Genetic Carrier Screening

Transcription

1 Genetic Carrier Screening Test Indications Carrier screening is a significant testing tool for inherited genetic conditions of prenatal care. The purpose of carrier screening is to identify couples at-risk for passing on genetic conditions to their offspring. The Genetic Carrier Screening Jewish Panel provides not only the currently recommended carrier screening tests, but also other important high-risk inherited genetic conditions. Identification of a pathogenic variant in one of these genes, or variants in genes associated with 43 high-risk diseases, can help health care providers and genetic counselors who wish to establish or confirm a diagnosis, predict the risk of having a child with a genetic disorder, or to guide patients management decisions. Considerations for Testing A health care provider or genetic counselor may determine if an individual is at-risk to have offspring with a genetic disorder by obtaining a family health history. The Carrier screening test should be offered if: an individual has a genetic disorder. an individual has a child who has a genetic disease. an individual has a family history of a genetic disorder. an individual belongs to an ethnic group that has a high carrier rate of genetic disorders (e.g., Jewish heritage). Furthermore, The American College of Obstetricians and Gynecologists (ACOG) recommends that carrier screening for cystic fibrosis should be offered to all women who are considering pregnancy or are currently pregnant (2). If a patient considering pregnancy is determined to be a carrier, testing is also recommended for their partner. Genetics The conditions listed in Table 1 can be caused by inherited genetic variants. The variants are inherited in an autosomal recessive pattern in which a gene variant is a recessive gene located on one of the non-sex chromosomes (non-x and non-y), or autosomes. One needs to inherit both copies of the pathogenic gene variant to be affected by the associated disorder. Autosomal recessive disorders are usually passed by two carriers, or individuals whose health is rarely affected but have one variant and one normal copy of the recessive gene. Two carriers have a 25% chance of having an affected child. Clinical Characteristics Abetalipoproteinemia is an autosomal recessive inherited disorder that affects the absorption of dietary fats, cholesterol, and fat-soluble vitamins. Patients with abetalipoproteinemia are unable to produce certain lipoproteins that help carry fats and fat-like substances in the blood. Symptoms including failure to gain weight and thrive, and occur in the first few months of life. Malfunctions of the nervous system and eye disorders may develop later in childhood. Pathogenic variants of the microsomal triglyceride transfer protein (MTTP) gene fail to produce beta-lipoproteins which are necessary for the absorption of fats and fat-like substances from the diet. A lack of beta-lipoproteins causes the associated nutritional and neurological problems in people with abetalipoproteinemia. Abetalipoproteinemia has been reported in approximately 100 cases worldwide. Alport syndrome: Also known as Collagen type IV alpha 3 (COL4A3), gene-related Alport syndrome has either autosomal recessive or dominant inheritance. This condition is featured by kidney malfunction, hearing loss, and eye abnormalities. Pathogenic variants of the COL4A3 gene encoding type IV collagen prevent the kidneys from filtering the blood properly, blocks transformation of sound waves into nerve impulses to the brain, and fails to maintain the shape of the lens and the normal color of the retina. Worldwide, Alport syndrome occurs in approximately 1 in 50,000 newborns, and is estimated to affect approximately 1 in 5,000-10,000 people in the general population in the United States. Arthrogryposis, Mental Retardation and Seizures (AMRS) is an autosomal recessive inherited disorder where newborns have multiple joint contractures in their arms or legs. Affected infants may have limb malformations, hypotonia, and microcephaly. Children with AMRS typically develop intellectual disability, autistic characteristics, as well as seizures. ARMS is caused by genetic changes in the solute carrier family 35 member A3 (SLC35A3) gene, which encodes a transporter found in the Golgi apparatus membrane. AMRS is a rare disorder, which is seen more commonly in those of Jewish descent, where it has a 1 in 453 carrier frequency. Bardet-Biedl Syndrome is an autosomal recessive disorder characterized by severe pigmentary retinopathy, obesity, polydactyly, renal malformation and mental retardation. Pathogenic variants of the Bardet-Biedl syndrome (BBS) genes are responsible for this disorder. Most cases of Bardet-Biedl syndrome result from pathogenic variations in BBS1, BBS2, and BBS10. The BBS genes are involved in cell movement and various chemical signaling pathways. They are also necessary for the perception of sensory input. Bardet-Biedl syndrome has a prevalence of 1 in 140,000 to 1 in 160,000 in newborns in most of North America and Europe. It is more common on the island of Newfoundland, Canada (1 in 17,000 newborns), and the Kuwait Bedouin population, affecting 1 in 13,500 newborns. Bloom Syndrome is an inherited condition following an autosomal recessive pattern. Patients with this condition are characterized by short stature, a skin rash that develops after exposure to the sun, and a greatly increased risk of cancer. Others features may appear in patients, including learning disabilities, a high risk of diabetes, chronic Medical Diagnostic Laboratories, L.L.C Upd: 5/2018

2 Table 1. MDL s Genetic Carrier Screening Panel Detected Diseases, Target Genes and Frequencies. Carrier Frequency- The chance of an individual with no symptoms having a disease-causing variant. Detection Rate- The percentage of carriers calculated to be detected by the Genetic Carrier Screening test. Residual Risk- The chance of being a carrier even if an individual is found negative for the disease-causing variants tested. Disease Abetalipoproteinemia Alport Syndrome, COL4A3-related Arthrogryposis, Mental Retardation, and Seizures (AMRS) Bardet-Biedl Syndrome, BBS1-related Bardet-Biedl Syndrome, BBS2-related Bardet-Biedl Syndrome, BBS10-related Bloom Syndrome Canavan Disease Carnitine Palmitoyltransferase II Deficiency Gene Population MTTP Caucassian Asian Finnish Mennonite Finland European Asian Hispanic French Canadian French Canadian Finnish European French Canadian COL4A3 SLC35A3 BBS1 BBS2 BBS10 BLM ASPA CPT2 Congenital Amegakaryocytic Thrombocytopenia MPL Congenital Disorder of Glycosylation Type 1a PMM2 *Cystic Fibrosis Dyskeratosis Congenita, RTEL1-related Ehlers-Danlos Syndrome Type VIIC CFTR RTEL1 ADAMTS2 Familial Dysautonomia IKBKAP Familial Hyperinsulinism: ABCC8-related ABCC8 Fanconi Anemia Type C FANCC * Fragile X Syndrome FMR1 Galactosemia, GALT-related GALT Gaucher Disease GBA Glycogen Storage Disease 1a G6PC Joubert Syndrome 2 TMEM216 Maple Syrup Urine Disease Type 1a BCKDHA Maple Syrup Urine Disease Type 1b BCKDHB Maple Syrup Urine Disease Type 3 Mucolipidosis Type IV Multiple Sulfatase Deficiency Nemaline Myopathy Niemann-Pick Disease, SMPD1-related DLD MCOLN1 SUMF1 NEB SMPD1 Phosphoglycerate Dehydrogenase Deficiency, PHGDH-related PHGDH Polycystic Kidney Disease, Autosomal Recessive PKHD1 Retinitis Pigmentosa 59 DHDDS Smith-Lemli-Opitz Syndrome DHCR7 * Spinal Muscular Atrophy SMN1 Tay-Sachs Disease HEXA Tyrosinemia Type 1 FAH Usher Syndrome Type III CLRN1 Usher Syndrome Type 1F PCDH15 Walker-Warburg syndrome FKTN-related FKTN Wilson Disease ATP7B Zellweger Spectrum Disorder, PEX1-related Zellweger Spectrum Disorder, PEX2-related Zellweger Spectrum Disorder, PEX6-related PEX1 PEX2 PEX6 2 Available for order as an individual test. * Included in Test 1274 Genetic Carrier Screening Panel. Carrier Frequency 1 in in in in in in in in in 55 1 in in in 45 1 in in in 57 1 in in 42 1 in 61 1 in in 25 1 in 94 1 in 24 1 in in in 31 1 in 52 1 in in 89 1 in in 58 1 in in in in in in 15 1 in in 71 1 in in 10 1 in in 97 1 in in in 89 1 in in in in in in in 70 1 in in in in 48 1 in 41 1 in 68 1 in 41 1 in in 53 1 in in 73 1 in 27 1 in in 66 1 in in in in 70 1 in 78 1 in in 80 1 in 90 1 in 67 1 in 90 1 in in in in 55 Detection Rate 75% 48% 44% 70% 29% 46% 97% 53% 70% 73% 65% 82% 76% 77% 89% 71% 94% 65% 97% 46% 96% 80% 97% 43% 30% 80% 80% 80% 60% 69% 87% 60% 48% 18% 5% 97% 20% 92% 60% 18% 19% 62% 81% 63% 94% 93% 47% 98% 90% 75% 47% 79% 53% 70% 68% 30% Residual Risk 1 in in in in 9,060 1 in 10,000 1 in 1,307 1 in 2,800 1 in in in 4,466 1 in 5,500 1 in in in in in 1,478 1 in 1,140 1 in 1,729 1 in in in in in in in 3,300 1 in in 4,675 1 in 2,500 1 in 3,100 1 in 1,733 1 in in 8,900 1 in in 1,160 1 in 5,000 1 in in in in in in in 7,100 1 in 2,200 1 in 10,000 1 in 67 1 in in 9,700 1 in in in 1,250 1 in in 5,580 1 in in in in 3,833 1 in in 9,060 1 in 6,250 1 in in in in 2,340 1 in in in in in in in 2,340 1 in in 1,350 1 in 2,880 1 in 1,320 1 in 14,300 1 in 2,400 1 in 78 1 in in in 1,600 1 in in in in in 1,563 1 in 22,700 1 in in 21,620

3 obstructive pulmonary disease, and mild immune system abnormalities. Bloom syndrome is caused by pathogenic variants of the bloom syndrome RecQ-like helicase (BLM) gene that produces RecQ helicases which maintain the stability of DNA during duplications. Without this helicase, the cell is less able to repair DNA damage caused by ultraviolet light, resulting in increased sun sensitivity. Cancers develop from this abnormal DNA repair process leading to cell division in an uncontrolled way. Bloom syndrome is a rare disorder. Approximately one-third of people with the disease are from Jewish descent. Roughly 1 in 48,000 Jews are affected by the disease. Canavan Disease is a rare autosomal recessive inherited disorder that damages the ability of neurons in the brain to send and receive messages. This disrupts the growth or maintenance of the myelin sheath, which is the covering that protects nerves and promotes the efficient transmission of nerve impulses. Neonatal and infantile Canavan disease is the most common and most severe form of the condition. Affected infants develop atonia of neck muscles, hypotonia, hyperextension of legs and flexion of arms, blindness, severe mental defects, megalocephaly, and death by 18 months on the average. Pathogenic variants of the aspartoacylase (ASPA) gene cause Canavan disease. The ASPA gene contributes making aspartoacylases. This enzyme normally breaks down a compound called N-acetyl-Laspartic acid (NAA), which is predominantly found in neurons in the brain. The enzyme may be involved in the transport of water molecules out of neurons. Pathogenic variants of the ASPA gene reduce the function of aspartoacylase, which prevents the normal breakdown of NAA. A buildup of NAA destroys existing myelin sheaths. Nerves malfunction without this protective covering, which disrupts normal brain development. While this condition occurs in people of all ethnic backgrounds, it is most common in people of Jewish heritage. Studies suggest that this disorder affects 1 in 6,400 to 13,500 people in the Jewish population. Carnitine Palmitoyltransferase II, (CPT II) Deficiency is an autosomal recessive condition that prevents the body from processing longchain fatty acids into energy, particularly during fasting. Carnitine palmitoyltransferase (CPT2) gene products are responsible for the long-chain fatty acid breakdown process. The symptoms of the lethal neonatal form of CPT II deficiency appear after birth and affect various organs including the liver, lungs, heart, kidney, and brain. Affected infants die within the first year of life. A severe infantile hepatocardiomuscular form of CPT II deficiency has similar characteristic symptoms as the lethal neonatal form, but these symptoms begin from 2 years of age. The myopathic form of CPT II deficiency is the most common type. Symptoms can begin at any time from childhood to one s 60 s. People with this form experience periodic attacks involving their muscles. Worldwide, carnitine palmitoyltransferase II deficiency occurs in approximately 1 in 10,000 to 1 in 100,000 births. Congenital Amegakaryocytic Thrombocytopenia (CAMT) is a rare inherited autosomal recessive disorder characterized by thrombocytopenia and an absence of megakaryocytes. It presents with bleeding recognized on day one of life, or at least within the first month. Thrombopoeitin (TPO) receptor encoded from the MPL gene is essential for the proliferation of megakaryocytes which produce platelets. Pathogenic variants of the MPL gene prevent the production of TPO receptors. The absence of platelets leads to the symptoms of this disorder. Genetic changes in the MPL gene may also result in blood cancers. Congenital amegakaryocytic thrombocytopenia is estimated to occur in 1 in 22,500 pregnancies amongst the Jewish population, and 1 in 57 of this population are carriers. Congenital Disorder of Glycosylation Type Ia is also known as PMM2-congenital disorder of glycosylation (PMM2-CDG), and is inherited in an autosomal recessive pattern. Signs and symptoms of the condition are usually developed during infancy. Affected infants may have hypotonia, abnormal distribution of fat, strabismus, developmental delay, and a failure to