Genetikk

Question 1

Genetic life time prevalence

Answer 1

3,4-7,3 %

Phenotype description, molecular basis known: 5625

Question 2

Allel

Answer 2

One of several alternative forms of a gene
In the 22 pair of autosomes, you have 2 alleles; one from mom and one from dad
A/a = Heterozygous
A/A or a/a = Homozygous

Question 3

Polymorphism

Answer 3

a genetic variant that is conserved in the population, and has an allele frequency of ≥ 1%.
Common genetic variants.
Over 340 mill known Single Nucleotide Polymorphisms (SNPs) in the human genome.

Question 4

Mutasjoner:

Most chromosome abnormalities (aneuploidies, translocations, deletions, duplications) have no previous family history.

Prevalence of chromosomal abnormalities
• Newborns: 0.5%
• Stillborns: 5%
• Spontaneous abortions, 1st trimester: 50%

• Robertsonian translocation (between acrocentric chromosomes). Balanced carriers: 1/1000

Answer 4

Kromosomal: Numerical alterations
e.g. monosomy, trisomy, triploidy

Regional: Deletions, duplications, translocations.

Gen:
- Punktmutasjoner; Deletion and insertion of one or more nucleotides. Missense, nonsense, splice site

Question 5

Missense mutation

Answer 5

One aa is replaces by another one

Question 6

Nonsense mutation

Answer 6

an aa codon is replaced by a stop codon

Question 7

Frameshift mutation

Answer 7

Deletion (or insertion) of a base resulting in a shift in all subsequent codons)

Question 8

Splice site = no splicing

Answer 8

Leads to «exon-skipping» and deletions at the mRNA

level.

Question 9

DNA analysis:

to detect changes that are not visible under an optical microscope

Answer 9

• PCR (Polymerase Chain Reaction)

• MLPA (Multiplex Ligation-dependent Probe Amplification) used for Deletion and Duplications.
Eks:
Duchenne muscluar dystrophy (deletion) Charcot Marie Tooth disease.

• SNP- array (single nucleotide polymorphism)

• DNA Sequencing. Can detect any point mutation
and small deletion/insertion.

Question 10

Classical Mendelian inheritance
• Autosomal recessive
• Autosomal dominant
• X-linked inheritance

Answer 10

Non Mendelian inheritance

• Mitochondrial inheritance, from mother

Question 11

Pedigree (genogram)

Answer 11

Is a family tree showing the pattern of inheritance for a specific phenotypic trait

Question 12

Autosomal (ikke kjønnsbundet) recessive diseases

• Healthy parents may have one or more affected children.
• Men and women are affected equally often.
• The parents of an affected child are obligate carriers.
• Each of their children has a 25% risk of inheriting the disease.
• Each healthy sibling of an affected child has a 67% risk of being a carrier.
  Mor og far må begge være bærere: 1 unge får sykdommer, 2 blir bærere, 1 blir frisk. Så 2/3 = 67% bærere.

• The affected gene resides on one of the 22 autosomal chromosomes.

Answer 12

Examples:

• Cystic fibrosis is a disease caused by “thick mucous”: mucoviscidosis; chronic lung infection !!
  pancreatic insufficiency – malabsorption, increased CHLORIDE concentration in sweat !!
  More than 1500 known mutations. Most frequent mutation is del F508 (Phe508).
• Phenylketonuria (PKU) (Følling’s disease)
  Neonatal screening Caused by mutations in the PAH gene (Phenylananine hydroxylase).
  Phenylalanine accumulates in the body fluid and damages the developing CNS.
  Therapy: Restricted dietary intake of phenylalanine.
• Spinal muskel atrofi: 1/30-40 er bærere
  SMA-1, onset før 6 måneder, death before 2 years
  SMA-2, onset 6-18 måneder, 70% overlever til 25 år.
  Behandling: Nusinersen: Spinraza

In total over 2000 autosomal recessive diseases, including many congenital metabolic diseases.

Question 13

Autosomal (ikke kjønnsbundet) dominant disorders

• Each child of an affected person has a 50% risk of inheriting the disease
• Occurs in each generation. Often late onset.
• Men and women are affected equally often. Male-to-male transmission occurs, which is an important way of distinguishing these disorders from X-linked recessive disorders.
• Penetrance; is the proportion of individuals with the mutation who exhibit clinical symptoms.
• Expressivity; is the variations in a phenotype among individuals carrying a particular genotype.

