ORIGINALARTICLE Apurifiedreconstitutedbilayermatrixshowsimproved outcomesintreatmentofnon-healingdiabeticfootulcers whencomparedtothestandardofcare:Finalresults andanalysisofaprospective,randomized,controlled, multi-centreclinicaltrial DavidG.Armstrong 1 |DennisP.Orgill 2 |RobertD.Galiano 3 | PaulM.Glat 4 |JarrodP.Kaufman 5 |MarissaJ.Carter 6 | LawrenceA.DiDomenico 7 |CharlesM.Zelen 8 1 DivisionofSurgery,KeckSchoolofMedicine,UniversityofSouthernCalifornia,LosAngeles,California,USA 2 DivisionofPlasticSurgery,BrighamandWomen'sHospital,Boston,Massachusetts,USA 3 DivisionofPlasticSurgery,FeinbergSchoolofMedicine,NorthwesternUniversity,Chicago,Illinois,USA 4 SurgeryandPediatrics,DrexelUniversityCollegeofMedicine,St.Christopher'sHospitalforChildren,Philadelphia,Pennsylvania,USA 5 DepartmentofSurgery,TempleUniversitySchoolofMedicineandMcGowanInstituteforRegenerativeMedicine,UniversityofPittsburgh, Pittsburgh,Pennsylvania,USA 6 StrategicSolutions,Inc.,Bozeman,Montana,USA 7 LowerExtremityInstituteforResearchandTherapy,Youngstown,Ohio,USA 8 ProfessionalEducationandResearchInstitute,Roanoke,Virginia,USA Correspondence CharlesM.Zelen,ProfessionalEducation andResearchInstitute,Inc.222Walnut Avenue,Roanoke,VA24016,USA. Email:cmzelen@peridedu.com Fundinginformation GeistlichPharma,Grant/AwardNumber: 003 Abstract Astheincidenceofdiabeticfootulcers(DFU)increases,bettertreatmentsthat improvehealingshouldreducecomplicationsoftheseulcersincludinginfec- tionsandamputations.Weconductedarandomizedcontrolledtrialcomparing outcomesbetweenanovelpurifiedreconstitutedbilayermembrane(PRBM)to thestandardofcare(SOC)inthetreatmentofnon-healingDFUs.Thisstudy included105patientswhowererandomizedtoeitheroftwotreatmentgroups (n = 54PRBM;n = 51SOC)intheintenttotreat(ITT)groupand80who completedthestudyperprotocol(PP)(n = 47PRBM;n = 33SOC).Thepri- maryendpointwasthepercentageofwoundsclosedafter12weeks.Secondary outcomesincludedpercentareareduction,timetohealing,qualityoflife,and costtoclosure.TheDFUsthathadbeentreatedwithPRBMhealedatahigher ratethanthosetreatedwithSOC(ITT:83%vs.45%,p = 0.00004,PP:92% vs.67%,p = 0.005).WoundstreatedwithPRBMalsohealedsignificantlyfaster thanthosetreatedwithSOCwithameanof42versus62daysforSOC Received:3March2024Accepted:27March2024 DOI:10.1111/iwj.14882 ThisisanopenaccessarticleunderthetermsoftheCreativeCommonsAttribution-NonCommercial-NoDerivsLicense,whichpermitsuseanddistributioninany medium,providedtheoriginalworkisproperlycited,theuseisnon-commercialandnomodificationsoradaptationsaremade. ©2024TheAuthors.InternationalWoundJournalpublishedbyMedicalhelplines.comIncandJohnWiley&SonsLtd. IntWoundJ.2024;21:e14882.wileyonlinelibrary.com/journal/iwj1of13 https://doi.org/10.1111/iwj.14882(p = 0.00074)andachievedameanwoundareareductionwithin12weeksof 94%versus51%forSOC(p = 0.0023).Therewerenoadverseeventsorserious adverseeventsthatwererelatedtoeitherthePRBMortheSOC.Incomparison totheSOC,DFUshealedfasterwhentreatedwithPRBM.Thus,theuseofthis PRBMisaneffectiveoptionforthetreatmentofchronicDFUs. KEYWORDS advancedwoundcare,advancedwoundmatrix,diabeticfootulcers,standardofcare,wound healing KeyMessages • Patientswithnon-healingDFUsrandomizedtotreatmentwithanadvanced woundmatrix,purifiedreconstitutedbilayermembrane(PRBM),demon- stratedasignificantlyimprovedhealingrateovera12-weekperiod. • PRBMdemonstratedastatisticallysignificantimprovementinpercentarea reductionover12weeksversusstandardofcare(SOC) • PRBMshowedastatisticallysignificantdecreasedtimetohealingversusSOC 1|INTRODUCTION Itisestimatedthat9%oftheworldwidepopulationisdia- betic,andthisisexpectedtoaffectover600millionpeo- pleincomingyears. 