A trial of quinidine in genetic epilepsy
A randomized, double-blind, placebo controlled, cross-over trial of quinidine in genetic epilepsy due to KCNT1 mutations
Principal Investigator – Dr Saul Mullen
Associate Investigators – Prof Ingrid Scheffer, Prof Samuel Berkovic,
Dr Patrick Carney,
Version 4
19th November, 2014
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Version 4, November 2014 A trial of quinidine in genetic epilepsy
Introduction
Epilepsy is defined by repeated, unprovoked seizures and is a major global health problem with a lifetime incidence of over 3% 1. Epilepsy is not, however, a uniform condition but rather a collection of syndromes with widely variable course, severity and underlying causes. Amongst these causes of epilepsy, inherited factors are prominent. The genetics of most inherited epilepsies is likely complex with multiple genes interacting with environmental factors to produce disease. An increasing number of monogenic epilepsies are however recognised. The majority of genes carrying epilepsy- causing mutations are neuronal ion channel subunits, thus leading to effects on either synaptic transmission or action potential firing.
At present, the vast majority of epilepsy therapies take little account of the underlying causes. Anti- epileptic drugs are largely developed and tested in broad cohorts of epilepsy with mixed aetiologies. This has led to drugs each individually usable in most patients but with modest efficacy at best. Tailored drug therapies are starting to emerge for genetic epilepsies. For instance, in tuberous sclerosis complex, genetic abnormalities of the functional cascade associated with Mammalian Target of Rapamycin (mTOR) protein lead to the disease. Treatment with a specific inhibitor of this pathway, everolimus, leads to improved outcomes both in terms of seizures and secondary development of tumours 2. Another example is the use of the orphan drug stiripentol in Dravet Syndrome, a catastrophic epilepsy of early life associated with usually de-novo mutations of the neuronal sodium-channel sub-unit gene SCN1A3, 4.
Here we propose a small trial to test a tailored drug therapy in a recently discovered genetic epilepsy, utilising the potassium-channel antagonist quinidine to treat a genetic disease caused by over-active potassium-channels.
KCNT1 as an epilepsy gene The gene KCNT1 codes for a potassium channel, variously known as KCNT1, Slo2, SLACK and KCa4.1, expressed mainly in the central and peripheral nervous systems 5, 6. The protein KCNT1, often in hetero-dimers with a related protein KCNT2 (or SLICK), is involved in adjusting the frequency and duration of the firing of neurons in response to a stimulus 7-9. KCNT1 also appears to interact with the Fragile X Mental Retardation Protein (FMRP), the critical protein affected to produce the common Fragile-X mental retardation syndrome 10. KCNT1 and FMRP appear to enhance each other’s function with the loss of FMRP reducing the KCNT1 currents recorded and activation of KCNT1 showing evidence of enhancing the RNA binding and regulation role of FMRP.
Recent genetic findings have demonstrated KCNT1 mutations to be a cause of inherited epilepsy. Our group reported in 2012 mutations in KCNT1 in three families with the inherited epilepsy syndrome autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE)11. In this dominantly
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Version 4, November 2014 A trial of quinidine in genetic epilepsy inherited syndrome frequent, often brief, focal seizures occur in sleep at a rate of multiple attacks per night 12. Although seizures in this syndrome are often refractory to treatment, the MRI is normal and intellect is usually preserved. ADNFLE was the first inherited epilepsy for which a gene was conclusively identified, the acetyl choline receptor subunit CHRNA4 13. Subsequently mutations in two other acetyl-choline receptor subunit genes were also identified, CHRNA2 and CHRNA2B 14-16. The families with KCNT1 mutations are notable for more severe phenotypes than individuals carrying acetyl-choline receptor gene mutations. In the 15 reported KCNT1 mutation carriers, penetrance was higher (100% vs. 70%), seizure onset earlier (median 6yrs rather than 10yrs), and additional neuropsychiatric disability was common with 6/15 having intellectual disability and a further 4/12 having psychiatric co-morbidity.
