Session 3 - Postnatal testing Flashcards
1
Q
Define Anticipation.
Give examples of 3 diseases that show anticipation
A
The phenomenon in which the severity of disease increases as it is passed down the generations.
FraX, DM, HD, DRPLA, SCA1, SCA3, SCA7.
2
Q
List possible mechanisms of repeat expansion in diseases showing anticipation.
A
Replication slippage.
Incorrect reassembly of Okazaki fragments
Formation of hairpin loops causing the replication fork to stall.
Unequal crossing over.
3
Q
Is anticipation observed in polyglutamine or polyalanine disorders?
A
Polyglutamine.
4
Q
What three biases can give a false impression of anticipation?
A
Preferential ascertainment of parents with late onset of disease.
Preferential ascertainment of children with severe early onset disease.
Preferential ascertainment of parent-child onset at the same time.
5
Q
Define Age-related mosaicism
A
The accumulation of somatic mutations over time.
6
Q
Give examples of age-related mosaicism.
A
X-chromosome aneuploidy/loss. Caused by premature centromere division during mitosis.
Cancer
7
Q
Define Variable expressivity
A
Phenotype is expressed to different degrees among individuals with the genotype
8
Q
Give examples of variable expressivity
A
Marfan syndrome - FBN1.
NF1
Waardenburg syndrome
9
Q
Define penetrance.
A
The likelihood of experiencing a disease if you have a pathogenic mutation.
10
Q
Give examples of penetrance
A
TOR1A - early onset dystonia - common mutation c,907_909delGAG present in >99% of individuals - only 30% express disease.
11
Q
Define sex-limiting
A
Phenotype is limited to a single gender - only expressed in males or females.
12
Q
Give an example of a sex-limited disease.
A
Familial precocious puberty in males. LHCGR mutations
13
Q
Define epistasis
A
The presence of a variant in one gene determines the expression of a variant in a non-allelic gene
14
Q
Give an example of epistasis.
A
Bombay phenotype ‘overrides’ the usual A/B dominance/co-dominance - if an individual is homozygous for the Bombay variant their blood group will be O.
Albinism overrides hair colour genes.
15
Q
Define pleiotropy
A
One gene causes mutliple, seemingly unrelated phenotypes.
16
Q
Give an example of pleiotropy
A
PKU - caused by mutations in PAH, require to turn phenylalanine into tyrosine. Lack of tyrosine = lack of downstream compound, melanin, hence the albinism.
17
Q
Define the following mutation types:
- Amorphic
- Hypomorphic
- Hypermorphic
- Antimorph
- Neomorph.
A
- No protein product.
- Partial loss of protein expression
- Partial gain in protein activity - GoF
- Mutation acting as antagonist to usual function - dominant negative.
- Mutation gives protein a new function.
18
Q
Give examples of X-linked dominant diseases
A
Non-lethal in males:
Alports (COL4A5 - deafness, RP, renal problems)
Hypophosphataemia (PHEX - loss of phosphate through kidneys = vitamin D3 deficiency and rickets)
FraX
Lethal in males:
IP - NEMO - can be in in 47,XXY
Rett syndrome - MECP2 (CDKL5, FOXG1) - can be seen in 47, XXY and in MECP2 duplication.
XLD - males unaffected
Lack of protein is not disease-causing - mutant protein is.
Or, only mutant protein is fine, WT + mutant causes problems. Craniofacial synostosis syndrome (EFNB1)
19
Q
Give examples of XLR diseases.
A
Dystrophinopathies - DMD/BMD Androgen Insensitivity Haemophilia A/B SBMA Fabry XL-RP Huner syndrome Colour-blindness.
20
Q
List clinical features of DMD
A
Calf hypertrophy Progressive symmetrical muscle weakness Joint contractures Gower sign Age of onset 2-5years Wheelchair bound by early teens. Death by 3rd decade
21
Q
List clinical features of BMD
A
Later onset muscle weakness
Death in mid-40s
Often remain ambulatory til 20s
22
Q
What serum metabolite can be used to diagnose DMD/BMD/DCM? What results would you expect to see for each?
