STOPprogression

PROTECT&REGROWremaining,ssues

REPLACEdamagedgenes&,ssues

ACureForWolframThreeSteps3.0

FumiUrano,MD,PhDJuly16,2016

Lossoffunc,onofWFS1

CalciumLeakagefromtheendoplasmicre,culumtothecytosol(ERstress)

Neuro/re,naldegenera,onBetacelldeath

Uranolabref:Nature(2002);JBiolChem(2006);CellMetab(2006);JClinInvest(2010);ProcNatlAcadSciUSA(2014);ScienceSig(2015)

Mechanisms

Calpain2ac,va,on(enzyme)

ADRUGtarge,ngtheUPSTREAM

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Dantrolene(FDAapproved)

1.TargetsryanodinereceptorintheER

2.Spas,city(capsules)Adults25mgoncedailyforsevendays,then25mgt.i.d.forsevendays,50mgt.i.d.forsevendays,100mgt.i.d.Children0.5mg/kgoncedailyforsevendays,then0.5mg/kgt.i.d.forsevendays1mg/kgt.i.d.forsevendays,2mg/kgt.i.d.

3.Malignanthyperthermia(intravenous)1mg/kg-10mg/kgun<lsymptomssubside.

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Dantrolenepreventscalpainac,va,on(celldeathenzyme)inaWolfram-neurdegenera,onmousemodel

i.p.,q.d.,20mg/kg,4weeks(ForSpas,cityinchildren:6mg/kg/day)

Dantroleneinbetacell-specificWFS1KOmice(Wolfram-diabetesmousemodel)

Group1:10WFS1betacell-specificknockoutmalemice,8weeksofage,6mg/kgofdantrolene,I.P.Injec,onfor5daysperweekupto18weeks(10mice).Group2:10WFS1betacell-specificknockoutmalemice,8weeksofagewith0.9%saline(10mice).Group3:10Lidermatecontrolmalemice,8weeksofagewith0.9%saline(10mice).

0.0 50.0

100.0 150.0 200.0 250.0 300.0 350.0 400.0 450.0 500.0

0 5 15 30 60 90 120

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Time in minutes after 2g/Kg dextrose injection

WFS1 -/- + 0.9% saline

WFS1 -/- + 18 week dantrolene

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!WFS1(-/-) mice + 0.9% saline

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Control mice β-WFS1 -/- mice Tx: saline

β-WFS1 -/- mice Tx: dantrolene

Bright field microscopy image of islets from Wolfram mice treated with dantrolene

UranoF.Diabetes(2014)

InducedPluripotentStemCellsfromWolframpa,ents

DantrolenepreventsapoptosisinWolframiPSC-derivedneuronalcells

TREATMENT

Dantrolene

1.  PreclinicalstudiesintheWolframneurodegenera,onmousemodel2.Orphandrugdesigna,on(ApprovedbyFDA,2016)3.Safetystudiesinpa,entswithWolframsyndrome

AProofofConceptClinicalTrialClinicalTrials.govIden,fier:NCT02829268

•  Anopenlabelandascendingdosestudyinchildrenandadultpa,ents

•  Toassessthesafetyandtolerabilityofdantroleneadministeredorallyatupperendoftherapeu,cdoserangefor6months.

•  Primaryendpoint§  SafetyandTolerability(Liverenzymes)

•  Secondaryendpoints§  Betacellfunc,on(C-pep,de,mixedmealchallengetest)§  Balancetest§  Visualacuity§  QOL,WURS,ERstress

! ! !

FIGURE: CLINICAL TRIAL DIAGRAM

Screening Visit 1 Dose Escalation Study Visits 2-5; Wks 0-3 Treatment Assessment Study Visit 6, Month1 Study Visits 7, 8; Months 2, 4 Study Visit 9 Month 6 Study Visit 10 Final Follow up

• Obtain informed consent & assent (age appropriate forms) • Screen subjects by criteria; medical history, Genetic testing, physical exam (PE), ECG,

safety labs, pregnancy test • Baseline beta cell functions, ophthalmology & neurological assessments, baseline lab

assessments • myelin basic protein and MANF assays (ELISA)

Administer Study med per dose escalation guidelines; Complete physical exam (PE), VS/Wt, and liver enzyme testing (ALT, AST, alkaline phosphatase) at each visit; adverse event monitoring; Evaluative labs visit 4, wk 2 (HbA1c, C-peptide (C-P), Proinsulin/C-peptide, blood glucose levels, serum osmolality (osm), Phosphorus (phos), Ammonia (Am), UA (micro/macro, glucose)

Continue Study med or placebo at determined dose. Adverse event monitoring. Quality of Life (QOL) questionnaire, Review medication diary, Complete PE, VS/Wt, pregnancy test, and Safelty labs: liver enzyme testing (ALT, AST, alkaline phosphatase), Eval labs: C-P, Proinsulin/C-peptide, blood glucose levels, osm, phos, Am, UA (micro/macro, glucose), myelin basic protein and MANF assays (ELISA)

Continue Study med or placebo at determined dose. Adverse event monitoring. Quality of Life questionnaire, Review medication diary, complete PE, VS/Wt, pregnancy test, Safety labs, myelin basic protein and MANF assays (ELISA), Month 4: Eval labs: C-P Proinsulin/C-peptide, blood glucose levels, osm, phos, Am, UA(micro/macro, glucose)!!

