STEM CELL THERAPY FOR CYSTIC FIBROSIS, research updates.pdf

Cystic fibrosis (CF) is a genetic disease that causes the body to produce thick, sticky mucus that builds up in the lungs and pancreas.

1. STEM CELL THERAPY FOR
CYSTIC FIBROSIS
SOUSAN
A274153121006
HG&MM

2. CYSTIC FIBROSIS
➢Anautosomalrecessivegeneticdisorder.
➢MutationsintheCysticFibrosisTransmembraneconductanceRegulator
(CFTR)geneencodeaproteinexpressedintheapicalmembraneof
exocrineepithelialcells.
➢TheCFTRgeneisfoundonthelong(q)armofhumanchromosome7.
➢Affectsmultipleorgans,butprogressiveremodelingoftheairways,
mucusaccumulation,andchronicinflammationinthelungresultinlung
diseaseastheprimarycauseofmorbidityandmortality.

3. CLASSIFICATION OF CYSTIC FIBROSIS
There are over 2000 different mutations in the CFTR gene that
can cause disease. These mutations are divided into five
classes:
i.Defective protein synthesis
ii.Defective protein processing
iii.Disordered regulation
iv.Defective chloride conductance
v.Accelerated channel turnover

4. TREATMENT FOR CYSTIC FIBROSIS
Thereisnocureforcysticfibrosis,buttreatmentcanease
symptoms,reducecomplications,andimprovequalityoflife.
Thegoalsoftreatmentinclude:
➢Preventingandcontrollinginfectionsthatoccurinthelungs
➢Removingandlooseningmucusfromthelungs
➢Treatingandpreventingintestinalblockage
➢Providingadequatenutrition

5. STEMCELLTHERAPYFORCYSTICFIBROSIS
➢Ifastemcelliscorrectedwithgeneeditingorgenetherapy,
everycellproducedbythestemcellwouldalsohavethe
correctgene.
➢Scientistsareexperimentingwithcellsknownasinduced
pluripotentstem(iPS)cells.iPScellscanbemadeina
laboratoryfromskin,blood,orothercellsfromanyperson
usingaprocesscalledreprogramming.
➢Inthefuture,itmightbepossibletore-implantthecorrected
iPScellsintothelungsofpeoplewithCFtomakehealthy
lungcellsorothercelltypeswithfunctionalCFTRprotein.

6. INTRODUCTION
➢TargetingdefectiveCFTRfunctioninepithelialcellsofthesmallestairways.
➢RatherthanfixingCFTR,cellreplacementtherapyaimstoreplacedefective
epithelialcells.
➢Potentialbenefitsincludeimprovedchloridetransportandreduced
inflammation.
➢Selectivetargetingofintactbutgeneticallydefectiveepitheliumforremoval.
➢ReplacementwithprogenitorcellscontainingcorrectedCFTR.
➢Analogoustohematopoieticstemcelltransplantationcreates"space"for
healthycells.

7. OVERVIEW OF CELL TYPES USED IN STEM CELL-BASED
THERAPEUTIC APPROACHES
➢Lung comprises 40+ distinct cell populations: epithelial, inflammatory, stromal, and endothelial.
➢Epithelial cells include alveolar and airway types like ciliated, mucous, Club, basal, and PNEC cells.
➢Low cell turnover in the lungs makes identifying stem cells difficult.
➢Techniques like cell proliferation assessment and lineage tagging aid in identifying
stem/progenitor cell niches.
➢Generating therapeutically applicable cell numbers from endogenous stem cells remains
challenging.
➢Lung regenerative studies often use bone marrow-derived MSCs.
➢MSC engraftment shows therapeutic effects in lung injury models.
➢Pluripotent cells are ideal for lung regeneration due to lung complexity.
➢ES and iPScells explored for generating lung epithelium.
➢Recent advances in directed differentiation show potential for ES/iPScells in lung injury
treatment.

