GENETIC DISORDERS

M. Kent Froberg, M.D.

2009

Objectives

• Learn the basic characteristics of cytogenetic, Mendelian and non-classical inherited disorders

• Be able to distinguish features of autosomal dominant, autosomal recessive and X-linked genetic disorders

• Understand the pathogenesis of the most common genetic diseases

GENETIC DISORDERS:METHOD OF STUDY

• Classical genetics: requires knowledge of gene

product to ID gene

• Reverse genetics: protein unknown, use marker

linkeage to locate gene of interest

DISORDERS

• Chromosomal: # or structure, ex. trisomy 21

– Good for cytogenetic ID

• Mendelian disorders: Mutant genes of large effect (SCD), often requires molecular techniques

• Multifactorial: polygenetic + environment (DM, HTN, CHD)

– Less amenable to genetic studies

CYTOGENETICS (1)

• Cytogentic disorders identified through karyotyping

• Karyotyping: visual image of chromosomes– NL karyotype 46 xx (xy) – 22 homologous pairs + sex chr.

• Method: – Cell culture (usually peripheral blood

lymphocytes)– Arrest in metaphase (colchicine)– Stain G banding (Giemsa stain)

CYTOGENETICS (2)

• Method (cont.):

– Light and dark bands, sort by size

– Banding pattern

– Centromere location:

• metacentric: middle

• submetacentric: off-center

• acrocentric: near end

• telocentric: not in humans

CYTOGENETICS (3)

• Limitations:

– Requires viable cells, visible changes (4 mil bp)

• Advantages:

– Good for aneuploidy, large structural changes, del, t, i

– Examples:

• 47, xy, +21

• 46, xx, del 5q-

• 46, xy, t(9;22)

MUTATIONS (1)

• Definition: heritable alteration in genome

• Types:

– Substitution: point mutation

– Insertion: alters reading frame

– Deletion: alters reading frame, i.e., frameshift

MUTATIONS (2)

• Consequences:

– Codon mutations:

• Missense: one AA for another

–Ex.: CTC to CAC, glu to val in SCD

• Nonsense: stop; CAG to UAG = stop

–Ex: thal

MUTATIONS (3)

• Consequences (cont.):

– Noncoding sequence:

• Promoter or enhancer: decreased or no transcription

• Introns: defective splicing, most common cause -thal

CYTOGENETIC DISORDERS

• 50% of early abortions, 5% stillbirths, .5-1% livebirths

• Aneuploid = < 46 >

– Secondary meiotic nondisjunction or anaphase lag

– Monosomy (autosomal usually fatal) and trisomy (some viable)

• Mosaicism = 2 or more cell lines in genome

– Secondary early mitotic errors

– Most common with sex chr. and Down syndrome

CYTOGENETIC DISORDERS

• Chromosome breakage syndromes– Autosomal recessive– Have increased risk of CA– Multi-organ system changes:

• Fanconi anemia: developmental problems, aplastic anemia, myelodysplasia, leukemia

• Bloom syndrome: short stature, photosensitivity, risk cancer

• Ataxia-telangectasia:

neurodegenerative disorder, risk ALL & lymphoma

CYTOGENETIC DISORDERS

• Rearrangements:

– Reciprocal translocation: usually compatible with life, but associated with CA or abnl offspring

– Robertsonian translocations: centric fusion of acrocentric chr. (rRNA from p of 13, 14, 15, 21, 22), abnl offspring

CYTOGENETIC DISORDERS

• Rearrangements (cont.):

– Isochromosome: one arm lost, other duplicated & fused to centromere, so two p or two q (iXq)

– Ring: deletion at both ends & fusion (2 breaks)

– Inversions: compatible with NL development

Origins and examples of

triploidy, trisomy and

monosomy

Most common

trisomies

CYTOGENETIC DISORDERS

• Trisomy 21 (Down Syndrome): most common chr disorder, causes MR

– Incidence ~1/800; 95% extra chr., 4% Robertsonian trans (familial), 1% mosaics (milder MR)

– Correlated with maternal age: incidence <20 ~1/1550, >45 ~1/25. Due to maternal meiotic non-disjct in 95%

CYTOGENETIC DISORDERS

• Trisomy 21 (Down Syndrome) (cont.):

– CSx:

• flat face, oblique palpebral fissures, epicanthal folds, severe MR ( IQ 25-50 in 80% ),

• congenital heart disease (40%), ALL & AML ~10-20x risk, Alzheimer Disease at ~40,

• abnl immune response (lung infections & thyroid disorders)

• 80% survive >30 years

Mosaic with subtle features of trisomy 21

CYTOGENETIC DISORDERS

• Other trisomies (cont.):