24 eligible subjects (12 adults and 12 children) Dantrolene PO

!

Safety labs, QOL, adverse event monitoring, complete PE, VS/Wt

Assessment of Final Study Outcome Measures Discontinue study med or placebo; Complete PE, VS/Wt, all safety labs and evaluative labs: C-P Proinsulin/C-peptide, blood glucose levels, osm, phos, Am, UA(micro/macro, glucose); ECG, beta cell functions, neurological and ophthalmology exam, QOL, myelin basic protein and MANF assays (ELISA)

!

Adultpa,entsVisit#2:25mgoncedailyforsevendays,thenVisit#3:25mgt.i.d.forsevendaysVisit#4:50mgt.i.d.forsevendaysVisit#5:100mgt.i.d.Pediatricpa,entsVisit#2:0.5mg/kgoncedailyforsevendays,thenVisit#3:0.5mg/kgt.i.d.forsevendaysVisit#4:1mg/kgt.i.d.forsevendaysVisit#5:2mg/kgt.i.d.

Doseescala,oninthefirstfourweeks

Screening

•  Dura,on:6-9months(Screening,Doseescala,on,6monthtreatment,andfollow-up)

•  24pa,ents(12adults,12children)•  Primaryendpoint

§  Safety(liverenzymes,labtest,physicalexams)§  Tolerability(Doseescala,onover4weeks)

•  Secondaryendpoints§  Betacellfunc,on(baselineCpep,de,Mixedmeal

boosttest)§  Balancetest(Mini-BESTest)§  Visualacuity§  QOL,WURS,Serumbiomarkers(MANF,MBP)

An,cipatedTimeline

•  February 2016:OrphanDrugDesigna,onApproved bytheUSFDA

•  June 2016: A clinical trial protocol submided to theWashUIns,tu,onalReviewBoard(IRB)

----------•  Fall2016:IRBapproval•  Winter2016:ScreeningandRecruitmentofPa<ents•  Spring-Summer2017:Commencementofthetrial----------Funding:SnowFounda,onandEllieWhiteFounda,onNa<onalIns<tutesofHealth(pending)PrivateFounda<ons,suchasJDRF(pending)

Stoptheprogression

FDA-approveddrugs

Noveldrugs

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TheTherapeu,csforRareandNeglectedDiseases(TRND)programatNCATS

•  Aims to encourage and speed the development of new treatments for rare and neglected diseases.

•  Optimization and pre-clinical testing of therapies, with the goal to generate sufficient-quality data to support successful Investigational New Drug applications to the Food and Drug Administration and first-in-human studies in limited circumstances.

NCGC00384450-01preventsWFS1-deple,on-mediatedbetacelldeath

NewDrugCandidatesdesignedforthetreatment

ofWolframsyndrome

1.Medicinalchemistry2.Pharmacokine,csandtoxicitystudies3.PreclinicalstudiesincellandanimalmodelsfortheInves,ga,onalNewDrug(IND)applica,on

An,cipatedTimeline

•  2014-2015:Screeningofdrugs•  2015:Tes,ngincellmodelsofWolfram•  2015-2016:Iden,fica,onoftwoleaddrugcandidates•  2016:ToxicityandPKstudiesoftwodrugcandidates----------•  2016-2017: Tes<ng of two lead drug candidates in

mousemodels•  2016-2017: Further modifica<on of two lead drug

candidates•  2017-2018: Toxicity and PK studies of modified

candidates•  2018-2020:Furtheranimalstudiesandhumanstudies

STOPprogression

PROTECT&REGROWremaining,ssues

REPLACEdamagedgenes&,ssues

ACureForWolframThreeSteps3.0

REGENERATIONFACTORSProducedinourbody

MANF:Regenera,onFactor

Producedinourbody

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An,cipatedTimeline

•  2016-2018: Genera,on of MANF-like drugs (receptormodulators)

----------•  2018-2020:Preclinicalstudiesincellandanimalmodels•  2020-:PreclinicaltoFirst-in-HumanStudies

STOPprogression

PROTECT&REGROWremaining,ssues

REPLACEdamagedgenes&,ssues

ACureForWolframThreeSteps3.0

Replace disease-causing DNA with the healthy one (CRISPR/CAS9)

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BrainCellsandEyecellsfromskincells

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An,cipatedTimeline

•  2016-2020:Earlystagepreclinicalstudies----------•  A^er2021:PreclinicaltoFirst-in-HumanStudies

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