A
Increased CK in the blood.
DMD 10x normal
BMD 5x normal
DCM ~normal
23
Q
What is the advantage of performing genetic testing for DMD/BMD/DCM?
A
Don’t need to take a muscle biopsy.
24
Q
Summarise the testing strategy for dystrophinopathies.
A
del/dup analysis
no variant then sequencing
no variant then western blot of tissue sample and IHC
25
What is special about the DMD gene?
It has three promoters - B (brain), M (muscle) and P (purkinje). These promoters produce a different transcript in different tissues. Each transcript has a unique exon 1, and the same 78 subsequent exons.
26
What is DMD-associated cardiomyopathy caused by?
Mutations in exon 1 associated with the M promoter. Skeletal muscle effects of the disease are not observed, as transcription of B and P transcripts in other tissues is increased to compensate.
27
Describe the mutation spectrum seen in DMD/BMD
Most are del/dups of >1 exon: DMD 60-65%, BMD 80-85%
Two deletion hotspots exist:
1. central region, exons 44-53 - 80% of DMD del, 20% DMD dup
2. 5' region, exons 2-20 - 20% of DMD del, 80% DMD dup.
28
What can be used to predict pathogenicity in DMD/BMD?
Frameshift hypothesis - out of frame deletions are most likely to cause DMD (more severe phenotype) than in-frame deletions.
29
What factors complicate molecular diagnostics in DMD/BMD?
High new mutation rate (1/3 cases)
Germline mosaicism in mothers of patients. Recurrence risk is 5%
High recombination rate - 10%.
30
What are possible treatments for DMD/BMD?
STOP codon read-through - Gentamicin treatment
| Exon skipping using antisense oligos to promote skipping of exon(s) and restoration of reading frame.
31
What causes Kennedy disease (SBMA)?
Repeat expansion in the AR gene (>38 repeats of the polyglut tract causes fully penetrant disease)
32
What are Haemophilia A and B caused by?
Mutations in F8 and F9, respectively.
33
What is the most common mutation, seen in 45% of haemophilia A patients?
inversion of intron 22.
34
What are the diagnostic sensitivities of testing methods used to diagnose Haem A and Haem B?
Haem A - 98%
| Haem B - 90%
35
Define haploinsufficiency.
The loss of half of the amount of gene product is sufficient to cause disease.
36
What type of genes show haploinsufficiency?
Highly expressed genes - lots of gene product needed, so one copy isn't enough.
Dosage sensitive genes - proteins that are part of a quantitative signalling system etc.
Imprinted genes
37
List examples of haploinsufficient genes in single gene disorders
Hypertrophic cardiomyopathy: MYH7, MYBPC3, TNNT2, TNNI3 - 4 gene assay will detect 50% of all cases of HCM
Alagille syndrome - JAG1 - multisystem phenotype
Aniridia (PAX6)
Autosomal dominant optic atrophy (OPA1)
HNPP/PMP22 - duplications cause CMT1A, mutations cause HNPP.
38
List examples of haploinsufficient genes in contiguous gene deletion syndromes
DiGeorge - TBX1
Williams - ELN
WAGR - PAX6
39
List examples of haploinsufficient genes in cancer
TP53
BRCA1
PTEN
It is now suspected that a single mutation in these genes may be sufficient to cause cancer. Two-hits not required.
40
What is a gain of function mutation?
A Hypermorph or Neomorph; result in an increase in gene expression or increase in activity of the gene product
41
Give some examples of Gain of Function mutations
PTPN11 - Noonan syndrome - activation of RAS/MAPK pathway
HTT - Huntington disease - protein aggregation
PMP22 - HMSN/CMT - Overexpression
BCR-ABL1 - CML - Chimeric protein
42
Where in HTT is the unstable CAG repeat?
Exon 1
43
What is a normal sized allele in HD?
Normal <36
44
What evidence is there that HD is a GoF disease?
Patients with chromosomal deletions do not manifest
Homozygotes are as affected as heterozygotes
Phenotypically normal individuals have been identified with a translocation with a breakpoint in HTT
HD levels of polypeptide from the normal and mutant alleles are the same.