In the same issue of the journal, a French group reported de-novo KCNT1 mutations in six patients of the twelve tested with the severe, early-onset epileptic encephalopathy of malignant migrating partial seizures of infancy (MMPSI) 17. MMPSI is characterised by the onset under 12 months of age of focal seizures that are continuous or near continuous in an infant without significant MRI abnormality 18. The foci for these seizures shift over time, migrating across both hemispheres, leading to a long-term status epilepticus that on EEG moves across the brain. The outcome is extremely poor with developmental regression, refractory seizures, mortality under 10 years in up to half of cases and, in those that survive, severe intellectual disability18, 19. The reported mutations in KCNT1 all appeared to be causative as they were de novo, affected conserved amino acids and led to significant changes in the function of the mutant protein 17. All mutations led to significant increase in current; they are gain of function mutations.
Mutations in KCNT1 thus lead both to a severe form of focal epilepsy and a catastrophic epilepsy of early childhood. This appears to be due to excess function of the KCNT1 channel. A specific therapy would be of great utility, particularly in MMPSI where the outcome is extremely poor.
Quinidine Quinidine is a now uncommonly used Class 1a anti-arrhythmic drug. Derived from a Cincha alkaloid, quinidine leads to slowing of the upstroke of the cardiac action potential as well as prolongation of the action potential duration and effective refractory period. These actions led to its’ use in re- entrant, particularly atrial tachycardias 20, 21. The cardiac action potential duration is the determinant of the ECG measure of QT interval, hence quinidine produces predictable prolongation of the QT interval with the consequent risk of R on T and induction of the ventricular tachycardia Torsade de Pointe 20-23. This prolongation of the QT has led to the main modern application of quinidine, the treatment of genetic short-QT syndromes such as Brugada Syndrome 24.
Quinidine and potassium channels Although the original conception of Class 1 antiarrythmic agents was as Na-channel blockers, in fact their action is likely due to blockade of a wide variety of K-channels 25. The cardiac action potential, unlike neuronal action potentials, has a pronged plateau of depolarization following the initial sodium channel dependent upstroke. This leads to sustained muscle contraction and determines the length of the QT interval. This plateau phase is caused by the opposition of depolarising calcium and Page 3 of 14
Version 4, November 2014 A trial of quinidine in genetic epilepsy to a lesser extent sodium currents and re-polarising (or rectifying) potassium currents. Gain-of- function mutations of the relevant K-channels increase this rectifying current leading to short-QT syndrome26. Quinidine acts to prolong the action potential by reducing these rectifying potassium currents 27.
Although quinidine has an effect on a variety of K-channels, it is a potent blocker of KCNT1 28. As mentioned above, recent work by our molecular collaborators has demonstrated that increased current is seen in all reported KCNT1 mutations when expressed in xenopus oocytes, with the increase greatest in those mutations associated with the most severe phenotypes29. This work also demonstrated that quinidine functioned as an effective blocker of those pathologically increased currents, reducing them to near-normal (figure 1). There is also at least suggestive, although not well controlled, animal evidence that quinidine reduces susceptibility to induced seizures in animal models overall 30. Thus quinidine is overall likely to be mildly anti-convulsant in effect with a potential to specifically reverse the effects of the faulty KCNT1 gene product.
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Figure 1
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Quinidine pharmacokinetics Quinidine is readily absorbed orally with a half-life in adults of 6-8 hours and 80-90% of blood quinidine being protein bound 22. Serum levels show good correlation with toxicity, particularly cardiac side effects. Excretion is predominantly via hepatic metabolism. Product information is attached.