9. Mesenchymal Stromal Cells (MSC)
➢Studiesindicatethatmesenchymalstemcells(MSCs)secretevarious
extracellularvesicles(EVs)-Secretomesthatcontributesignificantlytotheir
therapeuticeffects,includinganti-inflammatoryproperties.
➢RecentreportssuggestthatEVsreleasedbyMSCscouldbeapromising
therapeuticapproachforcontrollinginflammationinCF.ResearchontheIB3-
1CFcellline,aninvitromodelofCF,showedthatMSC-derivedEVs
downregulatedpro-inflammatorycytokinesandupregulatedanti-inflammatory
mechanisms.
➢TreatmentwithMSC-derivedEVsresultedinthedownregulationofpro-
inflammatorycytokinessuchasIL-1β,IL-8,andIL-6andtheupregulationof
PPARγmRNAexpression,atranscriptionfactorinvolvedinanti-inflammatory
andantioxidantmechanisms.ThiswasaccompaniedbyreducedNF-kBnuclear
translocationandincreasedHO-1expression,indicatingsuppressionofthe
inflammationcascade.

10. CLINICAL TRIALS
➢TherearecurrentlytwoongoingphaseIclinicaltrials(NCT02866721and
NCT03058068)investigatingMSCtherapyinCF.TheCEASE-CFtrialaims
toevaluatethesafetyandtolerabilityofallogeneichMSCinfusioninadults
withCF.Incontrast,theHAPItrialfocusesonassessingsafetyand
exploringpotentialimprovementsinCFsymptoms,includinglungfunction
andqualityoflife.
➢Whilepreliminaryresultsfrompreclinicalandsmallanimalstudiesare
promising,largerclinicaltrialsarenecessarytoassessthesafety,efficacy,
andlong-termbenefitsofMSCtherapyinCFpatients.Thesetrialswill
providevaluableinsightsintothefeasibilityandpotentialofMSC-based
treatmentsforCF.

12. TRIAL 1
➢Prospective,single-centerstudywith15clinicallystableCFpatients≥18yearsold.
➢ScreeningperiodoftwotosixweeksfollowedbyaBaselinevisitforintravenousinfusionof
upto5x10E6allogeneichMSCs/kg.
➢InfusionsperformedatUniversityHospitalsClevelandMedicalCenter'sDahmsClinical
ResearchUnit.
➢Monitoringforinfusion-relatedtoxicitiesfor24hourspost-infusion;subsequentvisitsat
Days7,14,28,Months3and6,withphonecallsonDays4(or5),21,56,andMonth12.
➢Evaluationofsafetyandtolerabilitythroughsubjectdiaries,history,physicalexams,
spirometry,andsafetylabs.
➢Focusondetectingpulmonaryexacerbations;explorationofinflammatorybiomarkersin
bloodandsputum.
➢SerumandsputummarkersdeterminedatBaseline,Days7and28.
➢Allsubjectsamplesarearchivedforfutureuse;anoptionaldiagnosticbonemarrowexamis
offeredfortranslationalstudies.

13. TRIAL 2
➢Primary objective: Demonstrate safety of intravenously administered
Mesenchymal Stem Cells (MSCs) in subjects with cystic fibrosis.
➢Secondary objective: Explore if MSCs can improve CF symptoms, including
lung function, pulmonary exacerbation rate, inflammation, and quality of life.
➢Safety Run-In: 3 subjects receive a single MSC infusion to evaluate safety.
➢Randomized phase: 15 subjects in 3 cohorts (1:1:1 ratio) receive infusions.
➢Total duration: 12 months post-infusion, with up to 2 additional months for
Screening and Baseline Visits.
➢Approximately nine total visits for each subject.

