HD II Flashcards
the goal of HD treatment
to increase the firing of the GABAergic Enk+ neurons (D2R-expressing) in the striatum to reduce dyskinesia
Pharmacological approaches to treat HD motor symptoms
- relief of dopamine inhibition by dopamine antagonists (classic antipsychotics such as haloperidol and chlorpromazine)
- Atypical antipsychotic (clozapine and olanzapine) are also used with less side effects
- TETRABENAZINE (presynaptic monoamine-depleting agent)
Gold standard treatment of motor symptoms in HD
TETRABENAZINE (presynaptic monoamine-depleting agent)
Tetrabenazine (TBZ): mechanism
Reversible inhibitor of vesicular monoamine transporter 2 (VMAT-2) –> depletes DA stores –> no DA release in striatum –> prevents too much movement
Also a weak inhibitor of D2 receptors
VMAT 2 role
Transports DA back into vessicles
inhibited by TBZ so that no DA is stored in vesicles and therefore no DA will be released in striatum
Tetrabenazine (TBZ): drug type/use
benzoquinolizine derivative used for the treatment of various diskinetic disorders
Tetrabenazine (TBZ): Side effects
Side effects are similar to those caused by anti-psychotics:
- depression (15% of patients)
- suicidal thoughts
- akathisia (restlessness)
- dizziness, sedation
- parkinsonism
TBZ vs classic antipsychotics (side effects)
Reduced incidence of complications such as parkinsonism and dyskinesia with respect to classic antipsychotics (due to much weaker D2R affinity)
______ is a major concern with TBZ treatment and patients need to be carefully monitored
Increased depression and suicidal thoughts are a major concern and patients must be carefully monitored for such complications.
This concern decreases the number of “treatable” patients.
patient with existing psychiatric issues should avoid TBZ
What limits the number of patients on TBZ
concerns regarding increased depression and suicidal thoughts
Deuterated tetrabenazine (Deutetrabenazine): what is it
Analogue of tetrabenazine with the hydrogen atoms in the two methoxy groups replaces with the isotope deuterium
Deutetrabenazine vs. TBZ
Deuterium substitution slows down the oxidative
metabolism of the methoxy groups –> decreased metabolism (longer half-life) –> Decreased dosage, greater tolerability and improved dosing regimen
Antisense oligonucleotide (ASO): what are they
short synthetic modified nucleic acids that:
- are delivered as a single strand
- Target RNA in cytoplasm and nucleus
- Can be taken up by cells in a naked form (+ enter nucleus)
- have poor passage through the BBB
ASOs goal
to silence mutant huntingtin
ASO mechanism simple
ASOs are able to recognize and bind to complementary mRNA through Waston and Crick Base Pairing then modulate the fate of that RNA
ASO mechanism: target mutations
Once bound, form duplex RNA-DNA recognized by RNAse H1 (enzyme) that degrades the RNA, leaving the DNA intact –> allows further degradation = decreased intracellular levels of mutant HTT
ASO mechanism: target splice sites
antisense nucleotide binding to intron/exon regions of preRNAs –> blocks RNA splicing –> decreased production of target protein
ASO mechanism: target translation start sites
target translation start sites –> prevent protein translation –> decreased production of target protein
ASO mechanisms
- target mutations
- target splice sites
- target translation start sites
= all end up decreasing pathological protein production
Small interfering RNA (siRNA): characteristics
- Delivered as a duplex
- Target mRNA in cytoplasm only
- Must be delivered with viral vectors (can’t get into cells themselves)
- Poor passage through the BBB
Goal of siRNA
to silence mutant huntingtin
siRNA: mechanism
Delivered as a duplex –> associates with AGO part of RISC –> passenger strand is removed –> guides AGO in search of of complementary sequence –> once complementary sequence is found RISC complex degrades the complement mRNA (if perfect match) OR represses/blocks it (if partially mismatched)
RISC
RNA-inducing silencing complex
Guide strand in siRNAs
Complementary to RNA targetted and is retained attached to AGO to help guide AGO to find complementary sequence –> once complementary sequence is found it is degraded (if perfect match) or blocked but not cleaved (if imperfect match)
T/F: ASOs are rapidly degraded by nucleases if their structure is not chemically modified
TRUE
need to modify their phosphate backbone increases ASOs stability
Phosphorothioate DNA (PS)–what is it used for and how does it work
Used to increased ASO stability
Substitution of a non-bridging oxygen in the phosphate backbone with a sulfur atom –> Resistance to exo- and
endonucleases, BUT Recognition and cleavage
of target sequence by RNase H preserved.