1 Anotableconsequenceofthisis likelytobeanincreaseintheincidenceofdiabeticfoot ulcers(DFUs),whichcurrentlyaffect19% – 34%ofdia- betics. 2 Asmostclinicianscanattest,themanagementof DFUscanbechallenging,astheinterruptionofthenor- malhealingcascadecanresultin30%ofthesewounds becomingchronic. 3 Sadly,the5-yearmortalityforDFUs iscomparabletomanycancers 4,5 andpatientswhose DFUsprogresstoamputationhavea5-yearmortalityof 55%. 6,7 Withsuchasubstantialburdenonpatients,and thehealthcaresystemingeneral,thereisanobvious demandfortherapiesthatcanamelioratethehealing process,thusleadingtowoundclosureandpreventing thecomplicationsthatleadtotheworstofthemorbid- itiesandsubsequentprematuremortality. InlightoftheconsequencesthatDFUscanhaveon patients'lives,aswellastheprognosisfortheincreased cases,thereisaclearneedformethodsofsuccessfully treatingthesewounds.Thisisreflectedinthecurrentrec- ognition,bytheCentersforMedicareandMedicaidSer- vices,of76skinsubstitutesofvariedmaterialproperties forthetreatmentofDFUs. 8 Thesignificanceisfurther underscoredbytheongoingclinicalresearch,with Clinicaltrials.govcurrentlylisting838studiesforthe treatmentofDFUs. 9 Whetherthegraftsarederivedfrom allogeneic,xenogeneic,orsyntheticsources,theyallhave thesamegoalsofprovidingalocalenvironmentthatfos- tersthehealingofthewound.Whilemanydevices purporttoenhancehealing,theresultsofthepeer- reviewedpublicationspointtoevidencegapsandthere- forethereisacallforbetterinformationonwoundheal- ingandrecurrencewhenusingskinsubstituteproducts. 8 Withclinicalnecessityformodalitiesthatwill improvethechancesofsuccessfulhealing,theintroduc- tionofaPurifiedReconstitutedBilayerWoundMatrix (PRBM,GeistlichDerma-Gide ® ,GeistlichPharmaAG, Switzerland)mayoffercliniciansanotheroption.This bilayermatrix,whichhasbeenprocessedtoremove cells,lipids,antigensandinactivatepotentialviruses, hasa3-dimensionalstructurethatissimilartothe humandermis.Importantly,thePRBMmayimpactthe physiologicalprocessesandthewoundenvironment. ThegrowthfactorsTGF- β 1,bFGF,andVEGFareessen- tialconstituentsinwoundhealing, 10 whilematrixmetal- loproteases(MMP)canimpedehealing. 11 Theinvitro dataforthisPRBMhasshownthatthematerialallows thebindingandproliferationofgrowthfactorswhile alsoappearingtomodulateMMPactivity. 12 Withpromisinginvitroresults,apilotstudyin patientswithchronicDFUsreportedrapidhealingin 90%ofthepatients. 13 Whilethiswasasmallstudy (n = 10)itprovidedareasontopursuefurtherclinical researchonthisdevice.Therecentinterimresultsofthis Randomizedcontrolledtrial(RCT),inwhichthePRBM wascomparedtoacollagenalginatedressing(Fibracol; 3MMaplewood,MN),showedthatwoundstreatedwith thePRBMhealedsignificantlyfasterandweremore likelytoclose. 14 Herewepresentthesafetyandefficacy resultsofthefinalcompletedRCTinwhichthetargeted enrolmentofover100subjectswasmet. 2of13ARMSTRONG ETAL . 1742481x, 2024, 4, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/iwj.14882, Wiley Online Library on [23/04/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License2|METHODS Amulti-centre,prospective,parallel-group,RCTwas designedtoevaluatethetreatmentoffull-thickness,non- infectednon-ischaemic(WagnerGrade1/Universityof Texas1A)DFUswithPRBMorstandardofcare(SOC). ThestudywasregisteredontheGermanclinicaltrialreg- ister(DRKS)withareferencenumberofDRKS00016754 andapprovedbyWesternIRB(Protocol#20190130)and conductedincompliancewithGoodClinicalPractice standardsandwasinconformancewiththeethical guidelinesoftheDeclarationofHelsinki.Priortoany studyactivities,patientsprovidedwritten,informedcon- sent.Thetrialwasperformedatmultiplespecialtywound carecentresbetweenFebruary2019andApril2023.The protocolscheduleisshowninTable1. 