– 18 (Edward Syndrome):

• 1/8000 births, overlapping fingers, MR, low set ears, heart defects

– 13 (Patau syndrome):

• 1/6000, heart defects, polydactyly, cleft lip & palate, microphthalmia, MR

Features of trisomy 13

Facies of trisomy 13

Trisomy 18

CYTOGENETIC DISORDERS

• Cri du chat (5p- syndrome):

– Severe MR

– Microcephaly

– Round face

– Some survive to adults

CYTOGENETIC DISORDERS

• Sex chromosome disorders:

– More common than autosomal, better tolerated because of lyonization & lack of genetic material of Y chr: lyonization (16th day embryonic life, Xist gene, secondary methylation).

• subtle/chronic problems sexual development & fertility

• often 1st recognized at puberty

• more Xs ~ more likely MR

CYTOGENETIC DISORDERS

• Klinefelter Syndrome: Male hypogonadism; 1 or more Ys & 2 or more Xs

– 82% = 47, XXY, ~50% from 1st meiotic paternal non-disjct, 15% mosaics, incidence 1/850 live male births

– Principal cause male infertility

– Testes: tubular atrophy, prominent Leydig cells

CYTOGENETIC DISORDERS

• Turner Syndrome: monosomy X, hypogonadism in phenotypic female

– 45, X (57%) but only 1% survive to birth

• del Xp, i (Xq), or partial del of Xp or Xq

• so variable phenotypes

CYTOGENETIC DISORDERS

• Turner syndrome (cont.):

– CSx: edema in infancy in most severely affected =

cystic hygroma,

– congenital HD esp. coarctation of aorta

– Later: #1 cause primary amenorrhea, short stature, webbed neck, broad chest, widely spaced nipples

– Streak ovaries secondary to accelerated loss of oocytes by 2 years

Cystic hygroma in female stillborn fetus

Edema of the hand in newborn with Turner Syndrome

Short stature and webbed neck of Turner Syndrome

CLASSICAL GENETIC DISORDERS (MENDELIAN)

• Expressed mutations in single genes of large effect (sickle cell anemia)

– Pleiotropism: multiple end effects

– Genetic heterogeneity: same effect by mutations at several genetic loci

TRANSMISSION PATTERNS

• Autosomal dominant:

– Manifested in heterozygous state

– Affects males & females

– Affected X unaffected = 50% transmission

– Some due to new mutations

– Clinical features modified by variable expressivity (seen in all with gene, but expressed differently) & reduced penetrance (i.e., 50% of those with gene express trait)

TRANSMISSION PATTERNS

• Autosomal dominant (cont.):– Age of onset may be delayed• Huntington Disease

– Often structural protein or transport protein• hereditary spherocytosis• Marfan syndrome• familial hypercholesterolemia• NF• tuberous sclerosis

TRANSMISSION PATTERNS

• Autosomal recessive:

– Single largest category Mendelian disorders

– Parents not affected

– Siblings 1 in 4

– Expression more uniform

– Penetrance usually complete

– Onset early in life, new mutations undetected

TRANSMISSION PATTERNS

• Autosomal recessive (cont.):

– Enzymes involved

– Heterozygote has 1/2 nl & 1/2 abnl

• inborn errors of metabolism

• CF, SCD, PKU, 1-antitrypsin deficiency, most storage diseases, Ehlers-Danlos

TRANSMISSION PATTERNS

• X-linked:

– All sex-linked are X-linked

– Most recessive

– Male is hemizygous for X so expressed in males

– Sons of affected male not affected

– Daughters carriers

– Sons of heterozygous female have 1 in 2 chance

TRANSMISSION PATTERNS

• X-linked (Cont.):

– Female may be affected because of skewed lyonization

– Usually partial expression (G6PD def.)

– Examples:

• DMD

• hemophilia

• Wiscott-Aldrich

• chronic granulomatous disease

BIOCHEMICAL BASIS OF MENDELIAN DISORDERS

• Direct effect on protein or indirect–consequences:– accumulation of substrate or abnl product

• Enzyme defects: – Decrease end-product: albinos - no melanin

secondary to deficiency of tyrosinase– Increased substrate: phenylalinine-PKU– Failure to inactivate toxic substrate

-antitrypsin deficiency

BIOCHEMICAL BASIS OF MENDELIAN DISORDERS

• Receptor or transport defect:

– Failure to cross cell membrane (familial hypercholesterolemia)

• Structural protein defect:

– Affects function (DMD, SCD)

• Adverse drug reactions:

– Oxidative injury (G6PD def)