45
Describe the molecular pathogenesis of HD
Mutant HTT forms abnormal protein structuers and is truncated by Caspase-6 cleavage to produce toxic N-terminal fragments. This mutant protein then interferes with gene transcription, particularly in the mitochondria, affecting metabolic processes and inducing oxidative stress.
46
Name other triplet repeat disorders thought to act by dominant GoF
SCA1,2,3,6,7,17
| DM
47
What is the most common cause of CMT?
PMP22 duplication - a 1.5mb region
48
What does deletion/LoF of PMP22 cause?
HNPP
49
List examples of diseases showing a dominant negative effect
```
Marfan syndrome
Non-syndromic hearing loss (Cx26/30)
OI - Col1a1 and COL1A2
TP53
Alzheimer's disease (APP)
```
50
How do mutations in GBJ2/6 (Connexin 26 and 30, respecitvely) cause deafness?
Mutation in one gene prevents interaction and formation of gap junctions in the cochlear membrane (Cx26 and Cx30 form a complex together). Also involved in neurotransmitter release. As signals cannot travel through the junctions the patient is deaf.
Often AR disease, but some dominant mutations in Cx26 form a full length structurally abnormal protein that interferes with junction formation.
51
What are the two type of Osteogenesis Imperfecta? What is the difference in the mutation spectrum for each?
OI type 1 - less severe, caused by loss of the COL1A1 gene function through PTC and nonsense mutation. Reduced amount of functionally normal COL1A1 is available.
OI type 2, 3 and 4 - severe, caused by missense mutations affecting glycine residues. These glycine residues are instrumental in holding the collagen helix structure together. This leads to the formation of abnormal protein.
52
How is TP53 considered a dominant negative gene?
TP53 usually forms tetramers (2x dimers) and acts to prevent passage through the cell cycle.
Mutations in the DNA binding domain prevent TP53 complex from binding the DNA. If the dimers contain abnormal TP53, they are are functionally abnormal then they will not bind the DNA
53
Describe the structure and function of CFTR
2 transmembrane domains, 2 nucleotide binding domains, a regulatory domain (R) and C-terminus.
It is a cAMP activated chloride channel present in the apical membrane of secretory epithelial cells
54
What controls the opening/closing of the CFTR protein?
The binding of protein kinase A to the R domain.
55
Summarise the phenotype of CF patients.
Prenatal:
Meconium ileus, Echogenic bowel (3% of EB cases are CF)
```
Postnatal:
FTT
Digestive problems
Respiratory infections
Pancreatic insufficiency
Chronic cough
Bronchiectasis
Infertility
Clubbing of the fingers
Sputum with staph or pseuodomonas infection.
```
56
What are the common mutation types in CF?
Most are point mutations. 10% are larger deletions.
57
What testing methods are available?
```
OLA
ARMS
MLPA
Sequencing
CF4 kit for newborn screening (G542X, G551D, Phe508del, c.489+1G>T)
UPD testing for UPD7
```
58
What is a problem with the OLA CF detection kit?
Presence of a p.Phe508Cys can hide mask a normal p.Phe508del allele, leading to potential misdiagnoses in the future.
59
What is detected in the neonatal blood spot test for CF? At what day is this taken, and what happens if there is an abnormal result?
IRT, blood spot taken at 5 days old.
If raised, test again at 28 days.
If still raised, send for CF4 test (Fdel508, G542X, G551D, c.489+1G>T)
60
What is the gold standard test for CF? How many patients test positive?
Sweat test. 90% positive.
61
What is the cut off for the sweat test?
60 meq/L - anything more and this is diagnostic. Two abnormal readings are needed.
62
Apart from the sweat test, what other biochemical test can be performed to diagnose CF?
Nasal potential different (NPD) test. Measures the transport of Na+ and Cl- in the epithelium of the upper respiratory tract.
63
Describe the 5 classes of CF mutation. Give examples of mutations and what phenotype they are associated with.