• Autosomal dominant disorders may be caused by new mutations (relatively rare) originating from germline mosaicism in one of the parents or from early embryonic mutations in the offspring.

Answer 13

Examples:

• Huntington’s disease; neurodegenerative
• Marfan syndrome; connective tissue disorder
• Colon polyps; origin of colon cancer
• Myotonic dystrophy; progressing muscular dystrophy

More than 2600 known autosomal dominant diseases

Question 14

X-linked recessive disorders

• Mainly affecting men!
• Diseases are transmitted by healthy* mothers
• Heterozygous women are generally unaffected. Exceptions: skewed X inactivation, monosomy X (Turner)
• All daughters of any man with the disease become carriers. Each of his daughters’ sons has a 50% risk of inheriting the disease
• Not transmitted from father to son.
• Can be transmitted unrecognized via several female carriers

Answer 14

Examples:

• Haemophilia type A and B

• Duchenne muscular dystrophy
1/ 3500 boys, very few girls

• Fragile X syndrome

Question 15

Mitochondrial inheritance

• Caused by mutations in mtDNA
• Inherited from mother to child; maternal inheritance (sperms do not transmit mitochondria)
• Low incidence
• Affects both sexes

Answer 15

Examples:

– Lebers’ inheritable optic neuropathy (LHON)

– Inheritable hearing impairment

• Most proteins in mitochondria are encoded by nuclear genes. Mutations in these genes may cause mitochondrial disease.

Question 16

The genetic consultation

Laboratory analyses

1) Chromosome analysis

I. Traditional chromosome analysis (G-band, living cells)

Answer 16

• Chromosomes only visible in dividing cells

• Living cells from different tissues cultured to divide
– Blood lymphocytes (Na-heparin sample)
– Skin fibroblasts (or achilles tendon biopsy)
– Amnion - amniocytes
– Chorionic villi biopsy - fetal cells

• Culture cells, make them divide
• Arrest cells, metaphase
• Stain with ie Giemsa = G-band preparation
• Microscope (11 cells or more)
• 2-3 weeks for result

Question 17

III. QF-PCR
Quantitative Fluorescence–Polymerase Chain Reaction

DNA based, RAPID test for aneuploidies
– 1-2 days
Amplifies amount of DNA in specific regions
– Quantifies chromosomes 13, 18, 21, X, Y

Answer 17

Indications for chromosomal analyses

• G-band analysis
– 3 or more spontaneous abortions or infertility.
– Suspected structural chromosomal aberration.
– Suspected sex chromosome abnormality (growth, pubertal development)

• QF-PCR including sex chromosomes
– Prenatal diagnostics
– Learning difficulty

• SNP array
– Intellectual disability, severe autism
– Congenital malformations – major, or several minor

Question 18

IV. Chromosomal microarrays

• Submicroscopic (< 5-7 Mb) chromosomal aberrations by DNA based methods.

• Several names/methods
– (Genomic) Copy number analysis («kopitallsanalyse»)
– Array CGH (comparative genome hybridisation)
– SNP array (Single Nucleotide Polymorphisms)
– «Matrisebasert kopitallsundersøkelse»

Answer 18

• Detects deletions, duplications
• NB! Normal variation
• Indications: ID, (multiple) malformations, syndromes,…
• Probably replacing most traditional G-band analyses
• Detects only unbalanced abnormalities

Question 19

Aneuploid = Extra/missing single/few chromosomes

• Monosomy (e.g. 45, X)
• Trisomy (e.g. 47, +21)
• Sex chromosomes or autosomes

Answer 19

Euploid = Extra set(s) of chromosomes

– Polyploidy (e.g. triploidy 69, XXX)
– Not compatible with life (but occurs at conception, or mosaic)

Question 20

Klinefelter syndrome (Høy)
- 47, XXY
(48, XXXY etc)

Turner syndrome, 45, XO (Lav)

Answer 20

Congenital anomalies:

• Major congenital anomaly: 2-3% newborns

Question 21

Huntington disease

• Irregular involuntary movements (chorea)
• Cognitive and mental changes
• Dementia
• Gradual weight loss
• Appears in adulthood
• Risk of early/serious illness if inherited from father
• Autosomal dominant inheritance

• CAG repeat expansion of exon 1 in HTT gene; polyglutamine tract
– normally: < 26
– Intermediate 27-35
– w/ reduced penetrance 36-39
– Full penetrance > 40
– Juvenile > 60 (rare, usually from father)

Answer 21

Myotonic dystrophy –> Maternal transmission!
• Atrophy (dystrophy): face, distal in upper and lower extremities
• Myotonia: inability of the muscles to relax after contraction (EMG)
• Cataracts
• Frontal balding
• Endocrine symptoms / gastrointestinal / cardiac
• Mental retardation can occur
• Autosomal dominant inheritance

• Type 1: CGG repeats ”in ” DMPK gene:
– Normal 5-34 
- premutation 35-49
– Mild 50-150 
- classic 100- 1000; RNA disturbance
– Congenital (from mother) > 1000

• Type 2: CCTG repeat expansion in intron 1 of CNBP
– No anticipation No congenital presentation
– Extreme somatic instability, RNA disturbance

Question 22

Genomic imprinting:

• Normal development requires the complementary presence of both maternal and paternal genetic material.
• An imprinted gene will be inactivated (turned off) and therefore not be expressed during gametogenesis in individuals of one sex, whereas in the opposite sex it will be expressed.

Answer 22

Examples of genomic imprinting:

• Prader-Willi
  • neonatal hypotonia ”floppy infant”
  • failure to thrive, feeding difficulties
  • later on; excessive eating (insatiable appetite, aldri mett)
  • slight mental retardation
  • MLPA Methylation testing: > 99%
  – paternal deletion 15q11-q13

- AngelMan syndromes
• Normal at birth
• Seizures w/ specific EEG pattern? 
• microcephaly – 50% by 12 months 
• Ataxia
• Mental retardation
• MLPA Methylation testing ~ 80%
– maternal deletion of 15q11-q13

Question 23

Duchenne MD (DMD):

• CK: >10x normal , almost complete lack of dystrophin in biopsy.
• Clinical onset at the age of 2-3 with progressive proximal muscle weakness (lower extremities).
• Historically premature death around the age of 20
– (cardiomyopathy, respiratory problems)

• Hypertrophy of calves (tykke legger)
• Gowers sign, holder seg til knærne når røyser seg opp.

Answer 23

BeckerMD (BMD):

• CK: >5x normal, reduced dystrophin in biopsy
• Greater clinical variation and later onset

Question 24

Hereditary cancer
• 5-10 % of all cancers

• Germline (mostly inherited) mutations in:
• Tumor suppressor genes (common cause):
• Negative regulate cell cycle
• «Loss of function» mutations
• Two hit.
• BRCA1/2, TP53, RB1, APC

• Proto-oncogenes (rare cause):

• Positive regulate cell cycle
• «Gain of function » mutations
• One hit.
• RET (MEN2B).

But cancer is common – > 40% of aging population

Answer 24

Knudson 2 hit hypothesis:

Both copies of the gene must be knocked out in order to cause malignancy.

Sporadic cancer: 2 acquired mutations

Hereditary cancer: 1 acquired mutation, 1 inherited

Familial cancer is caused by?
- A germline mutation plus a somatic mutation in affected tissue.
Heterozygosity of mutation (inherited via germline) in a tumour- suppressor gene.
A second hit on the other allele (in somatic tissue) is required for tumour formation.

Question 25

Proto-oncogenes

• Positively regulate the cell cycle
• Activating mutations

«One hit». «Gain of function».