2.1|TREATMENTS 2.1.1|Purifiedreconstitutedbilayer membrane(PRBM) Sterile,shelf-stablePRBM(Derma-Gide ® ;Geistlich PharmaAG,WolhusenSwitzerland)wasprovidedin individualdrypackaginginsizesfrom1.1to12cm 2 .The PRBMwasmanuallytrimmedtomatchthesizeofeach woundandplaceddrywiththeporouslowerlayerfacing down,directlyontothewoundbed,thusallowingfor uptakeofwoundfluids.IfthePRBMwasnotcompletely hydratedbywoundfluid,sterilesalinewasaddedin ordertoassurecompletehydration,allowingthegraftto conformtothewoundbedbeforetheapplicationofa routinedressing. 2.1.2|Standardofcare(SOC) Amoisture-retentive,conformablecollagenalginate dressing(FIBRACOLPlusDressing;3MMaplewood, MN)wastheprimarywounddressingintheSOCstudy arm.Thisdressinghasbeenrigorouslytestedwithfavour- ableresultsandwaschosenasawell-known,clinically acceptedSOCproductreadilyavailableinwoundclinics. 2.1.3|Studyendpoints Theprimarystudyendpointwasacomparisonofthepro- portionofindexulcershealedat12weeks.Secondary endpointsincludedcomparisonsoftimetohealat6and 12weeks,percentageareareduction(PAR)ofthewound at6and12weeks,patient-reportedqualityoflifeout- comes,safetyandcosttoclosure. 2.1.4|Patientscreening Afterobtaininginformedconsent,participantswere screenedovera14-dayrun-inperiodtodetermineeligi- bilityaccordingtoinclusionandexclusioncriteria (Table2).Therun-inprecededrandomizationtoelimi- natethosepatientsinwhomashortcourseofroutine therapywoulddemonstrateeffectivenessasmeasuredby TABLE1Thestudyprotocolschedule. Weeks1 – 2screeningphase • InformedConsent,inclusion/exclusioncriteriaassessment • MedicalHistory&Physical,VitalSigns&Labs • Assessmentofdiabeticwounds;DFUhistory • Assessmentofcurrentwoundtherapies • 10-pointmonofilamenttest • X-ray • ABI,SPP,TCOMTBImeasurementorarterialDopplerstudy • PatientcompletesWound-QoLandpainassessment. • Selectionofindexulcer;measurementofsurfacearea& digitalimaging • IndexUlcerAssessmentofexudateandinfection • TreatmentofindexulcerwithSOCprotocol • Woundimprovementover14days • Confirmeligibilitytocontinueenrolmentintostudy Weeks3 – 14treatmentphase • Medicalhistory&Physical,VitalSigns&BloodSugar • Assessmentofanyadverseevents • Pain&NeuropathyAssessments • IndexUlcerAssessment(exudateandinfection),Cleaning& Debridement • Measurementofsurfaceareaanddigitalimagingofindex ulcer • AssessmentofOffloading • Randomization(Week3) IfrandomizedtoSOC:ApplySOCtherapywithFibracol andouterdressing IfRandomizedtoPRBM:ApplySOCtherapywithPRBM andouterdressing • Weeklyassessmentofindexulcer,measurement,cleaning, debridementandrepeatdressings IfIndexulcerishealed,nofurthertreatment After6treatmentvisits,ifwound<50%healed,treatment phaseended;treatmentfailure Week15 – 16:Endofstudy/confirmationvisit • MedicalHistoryandPhysicalVitalsigns&Labs/Bloodsugar • Assessmentofanyadverseevents • Assessmentofoffloading • Assessmentofindexulcer(exudateandinfection);complete epithelializationorifwoundhasre-opened • Measurementofsurfaceareaanddigitalimagingofindex ulcer • Assessmentofinfection • Cleaning,debridementanddressingofindexulcerif applicable • PatientcompletesWound-QoLandpainassessment ARMSTRONG ETAL .3of13 1742481x, 2024, 4, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/iwj.14882, Wiley Online Library on [23/04/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons Licensea>20%reductioninwoundarea.Areviewofeach patient'smedicalhistoryandacompletephysicalexami- nationwereperformedincludingvisualassessmentofall footulcerswithattentiontosignsofinfection.Theindex ulcerwasselected,imaged,andmeasuredforareaand depth.Allwoundsweremanagedduringrun-inusinga standardprotocolincludingcleaning,appropriatesharp debridement,infectionmanagement,dressing,andoff- loadingusingadiabeticoffloadingbootorwhenthe patient'sfootcouldnotbeaccommodatedwiththeoff- loadingboot,atotalcontactcastwasused.Subjectswere instructedtokeepthewoundsitedryandinformedon theimportanceofoffloading.Theyreceivededucation oninfectionindicatorsandaskedtocontacttheclinic withconcerns.Patientscompletedthewoundquality-of life(Wound-QoL)questionnaireandscoredtheirpain intensityonascaleof0to10usingavisualanalogue scale(VAS).Afterfailingsufficientprogressduringthe2- weekrun-inperiod,subjecteligibilitywasreconfirmed, andalleligiblepatientsproceededtorandomization. 2.2|Randomizationandmeasuresto minimizebias SubjectswererandomizedtoeitherthePRBMarmor SOCarm.Toassureabalancedrandomization,envelopes werecreatedwitharandomallocationsequenceinblock sizesof10.Woundassessmentatconclusionoftreatment wasperformedbyaclinician,otherthantheinvestigator, whowasblindedtothetreatment.Additionally,confir- mationofwoundhealingwasoverseenbyanindepen- dentplasticsurgeonadjudicationcommittee. 2.3|Treatmentphase Regardlessofthestudyarm,woundsweremanagedwith acceptedroutineSOCpractices,includingweeklysharp debridementasindicated.PatientsrandomizedtoPRBM armweretreatedwithaPRBMgraftfollowedbyasili- conenon-adherentdressing(AdapticTouch,3MMaple- wood,MN)orequivalent,andthoserandomizedtoSOC armweretreatedwithcalciumalginatedressing(FIBRA- COLPlus3MMaplewood,MD).Allwoundsreceivedan outerdressingcomprisedofapadded3-layerdressing (Dynaflex,3MMaplewood,MN)orequivalent. Studyvisitswereperformedweeklyuntileithercom- pletehealingoftheindexulcerorfor12weeks,which- evercamefirst.Ateachvisit,thesubject'soverallhealth, glucosecontrol,andoffloadingwereassessed,andclo- sureoftheindexulcerwasgaugedbyablindedinvestiga- tor.Iftheindexulcerwasnotcompletelyreepithelialised, thewoundwasevaluatedforsignsofinfection,cleaned, imaged,andmeasured.Whentheindexulcerwas deemedtobe100%reepithelialised,furthertreatment ceased,andthepatientwasscheduledforawoundheal- ingconfirmationtovisit2weekslater.Finalwoundarea measurementandconclusiveimagingwereperformed, andpatientscompletedtheWound-QoLquestionnaireat TABLE2Inclusionandexclusioncriteria. InclusioncriteriaExclusioncriteria 1.Atleast18yearsold. 2.DFU,WagnerGrade1 3.Indexulcerhasbeen present ≥ 4weekspriorto screening,but ≤ 1year 4.Indexulcer ≥ 1.0and ≤ 25cm 2 5.Theindexulcerwillbe thelargestulceriftwoor moreDFUsarepresent withthesameWagner gradeandwillbetheonly oneevaluatedinthe study.Ifotherulcerations arepresentonthesame foot,theymustbemore than2cmdistantfrom theindexulcer. 6.Adequatecirculationto theaffectedfoot:TCOM orSPP ≥ 30mmHg,or ABIbetween0.7and1.3 within3monthsof screeningorabiphasic Dopplerultrasoundof dorsalispedisand posteriortibialvesselsat theleveloftheankleora TBIof>0.6. 7.Offloadingoftargetulcer ≥ 14daysbefore randomization. 8.Femalesofchildbearing potentialmustbewilling touseacceptable methodsofcontraception (birthcontrolpills, barriers,orabstinence) duringthecourseofthe studyandundergo pregnancytests. 9.Subjectunderstandsand iswillingtoparticipatein theclinicalstudyandcan complywithweekly visits. 1.Ulcer(s)deemedtobe causedbyconditionother thandiabetes. 