thrive. Pathogenic variants of the PMM2 gene are responsible for the condition. Without an enzyme called phosphomannomutase 2 (PMM2), encoded from the PMM2 gene, glycosylation cannot be processed to modify proteins, leading to the symptoms of PMM2CDG. PMM2-CDG accounts for 70% of the congenital disorders of glycosylation, which combined affect 1 in 50,000 to 100,000 births. Cystic Fibrosis (CF) is the most common life-threatening autosomal recessive condition in the non-hispanic, white population. It is a progressive, multisystem disease that primarily affects the pulmonary, pancreatic, and gastrointestinal systems by the buildup of a thick, sticky mucus that can clot the airways and block the intestine and ducts. The current median survival is approximately 37 years, with respiratory failure as the most common cause of death. Cystic fibrosis is caused by pathogenic variants of the CF transmembrane regulator (CFTR) gene, which provides instructions for making a channel that transports negatively-charged chloride ions into and out of cells. Pathogenic variants of the CFTR gene disrupt the function of the chloride channels, preventing the regulation of chloride ions and water across cell membranes. As a result, cells that line the passageways of the lungs, pancreas, and other organs produce mucus that is unusually thick and sticky. Two copies of pathogenic variants of this gene cause CF. Cystic fibrosis is a common genetic disease within the population in the United States. The disease occurs in 1 in 2,500 to 3,500 white newborns. Cystic fibrosis is less common in other ethnic groups, affecting about 1 in 17,000 African Americans and 1 in 31,000 Asian Americans. ACOG Recommendation: CF screening is important to be offered to women of reproductive age. It is becoming increasingly difficult to assign a single ethnicity to individuals. When one member of a couple is a carrier of CF, the other partner should be offered screening. Dyskeratosis Congenita, RTEL1-related: Dyskeratosis congenita has three major characteristics, including nail dystrophy, pigmentation, and oral leukoplakia. RTEL1-related Dyskeratosis congenita can be inherited in an autosomal dominant or autosomal recessive pattern. RTEL1 encodes a DNA helicase implicated in telomere-length regulation, DNA repair, and maintenance of genomic stability. This helicase acts as an anti-recombinase to counteract toxic recombination and limit crossover during meiosis. Pathogenic variants of RTEL1 result in a loss of function of DNA repair, leading to the signs and symptoms of Dyskeratosis congenita. Dyskeratosis congenita has an estimated approximately 1 in 1,000,000 worldwide. incidence of Ehlers-Danlos Syndrome: More than 10 recognized types of Ehlers-Danlos syndrome have been reported. Among these, the 3 Medical Diagnostic Laboratories, L.L.C

4 dermatosparaxis Ehlers-Danlos syndrome type VIIC is of autosomal recessive inheritance. The dermatosparaxis type of the condition is characterized by skin that sags and wrinkles. Redundant folds of skin may be present as affected children get older. A gene called a disintegrin-like and metalloproteinase with thrombospondin type 1 motif 2 (ADAMTS2) is associated with dermatosparaxis Ehlers-Danlos syndrome type VIIC. The ADAMTS2 gene provides instructions for making an enzyme that processes several types of procollagen molecules. Procollagens are the precursors of collagens, which are complex molecules found in the spaces between cells that add strength, support, and elasticity to various body tissues. Pathogenic variants of the ADAMTS2 gene greatly reduce the production or activity of the ADAMTS2 enzyme. As a result, collagen fibrils are not assembled properly. The resulting fibrils are disorganized, which weakens connective tissues and leads to the signs and symptoms of the disorder. The dermatosparaxis type is relatively rare among all types of EhlersDanlos syndrome. About a dozen infants and children with this condition have been reported. Familial Dysautonomia is an autosomal recessive disorder that affects the development and survival of certain nerve cells. The disorder not only disturbs cells in the autonomic nervous system, which controls involuntary actions such as digestion, breathing, and the regulation of blood pressure and body temperature, but also affects the sensory nervous system, which controls activities related to the senses, such as taste and the perception of pain, heat, and cold. Familial dysautonomia is caused by pathogenic variants of the inhibitor of the kappa light polypeptide gene enhancer in B-cells, kinase complex-associated protein (IKBKAP) gene, which is responsible for production of IκB kinase complex-associated protein (IKAP). This protein is found in a variety of cells throughout the body, including brain cells. IKAP plays a key role in transcription of proteins that affect the cell s cytoskeleton and cell motility. Critical activities in brain cells are probably disrupted by reduced amounts or the absence of IKAP protein, leading to the signs and symptoms of familial dysautonomia. Familial dysautonomia occurs primarily in people of Jewish descent. It affects about 1 in 3,700 individuals in Jewish populations. Familial dysautonomia is rare in the general population. Familial Hyperinsulinism can be an autosomal recessive or dominant inherited condition in which the pancreas releases inappropriately large quantities of hormone insulin, leading to hypoglycemia. When blood sugar drops to dangerously low levels, seizures and permanent brain damage may occur. Pathogenic variants of the ATP binding cassette subfamily C member 8 (ABCC8) gene are the most common cause of the disorder and account for approximately 40% of affected individuals. Pathogenic variants of the ABCC8 gene lead to over-secretion of insulin from pancreatic beta cells, resulting in glucose being rapidly removed from the bloodstream. A lack of glucose in the blood causes frequent states of hypoglycemia in people with familial hyperinsulinism. Familial hyperinsulinism affects roughly 1 in 50,000 Europeans. It is particularly common among people of Finnish and Saudi Arabian descent, where the disease may affect as many as 1 in 2,500. Approximately 97% of cases of familial hyperinsulinism in the Jewish population can be attributed to two ABCC8 founder variants (p.phe1387del and c g>a). Two additional founder variants in the ABCC8 gene (p.val187asp and p.glu1506lys) have been identified in the Finnish population. Fanconi Anemia Type C: Iherited in an autosomal recessive pattern, Fanconi Anermia Type C is a condition in which the body cannot properly produce a protein that protects DNA from