Modifications to ASO
- Phosphorothioate DNA (modify their phosphate backbone)
- Modifications of the 2’ position of the sugar
Modifying sugar of ASO
Modifications of the 2’ position of the sugar:
- enhance ASO potency by stabilizing target binding
- increase resistance to nucleases
- reduce nonspecific protein binding
Results in MOE (2’-O-Methoxyethyl)
Which is the more common modification to ASO
Phosphorothioate DNA more common that modifications to 2’
How ASO and siRNAs are delivered
- NOT suitable for oral delivery (poor intestinal absorption and poor BBB permeability)
- ADMIN: Intraparenchimal, intracerebroventricular or intratechal (not ideal–hard to do)
- Continuous or repeated administration over the course of the disease
siRNA and ASO in multiple Phase I/II trials:
- silencing of apoB in familial hypercholesterolemia (approved by FDA)
- silencing of XIAP in acute myeloid leukemia
- silencing of mutant SOD1 in familial ALS (ISIS-SOD1-Rx)
- Also in clinical trial for the silencing of LRRK2 in PD, tau in AD.
What should you silence for effect
Silencing of huntingtin should be specific for the mutant allele product to avoid loss of normal huntingtin
___% of HD patients have a Single Nucleotide Polymorphism (SNP) in the mutant HD allele (but not in the normal allele) allows…
65%; allows for specific targeting of the mutant allele with specific ASOs
ASOs efficacy
- Only 10-50% of ASOs effectively reduce expression of the target
- Potential off-target effects must be carefully evaluated
IONIS-HTTRx Phase I trial in HD patients: what is it
48 patients given a bolus of AOS intrathecally once monthly for 4 months –> then monitored for 4 more months
IONIS-HTTRx Phase I trial in HD patients: Results
~40% reduction in mutant HTT levels in patients receiving the drug.
GM1 works by
modulating HTT post-translational modifications
GM1 props
- Constitutes ~25% of the total brain ganglioside pool
- Enriched in membrane microdomains involved in cell
signaling - Modulates the activity of various membrane receptors and ion channels
- belongs to class of glycolipid called gangliosides
Components of GM1
- Gycan Headgroup – Protrudes towards the extracellular
space and makes connections with proteins and other molecules - Lipid tail – anchors the molecule to membranes (made of ceramide)
Gangliosides act as ________
“lipid chaperones”
How Gangliosides work
Inserted into bio-membrane can interact with membrane protein receptors with high specificity –> interact and help deliver them to signalling microdomains where they facilitate the interaction of the ligands with receptors and recruitment of downstream signalling complexes
Gangliosides have roles in:
- Modulation of receptor activity (PDGFR, FGFR, EGFR, IR, Trk)
- Neuritogenesis and axon sprouting
- Cell adhesion
- Myelin-axon interactions
- Cell-cell communication
Disruption of ganglioside biosynthesis leads to
__________
Neurodegeneration
Loss of function mutations in genes of the ganglioside biosynthetic pathway cause severe neurodegenerative disorders
Loss of function mutations in genes of the ganglioside biosynthetic pathway cause these disorders
- Infantile-onset symptomatic epilepsy syndrome (progressive brain atrophy, epilepsy and motor symptoms)
- Complex hereditary spastic paraplegia (epilepsy, cognitive impairment, motor symptoms and deafness)
- loss of GM1 observed in DA neurons in human PD in substantial nigra
Admin of ___ protects HD cells from apoptosis
GM1 (it’s neuroprotective!)
Addition of GM1 protective in both WT and HD
in HD cells levels of ____ are decreased if we provide ___ it ____ cells
gangliosides; GM1: protects
GM1 protects cells from stress and apoptotic stimuli (return to WT levels of apoptosis)
Administration of GM1 induces ____ of mutant HTT which suggests that GM1 can ____ and ameliorate ___ symptoms
phosphorylation; slow disease progression; motor
admin of GM1 induces phosphorylation of mutant HTT at serine 13 and 16
Why is it important that GM1 phosphorylates serine 13 and 16
phosphorylation of these residues is crucial to modulate mutant huntingtin toxicity (decrease toxicity)
Motor tests of HD models
cross a ladder
WT have no problem
HD mice have trouble
HD + GM1 look more like WT
Effects of GM1 on crossing narrow beam
W/in 14 days of GM1 treatment HD + GM1 mice retun to normal (WT-like) performance
GM1 on non-motor symptoms
- Administration of GM1 corrects non-motor symptoms of HD (depression, anxiety, cognitive problems)
- GM1 slows down neurodegeneration in HD mice (less degen of striatal neurons)
Depression in HD
HD animals have higher depression + anxiety than WT but GM-1 returns these behaviours to WT-levels
Administration of GM1 decreases ____ and ____
soluble mutant HTT (act on misfolded mutant HTT); insoluble aggregates (act on fibrils/aggregates)
Domino effect and GM1
- Restoring GM1 in the brain restores cellular functions
- BUT GM1 also has direct effects on mutant HTT (promotes phosphorylation, helps degradation of muHTT, prevents aggregation)
- leads to increased neuronal survival and therefore fewer symptoms
Challenges to move GM1 to clinical trials
- Modality of administration is challenging due to poor/unclear permeability of the BBB to GM1 (routes are not ideal–ex. intraventricularly)
- Mechanism of action still not completely clear and likely to be pleiotropic (makes it harder to translate into clinical use)