For the use proposed, however, the critical question is the levels reached in brain. Quinidine is clearly measurable in cerebrospinal fluid in humans after a single dose 31. The kinetics of quinidine in the CNS are, however, complex. Efflux of quinidine from the brain is via the multi-drug transporter ATP-binding cassette sub-family B member 1 (ABCB1), also known as multidrug resistance protein 1 (MDR1) or P-glycoprotein 1 (P-gp) 32. Interestingly, quinidine is also regarded as an inhibitor of ABCB1, implying the drug inhibits its own clearance from the central nervous system 33, 34. There is some controversy regarding this inhibition so that even if it is occurring, the strength of inhibition appears small 34, 35. What is clear, however, is that the time-course of brain quinidine levels is different to serum. Microdialysis experiments in rats following a single dose suggest that the peak level is delayed compared to free serum level 32, 35. Also, the total area under the curve of drug level vs. time is 30-50% greater than in the serum. Accurate brain levels are not available during or following chronic dosing.
This has two implications for planning a trial of quinidine. Firstly, there is good evidence that quinidine reaches the brain at levels likely to be clinically relevant. Secondly, the clearance of the drug from the brain is delayed compared to serum concentrations.
Considerations for trial design Both epilepsy phenotypes associated with KCNT1 mutations are rare but potentially severe. Finding an effective treatment for KCNT1 disease would be still of great value to those with mutations, particularly those with the universally devastating, indeed often fatal, malignant migrating partial seizures of infancy. In order to be demonstrated as clinically useful, a treatment for conditions such as these needs to have a very large effect size. For the treatment to be valuable, it needs in essence to be a specific remedy. After all, if effect cannot be demonstrated in the very small number of known cases, it cannot be demonstrated at all. In addition, quinidine has significant potential risks, particularly of cardiac effects.
For these reasons we propose an inpatient, placebo controlled, cross-over trial of quinidine in participants with ADNFLE due to KCNT1 mutations. Seizures are usually very frequent, daily or more, in those with active frontal lobe epilepsy. This means a standard, ten-day admission for VEM is likely to capture enough seizures for meaningful analysis. Utilizing video-epilepsy monitoring with constant EEG allows very accurate assessment of seizure frequency both with treatment and placebo. In addition, the inpatient setting allows the closest possible monitoring for adverse events, particularly cardiac events.
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The issue with a cross-over trial is the pharmacokinetics of quinidine. The serum half-life of 6-8 hours suggests around two days is required at the end of the admission for wash-out prior to ceasing cardiac monitoring. Also, at least 24 hours will be required to reach useful serum levels of quinidine. Wash-out from the brain is likely to be delayed. The magnitude of this delay is neither well documented nor easily measurable. To be double-blind, the order of drug and placebo in the cross- over treatment needs to be randomized. The risk is thus that if active drug precedes placebo the medication may not have cleared from the brain, despite being undetectable in serum. Overall, however, this effect is likely to lead to under-estimate of quinidine effect while a single-blind or unblinded study opens the chance for over-estimation. We have elected to maintain a double-blind design, with an additional day between treatments to maximize time for CNS wash-out.
Methods A double blind, placebo-controlled, crossover trial of quinidine in autosomal dominant frontal lobe epilepsy due to KCNT-1 mutations will be performed. The primary end-point is difference in seizure frequency with quinidine versus placebo. We hypothesis that quinidine will significantly reduce seizure frequency compared to placebo.
Participants Patients known to have autosomal dominant nocturnal frontal lobe epilepsy due to KCNT1 mutations will be recruited. These patients will be identified from participants in the long –running Austin Health study, Genetics of Human Epilepsy. Patients must have active, uncontrolled epilepsy with ongoing seizures. All patients will have been part of the original discovery cohort for this genetic disorder and known to the investigating team. Participants will be excluded if they have contraindications to quinidine (table 1). It is expected that five people will be available to participate. The trial will, as far as possible, be continued until five participants with sufficient data for analysis have completed the trial.
