Week 10: Diagnosis and Intervention Flashcards
Beta Thalasseamia
- Failure to thrive as a child
- Severe anaemia
- Swollen belly due to splenomegaly
- Bone deformity due to bone marrow expansion
- Carriers present mild anaemia
Gene replacement therapy for beta thalassaemia - Future treatment as it is currently in clinical trials
- In a treatment called Beti-cel, short for Betibeglogene autosomal, stem cells are collected from the patient’s blood and functional genes are added to their DNA via a viral vector. The patient’s body is cleared from abnormal stem cells by chemotherapy and the added stem cells are infused back
Bone marrow transplant
- Haematopoietic stem cell transplantation (HSCT)
- HSCT uses human leukocyte antigen (HLA)-matched stem cells derived from a donors bone marrow, peripheral blood, or umbilical cord blood, survival after allogeneic transplantation depends on donor-recipient matching and the graft-versus-host response.
Gene replacement therapy
- Replacing a mutated gene that causes a disease with a healthy copy of the gene
- Inactivating or ‘knocking out’ a mutated gene that is functioning properly
- Introducing a new gene into the body to help fight disease
Severe Combined Immunodeficiency (SCID)
- A group of inherited disorders characterised by defects in both B and T cell responses.
- One infant in every 50,000 births
- Individuals with this disease are severely immunocompromised
- Mutations in the IL2RG gene cause X-linked SCID, accounting for 25 - 40% of cases
- Another form of SCID is caused by a deficiency of the enzyme adenosine deaminase (ADA) normally produced by a gene on chromosome 20. It accounts for 5 - 10% of the recessive SCID cases
- Early detection and treatment is imperative
- Absent or impaired ADA function leads to the accumulation of the toxic metabolites
- In addition to severe lymphocytopaenia affecting T- and B- lymphocytes and NK cells, neurodevelopmental deficits, sensorineural deafness and skeletal abnormalities can also occur.
HSCT for SCID - HSCT uses human leukocyte antigen (HLA) matched stem cells derived from a donors bone marrow, peripheral blood, or umbilical cord blood. Survival after allogeneic transplantation depends on donor-recipient matching and the graft-versus-host response
- Patients require chemotherapy prior to transplantation
ADA SCID gene therapy
1. Cells are collected from the patient
2. Blood stem cells are collected and purified
3. A working copy of the gene is inserted into the cells
4. The patient is conditioned to receive treatment
5. The gene-corrected cells are infused into the patient
Spinal Muscular Atrophy (SMA) Type 1
- SMA affects 1 in 8,000 to 10,000 people worldwide with type 1 accounting for half of cases
- Weak respiratory systems/muscles and an abnormally shaped bell-shaped chest
- Difficult feeding and swallowing
- Muscle weakness
- Most individuals die of respiratory failure during early childhood
- Autosomal recessive disorder
- Caused by a loss of function mutation in survival of motor neurone 1 (SM1)
- Presentation can be modified due to copy number variation (CNV) in SM2
- Zolgensma is a one off gene therapy with a reported list price of £1.79 million per dose
- Adeno-associated virus (AAV) 9 based gene therapy that delivers a functional copy of the SMN1 gene to the nerve cells
Gene replacement therapy for RP and Leber’s Congenital Amaurosis type 2 (LCA2)
- Adeno-associated viral vectors are the most utilised vectors for ocular gene therapy
- People with inherited retinal dystrophies cause by gene mutations in the 65kDa retinal pigment epithelium gene (RPE65)
- 2% of RP linked to RPE65
- 6 - 16% of LCA cases linked to RPE65
- Gene therapy is suggested for individuals with confirmed biallelic RPE65 mutations and who have sufficient viable retinal cells
- Different changes within the same pathway or related pathways can produce similar or slightly different phenotypes
Haemoglobin gene expression during development
- The switch from foetal to adult haemoglobin relies on repression or silencing of the upstream Y-globin gene but identification of the transcriptional repressors that bind to the sites at which a cluster of naturally occurring variants associated with HPFH (hereditary persistence of foetal haemoglobin) are found to be elusive
- In healthy humans, a shift from Y-globin to Beta-globin gene expression around birth underlies the switch from foetal to adult.
Hydroxyurea
- Hydroxyurea is used to reduce the frequency of painful sickle cell crises and to reduce the need for blood transfusions in adults and children two years or older.