2.Suspicionofmalignancy intheindexulcer 3.Patientswith ≥ 2weeksof treatmentwithimmune- suppressants,cytotoxic chemotherapy,or applicationoftopical steroids ≤ 1-monthpriorto screening 4.PatientstakingCOX-2 inhibitors. 5.Patientsoninvestigational drug(s)ortherapeutic device(s) ≤ 30days 6.Historyofradiationatthe ulcersite 7.Indexulcerhasbeen previouslytreatedorwill needtobetreatedwith anyprohibitedtherapies. 8.Osteomyelitisofthe affectedfoot ≤ 30days priortorandomization. 9.Subjectispregnantor breast-feeding. 10.Diabeteswithpoor metaboliccontrol(HbA1c >12.0)within90daysof randomization. 11.Patientswithendstage renaldisease,serum creatinine ≥ 3.0mg/dL within6months 12.Ulcerhasreducedinarea by ≥ 20%14daysofSOC fromSV1tothe TV1/randomizationvisit. 13.Inabilitytocompletethis studyorahistoryofpoor adherencewithmedical treatment 4of13ARMSTRONG ETAL . 1742481x, 2024, 4, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/iwj.14882, Wiley Online Library on [23/04/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons Licensethefinalvisit.Subjectswhoseindexulcerdidnotimprove by50%after6weeksweredesignatedperprotocolasa treatmentfailureandwereallowedtoreceivealternative treatmentsoutsideofthestudy.DetailsspecifictoPRBM includingtrimmedsizeandhandlingcharacteristicswere noted.Anyadverseevents(AEs),whichweretracked usingCTCAEv5,wereidentified,investigated,andman- agedasclinicallyappropriate.Suspectedwoundinfec- tionswerediagnosedviawoundswabs,andappropriate systemicantibioticswereprescribed.Topicalantibiotics werecontraindicatedperprotocol. 2.4|Samplesizeandstatisticalanalysis Accordingtotheaprioripoweranalysis,groupsample sizesof50ineachgroupwouldhaveachieved88%power todetectadifferencebetweenthegroups.Powercalcula- tionsfortheprimaryendpointwerebasedonatwo-sided Z-testwithpooledvariance.Statisticaltestingbetween treatmentgroupsatbaselinewascarriedouttoexamine thesuccessofrandomization.Forcategoricalvariables, chi-squareorFisherexacttestswereperformedandfor continuousvariablesindependentt-testsorMann – Whit- neytestswereused(dependingonvariablenormality)to testforstatisticaldifferences. TheprimaryendpointwasevaluatedbyaFisherexact testusinganintent-to-treat(ITT)analysisofallrandom- izedpatients.Timetohealwithin6and12weekswas analysedusingKaplan – Meierlog-ranktest,whilePARat 6and12weeksanalysedusingtheMann – Whitneytest. Theprimaryandsecondaryendpointsweretestedhierar- chicallyinaconfirmatorymanner.Otherendpoints assessedinanexploratorymannerincludedtheWound- QoLandpainscorechangesfrombaselineto12weeks. Statisticaltestsweretwo-sidedandperformedatasignifi- cancethresholdof0.05.Thelastobservationcarriedfor- ward(LOCF)principlewasusedinregardtomissing dataatstudyvisits.StatisticalAnalysiswasperformed usingSPSSStatistics27(IBM,Armonk,NY). 3|RESULTS Atotalof123patientswerescreenedand105wereran- domizedtotreatmentwitheitherPRBMorSOC(Fig- ure1).InthePRBMgroup,5subjectswerewithdrawnor losttofollow-up:1subjectwithdrewconsentand 1patient'sindexulcerhadaPAR<50%atweek 6.Although3peoplewereremovedduetoanAEor SAE,thesewerenotconsideredtoberelatedtothe PRBM.IntheSOCgroup,17patientswerewithdrawnor losttofollow-up.Atotalof14patientswerewithdrawn duetotheindexulcerpresentingaPAR<50%atweek 6while1patient'sulcerwasfoundtohavereopenedat thehealingconfirmationvisit.Inaddition,twopatients wereremovedduetoAEorSAE,butasseeninthe PRBMcohort,therewerenoinstancesofacausalrela- tionshipbetweentheAE/SAEandthetreatment. Thestudycohortwasrepresentativeofthediabetic populationattheinvestigationalsites,witharangeof comorbiditiesthatistypicaltopatientsbeingtreatedfor DFUs.Patientdemographicsandwoundcharacteristics werewell-balancedbetweenthetwoarms,andthedemo- graphiccharacteristicsatbaseline(showninTable3) showednosignificantdifferencesbetweenthegroups. Thiswasalsoseenforthewoundcharacteristicsatbase- line(Table4). 