Mutation Specific Therapies & Pharmacogenomics Flashcards
What are antisense oligonucleotides?
Small ssDNA or ssRNA molecules that affect translation by binding mRNA and blocking translational machinery
How do ASO’s enter cells?
Endocytosis so must be stable
3 concerns r.e. ASOs
Delivery to target tissue
Repeat treatments required to sustain effect
Complete inhibition not possible if level of ASO decreased compared to mRNA levels
3 mechanisms for ASO therapies
Reading frame correction
Enhanced splicing
Downregulation of transcription
1 example of reading frame correction therapy
Eteplirsen (Exondys51) for DMD patients with deletions where exon 50 skipping will restore the reading frame (14%) e.g. 49-50
Where is reading frame correction therapy for DMD approved?
US only
Refused approval in Europe as main studies showed minimal effect and no long term studies showing lasting benefits to pts
$300k per year
Example of enhanced splicing ASO therapy
Nusinersen (Spinraza) in SMA
How does Nusinersen work?
Blocks SMN2 ISS-N1 site in exon 7, impairing access to negative splicing factors, stabilises U1snRNP machinery and promotes exon 7 inclusion. Increases full length SMN protein expression
Trial outcomes of Nusinersen
Original trial in 2017 showed outstanding response with cohorts meeting normal motor milestones and no ventilation support
Approved in 2019 for Types 1-3 SMA
2 other SMA therapies and brief mechanisms
Risdiplam - small molecule oral drug (pyridazine derivative) that modifies splicing
81% alive wo ventilation after 12m
(25% in untreated cohorts)
Zolgensma - viral vector delivered SMN1 transgene and synthetic promoter
2 examples of downregulation of transcription ASOs
IONIS-HTTRx (Tominersen) for HD
Baliforsen for DM1
What is the mechanism for Baliforsen?
Targets DMPK for degradation (through RNA-DNA heteroduplexes) and frees MBNL1 protein for alternative splicing function
How has delivery to muscle been improved for Baliforsen ?
Combine with antibody for transferrin receptor (highly expressed on muscle cells due to high iron levels)
Ab+ASO are endocytosed into cell and improves targeted delivery to muscle
What is readthrough therapy?
Aminoglycoside antibiotics allow ribosomes to readthrough premature termination codons
Interact with aminoacyl tRNA site = altered rRNA conformation = decreased accuracy
When is readthrough therapy most effective?
When low levels of functional protein can ameliorate disease phenotypes
Example readthrough therapy
effect on protein?
Ataluren/Translarna for DMD pts with nonsense mutations (15%)
10-20% increase in dystrophin expression which protects against some contractional damage
What do CFTR modulators promote?
2 examples
correct folding and transport
Ivacaftor, Lumacafter (together = Orkambi)
What CF variants are targeted by potentiator therapy?
Class 3 - mutant CFTr cannot transport Cl- through channel (decreased gating efficiency)
e.g. G551D
Ivacaftor binds CFTR to induce non-conventional gating
What CF variants are targeted by corrector therapy?
Class 2 - CFTR is incorrectly folded/processed in ER and targeted for degredation
e.g. Phe508del
Lumacaftor faciliates proper maturation of CFTR and delivery to membrane. increases Cl- transport by ~14% (10% required for clinical response)
What do pharmacogenomic tests look for?
Likelihood of altered responsiveness to drugs
2 types of genes looked at in pharmacogenomics
Enzymes that activate drugs. Variants = decreased activation and sensitivity
Enzymes that breakdown drugs. Variants = accumulation
What is the clinical utility of pharmacogenomics
Improve patient outcomes through personalised drug therapies and decrease healthcare costs. Improved drug safety and efficiency.
ADRs cause 2000 deaths & cost NHS £2billion annually
5-fluorouracil mechanism
Antimetabolite chemo that affects with DNA synthesis by blocking thymidine formation, affects rapidly dividing cells
Enzyme involved in 5FU inactivation
Dihydropyrimidine dehydrogenase DPYD
ADR associated with DPYD deficiency
Diarrhoea, vomiting, cardiac/CNS issues
DPYD testing
4 variants (3 SNV and 1 haplotype of 3 var in cis)
DPYD level depends on hetz, homoz or comp hetz status
Levels of DPYD that confers intermediate activity
1-1.5
reduce dose 50%
What is clopidogrel used for?
Anticlotting drug that prevents blood clot formation by making platelets less likely to bind
Used in pts at risk of clotting events e.g. strokes
enzyme that activates clopidogrel?
CYP2C19
Significance of CYP2C19 variants?
Decreased activity = decreased conversion of clopidogrel to active form = increased risk of clotting events
Clinical utility for testing CYP2C19 in stroke patients?
How to test?
Identified pts that require alternative clotting medications
2023 NICE draft guidelines recommend POCT test (more expensive but does not require sample transit to genetics lab and results quicker in acute settings)
Example of a mitochondrial pharmacogenomic variant
m.1555A>G in MT-RNR1
1 in 500 have this variant
What does m.1555A>G cause?
Predisposition to aminoglycoside induced ototoxity (hearing loss post antibiotic treatment - implications in neonatal sepsis)
Example of POCT pharmacogenomics
Development of rapid POCT for m.1555A>G in Manchester study. Device is able to genotype in ~26 min and does not disrupt normal clinical practice i.e. no delay in lifesaving treatment
What did the PREPARE study conclude?
Pharmacogenomic panel testing is feasible in diverse European healthcare systems