14. Induced Progenitor-Like Cells (iPL)
➢Transient reprogramming with the transcription factors Oct4, Sox2, Klf4, and c-Myc(OSKM) led
to the generation of iPLcells.
➢iPLcells are highly proliferative and possess epigenetic memory, allowing them to revert to
their original identity upon withdrawal of reprogramming factors. This characteristic
distinguishes them from fully reprogrammed iPScells.
➢Highly purified populations of Club cells from double transgenic mice were isolated, enabling
the expression of OSKM transcription factors. Controlled, transient activation of these factors
with doxycycline resulted in reprogramming towards iPScells without reaching the
commitment point to pluripotency.
➢iPLcells were derived from various cell populations, including Club cells from proximal
airways, alveolar type II epithelial cells (AEC-II) from distal lung parenchyma, and pulmonary
endothelial cells.
➢iPLcells derived from Club cells could significantly expand (up to 30-fold) while retaining the
ability to differentiate into ciliated cells with functional expression of CFTR, which is crucial in
cystic fibrosis (CF) therapy.
➢In animal models deficient in CFTR, injected iPLcells successfully integrated into the airways
and differentiated into both Club and ciliated cells, indicating their potential therapeutic
efficacy in CysticFibrosis.

16. Embryonic Stem Cells
(ESCs)
➢Derived from the inner cell mass of early-
stage embryos.
➢Pluripotent nature allows them to
differentiate into any cell type in the body.
➢Ethical concerns due to the destruction of
embryos for their derivation.
➢Used in research for studying development,
disease modeling, and regenerative
medicine.
➢Subject to strict regulations due to ethical
considerations.
Induced Pluripotent Stem
Cells (iPSCells)
➢Reprogrammed from adult somatic cells
using specific transcription factors.
➢Possess pluripotent properties similar to
embryonic stem cells.
➢Ethically less controversial than ESCs as
they do not involve embryo destruction.
➢Potential for personalized medicine as
they can be derived from a patient's cells.
➢Applications include disease modeling and
regenerative medicine.

18. Airway Epithelial Cells (AEC)
➢Airwayepithelialcellsconsistofvariouscelltypes,includingbasalcells,columnar
epithelialcells,glandmyoepithelialcells,andmulti-ciliatedcells.
➢Thesecellsareinvolvedinprocessessuchasmucociliaryclearance,barrierfunction,and
immuneresponsesintheairways.
➢Cysticfibrosistransmembraneconductanceregulator(CFTR)proteinisexpressedin
submucosalglands,multi-ciliatedcells,andpulmonaryionocytes.
➢Columnarepithelialcellsfrompseudostratifiedregionsdemonstrateplasticityby
dedifferentiatingintobasalcells.
➢Thelungcanregeneraterapidlyafterinjury,withstemcellsplayingacrucialroleinrepair
andregeneration.
➢Airwayepithelialcells,particularlyhumanbronchialandbronchiolarepithelialcells
(HBECs),areconsideredascandidatesforcell-basedtherapyincysticfibrosis.

20. Designer Pluripotent Cells
➢Designerpluripotentcellsofferapromisingavenuetoovercomethelimitationsofhuman
inducedpluripotentstem(iPS)cellsfortherapeuticapplications
➢Advancesingenomeeditingtechnologies,suchastheCRISPR/Cassystem,haveenabled
targetedintegrationoffunctionalDNAelementsintothehumangenome,expandingthe
researchandtherapeuticpotentialofiPScells
➢Oneofthemajorchallengesinusinghumanpluripotentstemcells(hPSC)istheir
tumorigenicpotential,oftenassessedbyteratomaformation.Variousstrategieshavebeen
exploredtomitigatethisrisk,includingchemicalablation,targetingpluripotent-specific
antigenswithcytotoxicantibodies,geneticmodificationoftumor-drivinggenes,insertionof
suicidegenes,andselectiveeliminationusingsmallmolecules
➢Liangetal.(2018)introducedanovelcelltherapysafetysolution,thesafe-cell(SF)suicide
system.Thissystemlinksadrug-induciblesuicidesystem,HerpesSimplexVirustype1
thymidinekinase(HSV-TK),totheCDK1celldivisionessentialloci,ensuringsafetyby
enablingtheeliminationofpotentiallytumorigeniccells.

