Genetics - Imperial College Union · Insertion- The adding of an extra DNA nucleotide that results in a shifting of ... Chromosomal abnormalities. 4. ... Delayed fertilisation after

Collection of Genetics Definitions

Alleles: Alternate forms of gene or DNA sequence at the same chromosome location (locus)

Deletion- The deletion of a single nucleotide that results in the formation of a malfunctioning protein.

Dominant anticipation- This is where in conditions such as Huntingdon's and Myotonic Dystrophy the age of onset decreases, the severity increases and can be due to an expansible, unstable triplet repeat.

Duchenne Muscular Dystrophy- X-linked disorder which results in muscular degeneration. Cases have a shorter survival time then Becker Muscular Dystrophy who's symptoms appear later in childhood.

Epigenetic- is the study of heritable changes in gene expression or cellular phenotype caused by mechanisms other than changes in the underlying DNA sequence

Epistasis – interaction between disease gene mutations and other modifier genes can affect phenotype

Homologous chromosomes are a matching (but non-identical) pair, one inherited from each parent

Imprinting- is a genetic phenomenon by which certain genes are expressed in a parent-of-origin-specific manner.

Insertion- The adding of an extra DNA nucleotide that results in a shifting of amino acids and thus potentially the formation of a malfunctioning protein.

Missense- A change in the DNA base code that results in the coding of an abnormal amino acid (shouldn't be there)

Mosaicism- Denotes the presence of two populations of cells with different genotypes in one individual who has developed from a single fertilized egg.

Mutation- Heritable change in the DNA sequence of a gene.

Nondisjunction- This is a failure of the chromosomes to separate correctly in meiosis/mitosis I or II.

Nonsense- A change in the DNA base code that results in a termination of a sequence and the shortening of a protein.

Osteogenesis Imperfecta- Autosomal dominant disorder

Penetrance – frequency with which symptoms are present in an individual who inherits a disease-causing mutation.

Phenocopy - disease with the same phenotype as a genetic disease, but non-genetic

Polymorphism- Presence of more than one phenotype within a population. >1%

Teratogen- Agent that interferes with normal foetal or foetal development.

Uniparental isodisomy- The process whereby non-disjunction in meiosis II causes a gamete to form with 3 chromosomes. This is subsequently restored to normal chromosome number by the loss of chromosome that was contributed singularly.

Variable expressivity – degree of severity in an individual who inherits a disease-causing mutation

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Stuart Taylor

Genetics23 November 201100:00

Stuart's Genetics Page 1

Congenital abnormalities- Abnormalities present at birth may be due to genetics or infection

Malformation – primary structural defect e.g. atrial septal defects, cleft lip. Usually involves a single organ showing multifactorial inheritance.

1.

Disruption – secondary abnormal structure of an organ or tissue e.g. amniotic band causing digital amputation. Caused by ischaemia, infection, trauma. Not genetic, but genetic factors can predispose.

2.

Deformation –abnormal mechanical force distorting a structure e.g. club foot, hip dislocation. Occurs late in pregnancy and has a good prognosis as the organ is normal in structure.

3.

Syndrome –consistent pattern of abnormalities with a specific underlying cause, e.g. Down syndrome. Chromosomal abnormalities.

4.

Sequence- multiple abnormalities initiated by primary factor e.g. reduced amniotic fluid leads to Potter's sequence. Could have genetic component as initial factor.

5.

Dysplasia –abnormal organisation of cells into tissue e.g. thanatophoric dysplasia

6.

Association –non-random occurrence of abnormalities not explained by syndrome. Cause is typically unknown. e.g. VATER association.

7.

Dysmorphism- an unusual or abnormal physical feature (sometimes as part of a genetic condition) e.g. hypertelorism

8.

Chromosomal Abnormalities

Numerical – aneuploidy, loss or gain

Rings- Chromosomes break and sticky ends join together forming a ring whilst fragments are lost.

Structural – translocations, deletions, insertions, inversions, rings

Mosaicism – Denotes the presence of two populations of cells with different genotypes in one individual who has developed from a single fertilized egg.

60% of early spontaneous miscarriages•

4-5% of still births•

7.5% of all conceptions, 0.6% of live births•

Chromosome abnormalities are present in:

Oligohydramnios – reduced volume of amniotic fluid due to failure to produce urineClassically, due to bilateral renal agenesis (Potter, 1946)

Caused by single gene defect, High recurrence risk for siblings/ offspring, 1:60,000 incidence, FGFR3 mutations, Short flat bones, small thorax, large head

Vertebral, Anal, Tracheo-Esophageal, Renal

Aneuploidy

Aneuploidy- Abnormality in the chromosome number causes by the loss or gain of one or two chromosomes.

Three types of aneuploidy

Monosomy- Loss of a chromosome, nearly always fatal1.Trisomy- Gain of one chromosome- can survive2.Tetrasomy- Gain of two chromosomes- still viable fetus3.