- Hydroxyurea is an inducer of HBF and can be used to reduce the frequency of sickle cell crises
- In another approach, CRISPR-Cas9 based genome editing to disrupt the production of Y-globin inhibitor BCL11A in progenitors of red blood cells increasing foetal haemoglobin levels
RNA interference
- Hereditary transthyretin (hATTR) amyloidosis, a disease in which abnormal proteins called amyloids build up in tissues around the body over time including around the nerves
- Normal transthyretin (TTR) transports vitamin A (retinol) throughout the body
- Hereditary amyloidogenic transthyretin (ATTRv) amyloidosis is an autosomal dominant, adult-onset systemic disease that usually presents as a progressive peripheral neuropathy and is caused by point mutations in the gene that encodes transthyretin (TTR)
- Clinical presentation is diverse, including length-dependent small-fibre polyneuropathy, all-fibre polyneuropathy, pseudo chronic inflammatory demyelinating polyneuropathy, upper-limb-onset neuropathy and motor neuropathy: half of patients have cardiac amyloidosis
- V30M is the most common mutation worldwide with early age of onset symptoms (<50 years) in some cases
*Median survival 4.7 years following diagnosis and reduced survival (3.4 years) for patients presenting with cardiomyopathy
Traditional treatment options - Liver transplant for early-stage disease
- TTR tetramer stabilisers
- Patisiran is a double-stranded small interfering RNA (siRNA) encapsulated in a lipid nanoparticle for delivery to hepatocytes
- It targets the conserved 3’ untranslated region of both the mutant and wild-type transthyretin mRNA causing a reduction of all products
- For use in adult patients in the first two stages of nerve damage
RNA regulation
- Nucleotide sequences that influence the lifespan of mRNA in eukaryotes reside in the untranslated regions (UTRs)
- Destabilising elements (DEs) and stabilising elements (SEs) in the 3’ UTR act as binding sites for specific RNA binding proteins (RBPs) that act to either reduce or increase the half-life of the RNA
- 3’ UTRs contain recognition sites for microRNAs that are involved in the repression of mRNA translation and sometime their degradation through interaction with the RNA-induced silencing complex (RISC)
- The 5’ UTR acts as an entry point for the ribosome and can adapt elaborate RNA secondary and tertiary structures that may regulate translation initiation and in a cap-dependent or cap-independent manner.
- The miRNA binds to a target mRNA. If bases are complementary, mRNA is degraded, if the match is less complete, translation is blocked
Phenylketonuria (PKU)
- Autosomal recessive disorder where individuals are deficient in an enzyme that breaks down the amino acid phenylalanine called phenylalanine hydroxylase
- Different variants have different levels of activity
- About 1 in 10,000 babies born in the UK have PKU
Symptoms - Behavioural difficulties
- Fairer hair, skin and eyes than siblings without the condition
- Eczema
- Recurrent vomiting
- Jerking movements
- Epilepsy
PKU: Diet - Choose to eat the most phenyl-free foods
PKU: Drug - Sapropterin dihydrochloride is a synthetic form of BH4
- BH4 is a cofactor for phenylalanine hydroxylase (PAH)
- Used to increase residual PAH activity in patients with some PAH activity
- Does not replace all dietary restrictions
Inhibiting function
- Around 85 - 90% of people with chronic myelogenous leukaemia (CML) have the philadelphia chromosome which is formed by the t(9;22)(q34;q11) translocation.
- Tyrosine-protein kinase ABL1 is involved in cell proliferation, differentiation, and migration and is tightly regulated
- The new fusion gene, called BCR-ABL1, promotes cell proliferation and blocks apoptosis but isn’t tightly regulated
Beta-Thalassaemia
- Anaemia treated with blood transfusions. However, excess iron due to blood transfusions
- Swollen belly due to splenomegaly
Cystic Fibrosis
- Medicine for lung problems
- Antibiotics
- Mucus thinner
- Bronchodilators
- Steroid medicine
- Exercise
- Airway clearance techniques
- Dietary and nutritional advice
- Lung transplants
Pre-natal testing
Down’s Syndrome (Trisomy 21)
Symptoms
* 1/800 live births
* Abnormal features
* Malformation: atresia of oesophagus, duodenum or anus, structural heart defects, hearing loss, hypothyroidism, eye abnormalities
* Respiratory infection
* Leukaemia
* Mental retardation
* Sterile
Amniocentesis
* 10 - 20ml of fluid is aspirated through the abdominal wall under ultrasound guidance
* Usually occurs at around 16 weeks
* Most cells in the amniotic fluid have been shed by the foetus allowing genetic investigations to be undertaken
Chronic villus sampling (CVS)
* Usually carried out between 11 to 12 weeks
* Transabdominal aspiration of chronic villus. This tissue is foetal origin.
**Quantitative fluoresecent PCR (QF-PCR)
* QF-PCR can be performed on amniotic fluid to detect aneuploidies in chromosomes 13,18,21 X and Y.
* Fluorescently ladled primers are used to analyse up to 5 short tandem repeats per chromosome.
Cell-free foetal DNA (cffDNA)
* Cell-free foetal DNA represents extracellular DNA which originates from trophoblastic cells. However, the vast majority of cell free DNA in maternal blood originates from the mother with cell-free foetal DNA representing only 3% of the total cell-free circulating DNA in early pregnancy rising to 6% in late pregnancy. A massive parallel shotgun approach is used to detect over representation of specific chromosomes.
Ultrasounds
* A number of indicators of Down’s Syndrome can be observed on 2nd trimester ultrasound. Dilated brain ventricles, increased thickness of the back of the neck, and an abnormal artery to the upper extremities increase the risk by three- to four-fold and an absent or small nose bone increases the risk by six- to seven-fold
* Nuchal thickening, an accumulation of fluid at the base of the skull is an indicator of potential chromosomal abnormality.