Participants requiring third party consent will also be included. The person responsible will be asked to consent for children aged 12-18 and those with intellectual disability. Participants unable to consent for themselves will be included for a number of reasons. Firstly, this is a severe illness with intellectual impairment forming part of the disease. By excluding those unable to consent for themselves, the results of this trial may not be generalisable to those with more severe disease who most need a new therapy. Secondly, in order to recruit sufficient participants with active epilepsy, those requiring third-party consent will need to be included.
Procedures This will be an inpatient trial. Patients will be admitted for Video Epilepsy Monitoring (VEM) on ward 6E or 2E of Austin Health for a routine 10 day admission for VEM then a further 2 day stay of EEG monitoring for treatment wash-out phase; 12 days total. VEM involves 24-hour electroencephalogram (EEG) monitoring with simultaneous video monitoring. Usual anti-epileptic
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Version 4, November 2014 A trial of quinidine in genetic epilepsy drugs will not be altered. In addition to intensive nursing supervision, continuous electrocardiogram (ECG) monitoring will be undertaken. Prior to commencing treatment, a 12-lead ECG will be recorded with QT-interval measured and corrected for rate. Study drug or placebo will administered for 4 days each with a 2-day wash-out period between. The first day of drug or placebo administration will not be included for seizure analysis. This day will allow active drug to reach effective levels, or a total of 3-days wash-out before placebo treatment to allow drug to leave the CNS. Their usual epilepsy medicines will remain at the usual doses throughout. Study procedures are outlined in table 2.
Preliminary use in two participants has suggested that even the lower end of the cardiac or malaria treatment dose (900mg per day and 600mg per day) is not sufficiently well tolerated for use in epilepsy. QT interval changes were seen on day 3 of treatment leading to early cessation. No symptoms or arrhythmia arose from this but such changes still represent significant cardiac side effects. Based on this we have elected to move to low dose treatment. The dose will be 300mg per day in three divided dose. If any further QT alterations are seen, a further dose reduction to 150mg per day will be undertaken for subsequent participants.
Table 1 – Inclusion/exclusion criteria Inclusion criteria Exclusion criteria
Autosomal dominant nocturnal frontal lobe History of cardiac disease epilepsy with KCNT1 mutation including ischaemic heart disease, structural heart disease, arrhythmia or undiagnosed Ongoing seizures palpitations
QT-length abnormality on ECG
Liver failure or cirrhosis
History of quinidine allergy or drug-induced lupus History of immune thrombocytopaenia or thrombotic thrombocytopaenia History of myasthenia gravis
Current use (last 4 weeks) of other drugs known to prolong QT interval including Class I and III anti-arrhythmics and the anti-epileptic drugs phenytoin or felbamate
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Table 2 – Summary of study procedures Day of Treatment Daily monitoring Daily blood tests study phase
Video-epilepsy Urea and electrolytes Placebo monitoring Calcium, phosphate, ECG - monitoring magnesium 1-4 Or 12-lead ECG Liver function Quinidine Clinical review for FBE side effects Quinidine levels
Video-epilepsy Urea and electrolytes monitoring Calcium, phosphate, ECG - monitoring magnesium 5-6 Wash-out 12-lead ECG Liver function Clinical review for FBE side effects
Video-epilepsy Urea and electrolytes Quinidine monitoring Calcium, phosphate, ECG - monitoring magnesium 7-10 Or 12-lead ECG Liver function Placebo Clinical review for FBE side effects Quinidine levels
Urea and electrolytes ECG - monitoring Calcium, phosphate, 12-lead ECG magnesium 11-12 Wash-out Clinical review for Liver function side effects FBE
Day 1 Day 2-4 Day 5-6 Day 7 Day 8-10 Day 11-12 Treatment 1 Wash-out Treatment 2 Wash-out Seizure frequency Seizure frequency Page 9 of 14 measured measured
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Any prolongation of QTc beyond normal limits or documented arrhythmia will lead to withdrawal from the trial immediately. Cardiology opinion will be sought in the case of any significant ECG abnormality.