SPECIFIC DISORDERS

• Marfan Syndrome: connective tissue disorder, 70-85% autosomal dominant, variable expression– Marfanoid appearance: tall, long fingers, lax

joints, dolicocephalic, frontal bossing, scoliosis, kyphosis, ectopia lentis, aortic cystic medial degeneration (dissection cause of death in 30-40%)

– Cause: defective fibrillin (extracellular matrix protein for elastin, mapped to 15q21.1)

Section of aorta with cystic medial

degeneration

Cystic medial degeneration

Smooth muscle cells of media (arrows) are separated by cystic spaces

and basophilic material

Lack of elastin staining

SPECIFIC DISORDERS

• Ehlers-Danlos Syndrome:

– Heterogeneous group of defects of collagen synthesis or assembly

– Inheritance by all three Mendelian patterns

– Variable phenotypes

– Largely affects skin & ligaments

RECEPTOR DEFECTS

• Familial hypercholesterolemia: due to mutation in gene for LDL receptor–loss of feedback control– Increased cholesterol– Early AS & MI – Mendelian disorder: multiple mutations– Heterozygous (1 in 500): increased cholesterol 2-3X, increased

AS & xanthomas– Homozygous: 5-6X increased cholesterol, early AS, MI in 20s

ENZYME DEFECTS

• Lysosomal Storage Diseases:

– Membrane-bound bags of hydrolytic enzymes, autophagy & heterophagy

– Enzyme defect leads to substrate accumulation in cell

– Mutations may affect enzyme activator, post-translational processing, substrate activator, lack of transport protein

GANGLIOSIDOSIS

• Tay Sachs (GM2) most common of 3 types affecting

hexosaminidase A, all with similar phenotypic effects

– Eastern European Jews (carrier rate 1 in 3)

– Gangliosides accumulate in all tissues but neurons in CNS & retina affected most (ballooning degeneration with oil red o-positive vacuoles)

GANGLIOSIDOSIS (Cont.)

• Tay Sachs (GM2) (Cont.):

– NL at birth–CSx by 6 months–mental and motor deterioration, blindness, cherry-red spot on macula

– Multiple mutations on Chr 15 (Sandoff & activator def on chr 5)

A. Nerve cell body distended with

gangliosides

B. EM of lysosomes with whorled

configurations in neuron of Tay-

Sachs Disease patient

NIEMANN-PICK DISEASE

• Heterogeneous group that lead to accumulation of sphingomyelin & cholesterol

• Two major types

• Type A accounts for 75-80% (def. sphingomyelinase)

– Severe infantile form, CSx by 6 months

– Viscera & CNS affected (hepatosplenomegaly)

NIEMANN-PICK DISEASE (Cont.)

• Type A (Cont.):

– Skin xanthomas

– Small uniform lipid vacuoles accumulate in mononuclear phagocytes and neurons with increase in size to 90

– Spleen to 10X NL

– Retinal cherry-red spot in 1/3 to 1/2

– Have FTT, generalized lymphadenopathy, progressive psychomotor deterioration, death 1-2 yrs

Spleen of patient with Niemann-Pick Disease. The red pulp is expanded with

lipid-laden histiocytes. Arrows indicate white pulp.

Phagocytes in splenic red pulp contain many small vacuoles filled with

sphingomyelin.

GAUCHER DISEASE

• Group of autosomal recessive disorders with defect in glucocerebrosidase

– Most common lysosomal storage disorder

– Cleaves glucose from ceramide

– Accumulates in macrophages

– Three clinical subtypes

GAUCHER DISEASE (Cont.)

• Type 1:

– 99%

– Chronic non-neuropathic form

– Accumulation esp. in spleen & bone marrow, esp. European Jews

GAUCHER DISEASE (Cont.)

• Gaucher cells in spleen, liver, bone marrow: fibrillary distended cytoplasm (PAS-positive)

• CSx: 1st appear in adult life, often pancytopenia

or thrombocytopenia secondary to hypersplenism

– May have bone pain or pathologic fractures, but compatible with long life

Gaucher cells in bone marrow have pale, granular cytoplasm

with a fibrillar appearance

Lymph node filled with pale, eosinophilic histiocytes in Gaucher

Disease

Gaucher cells fill alveoli in lungs of patient with long-

standing disease

PAS positive (left) and unstained histiocytes (right) in Gaucher Disease

MUCOPOLYSACCHARIDOSES (MPS)

• Group of deficiencies of degradation of glycosoaminoglycans, accumulate within macrophages and endothelium

– MPS I H: Hurler syndrome, def -L-iduronidase, most severe form of MPS

• NL at birth

• 6 mos hepatosplenomegaly

• skeletal deformities

• death in 6-10 yrs of CV complications

MUCOPOLYSACCHARIDOSES (MPS) (Cont.)