Explanation as to why this is so:

Loss of a chromosome gives a reduction of 50% of all fully expressed gene products whereas gain of one chromosome gives an increase of 33% of all fully expressed gene products.

Partial aneuploidy results in translocations

Page 39 Kumar and Clarks Clinical Medicine

Mosaicism- Occasionally, non-disjunction can occur during mitosis shortly after two gametes have fused. It will then result in the formation of two cell lines, each with a different chromosome complement. This occurs more often with the sex chromosome, and results in a ‘mosaic’ individual.

hypertelorism abnormal distance between two organs

Down's Syndrome

95% of all cases of Down's•90% maternal origin of extra chromosome•75% of non disjunction in meiosis I and 25% chance in meiosis II.•

Trisomy 21/ Meiotic Non-Disjunction:

4% of all cases•Occurs due to a translocation of long arms of acrocentric chromosomes (13,14,15,21,22)

•

Structural:

1% of all cases•Less severely affected•Caused by mitotic non-disjunction•

1 cell zygote

Normal disomy trisomy monosomy

1st mitotic division

2nd mitotic division non-disjunction

4 cell zygote

(33% mosaicism)

Mosaicism/ Mitotic Non-disjunction-

Nondisjunction- This is a failure of the chromosomes to separate correctly in meiosis I or II.

P is short arm

q is long arm

1 in 50=50 congenital abnormalities40% are due to genetics.

Acrocentric refers to satellite short arms containing genes for ribosomal RNA?

Self test answers- DNA and chromosomes

Robertsonian Translocation Breakage at the centromere of 2 acrocentric chromosomes with fusion of their long arms to form one new derivative chromosome.

Causes of non-disjunction:

Ageing effects on primary oocyte

Age-related decrease in immunological competence (allows survival of trisomic embryos)

Delayed fertilisation after ovulation occurs

Radiation

Each female has an inactive copy of the X-chromosome

Balanced translocation= Reciprocal

G1 Mrs. Jones30 November 201115:13


Name of Condition

Type of Condition Causes Chromosome Location

Live Birth Rate

Symptoms

Down's Syndrome

Autosomal Abnormality

Trisomy

Acrocentric translocation

Mitotic mosaicism

21 (47) 1 in 650-700

New-born period - severe hypotonia, sleepy, excess nuchal skin•

Craniofacial - macroglossia, small ears, epicanthic folds, upward sloping palpebral fissures, Brushfield spots

•

Limbs – single palmar crease, wide gap between first and second toes•

Cardiac - A and V septal defects•

Other - short stature, duodenal atresia•Relatively advanced social skills•

Patau's Syndrome


Trisomy 13 (47) 1 in 5000 Low-set ears, cleft lip and palate, polydactyly, microophthalmia, learning difficulties

•

Edwards Syndrome


Trisomy 18 (47) 1 in 3000 Low-set ears, micrognathia, rocker-bottom feet, learning difficulties•

Fragile X Syndrome

Sex Chromosome Abnormality

46, XX fra (X)46, XY fra (X)

1 in 2000 Most common inherited cause of learning difficulties predominantly in male.•Macro-orchidism•

Turners' Syndrome


Monosomy X (45 XO) 1 in 3000 Generalised oedema and swelling in neck region can be detected in 2nd trimester

•

Can look normal at birth or have puffy extremities and intra-uterine oedema•

Low posterior hairline, short 4th metacarpals, webbed neck, aorta defect in 15% of cases

•

Normal intelligence•TREATMENT- Oestrogen replacement for secondary sexual characteristics and prevention of osteoporosis

•

Polysomy X Sex Chromosome Abnormality

Polysomy X (47 XXX) 1 in 1000 10-20 point decrease in IQ•

No physical abnormalities•

95% have extra maternal X arising in meiosis I•

Normal fertility•

48,XXXX and 49,XXXXX karyotypes show mental retardation, infertility and amenorrhoea.

•

Klinefelter’s syndrome


Polysomy X(47XXY) 1 in 1000 clumsiness, verbal learning disability 10-20 pts•

taller than average (long lower limbs)•

30% - moderately severe gynaecomastia•

all infertile•

increased risk of leg ulcers, osteoporosis and breast carcinoma in adult life•

Double Y Syndrome


Trisomy 47 XYY 1 in 800 Tall, fertile, minor mental and psychiatric illness, high incidence in tall criminals.•

XX Males Homozygous Male

Translocation of male encoding SRY chromosome from Y chromosome to an X chromosome.

Phenotypically male, testes develop, but sterile because some genes on Y chromosome needed for spermatogenesis.