Abnormalities of blood electrolytes can predispose to cardiac arrhythmia. Urea and electrolytes (UEC) as well as calcium, phosphate and magnesium (CPM) will be measured on a daily basis and reviewed each day by the inpatient medical team. Any abnormality, particularly low magnesium or low potassium, will be corrected promptly by oral or intravenous therapy as appropriate. Quinidine has been reported to uncommonly lead to thrombocytopaenia, anaemia or self-limiting hepatitis. Liver function and haematological parameters will thus be monitored. Although measured daily, results for drug levels are not expected to be available in time to influence clinical care.
The elimination half-life of quinidine in adults is 6-8 hours. In order to maximize safety, participants will continue to be monitored for 48 hours (>5 half-lives) post cessation of active treatment in a wash-out phase. If quinidine is ceased due to adverse reaction or ECG abnormality, at least 48 hours of wash-out phase with full EEG, cardiac and blood test monitoring will be performed.
Clinical review for potential side effects will occur daily. As well as open ended questioning, directed questions for important side effects of quinidine will be asked as a check-list (table 3).
Table 3 –Side effect checklist
Palpitations Chest pain Cardiac Dyspnoea
Abdominal pain Nausea Gastrointestinal Anorexia Diarrhoea
Rash Dermatological Pruritis
Altered vision Headache Neurological Altered hearing Symptoms of cinchonism Vertigo
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Study medication Austin pharmacy will arrange for compounding of identical quinidine and placebo capsules. These will be dispensed to participants by the ward nursing staff as “study drug”.
Blood tests Blood will be collected via the routine inpatient methods. For routine tests a 9ml serum tube and 4ml EDTA tube will be collected per day. This is subject to usual hospital practice for these routine tests and may change. In addition morning, pre-dose levels of quinidine will be collected via a second, 9ml serum tube at the same time. A total of 22ml of blood is expected to be drawn per day on days 1-4 and 7-10 while 13 mls will be drawn on other days.
Outcome measures The primary end-point is difference in unequivocal focal seizure frequency with quinidine versus placebo. In ADNFLE seizures are usually very frequent with an average of multiple per day. Seizures will be assessed by VEM; the gold standard for seizure detection and diagnosis. Given the half-life of quinidine (6-8 hours) a stable therapeutic level is unlikely on the first day of treatment. Due to this, and to allow a full 3 day wash-out between treatments, only the final three of the four treatment days will be averaged to give the seizure frequency on medication; days 2-4 and days 7-10.
Seizures will be divided into paroxysmal arousals and unequivocal focal seizures. Paroxysmal arousals comprise short seizures (<20s) without EEG correlate, typically occurring many times per hour of sleep 36. Unequivocal focal seizures with prolonged dystonic movements (>20s) and EEG changes occur less frequently, typically in sleep but may also occur in wakefulness. Only this second type will be included in the primary analysis of seizure frequency.
Data will be analysed by paired t-test. The sample size is determined by the number of available participants rather than statistical considerations. The power is difficult to estimate as a reliable standard deviation for seizure frequency is not available. However, assuming the sample standard deviation is no more than 1/3 of the average seizure frequency for each person, 5 participants gives over an 80% power of detecting a halving of seizure frequency. Overall, for a trial such as this using a medication with significant side effects in a rare disorder, only a very large effect size would be of clinical relevance in any case, with 50% reduction being the absolute minimum of importance.
Secondary analyses will include rates of paroxysmal arousals, adverse event rates and tolerability, as well as 50% responder rate.
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Significance
This small but tightly controlled study, if positive, would give an initial signal of efficacy for quinidine in epilepsy due to KCNT1 mutations. This would be an important translational outcome of genetic research. Two open label trials would then be planned. Firstly the long term use of quinidine in ADNFLE at Austin Health. Secondly, and more importantly, an international trial of early intervention in infants with malignant migrating partial seizures of infancy would be set up. This second application holds the potential to prevent a catastrophic, often fatal epileptic encephalopathy. Even with narrow application, this is an important treatment goal.
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