22. MECHANISMS OF ACTION
1.DifferentiationintoFunctionalLungEpithelialCells:Stemcellscanbedirectedto
differentiateintofunctionallungepithelialcells,includingbronchialandalveolarepithelial
cells,affectedinCF.Thesenewlygeneratedcellscanpotentiallyreplacedamagedor
dysfunctionalcells,restoringnormalepithelialfunctionandimprovingrespiratory
outcomes.
2.ModulationofInflammationandImmuneResponses:CFischaracterizedbychronic
inflammationintheairways,drivenbytheaccumulationofthickmucusandrecurrent
infections.Stemcells,particularlyMSCs,possessimmunomodulatorypropertiesandcan
modulateinflammatoryresponses,reducingtissuedamageandpromotingtheresolutionof
inflammation.Thisanti-inflammatoryeffectcanhelpalleviatesymptomsandprevent
diseaseprogressioninCFpatients.
3.DeliveryofFunctionalCFTRGenes:Genetherapyapproachesinvolvingstemcellsaimto
deliverfunctionalCFTRgenestoaffectedcells,restoringnormalCFTRproteinexpression
andfunction.Thisapproachholdspromiseforcorrectingtheunderlyinggeneticdefectin
CFandofferingapotentialcureforthedisease.However,challengessuchasefficientgene
delivery,long-termgeneexpression,andimmuneresponsemustbeaddressedfor
successfulclinicaltranslation.

23. Identification of Suitable Regenerative Cells
With Differentiation Capacity
➢Mesenchymalstemcells(MSCs)duetotheirimmunomodulatoryproperties,butconcernsexist
regardingtheirlowengraftmentlevelsinthelung.
➢MSCsfunctiontransientlytoreduceinflammationviathesecretionofextracellularvesicleslike
exosomes.
➢iPSCscandifferentiateintoairwayepithelialcells(AECs)orbasalstemcells.
➢iPSC-derivedAECculturesareheterogeneousandlacksufficientbasalcellsforlong-term
engraftment,limitingtheirefficacyinCFcelltherapy.
➢Inducedbasalcells(iBCs)differentiatedfromiPSCsshowpromisebuthavelimitations,including
theabsenceofrarecelltypeslikeCFTR-expressingmonocytesandbrushcells.
➢VerificationofiBCs'phenotypeanddifferentiationcapacity,bothinvitroandinvivo,is
necessarybeforeclinicaltranslationofiPSC-basedCFcelltherapy.
➢Basalcells(Heterogeneity)havebeenidentifiedastheairwayepithelium'sstemorprogenitor
cell.TheycandifferentiateintoCFTR-expressingionocytes.

25. ➢First-ever clinical trial of human mesenchymal stem cells (hMSCs) as a
potential treatment for people with cystic fibrosis (CF).
➢Asingle, allogeneic intravenous infusion of hMSCswas safe and well-
tolerated in 15 adult CF patients.
➢Claiming No dose-limiting toxicities, deaths, or life-threatening adverse
events were observed,
➢Human MSCs, agnostic as a therapeutic CFTR modulator
➢Data from pre-clinical studies suggest that allogeneic bone marrow-
derived hMSCsmay provide a new treatment for CF lung disease by
attenuating pulmonary inflammation while decreasing bacterial growth and
enhancing antibiotic efficacy.

26. Airway epithelial stem cell chimerism in cystic
fibrosis lung transplant recipients
➢Thestudyinvolvedtencysticfibrosislungtransplantrecipientswhounderwentflexible
bronchoscopywithairwaymucosalsampling.
➢Mucosalsamplingwasconductedusingaprotectedbronchialcytologybrushatpre-
determinedsamplingsitesinthebronchialairways.
➢ThestudyintroducedtheAirwayMucosalInjuryIndextoquantifyairwaymucosalinjury,
addressingagapinexistingscoringsystems.
➢Threepulmonologistsscoredimages,averagingthescorestoassessmucosalinjuryinthe
airways.
➢Imagesfrompatientsillustratedvaryingdegreesofmucosalinjury,includinghyperemic
airways,edema,submucosalhemorrhage,andsloughingofmucosaltissue.
➢Analysisofairwayepithelialstemcellchimerismshowedlessthan1%variationbetween
duplicatecultures,indicatingconsistentresults.