•

XY Females Heterozygous Female

Mutations or deletions of SRYgene

Infertile•

Di George Syndrome

Recurrent Microdeletion Disorder

Hemizygous Microdeletion of 1.5-3 Mb of 22q11

1 in 4000 Velocardiofacial (VCFS)/Sedlackova syndrome•

Congenital Heart Disease•

Palatal abnormalities•

Thymic/Parathyroid Hypoplasia•

Characteristic Facies•

Learning Difficulties•

Variable symptoms•

Commonest microdeletion disorder•

Cri du Chat Recurrent Microdeletion Disorder

Deletion varies from 5p15.2 to whole of 5p

1 in 50,000

Microcephaly, Hypertelorism, Micrognathia, Epicanthal folds, Low-set ears, Hypotonia

•

Characteristic Facies:•

Severe psychomotor and mental retardation•

Characteristic cat-like cry in newborns•


Types of Genetic Disorders

Monogenic disorders

• familial

• mode of inheritance

• common and rare

Eg. Huntington disease

Cystic fibrosis

Haemophilia

Complex disorders

• sporadic/familial

• environment factors

• common disorders

Eg. Type 2 diabetes

Obesity

Parkinson’s disease

Mutations vs. polymorphisms

Mutation- Heritable change in the DNA sequence of a gene.

Polymorphism- Presence of more than one phenotype within a population.

Types of mutation

Missense- A change in the DNA base code that results in the coding of an abnormal amino acid (shouldn't be there)

Nonsense- A change in the DNA base code that results in a termination of a sequence and the shortening of a protein.

Both of these are known as point mutations

Insertion- The adding of an extra DNA nucleotide that results in a shifting of amino acids and thus potentially the formation of a malfunctioning protein.

Deletion- The deletion of a single nucleotide that results in the formation of a malfunctioning protein.

Both can be frame shift mutations

Why take a genetic family history?

Identify diseases present and their inheritance•Identify relatives at risk.•Help aid a diagnosis•To help manage the condition of those affected.•

Pedigree Diagram symbols

male

female

sex unknown

affected

unaffected

carriers

deceased

consanguineous marriage

twins

Mendelian Inheritance Patterns

50% of children affected can be transmitted by either sex•Vertical Transmission•

In Huntingdon's the faulty gene on chromosome 4 codes for toxic form of huntingtin protein which clumps in basal ganglia and causes symptoms.

○

Caused by expansion of CAG- Glutamine repeat- Polyglutamine○

Huntingdon's disease is an example

Autosomal Dominant:

Dominant anticipation- This is where in conditions such as Huntingdon's and Myotonic Dystrophy the age of onset decreases, the severity increases and can be due to an expansible, unstable triplet repeat.

Usually no family history•Often presents without parents having the condition•25% chance of affected child if both parents are carriers.•

Chronic, life threatening condition.○

Thick mucus in lungs causes persistent infections○

Blockage of pancreatic duct into the duodenum results in digestive problems○

Faulty gene on chromosome 7(7q31) produces mutated copy of CFTR (CF transmembrane conductance regulator) which affects chloride transport in wet epithelia thus resulting in sticky mucus.

○

3bp deletion of Delta F508 is the major cause of over 70% of cystic fibrosis in Caucasians.

○

Cystic fibrosis is an example:

Caused by same gene.○

This is where the Vas Deferens (tube from epididymis to urethra) fail to form.○

Congenital Absence of the Vas Deferens (CAVD):

Autosomal Recessive:

Can be no affected parents•50% males affected•50% females carriers•

Condition causes people to bleed more easily and for longer.○

Blood clotting disorder○

Treatment by injection of clotting factor ○

Two types A and B○

Haemophilia is an example:

X linked:

Haemophilia

Type A is caused by faulty F8 gene on the X chromosome and thus clotting factor VIII•Type B is rarer and results from F9 gene and clotting factor IX.•Symptoms in both are similar. •

Mendelian Complications and definitions

Penetrance – frequency with which symptoms are present in an individual who inherits a disease-causing mutation.

Variable expressivity – degree of severity in an individual who inherits a disease-causing mutation.

Phenocopy - disease with the same phenotype as a genetic disease, but non-genetic.

Epistasis – interaction between disease gene mutations and other modifier genes can affect phenotype.

Mechanisms of Genetic Disease

Dominant Conditions- These usually result from a toxic protein being able to mask the effects of the normal protein that is present and thus cause the disease.(HD)

Recessive Conditions- Predominantly happen when a protein is not present, thus two copies of the faulty gene are required so that normal proteins is not present.(Haemophilia and CF)

Co-dominant Conditions- Effects of both mutated and normal genes apparent in people with both, eg. sickle cell trait

Treatments

Dominant conditions – need to counter effects of, or neutralise toxic protein, or ‘switch off’ mutant gene

•

Recessive conditions – need to restore activity of missing protein, by replacing genes, protein or affected tissues

•All others are characteristics of an ideal marker. A possible pneumonic is 'HiRES': Highly polymorphic, Randomly distributed, Easily assayable, Stable.