Answer 25

Tumor-suppressor genes (TSGs)

• Negatively regulate the cell cycle
• Inactivating mutations

Question 26

HEREDITARY CANCER

• Germline mutation (1. hit)
• Somatic mutations (2. hit)
• Bilateral
• Early onset
• Multiple tumors
• Combination of genetically «related» cancers on the same side of the family
• Rare tumors

Answer 26

SPORADIC CANCER

• Somatic mutation (1. hit)
• Somatic mutations (2. hit)
• Unilateral
• Later onset
• Single tumors

Question 27

Diagnostic, incident testing; by treating clinicians

• Female breast cancer < 60 years of age
• Male breast cancer regardless of age
• Ovarian cancer regardless of age

Answer 27

Healthy persons with 1st degree relative (or 2nd degree with a male between) with one of the following:

• Female breast cancer < 50
• Male breast cancer (any age)
• Female breast cancer, triple negative < 60
(østrogen, progersteron, herceptin-2)
• Female breast cancer, bilateral < 60
• Ovarian cancer
• 2 first degree relatives mean age < 55 (breast / prostate) • 3 first degree relatives with breast cancer (any age)

Question 28

Lynch syndrome (HNPCC) hereditary non-polyposis colorectal cancer)

• Autosomal dominant, reduced penetrance (=risk).
• Increased cancer risk;
- Colorectal cancer (CRC) – up to 70 %
- Endometrial cancer – up to 50%

• MMR (mismatch repair): MLH1, MSH2, MSH6, PMS2, EPCAM +
Better prognosis of cancer disease, often cancer surviviors and more than one cancer diagnosis.

Answer 28

Surveillance:

• Colonoscopy every second year from age 25.
• Gynecological examination (endometrialbiopsy if indicated) yearly from age 30.

Risk reducing surgery?
• Prostate examination + PSA from age 40
• Urine stix from age 30 (screening for hematuria)

Question 29

Familial Atypical Multiple Mole Melanoma Syndrome

• Autosomal dominant, reduced penetrance (=risk).
• Increased cancer risk;
• Malignant melanoma (MM)
–30-90%
• Pancreatic cancer
–15-25%
• CDKN2A, CDK4 + more
• Risk increases with UV light exposure, and depend
on place of residence and family history.

Answer 29

Surveillance:

• Dermatologist examination yearly from 18-20, or earlier based on age of onset MM in the family.
• Pancreatic surveillance NOT effective/available.
• SUN protection!

Question 30

FAP (familial adenomatous polyposis)

• > 100, often thousands adenomatous colon polyps at a young age.
• Autosomal dominant, 20 % de novo; APC- gene
• High risk of early onset colorectal cancer (near 100 % risk by 50 years)
• Risk of: duodenal cancer, thyroid cancer, desmoid tumors, CHRPE…. ++++

Answer 30

Prevention/screening:

• Rectoscopy / coloscopy from the age of 10
• Total colectomy (from late teens)

• Polyps also occur in the stomach and the duodenum

Question 31

Prenatal screening:

large populations, low risk, affordable
non-invasive methods.

Answer 31

Prenatal diagnosis:

selected groups, high risk, severe conditions
invasive procedures.

Question 32

Screening - various definitions:

• Sequential (stepwise) screening = provision of an initial set of tests followed by another set usually for those at LOW or intermediate risk
• Contingency screening = provision of an initial set of tests followed by another set only for those initially found to be at INTERMEDIATE risk
• “Secondary” screening = a follow-up screening test for those women who were POSITIVE by an initial primary test

Answer 32

Detection rate (sensitivity)

False positive rate (1-specificity)

Odds of being affected given a positive result (OAPR)
The ratio of affected to unaffected cases (usually 1:n)
TP:FP (true positive / fake positive)

Question 33

Invasive procedures

• Amniocentesis (AC): amniotic fluid withdrawal
- from 15th week of pregnancy

• Chorionic villus sampling (CVS): placental
biopsy
- From 10th week of pregnancy

Other invasive methods
• Cordocentesis: blood sample from the umbilical cord
– 18th week of pregnancy

• Foetoscopy
• Fetal skin-biopsy, liver biopsy

Answer 33

Amniocentesis:
• Aseptic
• Fetus and placenta located using ultrasound
• Transabdominal
• 15 ml amniotic fluid
• Cytogenetic investigations
– RAD: rapid analysis of selected chromosomes
(13, 18, 21): 2-3 days (QF-PCR)
– CMA (chromosomal microarray): 10 days
• Procedure rel. risk of miscarriage < 0.5%