damage. People with this condition may develop bone marrow failure, physical abnormalities, organ defects, and an increased risk of certain cancers. Approximately 90% of people with Fanconi anemia type C have impaired bone marrow function that leads to a decrease in the production of all blood cells. Affected individuals experience fatigue, frequent infections, and clotting problems. Another symptom of patients with Fanconi anemia type C is physical abnormalities such as spotted skin or malformations of organs. People with this condition are at higher than average risk of cancer development due to the loss of function of proteins that repair damaged DNA. The Fanconi anemia complementation group C (FANCC) gene is one of the critical genes responsible for the Fanconi anemia pathway, which triggers DNA repair processes when DNA damage is detected. Pathogenic variants of the FANCC gene result in either abnormal cell death or uncontrolled cell growth due to an inability to make new DNA molecules and a lack of the necessary DNA repair processes. Fanconi anemia type C affects approximately 1 in 100,000 people. This condition is more common among people of the Roma population of Spain, black South Africans, and, especially, those of Jewish descent, where 1 in 89 are carriers and 1 in 32,000 have the actual disease. Fragile X syndrome is the most common inherited form of mental impairment. The syndrome is inherited in an X-linked dominant pattern and occurs in approximately 1 in 3,600 males and 1 in 4,000 6,000 females of various ethnicities. Fragile X syndromerelated impairment ranges from borderline, including learning disabilities, to severe, presenting with cognitive and behavioral disabilities, including autism. Signs of Fragile X syndrome are subtle in newborns, making Fragile X syndrome difficult to diagnose based on clinical findings alone. Fragile X syndrome is caused by expansion of a repeated trinucleotide segment of DNA, cytosine guanine guanine (CGG), that leads to altered transcription of the Fragile X mental retardation 1 (FMR1) gene. The FMR1 gene provides instructions for making a protein called FMRP, that helps regulate the production of other proteins and plays a role in the development of synapses, which are specialized connections between nerve cells. A person with less than 45 repeats is regarded as unaffected. When more than 200 repeats are present, an individual is said to have a full mutation resulting in the full manifestation of Fragile X syndrome. This condition causes the FMR1 gene to become methylated. ACOG Recommendation: Genetic counseling and fragile X syndrome carrier screening is recommended testing to the following: Women with a family history of Fragile X-related disorders, unexplained mental retardation or developmental delay, autism, or premature ovarian insufficiency are candidates for genetic counseling and Fragile X carrier screening. If a woman has ovarian insufficiency or failure or an elevated follicle-stimulating hormone level before age 40 without a known cause, Fragile X carrier screening should be considered. Women who request Fragile X carrier screening, regardless of family history, should be offered FMR1 DNA genetic analysis after genetic counseling about the risks, benefits, and limitations of screening. All identified carriers of a Fragile X pre-mutation (or Medical Diagnostic Laboratories, L.L.C

5 full mutation) should be referred for follow-up genetic counseling to discuss the risk to their fetuses of inheriting an expanded full mutation Fragile X allele and to discuss Fragile X associated disorders (premature ovarian insufficiency and Fragile X tremor ataxia syndrome). Prenatal and preimplantation diagnoses and donor eggs should be discussed. Galactosemia is a disorder that prevents the effective breakdown of galactose and energy production. Affected infants fail to gain weight and thrive. They may also develop jaundice, lethargy, liver damage, and abnormal bleeding. Affected children may also have delayed development, intellectual disability, and vision abnormality. Genetic changes of the galactose-1-phosphate uridylyltransferase (GALT) gene affect the activity of an enzyme encoded from it, which leads to the signs and symptoms of this disorder. Galactosemia affects 1 in 30,000 to 60,000 newborns. Gaucher Disease is an autosomal recessive disorder. The signs and symptoms of this condition vary widely among affected individuals including anemia, thrombocytopenia, bone abnormalities, brain damage, skin abnormalities, heart problems, and hepatosplenomegaly. Gaucher disease is caused by pathogenic variants of the glucosylceramidase beta (GBA) gene, which produces an enzyme called beta-glucocerebrosidase. This enzyme breaks down the fatty substance glucocerebroside into glucose. Pathogenic variants of the GBA gene greatly decrease or eliminate the activity of betaglucocerebrosidase. As a result, glucocerebroside can accumulate to toxic levels within cells, causing the characteristic features of Gaucher disease. Gaucher disease occurs in 1 in 50,000 to 100,000 people in the general population. The Type 1 non-neuronopathic form is the most common form of the disorder and occurs more frequently in those of Jewish heritage from 1 in 500 to 1,000 people, with 1 in 15 of this population as carriers. Glycogen storage disease type I is an autosomal recessive inherited disorder characterized by the accumulation of glycogen that, in turn, impairs the normal function of certain organs and tissues, especially the liver and kidneys. Symptoms typically occur around the age of 3 months, when babies start to sleep through the night and do not eat as frequently as newborns. Infants with the condition may have hypoglycemia which can lead to seizures. Patients can also develop hyperuricemia and hyperlipidemia. Pathogenic variants of the glucose-6-phosphatase catalytic (G6PC) gene prevent the effective breakdown of glucose-6-phosphate, leading to glycogen accumulation within cells. A buildup of excessive glycogen causes the symptoms of glycogen storage disease Type I. The overall incidence of glycogen storage disease Type I is 1 in 100,000 people. Approximately 1 in every 20,000 to 25,000 babies in the U.S. and Europe are born with glycogen storage disease. Joubert Syndrome 2 is an autosomal recessive disorder characterized by a specific hindbrain malformation, hypotonia, developmental delay, oculomotor apraxia, and breathing abnormalities that occur from infancy to early childhood. Additional characteristic features include retinal anomalies, polydactyly, hepatic fibrosis, and renal disease. This disorder is associated with pathogenic variants