3.1|Outcomes WoundclosureratewassignificantlyhigherinthePRBM armthantheSOCarm.UsinganITTapproach,at 12weeksoftreatment,83%ofthepatientsinthePRBM arm(45/54DFUs)presentedwithhealedwounds,com- paredto45%intheSOCgroup(23/51DFUs).Thiswas significantatp = 0.00004.Whenthisendpointwasevalu- atedperprotocol(PP),92%ofpatients(43/47)inthe PRBMarmdemonstratedwoundclosurecomparedto 67%(22/33)intheSOCarm(p = 0.005). Thesecondaryendpointsalsoshowedthatulcers healedsignificantlyfasterforthosepatientsinthe PRBMgroup.FortheITTcohortthetimetoheal within12weekswasameanof42days(95%CI:35 – 49, median42days)amongPRBMpatientswhileintheSOC groupthiswasameanof62days(95%CI:55 – 70, median77days),whichwasstatisticallysignificant (p = 0.00074).TheKaplan – Meierplotofhealing,shown inFigure2,illustratesthisearlydivergenceinthehealing probabilitybetweenthegroups.Thisoutcomewasalso statisticallysignificantinthePPanalysis,withPRBM showingameantimetohealof40days(95%CI:32.1 – 46.8)whileSOCdemonstratedameanof55days(95% CI:44.6 – 64.4),whichwasasignificantdifference (p = 0.035). Analysisoftimetohealwithin6weeksfortheITT cohortalsoshowedasignificantdifferencebetweenstudy arms.ThePRBM-treatedwoundshealedinameanof 31days(95%CI:27 – 34)comparedwithamean of66.5days(95%CI:59 – 74)forwoundsintheSOCarm (p = 0.007).ThehealingrateinthePRBMarmatthe6- weekmid-studypointwas56%comparedwith29%inthe SOCarmandwasalsosignificant(p = 0.0098). ARMSTRONG ETAL .5of13 1742481x, 2024, 4, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/iwj.14882, Wiley Online Library on [23/04/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons LicenseThePARvaluesfortheITTcohortalsoshowedadra- maticdifferencebetweenthetreatmentarms.Atweek 6,thePRBMgrouphadameanPARof90.3(SD:17.42) whileintheSOCthiswas52.6(SD:60.14)(p = 0.00074). Likewise,theweek12meanvaluesforPARwere93.6 (SD:18.83)inthePRBMcohortcomparedto50.5(SD: 69.07)intheSOCcohort(p = 0.0023).Thedifference betweengroupsinthePARvaluesisshowninFigure3. WithregardtotheexploratorymeasuresofWound QoL,fortheITTcohortthepatientsinthePRBMcohort hadpresentedwithameanscoreof1.2(SD:0.90)at baseline,whichdecreasedto0.8(SD:0.73)attheendof study,andthiswassimilartotheSOCpatients,witha changefrom1.4(SD:1.08)to1.0(SD:0.91),thustherewas nosignificantdifference(p = 0.89).Similarly,thechange intheVASforpainfrombaselinetoendoftreatment, werequitecomparableforbothgroups:1.2(SD:2.26)at baselineand0.8(SD:1.67)attheendofstudyforPRBM versus1.3(SD:2.31)forSOCatbaselineand0.9(SD:1.96) attheendofstudy. Specifictosafety,therewere13SAEs,sixinthe PRBMgroup(fivepatients)andsevenintheSOCgroup FIGURE1CONSORTflowdiagram. 6of13ARMSTRONG ETAL . 1742481x, 2024, 4, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/iwj.14882, Wiley Online Library on [23/04/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License(sixpatients).ThesearepresentedinTable5.Therewere noinstancesofacausalrelationshipbetweentheSAE andeitherthePRBMortheSOC.NotallSAEsrequired removalfromtheclinicaltrial. RepresentativeCasesdocumentthehealingwiththe useofPRBMinFigure4. 4|DISCUSSION TheincreasingincidenceofDFUs,withitsattendant impactonclinicalloadandpatientwell-being,pointstoa needforeffectivetreatmentstrategies.Ourstudy,in which105patientswererandomizedtoeitherPRBMor SOC,demonstratedasignificantlyshortertimetohealing amongthosepatientswhohadbeentreatedwithPRBM, ascomparedtoSOC,andmirroredtheinterimresults. 