27. CHALLENGES
➢Difficultygeneratingsufficientnumbersofdifferentiatedairwayepithelial
cellsfromiPSCs.
➢Safetyconcernsregardingtumorigenicityduetogeneticinstabilityand
mutationrisk.
➢Non-integratingepisomal-basedmethodspreferredforderivingclinical-
gradeiPSCs.
➢Needforpurepopulationsoflungepithelialcells.
➢Ongoingresearchusingtechniqueslikesingle-cellRNAsequencingtoaidin
cellpurification.
➢Uncertaintyregardingoptimaldonorcellnumbersforeffectively
repopulatingdiseasedairwayepithelium.
➢Engraftmentremainsthefinalhurdleforsuccessfulcell-basedtherapyinCF.

28. Researchers discover disease-causing stem cells in
the lungs of cystic fibrosis patients
➢TwoexpertsfromtheUniversityofHouston,WaXianandFrankMcKeon,reportthatfivelung
stemcellvariantsdominatethelungsofpatientswithadvancedcysticfibrosis(CF),drivingkey
aspectsofCFpathologysuchasinflammation,fibrosis,andmucinsecretion.
➢CFTRmodulatordrugshaveshownremarkableimprovementsinthelungfunctionofCF
patientsbyrescuingfunctiontothemutantCFTRgene.However,lunginflammationpersistsin
patientswithestablishedlungdiseasedespitetreatmentwithCFTRmodulators.
➢XianandMcKeon'slaboratoryutilizedsingle-cellcloningtechnologytoidentifyfivestemcell
variantscommoninthelungsofadvancedCFpatients,threeofwhichexhibit
hyperinflammatorygeneexpressionprofilesanddriveneutrophilicinflammationupon
xenograftingtoimmunodeficientmice.
➢CFTR-modulatingdrugsdonotsuppresstheproinflammatoryactivityorgeneexpressionofthe
threeCFvariantsdrivinginflammation,raisingthepossibilitythattheseinflammatorystemcell
variantsarethesourceofpersistentinflammationinCFpatientstreatedwithCFTRmodulators.
➢ThesefindingshighlighttheimportanceofunderstandinglungstemcellheterogeneityinCF
pathologyandsuggestapotentialtargetforaddressingpersistentinflammationinCFpatients
despiteCFTRmodulatortreatment.

29. CONCLUSION
➢Stemcelltherapy,particularlyinducedpluripotentstemcells(iPS)andembryonicstemcells,
offerstheregenerativepotentialtoreplacedamagedordysfunctionallungepithelialcellsinCF
patients.
➢Thesestemcellscanbedifferentiatedintoproximalairwayepithelialcells,whichiscrucialfor
addressingtherespiratorycomplicationsassociatedwithCF.
➢Theuseofpatient-specificinducedpluripotentstemcellsallowsforpersonalizedtreatment
approachesinCF,potentiallycorrectinggeneticdefectsandrestoringnormalairwayepithelium.
➢AutologousiPScelltherapyinvolvesreprogrammingapatient'scellstogeneratecorrected
airwayepithelialcellsfortransplantation,reducingtheriskofimmunerejection.
➢Whilestemcelltherapyshowspromise,challengesremaininoptimizingdifferentiationprotocols,
ensuringcellpurityandyield,addressingsafetyconcerns,andovercomingtumorigenicrisks
associatedwithpluripotentstemcells.
➢Continuedresearcheffortsareneededtoadvancestemcell-basedtherapiesforCF,focusingon
enhancingtheseapproaches'efficacy,safety,andclinicalapplicability.

30. THANKYOU
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