Pathobiology: Genetic Diseases (Poulik)
GENERAL PRINCIPLES OF GENETIC DISEASES AND SINGLE GENE DISORDERS:
· Genetic Mutations:
- Definition: stable, heritable alteration in DNA
- Different Types of Mutations:
o Chromosomal Level: result in cytogenic or karyotypic abnromalities
§ Genome mutation: loss or gain of a whole chromosome (ie. trisomy)
§ Chromosome mutation: structural changes in chromosome that gives rise to visible structure changes in the chromosome (ie. translocations, deletions)
§ Trinucleotide repeats: although gene mutations, may be visible by karyotype when cells cultured in different media (ie. Fragile X syndrome visible in foalte deficient media)
o Gene Level: most are NOT visible by karyotype (some trinucleotide repeats are the exception) and therefore require molecular genetics techniques to define mutation (ie. DNA sequencing)
§ Point Mutations: single base substitution
Ø In exon: protein coding sequence of enzyme, structural or regulatory protein
Ø In intron: in non-protein coding sequences (promoter, enhancer etc.)
o May lead to reduction or loss of transcription
§ Frameshift Mutations: caused by deletions or insertions in coding sequence and alterations in reading frame of DNA
§ Trinucleotide Repeats
- Pathological Significance of Genetic Alterations: may do a number of things
o Directly cause disease (ie. Tay Sachs)
o Predispose to disease (ie. X-linked agammaglobulinemia)
o Alter response to another disease (sickle cell anemia protection against malaria)
- Manifestations of Genetic Disease:
o Abnormal metabolism or physiology
o Abnormal development that may result in major or minor malformations
o Spontaneous abortion or stillbirth (with or without malformations)
o Asymptomatic or subclinical (ie. female carrier of X-linked trait)
· Single Gene Disorders with Mendelian Inheritance:
- Definition: diseases resulting from a mutation in a single gene of large effect, inherited according to Mendelian patterns
- Inheritance Patterns: most are recessive*
o Autosomal Dominant: clinical phenotyp occurs with single copy of mutant allele
§ Usually non-enzymatic proteins
§ 1 parent affected (if not affected, de novo mutation)
§ Both males and females affected
§ 1 in 2 chance of transmitting disorder
§ Clinical features modified by reduced penetrance and expressivity
o Autosomal Recessive: clinical phenotype occurs only when both alleles are defective (although defect in both alleles does not have to be the same)
§ Usually enzymatic proteins
§ Trait usually does not affect parents
§ Both males and females affected
§ 1 in 4 chance of transmitting disorder
§ More uniform expression of defect than AD disorders (complete penentrance common)
§ Onset usually early in life
o X-linked: more common in males (only 1 X cs)
- Mechanisms of Pathogenesis of Single Gene Disorders:
o Mutations:
§ Enzymes or enzyme inhibitors
§ Receptors
§ Transport or structural proteins
o Enzyme Defects: result in reduce or absent enzyme leading to specific block in metabolism (usually of a catabolic pathway) leading to
§ Abnormal accumulation of metabolites (substrates)
§ Decreased amount of end product that is necessary for normal function
§ Failure to inactivate a tissue damaging substance
ENZYME DEFECTS- LYSOSOMAL STORAGE DISEASES:
· Lysosomal Storage Diseases- General:
- Lysosomes: “intracellular digestive tract” involved in turnover of macromolecules within the cell
o Contain degradative enzymes called hydrolases
o Function in acid environment of the lysosome
o Made in the ER, sent to the Golgi, undergoes post-translational modification to target it to lysosomes
- Lysosomal Storage Diseases: due to lack of any protein essential for normal function of lysosomes
