologies, offering improved diagnostic results, unexpected findings have also been delineated in some cases. Many groups have addressed this issue by providing * Corresponding author.