of the transmembrane protein 216 (TMEM216) gene. Joubert Syndromes are estimated to occur in between 1 in 80,000 and 1 in 100,000 births. However, Joubert Syndromes have a large range of possible manifestations and are likely underdiagnosed. Maple Syrup Urine Disease is an autosomal recessive disorder characterized by a distinctive sweet odor of urine in affected infants. Additional symptoms include poor feeding, vomiting, lethargy, and developmental retardation. Malfunction of a protein complex called branched-chain alphaketo acid dehydrogenase, or BCKD, causes maple syrup urine disease. This complex functions to break down amino acids and produce molecules that can be used for energy. BCKD is produced from branched-chain keto acid dehydrogenase E1, alpha and beta polypeptides (BCKDHA, and BCKDHB) genes. Pathogenic variants in these genes cause accumulation of amino acids (leucine, isoleucine, and valine) and their byproducts, leading to serious health problems, known as maple syrup urine disease type Ia and Ib. Pathogenic variants of the third component dihydrolipoamide dehydrogenase (DLD) cause maple syrup urine disease type III, referred to as dihydrolipoamide dehydrogenase deficiency. Maple syrup urine disease type Ia and Ib affects 1 in 185,000 infants in the general population. It is most common among the Old Order Mennonite population, where about 1 in 385 infants are affected by the disease. Type Ia and Ib are also common among Jews, with roughly 1 in 50,000 people, and 1 in 35,000 to 48,000 individuals in type III. Mucolipidosis Type IV is a rare autosomal recessive inherited condition that affects the development of the nerves. It also causes existing nerves to degenerate. Patients with mucolipidosis type IV show delayed development in mental and motor skills. Affected infants and children are unable to sit up, crawl, walk or speak properly. Mucolipidosis type IV also leads to poor vision caused by cloudy corneas and degeneration of the retina. Mucolipin 1 protein, produced by the mucolipin 1 (MCOLN1) gene, is located in the membranes of lysosomes and endosomes and plays a role in trafficking lipids and proteins within the cell. Pathogenic variants in the MCOLN1 gene cause the accumulation of these substances in lysosomes, leading to the symptoms of mucolipidosis type IV. Mucolipidosis type IV is estimated to occur in 1 in 40,000 people. About 70% of affected individuals have Jewish ancestry. It is reported that roughly 1 in 89 Jews are carriers. Multiple Sulfatase Deficiency is a condition inherited in an autosomal recessive manner with signs and symptoms occurring from birth to young adulthood. Affected individuals have deterioration of tissue in the nervous system, causing developmental delays, movement problems, and slow growth. Pathogenic variants in the sulfatase modifying factor 1 (SUMF1) gene contribute to the disorder. The SUMF1 gene produces formylglycinegenerating enzyme (FGE) that functions in protein modification. Malfunction of FGE leads to cell death, especially in the brain, skeleton and skin, causing the symptoms of multiple sulfatase deficiency. Multiple sulfatase deficiency affects approximately 1 per million individuals worldwide. Nemaline Myopathy is usually inherited in an autosomal recessive pattern but sometimes in an autosomal dominant pattern. This disorder results from a lack of nebulin protein encoded by the NEB gene. Nebulin plays a significant role in generating the mechanical force needed for muscle contraction. People with nemaline myopathy have muscle weakness. They have difficulties in feeding and swallowing during infancy. As the condition progresses, the breathing muscles are affected and may lead to a lethal condition. Nemaline myopathy has an estimated incidence of 1 in 50,000 in the general population. 5 Medical Diagnostic Laboratories, L.L.C

6 Niemann-Pick Disease is an autosomal recessive inherited condition which is divided into four types. Among them, type A and type B are the most common. Infants and children with Niemann-Pick disease type A usually develop hepatosplenomegaly, respiratory failure, mental and physical disabilities, and eye abnormalities. NiemannPick disease type B, whose symptoms are similar to, but not as severe as type A, usually present in mid-childhood. Smith-Lemli-Opitz Syndrome, or SLO syndrome, is an autosomal recessive inherited developmental disorder in which the body loses the ability to make sufficient cholesterol due to pathogenic variations in the 7-dehydrocholesterol reductase (DHCR7) gene. The features of this disorder vary widely, including microcephaly, intellectual disability, malformation of various organs, muscle hypotonia, and syndactyly and polydactyly. Niemann-Pick disease types A and B are caused by pathogenic variants of the sphingomyelin phosphodiesterase 1 (SMPD1) gene. This gene produces an enzyme called acid sphingomyelinase which localizes to lysosomes. This enzyme is responsible for breaking down lipid molecules. Pathogenic variants in SMPD1 in people with Niemann-Pick disease types A and type B cause lipid accumulation in cells leading to cellular malfunction and death. This leads to a loss of function of various tissues and organs including the brain, lungs, spleen and liver. Smith-Lemli-Opitz syndrome affects roughly 1 in 20,000 to 60,000 newborns. This condition is most common in s of European ancestry, particularly people from Central European countries such as Slovakia and the Czech Republic. Smith-Lemli-Opitz syndrome is very rare among African and Asian populations. Niemann-Pick disease type A and type B affect 1 in 250,000 in the general population. The incidence within the population is roughly 1 in 40,000 individuals. Phosphoglycerate Dehydrogenase Deficiency is a condition characterized by microcephaly, impaired movements and recurrent seizures. It follows an autosomal recessive pattern of genetic inheritance. Pathogenic variants of the phosphoglycerate dehydrogenase (PHGDH) gene are responsible for the disorder. The PHGDH gene encodes an enzyme that is involved in the production of serine, which is essential for the development and function of the brain and spinal cord. Pathogenic variants of the PHGDH gene affect the enzyme by decreasing its activity. As a consequence, the formation of myelin and the production of neurotransmitters in the brain can be impaired. Phosphoglycerate dehydrogenase deficiency is a rare disorder. Only 15 cases have been reported in the scientific literature. Polycystic Kidney Disease, Autosomal Recessive, PKHD1-Related: Patients with polycystic kidney disease always develop clusters of fluid-filled sacs (cysts) in the kidneys and