Incest is best

G2 Transmission02 January 201212:34


Learning Objectives

Understand the imprinting disorders Prader-Willi and Angelman Syndrome

Understand Uniparental Isodisomy

Understand the mitochondrial disorders MELAS and LHON

Understand Phenylketonuria and MCAD.

Understand the imprinting disorders Prader-Willi and Angelman Syndrome

Parental origins of chromosome are important.•

Maternal- Ovarian Teratoma○

Paternal- Hydatidiform Mole○

46 XX where genome is only from one parent•

The genome carries an imprint of its parental origin•Imprinting is a reversible epigenetic effect•DNA methylation is the mechanism•

Prader-Willi/Angelman Syndrome

Same chromosome region involved on Chr15•

Paternal= Prader-Willi○

Maternal- Angelman ○

Match the start of the word Paternal and Prader-Willi.○

Also remember its messed up you'd think Paternal would be Angelman○

Result from loss of function of one of the two parental chromosomes•

Prader-Willi Syndrome:

Muscle hypotonia•Hyperphagia•Obesity/Diabetes•Mental retardation•

1:10,000 to 1:25,000 live births•Hyperphagia is managed by diet restriction•Exercise to increase muscle mass•Growth Hormone treatment for short stature•Hormone replacement at puberty.•

Genetic Mechanism of PWS

70% deletion on paternal chromosome○

25% inheritance of two maternal copies by uniparental isodisomy○

5% due to translocation, point mutation.○

Lack of a functioning paternal copy of PWS critical region on 15q11-q13•

Angelman Syndrome

Severe developmental delay•Poor or absent speech•Gait ataxia•"Happy demeanour"•Microcephaly•Seizures•

Prevalence is 1:10,000•Treatment is with anti-convulsants, physiotherapy,•communication therapy.Normal lifespan is observed•

Short stature•Small hands and feet•Delayed/incomplete puberty•Infertile•

Understand Uniparental Isodisomy

Non-disjunction in Meiosis II•Fertilisation of normal monosomic gamete•Loss of chromosome from parent contributing the single chromosome.•

Uniparental isodisomy- The process whereby non-disjunction in meiosis II causes a gamete to form with 3 chromosomes. This is subsequently restored to normal chromosome number by the loss of chromosome that was contributed singularly.

Understand the mitochondrial disorders MELAS and LHON

Transmission is exclusively through females.•Disease can be variable because of heteroplasmy- Mixture of more than one type of organelle genome.

•

Mitochondrial Genome

~16kb•37 genes•13 for respiratory chain complexes- Unlucky if your ATP gets messed with.•22 for tRNA- Think of the t's Twenty Two•2 for rRNA- R is for Rubbish so it hasn’t got many.•2-10 copies per mitochondria.•

MELAS

Mitochondrial myopathy, Encephalopathy, Lactic Acidosis and Stroke.•

Muscle weakness○

Episodic seizures and headache○

Hemiparesis- Weakness on one side of body.○

Vomiting○

Dementia○

Progressive neurodegenerative disorders•

Estimated prevalence of 1:13,000•Diagnosis by muscle biopsy•

Understand the inborn errors of metabolism disorders Phenylketonuria and MCAD.

"One gene- one enzyme" concept•More than 200 disease known•Mostly autosomal recessive or X-linked•A few are dominant (rate-limiting step or part of a multimeric complex)•The defective proteins are usually enzymes•

UK Newborn Screening Programm

Phenylketonuria1.Congenital Hypothyrodism (cretinism)2.MCAD3.Cystic Fibrosis4.Sickle Cell anaemia 5.

Phenylketonuria

Severe mental retardation and convulsions•Blond hair/ blue eyes; eczema•Phenylalanine hydroxylase deficiency•Phenylalanine accumulates and is converted to phenylpyruvic acid which is excreted in urine.

•

Tyrosine deficiency- reduced melanin.•

G3 Imprinting, Mitochondrial + Metabolic Disorders04 April 201210:52


Vomiting○

Dementia○

Estimated prevalence of 1:13,000•Diagnosis by muscle biopsy•

Single point mutations in several genes○

MTTL1- Phe instead of Leu○

MTND1, MTND5- NADH dehydrogenase complex ○

Genetics•

LHON

Leber's Hereditary Optic Neuropathy•Much more common in males- reason unknown.•Average age is mid-twenties to mid-thirties•Age range is wide though (6-62 years)•Bilateral, painless, loss of central vision and optic atrophy•Most will become blind•

Estimated prevalence of 1:50,000•Diagnosis on the basis of opthalmological findings and a blood test for mtDNA mutations.

•

MTND (1,4,5,6) and MTCYB○

NADH dehydrogenase sub units 1,4,5,6○

Cytochrome B.○

>90% of mutations are in•

Blond hair/ blue eyes; eczema•Phenylalanine hydroxylase deficiency•Phenylalanine accumulates and is converted to phenylpyruvic acid which is excreted in urine.