o Distribution of non-degraded material (ie. organ affected) due to an LSD is determined by:
§ Tissue where most of the degraded material is found
§ Location where most of the degradation usually occurs
o Rare (1/8000 births)
o Most are autosomal recessive
§ Exceptions: both are X-linked recessive
Ø Fabry Disease
Ø Hunter Syndrome
o Grouped based on type of macromolecule undergoing degradation:
§ Mucopolysaccharides
§ Sphingolipids
§ Sulfatidoses
§ Cerebrosides
· Mucopolysaccharidoses (MPS):
- General:
o Mucopolysaccharides: group of macromolecules composed of glycosaminoglycans (GAGS), which are long chain carbohydrates with disaccharide repeating units
§ GAG Examples:
Ø Dermatan sulfate (heart, blood vessels, skin)
Ø Heparan sulfate (lung, arteries, skin)
Ø Keratan sulfate (cartilage, cornea, intervertebral discs)
Ø Chondroitin sulfate
Ø Hyaluronic acid
§ Proteoglycans: GAG + protein (secreted by GAG synthesizing cells)
§ Function: involved in structural integrity of the ECM
§ GAG Synthesizing Cells: fibroblasts, endothelial cells, leukocytes
Ø Secrete proteoglycans
Ø GAGs that are not secreted are degraded by lysosomal hydrolases (enzyme deficiency results in accumulation in lysosomes and free GAGs in urine)
o Mucopolysaccharidoses (MPS):
§ Usually progressive disorders
§ Classified numerically from MPSI-MPSIV
§ Share similar clinical features:
Ø Multi-system involvement
Ø Organomegaly
Ø Abnormal facies
Ø Joint stiffness and deformity
Ø Mental retardation
§ Diagnosis:
Ø Presence of GAGS:
o Urinary: age-dependent (present in normal infants up to a year old)
§ Look for larger than normal amounts of heparan and dermatan sulfate (normal urine contains mostly chondroitin sulfate)
o Amniotic fluid
Ø Enzyme Assays:
o Prenatal:
§ Cultured cells from amniotic fluid
§ Chorionic villus biopsy less suitable (have low enzyme activity)
o Postnatal: measure enzyme activity in plasma, leukocytes or skin fibroblasts
- Hurler Syndrome (MPS I):
o Inheritance Pattern: autosomal recessive
o Deficiency: α-L-idurondase enzyme activity
o Onset: 6-8 months
o Common Features: severe mental and motor regression with death usually before 10 years of age
§ Respiratory disease
Ø Storage in airway epithelium and bone
Ø Small ribcage (oar-shaped ribs) and stiff joints
Ø Decreased expansion due to hepatomegaly
Ø Upper airway obstruction due to storage in tongue, lymphoid tissue, airway epithelium, pharyngeal soft tissue
§ Coarse facial features
§ Ophthalmic disease with early corneal clouding (retinal disease, glaucoma)
§ CV disease (valve disease, CAD, congestive heart failure)
Ø Accumulation of GAGs in histiocytes, mycocardial cells, heart valves, coronary arteries and aorta
§ Dwarfing
Ø Accumulates in bone (growth plates) and prevents linear growth
§ Stiff joints
§ Hepatosplenomegaly
Ø Storage in hepatocytes and Kupffer cells of lvier
Ø Storage in histiocytes in the spleen
§ CNS disease (mental retardation, hydrocephalus, spinal cord compression)
Ø Storage in neurons, macrophages and meninges
§ Deafness
§ Alder-Reilly anomaly (accumulation of MPS in PMNs- granular appearance)
o Diagnosis: urinary excretion of dermatan and heparan sulfate
- Hunter Syndome (MPS II):
o Inheritance Pattern: X-linked recessive
o Deficiency: iduronate sulfatase
o Onset: in early infancy or childhood
o Common Features: similar clinical features to Hurler, but LESS severe (mental deterioration with varying degrees of neurological involvement)
§ Coarse facial features
§ Dwarfing
§ Stiff joints
§ Progressive deafness
§ NO corneal clouding*
o Diagnosis: urinary excretion of dermatan and heparan sulfate
· Sphingolipidoses:
- General:
o Cause: defect in metabolism of sphingolipids