their ability to filter waste products from the blood is impaired. Frequent complications of polycystic kidney disease include severe hypertension, hematuria, recurrent urinary tract infections, kidney stones, and heart valve abnormalities. The signs and symptoms of this condition are usually apparent at birth or in early infancy. Pathogenic variants of the PKHD1 gene are related to polycystic kidney disease. This gene encodes fibrocystin that is involved in various cellular signaling pathways, cellular adhesion, and cellular proliferation. PKHD1-related autosomal recessive polycystic kidney disease occurs in an estimated 1 in 20,000 to 40,000 people worldwide. Retinitis pigmentosa: Pathogenic variants of the dehydrodolichyl diphosphate synthase (DHDDS) gene are responsible for retinitis pigmentosa disorder, which is an autosomal recessive inheritance. In people with this condition, vision loss occurs as the light-sensing cells of the retina gradually deteriorate. A loss of night vision usually appears as the first symptom in childhood. Later, this disease causes blind spots in the peripheral vision. Over time, the disease progresses and eventually affects central vision. In adulthood, many people with this condition become blind. Retinitis pigmentosa is one of the most common inherited diseases of the retina. It occurs in 1 in 3,500 to 4,000 people in the United States and Europe. Spinal Muscular Atrophy (SMA) is an autosomal recessive neurodegenerative disease that results from the degeneration of spinal cord motor neurons leading to atrophy of skeletal muscle and overall weakness. Affected children have a deficiency in movement, possibly requiring wheelchair assistance. A frequent cause of death of patients with SMA is respiratory failure. There is no effective treatment for the disease. The disorder is caused by a variation in the survival motor neuron gene (SMN1), which is responsible for the production of a protein essential to motor neurons. More than 98% of patients with SMA have an abnormality in both copies of the SMN1 gene, causing a deletion or other pathogenic variation. The occurrence of SMA is approximately 1 in 10,000 live births and it is reported to be the leading genetic cause of infant death. ACOG Recommendations: Genetic counseling and SMA carrier screening should be offered to the following patients or couples: Those with a family history SMA or SMA-like disease Those who request SMA carrier screening and have completed genetic counseling that included discussion of the sensitivity, specificity, and limitations of screening. Tay-Sachs Disease (TSD) is inherited in an autosomal recessive pattern, and is a lysosomal storage disease in which GM2 gangliosides accumulate throughout the body. The accumulation of these gangliosides in the central nervous system results in a severe progressive destruction of nerve cells in the brain and spinal cord that causes death in early childhood. Affected infants lose motor skills such as turning over and sitting. Along with the progression, patients experience seizures, vision and hearing loss, and intellectual disability. Pathogenic variants of the HEXA gene disrupt the activity of betahexosaminidase A, which prevents the enzyme from breaking down GM2 ganglioside. As a result, this substance accumulates to toxic levels, particularly in neurons in the brain and spinal cord. The TSD carrier rate in Jewish individuals of Eastern European descent () is 1 in 29; the carrier rate for non-jewish individuals is 1 in 288. It has been determined that individuals of French Canadian and Cajun descent also have a greater carrier frequency than the general population. ACOG Recommendations: Screening for TSD should be offered before pregnancy if both members of a couple are of Jewish, French Canadian, or Cajun descent. Those with a family history consistent with TSD also should be offered screening. When one member of a couple is at high-risk (i.e., of Jewish, French Canadian, or Cajun descent or has a family history consistent with TSD), but the other partner is not, the high-risk partner should be offered screening. This is particularly important if there is uncertainty about ancestry or if there is a family history 6 Medical Diagnostic Laboratories, L.L.C

7 consistent with TSD. If the high-risk partner is determined to be a carrier, the other partner should also be offered screening. If the woman is already pregnant, it may be necessary to offer screening to both partners simultaneously to ensure that results are obtained promptly and that all options are available to the couple. If TSD biochemical screening is performed in women who are pregnant or taking oral contraceptives, leukocyte testing must be used. If both partners are determined to be carriers of TSD, genetic counseling and prenatal diagnosis should be offered. Tyrosinemia type 1 is an autosomal recessive inherited metabolic disorder characterized by disruptions in the process that breaks down tyrosine. Accumulation of tyrosine and its byproducts in tissues and organs can cause severe symptoms. Affected infants fail to gain weight and thrive. They may also develop jaundice. This condition is caused by pathogenic variations of the fumarylacetoacetate hydrolase (FAH) gene, which produces an enzyme involved in the last step of the catabolic pathway of tyrosine. Tyrosinemia type I affects about 1 in 100,000 individuals in the general population. It is more common in Norway where 1 in 60,000 to 74,000 individuals are affected. This disease is even more common in Quebec, Canada where it occurs in about 1 in 16,000 people. Usher Syndrome is a condition of autosomal recessive inheritance characterized by partial or total hearing and vision loss that worsens over time. Individuals with Usher syndrome type 1F are born with severe hearing loss. Progressive vision loss becomes apparent in childhood. Children with this condition have difficulty in balance resulting from vestibular abnormalities. Different from type 1F, people with Usher syndrome type 3 have normal hearing at birth, but experience hearing and vision loss later in life. problems. The most characteristic symptom of patients with Wilson disease is the Kayser-Fleischer ring, which surrounds the colored part of the eye resulting from a copper deposit in the front surface of the cornea. Abnormalities in eye movements may also occur. Wilson disease is caused by pathogenic variants of the ATPase copper-transporting beta polypeptide (ATP7B) gene that encodes a polypeptide that acts as a plasma membrane copper-transport protein. Malfunction of this protein leads to copper accumulation in the body to a toxic level causing