•

Tyrosine deficiency- reduced melanin.•

ACETOACETIC ACID

C02 + H20

THYROXINEDIET

PHENYLALANINE TYROSINE DOPA

MELANINPHENYLPYRUVICACID IN URINE

1

1 - PKU

2

2 - THYROXINEDEFICIENCY

3

3 - ALBINISM

HOMOGENTISICACID IN URINE

4

4 - ALKAPTONURIA

Screening for elevated levels of Phe and treatment involves removing it from the diet.

•

Aspartame contain Phe.•

MCAD Deficiency

Medium Chain Acyl Co-Enzyme A Dehydrogenase Deficiency•Commonest disorder of fatty-acid oxidation.•Episodic hypoketotic hypoglycaemia.•Commonly present after 3 months.•Can present as coma, metabolic acidosis, encephalopathy•Sudden death can occur with a 25% mortality rate in undiagnosed cases•Frequency of 1:8000- 1:15000•Avoidance of fasting (<12 hrs in adults) to prevent fatty acid oxidation.•


Learning Objectives

Understand why genetic changes cause cancer and describe the 2 main classes of cancer gene

Understand the contribution of chromosome rearrangements to the formation of gene fusions and their contribution to oncogenesis.

Explain the difference between somatic and germline mutations

Discuss how inherited mutations in BRCA1 and BRCA2 genes influence risk of breast and ovarian cancer

Outline how defects in cell division or DNA repair influence risk of colorectal cancer

Explain, using an example, how chromosome translocations are used to quantify residual disease in some leukaemias.

Explain with examples what is meant by a “pharmacogenomic marker”

Understand why genetic changes cause cancer and describe the 2 main classes of cancer gene

Cancer is driven by accumulation of genetic changes that lead to altered levels of transcription or aberrant gene transcripts.

•

The resulting protein changes activate signal transduction pathways that confer a selective advantage to the cell.

•

Commonly affect pathways include those that control the cell cycle, proliferation. apoptosis and adhesion.

•

The 2 main classes of cancer gene are Oncogenes and Tumour Suppressors.•Proto-oncogenes promote growth and proliferation via growth factors, transcription factors and tyrosine kinases EG.

•

Tumour suppressor genes- regulate cell division, are DNA damage checkpoints (damage= no division), Apoptosis.

•

Most tumour suppressor genes require damage to both alleles for tumorigenic effect.•Hit 1 is often a mutation and reduces transcript/protein level but is insufficient to cause phenotypic effect. Inactivation of the second allele Hit 2 by a deletion causes total loss of transcription for malignant phenotype to be conferred. "2 hit model"

•

A few TS genes require damage to only one. The single hit reduces transcription and in level of protein product which for these genes is sufficient enough to have a biological effect via haploinsufficiency- expression of only one normal allele is not enough

•

Loss of heterozygosity (LOH)

This is a historical method of assessing the stability of cancer and looking for TS genes.•It is another term for deletion.•Unmasking of the mutated copy of a TS gene.•

LOH- Describes a region of apparent homozygosity, probably a deletion, in cancer tissue that may mark the location of a TS gene.

•

Chromosome 3

Mat

ern

al

Pate

rnal

Allele A

Allele B

Mat

Pat

genes

Genes

Mutated TS

Deleted

DEL

ETIO

N (

cau

ses

LOH

)

Hit 1

Understand the contribution of chromosome rearrangements to the formation of gene fusions and their contribution to oncogenesis.

Gene copy number change is brought about by duplications or deletions.•Gene fusion is brought about by inversion and translocation•Gene fusions occur at the DNA junction of chromosome rearrangements (translocation or inversion)

•

Cytogenetic changes= Global, microscopy based low res= Translocation, Deletion + Duplication

•

Change in amino acids can cause faulty protein.•Molecular changes= Targeted, biochem based, high res.•

Explain the difference between somatic and germline mutations

99% of cancers are sporadic (non-inherited) caused entirely by acquired changes in the somatic tissue.

•

1% have an inherited (germline) component which are initiated by the inheritance from one parent (or occasionally both) of a mutation in "germline" tissue, usually in a tumour suppressor gene (hit 1).

•

Inherited does not necessarily mean you have a 100% chance of getting cancer. The germline inheritance causes the 1st hit but in most cases you still need a 2nd somatic hit to initiate a cancer malignancy.

•

Discuss how inherited mutations in BRCA1 and BRCA2 genes influence risk of breast and ovarian cancer

2-4%/10? of breast cancer cases are caused by germ-line mutation of BRCA1 or BRCA2.

•

60% risk of developing breast cancer by ago of 90 and earlier age of onset.•Also convers increased risk of ovarian cancer.•BRCA2 predisposed men to breast cancer.•

Truncated non-functioning protein resulting from point mutations, several base pair deletions and whole exon deletions and amplifications.