14 Inadditiontotheproportionofwoundshealedat 12weeksinthePRBMcohortbeingclosetotwicethatof SOC(83%vs.45%),thetimetohealwithin12weekswas remarkablyshorterinthePRBMcohort,42daysas opposedto62inSOC,whilethePARvaluesat12weeks wasameanof93.6(SD:18.83)forthePRBMpatientsin contrastto50.5(SD:69.07)forSOC.Theseremarkable differencesinclinicaloutcomessupporttheuseofPRBM asaneffectiveandsafemodalityforthetreatment ofDFU. WiththeimpactthatDFUsexertonpatients,andthe treatmentdifficultiesasevidencedbyboththeriskof recurrenceandthe5-yearmortality, 4 thereisanacute requirementforeffectivetreatment.Theplethoraofprod- uctscurrentlyavailableandtheamountofpublisheddata pointtotheawarenessofthisdemand. 15 – 18 Despitethis, thetreatmentofDFUsremainsachallenge.Theseare difficultwoundstoheal,oftenbecomingchronicdueto thehealingprocessbeinginterruptedorstalled. 19 Wound TABLE3Keydemographic variablesatbaseline. VariablePRBMSOC p value Age62.5(12.01)63.8(11.03)0.57 BMI31.5(6.73)33.2(8.26)0.46 Gender Male31(57)32(63)0.58 Female23(43)19(37) Race/ethnicity Caucasian54(94)50(96)0.62 AfricanAmerican3(6)1(2) Hispanic0(0)1(2)0.49 Smoking Never23(48)30(59) Former19(35)17(33)0.33 Current9(17)4(8) Patientage,1stDFU54.7(12.8)58.0(10.8)0.16 Significantfootdeformities17(32)11(22)0.25 Charcot15(28)8(16)0.13 Hammertoes2(4)3(6)0.67 Bunions2(4)3(6)0.67 Other1(2)2(4)0.61 Amputation Studyfoot,minor19(35)10(20)0.07 Contralateralfoot,minor8(15)9(18)0.69 Contralateralfoot,major4(7)3(6)1.0 HbA1cscreening7.3(1.37) Median6.7(IQR1.5) 7.0(1.48) Median6.7(IQR1.5) 0.14 Creatinine1.2(0.52) Median1.1(IQR0.5) 1.1(0.33) Median1.0(IQR0.4) 0.13 ARMSTRONG ETAL .7of13 1742481x, 2024, 4, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/iwj.14882, Wiley Online Library on [23/04/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons Licensehealingitselfisacomplexphysiologicalphenomenon requiringaninterplayofphysiologicprocess, 10,20 which maybewhythehealingofchronicwoundscanbeunpre- dictable.Therefore,theinteractionofagraft'scharacter- isticsanditsphysiologicmechanismsareessential componentsofsuccessfulclinicaloutcomes. Inconsideringtheoverlappingphasesofwoundheal- ing,fromhaemostasistoinflammation,proliferation,and finallyremodelling,thereisacomplexanddynamic sequenceofcellularandbiochemicaleventsthatrestore skinintegrityandfunctionalityaftertrauma. 21,22 Among thefactorsthathavebeennotedtoimpactwoundhealing aretransforminggrowthfactor(TGF- β ),vascularendo- thelialgrowthfactor(VEGF)andfibroblastgrowthfactor (FGF). 10 IthasbeenreportedthatthePRBMthatweused inthetreatmentofthesesubjectsintheclinicaltrialhas, inaninvitroexperiment,boundthegrowthfactorsTGF- β 1,bFGFandVEGF. 12 Thenotablyimprovedhealing thatweobservedislikelyrelatedtothegraftsabilityto bindtheseendogenousgrowthfactors,whichareessen- tialcomponentsofwoundhealing. 23,24 Furtherwiththe invitrodatashowingthatthePRBMcanbindandpre- servetheactivityofTGF- β 1,bFGF,andVEGF, 12 itseems logicalthattheretentionoftheseendogenousgrowthfac- torswouldencouragewoundhealing,thusleadingtothe earlyclosurethatwehaveobserved. Whilegrowthfactorsareundoubtedlycrucialin woundhealing,thelocalenvironmentcanalsobeinflu- encedinordertocreatemoreoptimalconditions.One biochemicalfactorthatcaninfluencehealingisthepHof TABLE4Woundrelatedcharacteristicsatbaseline. VariablePRBMSOC p value Woundarea(cm 2 )2.8(2.46)4.0(3.59)0.059 