damage to tissues and organs. Approximately 1 in 30,000 people have Wilson disease worldwide. It is most common in China, Japan, and Sardinia, where it may affect as many as 1 in 10,000 people. Zellweger Spectrum Disorder includes Zellweger syndrome, neonatal adrenoleukodystrophy (NALD) and infantile Refsum disease. Zellweger syndrome is inherited in an autosomal recessive pattern. Individuals with Zellweger syndrome, the most severe form of the spectrum, develop signs and symptoms during the newborn period. These infants experience hypotonia, feeding problems, hearing and vision loss, and seizures. These problems are caused by the breakdown of myelin, which is the covering that protects nerves and promotes the efficient transmission of nerve impulses. People with NALD or infantile Refsum disease, the less severe form of the spectrum, usually do not develop signs and symptoms until late infancy. They may have similar characteristics as Zellweger syndrome, but their condition progresses more slowly. Zellweger spectrum disorder is caused by pathogenic variations in a group of genes which provide instructions for production of peroxins. These proteins are essential for the generation of lipids used in the nervous system. Pathogenic variants of the PEX1 gene are the most common cause of Zellweger spectrum disorder and are found in nearly 70% of affected individuals. The protocadherin 15 (PCKH15) gene was found to contribute to Usher syndrome type 1F, while the clarin 1 (CLRN1) gene is associated with Usher syndrome type 3. Both of these genes are involved in normal hearing, balance, and vision. Zellweger spectrum disorder is estimated to occur in 1 in 50,000 individuals worldwide. Usher syndrome is estimated to affect 1 in 20,000 people. Usher syndrome type 3 occurs more frequently in the Finnish population, where it accounts for about 40% of cases. Usher syndrome type 1F occurs more frequently in the Jewish population, where approximately 1 in 50 people are carriers. This disorder is also the leading cause of deafblindness among the Jewish population. Genomic DNA is extracted from a blood, mouthwash, OneSwab or ThinPrep sample. High-throughput Next Generation Sequencing is performed to examine over 1,300 DNA variants associated with 41 diseases. Some pathogenic variants are more severe than others. These variant regions are sequenced to high coverage and the sequences are compared to standards and references of normal variation. All the reported variants are confirmed by the gold standard Sanger sequencing. In addition, some of the variants in the panel may be partially subjected to Sanger sequencing to ensure adequate sequencing. Walker-Warburg Syndrome is an autosomal recessive inherited condition characterized by symptoms of muscle weakness, vision impairment, brain structure abnormalities, and severe developmental delay. Pathogenic variants of the fukutin (FKTN) gene account for this disorder. FKTN encodes the fukutin protein which helps to anchor the structural framework of the cytoskeleton and extracellular matrix. It also helps stabilize muscle fibers in skeletal muscles and direct migration of neurons in the brain. Walker-Warburg syndrome is estimated to affect 1 in 60,500 newborns worldwide. Approximately 1 in 150 individuals of Jewish descent are carriers. Wilson Disease is of autosomal recessive inheritance and causes accumulation of copper within the body. Copper that is retained in the liver, brain, kidneys and eyes causes tissue damage, organ failure and death. This initial feature of Wilson disease is usually a liver problem among affected children and young adults, while individuals diagnosed in adulthood often develop nervous system or psychiatric Test Methodology FXS methodology: The MDL Fragile X Syndrome testing was designed to provide accurate sizing of alleles up to 200 CGG, identification of full mutation alleles >200 CGG and a characteristic product peak profile that resolves zygosity in female samples. The MDL Fragile X Syndrome testing uses a regular gene specific PCR and a Triplet Repeat-Primed PCR (TRP PCR) from purified genomic DNA and fragment sizing on an Applied Biosystems Genetic Analyzer. Triplet repeat primed PCR (TRP PCR) allows rapid detection of PCR products formed by a chimeric primer binding inside a triplet-repeat region. In TRP PCR for fragile X, one primer is anchored completely outside of the CGG repeat region, whereas the other overlaps the CGG repeat and the adjacent non-repeated sequence. This TRP PCR will increase the amount of full-length product from the largest CGG-repeat allele and in MDL fragile X assay enables accurate sizing of alleles up to 200 CGG repeats. 7 Medical Diagnostic Laboratories, L.L.C

8 Risk assessment and clinical interpretation of FXS and related disorders are defined by the number of CGG repeats and methylation status of the gene. Based on the number of CGG repeats it is possible to distinguish four types of alleles: unaffected or normal alleles (<=44CGG), intermediate or gray zone(45-54 CGG), premutation ( CGG) and full mutation (>200 CGG). References SMA methodology: The MDL Spinal Muscular Atrophy test uses Multiplex Ligation-dependent Probe Amplification (MLPA) technique. The principle of MLPA is based on the amplification of up to 60 probes, each of which detecting a specific DNA sequence of approximately 60 nucleotides in length. After denaturation of the sample DNA, a mixture of MLPA probes is added to the sample. Each MLPA probe consists of two oligonucleotides that must hybridize to immediately adjacent target sequences in order to be ligated into a single probe. Each probe in an MLPA probe mix has a unique amplicon length, typically ranging between nucleotides. During the subsequent PCR reaction, all ligated probes are amplified simultaneously using the same PCR primer pair. One PCR primer is fluorescently labelled, enabling the amplification products to be visualized during fragment separation. The relative height of each individual probe peak, as compared to the relative probe peak height in various reference DNA samples, reflects the relative copy number of the corresponding target sequence in the sample. Five probes used in the MDL Spinal Muscular Atrophy test detect SMN1 exon 7, SMN1 exon 8, SMN2 Exon 7 and two probes that detect the rare allele of two polymorphisms that may be present in the SMN1 gene. More than of SMA patients show homozygous deletion of at least exon 7 of the SMN1 gene. The great majority of SMA carriers can be identified by the presence of only a single SMN1 exon 7 copy. 3. Variant Classification System: The MDL variant classification system is based on the 5-tier system recommendations for the interpretation of sequence variants proposed by the American College of Medical Genetics and Genomics (ACMG) and complies with the standards and guidelines for the interpretation of sequence variants by ACMG and the Association for Molecular Pathology (AMP). To classify each variant, MDL assigns weight to each piece of available evidence, including literature review, reputable database reports, population frequencies, and computational evidence and prediction. 