1. Loss of transcript, splicing aberrations. G542X. Classical CF
2. protein retained in the ER as it is misfolded. Phe508del. Classical CF
3. Problems with channel gating (G551D) Mild CF
4. Problems with channel conductivity (R117H) Infertility
5. Mutation in regulatory gene or related protein (ORCC, ENaC) Atypical CF
64
What modifiers can affect CF phenotype?
poly T (5,7,9) and poly TG (11,12,13) tracts.
65
How do the poly-T/TG tracts affect CF phenotype?
They result in exon 9 skipping in 90% of transcripts (5T, 12TG and 13TG). They are present at the splice acceptor site of intron 8.
66
What mutation is 5T commonly seen in cis with, and what affect dos this have?
R117H. It enhances the effect of the R117H mutation to produce a more severe phenotype.
67
List some other CF modifier genes.
MBL, ORCC, ENaC
68
What treatments are available for CF patients
Enzyme supplementation
Dietary supplementation
Physio
Fertility treatment
Inhalation of compounds to break down mucus
Potential gene therapy - in a lentivirus vector, can increase expression at surface.
CRISPR/Cas9 treatment to correct mutation - only performed in vitro so far.
69
List some treatments for the different mutation classes.
1 Nonsense mutation - PTC124 to promote read through. Gentamicin also promotes read-through. Antisense oligos can be used to treat splice variants
2 Correctors to promote correct folding of protein (Lumacaftor)
3 Ivacaftor acts to increase the functional presence of CFTR at cell surface
4 Ivacaftor can also be used
5 Compounds enhancing CFTR regulation and anchoring
70
What are the three disorders associated with FRM1 mutations?
FRAX, FXTAS, POI.
71
Where in FMR1 is the poly-CGG repeat? What does expansion cause?
5'UTR. Expansion causes methylation of a CpG island 250bp downstream of the repeat site to prevent transcription.
72
What is the function of FMRP?
An RNA binding protein involved in transporting mRNA from the nucleus to other organelles
73
What is the smallest known repeat size to expand to a full mutation in a single generation?
56
74
What are the range of repeat sizes seen in FRAX?
6-46 Normal
47-54 Intermediate
55-200 Premutation
200+ full mutation
75
What is the pathogenic mechanism behind FXTAS and POI
RNA gain of function.
76
How many women with a premutation experience POI?
20%
77
What proportion of men and women premutation carriers experience FXTAS?
45% men
| 16% women
78
When would a southern blot be performed for FRAX?
Single normal allele in female, absent allele in males
If moscaism is suspected
Prenatal testing
79
What is the advantage of performing the double-digest in FRAX blotting?
Allows determination of methylation status. Some individuals have a full expansion, but it remains unmethylated. These individuals may have a less severe phenotype.
80
What types of mosaicism can be seen in FRAX?
Repeat size, due to large repeats being unstable, and methylation.
81
What 5 mechanisms lead to imprinting defects?
```
UPD
Mutation of the imprinting centre
Deletion of an imprinted region
Duplication of an imprinted region
Epimutation
```
82
Where are the 6 imprinted regions in the human genome?
```
6q24.7
7
11p15
14q32
15q11.2
20q13.2
```
83
What has been associated with an increase in the number of imprinting defects? Why might this be so?
ART - methylation is set during early embryogenesis, when the embryo is still in vitro.
84
Give an example of whole genome UPD
Hydatidiform mole. No maternal imprint is set - all paternal.
85
What is special about BWS imprinting?
The region on 11p15.5 has two imprinting centres/differentially methylated regions: H19DMR and KvDMR
86
Which genes are involved in the pathogenesis of BWS, which parent are they expressed from and which IC/DMR are they found in?
```
IGF2 - paternally expressed (H19DMR)
H19 - maternally expressed (H19DMR)
CDKN1C - maternally expressed (KVDMR)
KCNQ1 - maternally expressed (KVDMR)
KCNQ1OT1 - paternally expressed (KVDMR)
```
H19DMR is unmethylated on the maternal chromsome
KVDMR is unmethylated on the paternal chromosome
87
How can mutations in the BWS imprinted gene cluster cause sporadic (85%) BWS?