•

BRCA1 and 2 are DNA repair genes, specifically of a process called homologous recombination. Mistakes or damage go uncorrected.

•

Outline how defects in cell division or DNA repair influence risk of colorectal cancer

Inheritance of one mutated allele of a TS gene.•

Characterized by the growth of 1000s of intestinal polyps, one or more of which is likely to become cancerous.

○

>1% of all cancers.○

Mutation of APC (adenomatous polyposis coli) gene.○

APC controls cell division.○

Virtually 100% lifetime risk of cancer.○

FAP- Familial adenomatous polyposis1.

3% of all cases○

Mutation of MLH1 or MSH2 (DNA repair genes)

Most common inherited from:○

Lifetime risk of cancer 80%○

Hereditary non-polyposis colorectal cancer (HNPCC or Lynch Syndrome)2.


Our knowledge of contribution of gross chromosomal rearrangements to cancer pathogenesis comes from cytogenetic investigations of malignant tissue.

•Explain with examples what is meant by a “pharmacogenomic marker”

Pharmacogenomics is an emerging branch of pharmacology which deals with •

G4 Cancer04 April 201212:42



Our knowledge of contribution of gross chromosomal rearrangements to cancer pathogenesis comes from cytogenetic investigations of malignant tissue.

•

Generally leukaemia genomes are more stable than those of solid tumours-therefore easier to pinpoint pathogenetic changes driving disease.

i.

Relative ease of performing cytogenetics on haemtopoeic circulating cells.ii.

This knowledge is most extensive in haematological malignancies (leukaemia and lymphomas) for 2 main reasons:

•

Chronic Myeloid Leukaemia

A clonal myeloproliferative disorder of the pluripotent haematopoeic stem cell leading to an overproduction of mature granulocytes.

•

1-2 cases per 100,000•15% of all adult leukaemias.•Triphasic- Indolent chronic phase, accelerated and terminal acute phase.•Consistent pathogenomic marker is a translocation between chromosomes 9 and 22resulting in a fusion gene called BCR-ABL1

•

Treatment is by a tyrosine kinase inhibitor.•

Acute promyelocytic leukaemia

Abnormal accumulation of immature granulocytes called promyelocytes.•Characterized by a chromosomal translocation involving the retinoic acid receptor alpha gene on chromosome 17 and the promyelocytic leukaemia gene on chromosome 15.

•

Retinoic acid is a ligand and is a form of Vitamin A that regulates DNA transcription.•

Explain with examples what is meant by a “pharmacogenomic marker”

Pharmacogenomics is an emerging branch of pharmacology which deals with the influence of genetic variation on drug response.

•

Pharmacogenetic tests are used to identify which patients are more likely to respond to cancer drugs based on the presence or absence of particular somatic mutations.

•

Examples include:

KRAS mutations= less likelihood of response○

KRAS test for colorectal cancer. •

Mutation= greater likelihood of response○

EGFR test for non small cell lung cancer•

This mutation= unlikely to respond.○

BCR-ABL1 for chronic myeloid leukaemia.•


Learning Objectives

Indications for Prenatal Diagnosis

Antenatal Screening for Aneuploidy (Down Syndrome)

Amniocentesis○

Chorionic Villus Sampling○

Fetal Blood Sampling○

Elective late karyotyping○

Prenatal Testing

Cytogenetic Techniques

Management options

Indications for Prenatal Diagnosis

High risk of Down Syndrome○

Previous aneuploid fetus○

Maternal request e.g. Age○

High risk of aneuploidy1.

Achondroplasia○

Cystic Fibrosis○

Haemoglobinopathies○

X linked disorders○

Parental balanced translocation○

Known genetic disorder2.

Structural anomaly detected in fetus on routine ultrasound screening.3.

Antenatal Screening for Aneuploidy (Down Syndrome)

Most common form of mental retardation 1 in 700 pregnancies, not inherited.•Often associated with birth defects.•

Standard trisomy 21=21 95%•Robertsonian translocations= 4%•Mosaicism= 1%•

Due to extra chromosome:•

Risk increases with woman's age•

11-13 weeks, measurement of the fold of the skin on the back of the foetal neck together with maternal age.

•

Increasing NT is indicative of Down's syndrome.•

Nuchal translucency•

99.5% of euploid fetuses have nasal bones that are present. Decrease 4 fold chance if bones are detected.

•

73% of aneuploid fetuses' nasal bones are absent. 127 fold increase in chance of aneuploidy if this is present.

•

85% sensitivity with only 1% false positive rate.•Bones absent in 8.8% of Afro-Caribbean population- test is currently of little value in T21 risk assessment.

•

Nasal bones•

Unconjugated estriol, AFP, Inhibin, Free beta hCG.•

14-21 weeks: AFP, unconjugated oestriol (uE3), and hCG together with maternal age.

○

AEG- 3 round burst - memory aide○

Triple test•

14-21 weeks: AFP, uE3, free beta hCG and inhibin- A together with maternal age.