20. Any detected variants that are a recognized cause of the disease (Pathogenic) will be reported. In addition, variants that have not previously been established as a recognized cause of disease may be identified. In these cases, only variants classified as likely pathogenic are reported. Benign variants, likely benign variants and variants of uncertain significance, and variants not directly associated with the intended disease phenotype are not reported. MDL variant results are reported using numbering and nomenclature recommended by the Human Genome Variation Society (HGVS All results are reported in reference to Human Genome 19, Human Build 37. Turnaround Time 14 to 21 days Specimen Requirements 1. Whole Blood (Yellow top tube-acd A) Mouthwash OneSwab ThinPrep Azimi M, Schmaus K, Greger V, et al Carrier screening by next-generation sequencing: health benefits and cost effectiveness. Mol Genet Genomic Med (4)3: Edwards JG, Feldman G, Goldberg J, et al Expanded Carrier Screening in Reproductive Medicine - Points to Consider: A Joint Statement of the American College of Medical Genetics and Genomics, American College of Obstetricians and Gynecologists, National Society of Genetic Counselors, Perinatal Quality Foundation, and Society for Maternal-Fetal Medicine. Obstet Gynecol 125(3): German J, Sanz MM, Ciocci S, et al Syndrome-causing mutations of the BLM gene in persons in the Bloom s Syndrome Registry. Hum Mutat 28(8):743. James C, Kapoor RR, Ismail D, et al The genetic basis of congenital hyperinsulinism. J Med Genet 46(5): Malfait F, De Paepe A. The Ehlers-Danlos syndrome. Progress in Heritable Soft Connective Tissue Diseases. Springer Netherlands. 2014: Surendran S, Michals-Matalon K, Quast MJ, et al Canavan disease: a monogenic trait with complex genomic interaction. Mol Genet Metab 80(1): Aggarwal A, Bhatt M Update on Wilson disease. Int Rev Neurobiol 110: Beales PL Lifting the lid on Pandora s box: the Bardet-Biedl syndrome. Curr Opin Genet Dev 15(3): Simon E, Flaschker N, Schadewaldt P, et al Variant maple syrup urine disease (MSUD) the entire spectrum. J Inherit Metab Dis 29(6): Accurso FJ Update in cystic fibrosis Am J Respir Crit Care Med 173(9): Saihan Z, Webster AR, Luxon L, et al Update on Usher syndrome. Curr Opin Neurol 22(1): Kruegel J, Rubel D, Gross O Alport syndrome - insights from basic and clinical research. Nat Rev Nephrol 9(3): Deschauer M, Wieser T, Zierz S Muscle carnitine palmitoyltransferase II deficiency: clinical and molecular genetic features and diagnostic aspects. Arch Neurol 62(1): Jira PE, Waterham HR, Wanders RJA, et al Smith-Lemli-Opitz Syndrome and the DHCR7 Gene. Ann Hum Genet 67(3): Daiger SP, Bowne SJ, Sullivan LS Perspective on genes and mutations causing retinitis pigmentosa. Arch Ophthalmol 125(2): Bergeron A, D Astous M, Timm DE, et al Structural and functional analysis of missense mutations in fumarylacetoacetate hydrolase, the gene deficient in hereditary tyrosinemia type 1. J Biol Chem 276(18): Deakyne JS, Mazin AV Fanconi anemia: at the crossroads of DNA repair. Biochem 76(1): Koukoui SD, Chaudhuri A Neuroanatomical, molecular genetic, and behavioral correlates of fragile X syndrome. Brain Res Rev 53(1): Chou JY, Mansfield BC Mutations in the glucose-6-phosphatase-α (G6PC) gene that cause type Ia glycogen storage disease. Hum Mutat 29(7): Timson DJ The molecular basis of galactosemia - Past, present and future. Gene, 589(2): Beutler E Gaucher disease: multiple lessons from a single gene disorder. Acta Paediatrica 95(S451): Fernandes Filho JA, Shapiro BE Tay-sachs disease. Arch Neurol 61(9): Axelrod FB Familial dysautonomia. Muscle Nerve 29(3): Altarescu G, Sun M, Moore DF, et al The neurogenetics of mucolipidosis type IV. Neurol 59(3): Ballmaier M, Germeshausen M, Schulze H, et al C-mpl mutations are the cause of congenital amegakaryocytic thrombocytopenia. Blood 97(1): Pons V, Rolland C, Nauze M, et al A severe form of abetalipoproteinemia caused by new splicing mutations of microsomal triglyceride transfer protein (MTTP). Hum Mutat 32(7): Yin X, Pu CQ, Wang Q, et al Clinical and pathological features of patients with nemaline myopathy. Molec Med Rep 10(1): Braverman NE, D Agostino MD, MacLean GE Peroxisome biogenesis disorders: Biological, clinical and pathophysiological perspectives. Dev Disabil Res Rev 17(3): Crane DI, Maxwell MA, Paton BC PEX1 mutations in the Zellweger spectrum of the peroxisome biogenesis disorders. Hum Mutat 26(3): Krause C, Rosewich H, Thanos M, et al Identification of novel mutations in PEX2, PEX6, PEX10, PEX12, and PEX13 in Zellweger spectrum patients. Hum Mutat 27(11): Tabatabaie L, De Koning TJ, Geboers AJJM, et al Novel mutations in 3-phosphoglycerate dehydrogenase (PHGDH) are distributed throughout the protein and result in altered enzyme kinetics. Hum Mutat 30(5): Adeva M, El-Youssef M, Rossetti S, et al Clinical and molecular characterization defines a broadened spectrum of autosomal recessive polycystic kidney disease (ARPKD). Med 85(1):1-21. Grünewald, S The clinical spectrum of phosphomannomutase 2 deficiency (CDG-Ia). Biochim Biophys Acta - Mol Basis Dis 1792(9): Baran I, Nalcaci R, Kocak M Dyskeratosis congenita: clinical report and review of the literature. Int J Dent Hyg 8(1): Edvardson S, Ashikov A, Jalas C, et al Mutations in SLC35A3 cause autism spectrum disorder, epilepsy and arthrogryposis. J Med Genet 50(11): Shi L, Webb BD, Birch AH, et al Comprehensive population screening in the Jewish population for recurrent disease-causing variants. Clin Genet 91(4): Lorson CL, Hahnen E, Androphy EJ, et al A single nucleotide in the SMN gene regulates splicing and is responsible for spinal muscular atrophy. Proc Natl Acad Sci 96(11): Dalal PG, Coleman M, Horst M, et al Case Report: Genetic analysis and anesthetic management of a child with Niemann-Pick disease Type A. F1000Research 4. Cosma MP, Pepe S, Parenti G, et al Molecular and functional analysis of SUMF1 mutations in multiple sulfatase deficiency. Hum Mutat 23(6): Edvardson S, Shaag A, Zenvirt S, et al Joubert syndrome 2 (JBTS2) in Jews is associated with a TMEM216 mutation. Am J Hum Genet 86(1): Medical Diagnostic Laboratories, L.L.C