Hypomethylation of KVDMR on the maternal chromosome (50-60%)
Hypermethylation of H19DMR on the maternal chromosome (2-7%)
Mosaic pat UPD (hypometh of KVDMR; hypermeth of H19DMR) - 20%
CDKN1C LoF mutation on maternal chromosome (5-10% of sporadic cases, 40% of familial cases)
Cytogenetic abnormalities in the region.
88
What is the etiology of Russell Silver syndrome?
Mat UPD7 - 7p11.2-13; 7q31
| Mat UPD 11p15.5/Mat imprinting
89
What imprinting abnormalities can cause RSS?
Hypomethylation of H19DMR on the paternal allele - 30-50%
Biallelic expression of H19 and biallelic silencing of IGF2 (Mat UPD11) - v rare
Duplications of 11p15 - rare
Mat UPD7 - 7-10%
90
What imprinting disorder is associated with imprinting problems on chromosome 6q24.7?
Paternal UPD causes transient nenoatal diabetes mellitus type 1
91
What disease is caused by Mat UPD14?
Temple syndrome - similar phenotype to PWS
92
What disease is caused by Pat UPD14?
Pat UPD14 - bell shaped thorax, placentomegaly and polyhydramnios, abdominal wall defects, facial dysmorphism
93
What is caused by Chromosome 20q13 UPD?
GNAS cluster loss of function disorders:
Pseudohypoparathyroidism (PHP) types 1a and 1b.
Pseudopseudohyopparathyroidism (PPHP)
94
What clinical features are associated with 20q13 UPD, and what parent of origin is each derived from?
PHP 1a - Albright hereditary osteodystrophy plus hormone resistance (PTH, TSH) - MATERNAL
PHP 1b - renal PTH resistance without AHO - MATERNAL
PPHP - AHO, no hormone problems - PATERNAL
95
Prader-Willi syndrome Which genes are expressed from the paternal allele, and which genes are expressed from the maternal allele? How is expression controlled?
Paternal: SNRPN, MKRN3, MAGEL2, NDN. SNRPN/SNURF may transcriptionally silence UBE3A on the paternal allele through an antisense mechanism.
Maternal: UBE3A, ATP10C
PWIC - includes the SNRPN promoter and exon 1.
ASIC.
UBE3A is biallelically expressed in some tissues, but preferentially maternal in the brain.
96
What are the different pathogenic mechanisms resulting in AS? Give proportion of cases and recurrence risk
Mat de novo deletion of 15q11-13 - 70-80% - <1%
Pat UPD - 3-7% - <1%
IC defect - 2-3% - <1%
IC mutation - 0.2-0.3% - up to 50%
UBE3A mutation - 10% - up to 50% if maternally inherited
Other - 10% - up to 50%
97
What are the different pathogenic mechanisms resulting in PWS? Give proportion of cases and recurrence risk
Pat de novo deletion of 15q11-13 - 75-80% - <1%
Mat UPD - 20-25% - <1%
IC defect - 1% - <1%
IC deletion - 0.1% - up to 50% if present in father.
98
If there is an absence of a methylation defect, what action can be taken?
PWS - consider differential diagnoses (BBS, Cohen syndrome, SMA, DM, MatUPD14)
AS - sequence UBE3A, consider blot to detect mosaicism for mat/pat alleles.
Consider differential diagnoses - Mowat-Wilson, Rett, ATR-X, CDKL5 mutation, Christianson syndrome.
99
How could Crispr/CAS9 be used to treat AS?
Target SNRPN locus and allow transcription of UBE3A from the paternal allele.
100
Give examples of cases where a single gene can cause multiple disorders.
RET: Somatic mutation - cancer; Gain of function - MEN2A, MEN2B - medullary thyroid cancer; Loss of Function - Hirschprung's disease
COL2A1 - DNE mutations - phenotype dependent on mutation: Achondrogenesis, Hypochondrogenesis, Stickler syndrome, SEDC.
PMP22: Loss of function/Haploinsufficiency (deletions) - HNPP; Gain of function (duplications) - CMT1A.
AR gene: Kennedy syndrome - expansion >36 poly-glut repeats in exon 1 of AR (toxic RNA accumulation, transcriptional dysregulation). AIS - LoF mutations, phenotype depends on retained function.