○

Quadruple test

10-13 weeks: NT measurement with free beta hCG, PAPP-A and maternal age.

○

Combined test

Integration of measurement of NT measurement and PAPP-A in the first trimester with serum AFP, beta hCG, uE3 and inhibin A in the second.

○

Integrated test

Serum screening.•

AFP is Alpha Fetoprotein and low levels of this indicate Down's Syndrome

Prenatal Testing

Through both a chorionic villi and amniotic fluid sample one is able to determine the karyotype, perform a DNA and biochemical analysis.

•

AmniocentesisPerformed >15 weeks and is an aseptic technique with gloves and no touch.•USS guided•A 22G needle with a stylet is used and has to avoid the placenta.•The first 2ml has to be discarded, the next 15-20ml can be aspirated.•

Complications:There is a 1% pregnancy loss rate. •Rh sensitization, all Rh Negative women get Anti D within 72 hours.•

Cytogenetic AnalysisFetal cells concentrated in centrifuge (skin, pulmonary, urogenital, extra-embryonic membrane cell)

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Detection of mosaicism has to be present in >2 cultures to be significant because it could just be due to a culture artefact.

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Cells cultured in multiple cultures for 14 days able to detect mosaicism. •

Chorionic Villus Sampling11 weeks onwards.•Transabdominal or transcervical- USS guided.•Short term culture gives count in 48 hours.•Ideal for DNA analysis.•

ComplicationsRh sensitization.•Bleeding/ROM/Infection rare.•Fetal anomaly.•

Cytogenetic analysisSynctiotrophoblast/Cytotrophoblast cells already dividing- Direct culture possible in 72 H.

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Primarily fibroblasts are cultured as they are from inner cell mass and therefore more representative of fetus.

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1:500 culture failure.•1:200 mosaicism.•

Fetal Blood SamplingTransabdominal USS guided.•>18 weeks.•Aseptic conditions, primarily used for assessing fetal anaemia as CVS and amnio are better for processing karyotype.

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Transplacental into umbilical cord insertion into placenta.•Or transamniotic into Intrahepatic vein.•

Elective late karyotyping

Cytogenetic Techniques

Rapid karyotyping: •FISH- Chromosome specific, fluorescence labelled, DNA probes.•

Management options

G5 Prenatal04 April 201215:59


Termination of pregnancy.•

Support parents decision.•Offer continued USS monitoring.•

Mode of delivery○

Monitoring in labour○

Neonatal resuscitation○

Postmortem○

Postnatal care○

detailed plans need to be made for:•

Continuation of pregnancy•

Risk of recurrence•Management of future pregnancy•Implication on other family members.•

Genetic counseling•

All others are characteristics of an ideal marker. A possible pneumonic is 'HiRES': Highly polymorphic, Randomly distributed, Easily assayable, Stable.


Uses of fat

Insulation1.Storage of food and water2.Support and protection of vital organs3.Sources of hormones4.Regulator and fuel for the immune system5.

Infertility1.Miscarriage2.Death from infections3.Osteoporosis4.

Males more likely to deposit fat in organ systems rather than subcutaneously•

BMI= 77%○

Waist Circ= 77%○

There is a genetic element to obesity- heritability in children•

30%=non genetic•Low SPA (sporadic physical activity)- fidgeting (leg gyration)= 10%•

Syndromic Obesity

There are around 30 known syndromic forms of obesity (i.e. usually accompanied by mental retardation and particular dysmorphic or clinical features)

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Prader-Willi syndrome deletion of paternal chromosome 15q11-13.•Also BDNF (brain derived neurogenic factor) in WAGR syndrome•And cilia in Bardet-Beidl syndrome•

Monogenic Obesity

leptin the first obese gene.1994-Leptin is produced by fat cells goes across the blood brain barrier to your hypothalamus and tell the brain how much fat there is in the body.

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Leptin deficiency- Hunger, obesity, no puberty, poor growth low thyroid and immune problems.

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Most fat people have lots of leptin because they become insensitive to it.•There is also leptin receptor deficiency which causes obesity.•The leptin-melanocortin "adipostat" gene regulates body fat levels.•Pro-opiomelanocortin gene deficiency causes hunger, obesity, red hair and low adrenal activity.

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PC1 is a recessive obesity•MC4R is the most common form (is dominant) and causes 2-6% of obesities. All affect appetite regulation.

•

Environment

Genes

Common obesity Single gene(dominant)

MC4-R(up to 6%)

Single gene(recessive)

LEPLEPRPOMCPC-1

Monogenic Forms of Obesity

Lack of fat

Common obesity

Common disease, common variant•Up to 1M SNPs typed in one experiment (single nucleotide polymorphism)•Over 38 genes can cause obesity polygenic condition?•SNPs explain only a small proportion of common obesity risk.•COMMON VARIANTS OF SUBTLE EFFECT NOT RELEVANT TO SEVERE FORMS OF OBESITY

•

Rare variants○

Epigenetics○

Genomic structural variation○

Missing heritability of complex diseases may be due to:•

Patients with syndromic obesity have more GSV's i.e. 3fold more deletions.•Examples include deletions, extra copies of certain regions (duplications or amplifications), inversions or translocations.