o Sphingolipids: long-chain amino alcohols (sphingosine) attached to a fatty acid to produce a complex lipid (ceremide)
§ Membrane Lipids:
Ø Sphingomyelin
Ø Glycosphingolipids:
o Cerebrosides (add sugar to ceremide)
o Sulfatides
o Globosides
o Gangliosides (add polysaccharide + N-acetylnueramic acid to ceramide)
- Niemann-Pick Disease:
o General:
§ Type I (A and B):
Ø Cause: deficiency of sphingomyelinase enzyme, leading to progressive accumulation of sphingomyelin (ubiquitous component of cellular organellar membranes)
§ Type II (C and D):
Ø Cause: defect in cholesol esterification causing lysosomal accumulation of unesterified cholesterol
o Type I:
§ Type A (Severe Infantile):
Ø Most common of type I: 70-80% of all cases (often in Eastern European Jews)
Ø Presentation: within the first weeks of life (death occurs by age 3 or 4)
o Severe neurologic impairment (hypotonia, progressive psychomotor retardation)
o Failure to thrive
o Hepatosplenomegaly (marked)
o Macular cherry red spot (50% of patients)
o Accumulation of foamy lipid macrophages in many tissues (liver, lung, spleen, LNs, kidneys, bone marrow, peripheral and central neurons)
o Atrophy of the brain
§ Type B (Chronic Visceral):
Ø Less common than Type A: also often seen in Eastern European Jews
Ø Presentation: infancy or childhood (typically survive into adulthood)
o Organomegaly (often present with splenomegaly and then develop generalized visceral involvement)
o Generally NO CNS involvement
o Type II:
§ Type C:
Ø Most common: more common than A and B combined
Ø Cause: defect in NPC-1 (95%) or NPC-2 gene that encodes for a protein involved in cellular trafficking of exogenous cholesterol (normally brings free cholesterol from the lysosome to the cytoplasm)
Ø Clinical Features: variable
o Classic Phenotype (Neurovisceral): presents in childhood (2-4 years) with death typically occurring between 5-15 years of age
§ Variable hepatosplenomegaly
§ Vertical supranuclear opthamoplegia (supranuclear palsy)
§ Progressive ataxia
§ Seizures
§ Psychomotor regression (accumulation of cholesterol in neurons is lethal to those cells)
§ Bone marrow contains Niemann-pick cells (foamy) and sea blue histiocytes
o May present at birth with hydrops fetalis and still birth
o Fatal neonatal liver disease (giant cell hepatitis)
o If live into adulthood, may present with dementia and psychosis
§ Type D:
Ø Rare variant of type C: found in Nova Scotia (less severely affected)
- Gangliosidoses:
o GM1 Gangliosidoses:
§ Deficiency: deficiency in β-galactosidase enzyme that results in accumulation of ganglioside in neurons (clinical variability depends on amount of residual enzyme activity)
Ø Β-Galactosidase Enzyme: 3 isoenzymes (A,B,C)
o Transcription of this enzyme requires an activator that may cause similar features if deficient
§ Type I (Generalized/Infantile):
Ø Deficiency: virtual absence of all 3 isoenzymes of B-galactosidase
Ø Onset: birth to 6 months (death often before age 2)
Ø Clinical Features:
o Progressive neurologic deterioration (seizures)
o Coarse facial features
o Hepatosplenomegaly (accumulation of gangliosides in liver, spleen, renal tubular epithelium)
o Macular cherry red spots (50% of patients)
o Skeletal deformities (dystosis multiplex)
o Has features of both neurolipidoses and MPS, leading to a “pseudo-Hurler phenotype”
§ Type II (Juvenille):
Ø Deficiency: absence of A and B isoenzymes only
Ø Onset: juvenile (1-2 years old) with death occurring by age 3-10
Ø Clinical Features:
o Slower psychomotor retardation
o Less visceromegaly
o Milder skeletal disease
§ Diagnosis:
Ø Measurement of B-galactosidase activity in peripheral blood leukocytes:
o Type I: virtually no activity
o Type II: 5-10% activity
Ø Peripheral Blood Smear: vacuolization of lymphocytes (crude method due to the fact that many LSDs have this feature)
o GM2 Gangliosidoses:
§ Cause: inability to catabolize GM2 ganglioside (required 3 polypeptides encoded by 3 separate loci)
Ø Tay Sachs Disease: defect in α subunit of Heaminidase A
Ø Sandhoff’s Disease: defect in β subunit of Hexaminidase A
Ø GM2 Activator Deficiency Gangliosidosis: defect in GM2 activator
§ Clinical Features: similar because they all result in GM accumulation
Ø GM2 accumulates in many tissues, but CNS and retina are the dominant features of the disease
o Neurons are swollen and contain cytoplasmic vacuoles
o Lysosomes contain whorled material by EM
o Brain first enlarges, and then becomes atrophic
Ø Infants appear normal at birth, followed by rapidly progressive neurodegenerative disease with seizures, dementia and blindness
o Loss of motor skills at 3-6 months
o Death by 2-4 years of age
Ø Macular cherry red spot
§ Ethnic Risk: 1/30 carrier rate in the Askenazi Jewish population
- Sulfatidoses:
o Metachromatic Leukodystrophy:
§ Cause: deficiency of arylsulfatase A enzyme, leading to the accumulation of non-degradeable galactocerebroside sulfate in the white matter of the brain, peripheral nerves, liver and kidney
Ø Results in breakdown of myelin sheath (demyelination and gliosis)
Ø As a result, predominantly a neurodegenerative disorder
Ø Arylsulfatase A also requires presence of saposin B (solubilizes hydrophobic lipid to allow it to be accessible to the enzyme) and therefore, absence of SAP will cause similar disease
§ 3 Forms:
Ø Late Infantile: most common (diagnosed by age 2; death before age 5)
o Regression of motor skills (hypotonia, muscle weakness)
o Mental deterioration (loss of milestones)
o Rigidity and convulsions
o Unusual loss of white matter on CNS imaging
Ø Juvenile: diagnosed between 3-16 years of age; death 6-8 years after diagnosis
o Changes in gait and cognitive skills
o Progressive regression of all skills
Ø Adult Onset: diagnosed after 16 years of age
o Psychiatric or cognitive symptoms occur first
o Motor symptoms (neurologic) appear later
§ Diagnosis:
Ø Urine: spot screening test (shows metachromasia due to presence of sulfatide)
Ø Imaging: usual loss of white matter due to demyelination
Ø Biopsy: usually of a sural nerve (demyelination, metachromatic granules and unusual cytoplasmic inclusions)
o Stains: PAS +, Alcian blue, acidified Cresyl violet
Ø Arylsulfatase A activity levels
Ø Genetic testing: gene located on long arm of cs 22
o Multiple Sulfatase Deficiency:
§ Cause: reduction in activities of several sulfatidases (arylsulfatidase A,B,C) resulting in the accumulation of sulfatides, sulfated GAGs, sphingolipids and steroid sulfates
§ Clinical Presentation: combines clinical features of metachromatic leukodystrophy and MPS
- Cerebrosidoses:
o Gaucher Disease: most common lysosomal storage disease*
§ Defect: glucocerebrosidase enzyme (results in accumulation of glucocerebrosides and other glycolipids)
Ø Glucocerebrosides: derived from breakdown of the membranes of senescent leukocytes and RBCs
Ø Accumulation: incompletely metabolized substrate stored in monocytes and macrophages (stains positively with PAS)
o Gaucher Cell: crumpled paper appearance to cytoplasm with eccentrically displaced nucleus
§ Type 1 (Chronic Non-Neuronopathic Form):
Ø Cause: reduced levels of glucocerebrosidase
Ø Most common type: 99% of cases
Ø Population affected: children and adults (common in Ashkenazi Jews)
Ø Clinical features: accumulation of glucocerebrosidase limited to mononuclear phagocytes throughout the body, WITHOUT brain involvement