101
Give examples of a single disorder being caused by multiple genes.
Alports - COL4A3, COL4A4, COL4A5
Bardet-Biedl - BBS1-10
HSP - >52 regions/loci
102
What is Friedreich's ataxia caused by?
GAA repeat expansion in Intron 1 of FXN.
103
How is FA inherited?
AR, Loss of function.
104
What is the carrier rate of FA?
~1/60
105
List some clinical features of FA
```
Ataxia
HCM
wheelchair bound by teens
loss of vibration sense
Death at ~37
Dysarthria
loss of bladder control
Abnormality of eye movements
```
106
What sorts of variants cause FA?
Expansion of GAA repeat - homozygous in 98%
Missense LoF mutation (common c.Gly130Val)
Deletions
Nonsense/FS
107
what is the clinical phenotype of FA dependent on?
Level of mosaicism
Repeat length
Mutation type
108
What 4 classes of FXN repeat expansion are there? What are their sizes?
Normal 5-33
Premutation 34-65
Intermediate 44-65
Full mutation 66-1700
109
Do FXN expansions expand on maternal or paternal transmission?
Maternal.
110
How is FXN tested for?
F-PCR, TP-PCR, Southern blot.
| Sequencing if second mutation suspected, but beware of carriers.
111
How does the expansion of FXN cause loss of FXN protein?
Expansion causes formation of secondary structures in the DNA. This prevents access of TFs and RNA pol during transcription, so less/no FDRA is produced.
Expansion is also associated with increased histone methylation in the region.
112
How does lack of FXN protein loss cause disease?
FXN is a nuclear encoded mitochondrial gene.
Involved in iron transport/chaperone
Lack of FXN causes iron accumulation in the mitochondria - this interferes with oxdiative phosphorylation pathways and results in increased amount of reactive oxygen species produced/oxidative stress.
Affects neurons and cardiac cells as these cells have high energy demand.
113
Due to the disease mechanism, what diseases is FDRA similar to?
MELAS, MERFF.
114
What potential treatments are available for FDRA patients?
Reduce HDAC activity with HDACi(nhibitors). This will open up the chromatin and allow transcription.
IFN-gamma treatment
Antioxidant treatment to reduce oxidative stress (Co-10q)
Gene therapy
115
Name some Gain of function triplet repeat disorders.
Split into categories for those affecting Protein function, and those affecting RNA function
Type 1: protein function affected: HD, DRPLA, SBMA
Type 2: RNA function affected: DM1, DM2
116
List the repeat expansions diagnostic of HD
```
0-27 Normal
27-35 - Premutation
>36 affected
36-39 reduced penetrance
40+ full mutation
```
117
Where is the HD repeat tract found?
Exon 1 of HTT
118
Which parent of origin promotes expansion for HD?
Paternal
119
What theories are there to explain pathogenesis of PolyQ GoF repeat disorders?
1. Aggregation theory: protein aggregates are found in cell cytoplasm and can disrupt normal cellular function
2. Toxic fragment hypothesis: HTT is cleaved by Caspase 6 to form toxic N-terminal fragments
3. Transcript dysregulation hypothesis: PolyQ expansions are found in the nucleus and disrupt gene transcription and TF binding
4. Cytoskeletal defect and axonal transport: disruption of axonal transport by affecting microtubule complexes
5. Effects on non-neuronal cell types
120
List the repeat expansions diagnostic of DM1. Where is the expansion located?
<36 N
37-50 Premutation
51-150 Affected
Expansion located in 3'UTR of DMPK
121
Expansions in which gene cause DM2?
CNBP
122
Which three models are proposed as disease mechanisms causing RNA mediated GoF repeat disorders?
1. Haploinsufficiency
2. Affect chromatin structure, affecting transcription of nearby genes.
3. RNAs form hairpins that are not transported from the nucleus and interrupt normal nuclear function.
123
Give an example of a poly-alanine repeat disorder
FSHD1 and FSHD2
124
What is the main difference between polyA and polyQ disorders?