CVS:
• Aseptic
• Fetus and placenta located using ultrasound
• Transabdominal (rarely transcervical)
• 5-10 mg placental tissue
• Karyotype: direct (– under 24 hrs)
• Long-term culture: 1-2 weeks
• Procedure related risk of miscarriage: 0,2-0,7 % (1. trimester)
• Fetomaternal bleeding (anti-D prophylaxis)
• Suitability for DNA analysis

Question 34

Preimplantation Genetic Diagnosis (PGD)
• when one or both genetic parents has a known genetic abnormality and testing is performed on an embryo to determine if it also carries a genetic abnormality.
• look for spesific diseases

Testing:
• technique used to identify genetic defects in embryos created through in vitro fertilization (IVF) before pregnancy.
• the procedure carries a risk of wrong diagnosis (technical, mosaicism).

Answer 34

Preimplantation genetic diagnosis (PGD)

is NOT allowed performed in Norway.

Question 35

Aneuploidy screening tests / Non-Invasive Prenatal Testing = NIPT

= Risk asessement
• Ultrasound scan (early gestational age: Nuchal translucency. 
NT > 2,8 --> økt risiko for Downs)
• Maternal biochemical serum markers 
(double test: B-hCG og PAPP-A). Combined tests (CUB) w/ USS.
• Analysis of cell free DNA (fetal DNA
circulating in maternal blood)
• Cell-based NIPT

 False positives
• Few but important!
- Confined Placental Mosaicism
- Vanishing twin (when used in early pregn) 
- Maternal chromosomal abberations
- Maternal cancer

Answer 35

First trimester combined test (CUB)

• The hCG and PAPP-A levels, along with the maternal age and size of the nuchal translucancy gives a risk assessement of the common aneuploidies
(trisomy 21, 18 and 13).
• Performed in week 11/12-13+6 of pregnancy (CRL 45-84 mm)

CUB- performance Trisomi 21
• Detection rate : 82-87 %

NIPT- performance T21, T13 and T18
• PPV T21: 98% !
• PPV T18: 92%
• PPV T13: 69%
• Sensitivity : 97,3-99,5%
• Spesificity : 99,7-99,9% very few false negatives!

USS: Ultrasound scan
• Structural malformations (isolated/multiple) 
• ”Soft signs” mainly indicative of trisomy (?)  
Nuchal Translucency measurements  
Nasal Bone presence
Nasal Bone Length
Short Femoral Length  
Echogenic Intracardiac Focus   
Hydronephrosis
SUA..etc

Question 36

Cell free (cf) fetal DNA in maternal plasma

• First discovered by Dennis Lo et al (1997)
• Circulating free DNA is either maternal or fetal
• Originates from placenta (apoptotic trophoblast)
• Detectable from 4th week
• Generally 10-20% of total cfDNA present in a sample (variable)
• Amount approximately constant in the late first trimester through the second trimester
• % increases with gestation

Answer 36

Differences in testing policies for laboratories offering cf-DNA screening

• Minimum fetal fraction (e.g. >4%, >2.8%, or not measured)
• Gestational age (>9 weeks, >10 weeks)
• Testing for DZ twins (including or excluding sex chromosome abnormalities)
• Surrogate mother
• Inclusion of diandric triploidy, molar pregnancy
• Inclusion of maternal age in the algorithm
• Reporting differences; use of a “patient specific risk score”, “positive” “aneuploidy detected,” “aneuploidy suspected (borderline value),” or “personalized risk’’.
• NIPT screening also for other disorders ! (some microdeletion syndromes, other rare autosomal trisomies, partial imbalances)

Cell-free DNA screening tests for microdeletions have NOT been validated clinically and are
NOT recommended at this time “ ACOG 2016

Question 37

Quadrupel screen: Utenom PAPPA-A

• Beta-hCG (høy ved Downs)
• AFP (lav ved Downs, høy ved Spina bifida)
• Estriol (lav ved Downs)
• Inhibin-A (høy ved Downs)

Second trimester Quadruple test = AFP, hCG, uE3, INH-A at 15-20 weeks

Triple test (rarely used in Norway)
• Performed in week 15-17 of pregnancy
• Blood test :AFP, hCG and estriol (Ikke Inhibin A)

Answer 37

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