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Deletion at 16p11.2 estimated to be 546kb-700kb.•Phenotype is moderately severe mental retardation, poor speech, hypertelorism, large protruding ears.

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Highly penetrant if you have this it is very very very likely you will get fat.•

G6 Obesity06 April 201210:58


Outline

‘Next generation’ DNA sequencing

Finding the causes of monogenic disease

Pharmacogenetics

Risk of common, complex diseases

Advances in genomic medicine

Direct-to-consumer genetic testing

Ethical issues

Future perspectives

Personalised healthcare

Preimplantation genetic diagnosis (PGD)

Uses and limitations

Ethical issues

Embryo testing

‘Next generation’ DNA sequencing

Cost of sequencing entire genome has decreased massively to around $10,000.•Speed an efficiency has increased•

Finding the causes of monogenic disease

Next-generation sequencing already used to identify novel gene mutations in monogenic disease.

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Miller syndrome- multiple malformation syndrome, characteristic facial features, cupped ears and absent toes results from DHODH gene mutations.

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Schinzel-Giedion syndrome- severe mental retardation, life limiting, characteristic facies. Caused by SETBP1 gene.

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Both genes identified by sequencing exomes of just 4 individuals.○

Whole exome sequencing (sequencing of all the exons)•

Pharmacogenetics

Right drug, right person right dose.•Variant in TPMT gene effect metabolism of 6-mercaptopurine.•Type 1 diabetes vs Mature Onset Diabetes of the Young. (insulin vs metformin which is a sulphonylurea).

•

Risk of common, complex diseases

Many genetic variants that affect risk of common, complex traits and diseases identified through:

•

E.g. Type 2 diabetes, schizophrenia, obesity, blood pressure.○

Genome Wide Association Studies (GWAS)•

Advances in genomic medicine

Direct-to-consumer genetic testing

Direct to consumer (DTC) genetic tests

For monogenic disease

Can provide carrier status information

Can detect rare serious conditions in newborn

Essential that service includes genetic counselling

For complex disease

Often have limited clinical utility

May cause undue alarm

May offer false reassurance

Data privacy concerns

Personalised healthcare

Monogenic diseases

To determine carrier status of recessive diseases such as Tay Sachs and cystic fibrosis.•To determine risk of later onset disease e.g. hereditary breast cancer•To determine presence of serious, treatable disease in newborn- MCAD etc.•

Complex/ common diseasesType 2 diabetes- caused by complex interaction of genetic and non-genetic factors. Sibling relative risk is ~3 in Europe.

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38 confirmed T2D genes so far which explain just ~10% of estimated genetic contribution.•TCF7L2 gene.•

Ethical issues

Commercial genetic testing for disease risk based on incomplete information.•Right not to know (particularly children)•Protection of data i.e. right to genetic privacy•Equality of access to genetic information.•

Future perspectivesLimited clinical utility at present but may become more in future as we understand the genome more and more.

•

Preimplantation genetic diagnosis (PGD)

PGD- Is a genetic test carried out on IVF embryos, usually to ensure that only embryos free from a particular genetic condition are returned to the woman's womb.

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PGD is an option for some families at risk of serious genetic disorders and is offered as an alternative to prenatal testing.

•

Uses and limitations

Most common method of carrying this out is via a blastomere biopsy.•Test after 3 days whilst all 8 cells are still totipotent.•

Fluorescent in situ hybridisation- To detect chromosome abnormalities.•Polymerase chain reaction and DNA sequencing- Used to detect mutations in single genes.

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What is allowed in the UK?

Severe early onset genetic diseases e.g. Tay-Sachs1.Severe late onset conditions e.g. Huntingdon's2.Disease with incomplete penetrance e.g. hereditary breast cancer.3.To choose a tissue matched baby that can provide umbilical cord blood to treat a sick sibling.

4.

NOT ALLOWED TO CHOOSE BOY/GIRL NON-MEDICAL REASONS5.

Limitations:It requires use of IVF which is physically and emotionally demanding.•Only suitable for conditions where genetic/chromosome abnormality is known.•Can only select for traits that already exist- NO DESIGN POSSIBLE.•

Ethical issues

Embryo testing

G7 Genomics06 April 201210:58


Involves discarding unused embryos.•Disability rights arguments.•Slippery slope- designer babies.•Eugenics.•


Documents

Genetics - Imperial College Union · Insertion- The adding of an extra DNA nucleotide that results in a shifting of ... Chromosomal abnormalities. 4. ... Delayed fertilisation after