PolyA repeats are pathogenic at much smaller sizes - 33rpts in PHOX2B is sufficient to cause CCHS.
125
List some phenotypic features of FSHD
Asymmetrical facial weakness
Weakness to upper arm, shoulder, hip girdle and lower leg
Scapular winging
Onset in the 2nd/3rd decade
126
What do both forms of FSHD centre around?
Expression of DUX4 transcript.
127
What genes are 'mutated' in FSHD types 1 and 2?
Type 1: contraction of polyA repeat in D4Z4 - chromatin relaxes and DUX4 produced. Normal range 11-100, mutated <10
Type 2: DUX4 expression caused by hypomethylation at 4q35 - this hypomethylation is caused by mutation in SMCHD1 (deletions, missense, splicing)
Type 1 is AD, type 2 is digenic.
128
What does DUX4 expression cause?
apoptosis of myocytes, leading to the symptoms of FSHD.
129
Give examples of chromosome breakage syndromes. List the genes involved, phenotype, cancer risk and method of testing.
Ataxia telangiectasia - ATM, ataxia, breast cancer, immuno deficiency, hypogonadism. Radiation sensitivity. No SCE - sequencing ~90% of mutations. ALL and lymphoma
Fanconi Anaemia - pancytopaenia, aplastic anaemia, ID, hypoplastic thumbs, onset age ~8. Multiple genes involved. Exposure to alkylating agents (MMC, DEB) to induce chromosomal breaks. No SCE
Bloom syndrome - BLM, UV sensitivity, hypopigmentation, butterfly rash, characteristic facial features, short stature leukaemias and solid tumours. UV and radiation sensitivity. SCE.
Nijmegan breakage syndrome - NBN1, characteristic facial features, dev del. Increased risk of NHL, burkitt's and other cancers. Radiation
XP - multiple genes, UV sensitivity, skin blistering, increased rick of melanoma
130
What is a typical cytogenetic finding in individuals with Bloom syndrome?
Quadriradial configuration.
131
How can antisense oligonucleotides be used to treat genetic disease?
Prevent transcription of mutant transcript in DNE diseases (e.g. Alports)
Suppress a cryptic splice site (a-globin in a-thalassaemia)
Enhance alternative splice site to include/exclude exon(s) - restore reading frame in DMD - mutation specific, Exon 51 skipping would benefit 13% of patients
Enhance a ESE or ESS - SMN1, to promote inclusion of a skipped exon.
Promote readthrough of PTC in CF - Gentamicin, PTC124
Knockdown of toxic transcripts by mutation targeted introduction of a PTC > NMD
Promote expression of a methylated allele - e.g. UBE3A expression from the paternal allele in Angelman syndrome
132
What are the problems with using ASO to correct mutations?
Delivery to target tissue
Longevity
Difficult to achieve complete inhibition due to large volume of mRNA molecules.
133
How can siRNAs be used to inhibit gene function?
Isoform-specific - target specific transcripts - VEGF
| Mutation specific - target specific mutations, e.g. HD
134
What are the limitations of using RNAi?
Delivery to target site
Non-specific off-target effects
May elicit immune responses.
135
Name some drugs that allow read through of PTCs
Gentamicin - makes the RNA pol less fussy.
PTC124 - CF.
136
How is a PTC defined?
By its position and sequence context.
137
Name a molecule that prevents misfolding of proteins.
Lumacaftor for CF class II mutations (Phe508del).
138
Name a molecule that aids ion channel gating function
Ivacaftor for CF class III mutations (G551D). IT makes the ion channel easier to open.
139
List some known microdeletion/duplication syndromes. State their location and how they are formed.
```
WHS - 4p-
CDC - 5pter-
Williams syndrome (ELN) 7q11.23
CMT1A/HNPP (PMP22) 17p11.2 - NAHR and LCR
SMS/PTL - 17p11.2 - NAHR and LCR
PWS/AS - 15q11-13 NAHR and LCRs
DiGeorge/VCFS - 22q11.2 - NAHR and LCRs
Miller-Dieker - 17p11.3
Langer-Giedion - 8q24
WAGR - 11p13 del
```