– MPS II H: Hunter syndrome

• X-linked

• milder CSx

• no corneal clouding

GLYCOGEN STORAGE DISEASES

• Glycogenoses: deficiency of one of enzymes of glycogen synthesis or degradation, liver and muscle target organs, Types I-VIII

– Hepatic forms: von Gierke disease (Type I), def. G-6-phosphatase

• hepato-renomegaly: glycogen in cortical tubular epithelium, hypoglycemia, FTT, bleeding tendency, 50% mortality

GLYCOGEN STORAGE DISEASES (Cont.)

– Myopathic forms: McArdle syndrome (Type V), def. muscle phosphorylase

• accumulation of subsarcolemmal glycogen

• painful cramps after exercise without increase in venous lactate

• normal longevity

CELL GROWTH GENE DEFECTS

• Most in somatic cells–CA, ~5% transmitted through germline, proto-oncogenes and tumor-suppressor genes

– Neurofibromatosis-1 (von Recklinghausen disease): 1/3000, 50% autosomal dominant, penetrance 100% but variable expressivity. Have:

• multiple neural tumors

• numerous pigmented skin lesions (90%)–”cafe au lait” spots

• iris hamartomas–Lisch nodules

CELL GROWTH GENE DEFECTS (Cont.)

– Neurofibromas:

• Arise from nerve trunks in skin

• Soft, multinodular tumors 1 cm to > 20 cm (plexiform)

• Contain all elements, i.e., proliferating neurites, Schwann cells and fibroblasts within loose myxoid stroma

• Mal transformation 3%

CELL GROWTH GENE DEFECTS (Cont.)

– NF-1: Gene mapped to chr 17q11.2, variable expression, pts also have bone cysts, scoliosis, meningiomas, optic gliomas

– NF-2: Bilateral acoustic schwannomas, may not have skin tumors, no Lisch nodules, mapped to chr 22

MULTIFACTORIAL INHERITANCE

• Increased risk if parents severely affected

• Greatly modified by environment

• Risk in sibling of affected child same (2-7%)

• Ex: DM, cleft lip and palate, CHD, HTN

NONCLASSIC INHERITANCE

• Triple repeats

• Genomic imprinting

• Mutations in mitochondrial genes

NONCLASSIC INHERITANCE:TRIPLE REPEAT MUTATIONS

• Fragile X, HD, myotonic dystrophy

– Fragile X: One of most common forms of familial MR, X-linked, IQ 40-70, macro-orchidism in 80%, long face, large mandible, large everted ears

• 20% of affected males are carriers, 30% of carrier females have MR, anticipation = clinical features worsen with successive generations

NONCLASSIC INHERITANCETRIPLE REPEAT MUTATIONS

– Fragile X (Cont.):

• Mechanism: nl to have 6-54 tandem repeats of CGG at Xq27

• “Premutations”: 52 - 200 CGG repeats

• “Full mutations”: 250 - 4000 repeats, amplified in oogenesis

Trinucleotide repeat blot showing pre- and full mutations.

NONCLASSIC INHERITANCE:GENOMIC IMPRINTING

• “Parent of origin” effects, one parent expressed for given gene– Prader-Willi & Angelman syndromes: same

area deleted; del15q11-13– PWS: MR, short stature, hypogonadism, FTT

early, obesity later, all cases del is in paternal chr 15

– AS: MR, ataxic gait, SZ, inappropriate laughter = “happy puppets,” have deletion on maternal chr 15

NONCLASSIC INHERITANCE:GENOMIC IMPRINTING (Cont.)

– Mechanism: methylation of DNA

– Some cytogenetically nl have two copies of 15 from one parent = uniparental disomy

Genomic Imprinting as seen in Pader-Willi and Angelman Syndromes

mat.

pat.

Deletion

pat.

mat.

mat.

mat.

Prader-Willi syndrome Angelman syndrome

pat.

pat.

Uni-parentaldisomy

mat.

pat.

PWS AS PWS AS

Normal

PWS AS

= active gene = inactive gene

Angelman Syndrome

MOLECULAR DIAGNOSIS

• Detection of inherited mutations

• Detection of acquired mutations in neoplasia

• Dx & classification of hematopoietic neoplasms

• Dx of infectious diseases (TB & HIV)

• Determination of relatedness & identity in transplantation, paternity testing and forensic medicine

MOLECULAR DIAGNOSIS (Cont.)

• Advantages:

– Very sensitive (can be disadvantage)

– Cells need not be alive

• any cell works (except mature RBCs)