Woundage(weeks)14.7(11.01)15.6(11.39)0.68 Prioroffloadingtimeatscreening(weeks)12.9(11.9)15.5(16.8)0.59 DFUside Left23(43)22(43)0.96 Right31(57)29(57) DFUposition Medial35(65)33(65)0.99 Lateral19(35)18(35) DFUlocation Plantar40(74)37(73)0.86 Dorsal14(26)14(27) DFUposition Toe17(32)8(16) Forefoot10(19)18(35) Midfoot16(30)13(25)0.27 Hindfoot3(5)2(4) Heel4(7)6(12) Ankle4(7)4(8) Indexulcerisarecurrence24(44)21(41)0.74 ConcurrentDFUs8(15)13(26)0.17 LifetimeDFUcount5.0(4.92) Median3.5(IQR6) 4.7(4.72) Median3(IQR4) 0.81 Offloadingtype(primary) Shoe11(20)9(18) Camboot/crowboot/walker39(72)37(72) TCC1(2)3(6)0.71 Wheelchair1(2)0(0) None2(4)2(4) 8of13ARMSTRONG ETAL . 1742481x, 2024, 4, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/iwj.14882, Wiley Online Library on [23/04/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons Licenseawound,andithasbeenreportedthatchronicwounds haveanalkalinepHwhereasmorenormaltissuehasa slightlyacidicpH. 25 Inthisregard,thePRBMthatwe appliedhasalsobeendocumentedtoamelioratethelocal environmentwithregardtopH. 12 Thiswouldcertainly beconsistentwithpublicationsstatingthatwoundpHis apotentfactorforthehealingprocess,withdifferentpH rangesbeingessentialinthephasesofwoundhealing. 26 Furtheramurineinvivomodelhaddemonstratedthat woundacidificationpromotedanumberofcellular FIGURE2Kaplan – Meierplotofprobabilityofwoundhealingbytreatmentgroup.Unadjustedtimedepictedafterrandomization. Censormarksindicatesubjectexitpriorto12weeks.AsuperiorhealingtrajectoryisdemonstratedinthePRBMtreatmentgroupwitha divergenceapparentafterabout1week. FIGURE3WeeklyPAR valuesforbothtreatment groups,basedontheITTdata. ARMSTRONG ETAL .9of13 1742481x, 2024, 4, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/iwj.14882, Wiley Online Library on [23/04/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons Licenseresponses,suchasanincreasedrateofre-epithelializa- tionandcollagensynthesis,thusinducingfasterwound healing. 27 Anadditionalconsiderationwithregardtowound healingwouldrelatetotherolethatmatrixmetallopro- teinases(MMP)playinhealing.Itisknownthatthese proteasesplayakeyroleinthedegradationandremodel- lingoftheextracellularmatrix(ECM)duringwound healing. 28 Thewoundhealingprocesscanthusbehin- deredbyanumberofbiochemicalorphysiologicalpro- cesses,whetherprolongedinflammation,imbalancein ECMsynthesisanddegradation,poorneovascularization, orimpairedmacrophageactivity. 29 TheMMPsare involvedinalltheseprocessesand,unfortunately,high levelsofMMPareassociatedwithpoorhealingandder- malgraftfailureinDFUpatients. 30 Asamechanismof healing,thePRBMthatweappliedhasalsobeenshown, invitro,tomodulateMMPactivity. 12 Inconjunctionwith thephysiologicalmechanismsthatwehavepreviously noted(i.e.,modifiedpHandtheretentionofgrowthfac- tors),themodulationofMMPactivityislikelyanother contributortotheimprovementinhealingthatwe observedinthisstudy. Whileourresultsshowedanimprovedclinicalout- comewithshorterhealingtimes,thecost-effectivenessof TABLE5AlistoftheSAEthatwereexperiencedbypatients. NoneoftheseSAEwererelatedtothePRBMortotheSOC. Derma-GideStandardofCare • Necrotizingfasciitisto alegvenousulcer, required hospitalization followedbyseptic shock(2SAEs) • Fractureintheleft anklethatrequired surgery • Cellulitisassociated withanon-indexulcer • Acutekidneyfailure • Acute,non-infective gastroenteritis • Deepveinthrombosis; cellulitisleukocytosis(2SAEs) • Pulmonaryhypertension • Plantarwoundinvolvingacute osteomyelitis(non-index ulcer),requiredextensive work-upincludingabone biopsyandNPWT • Acuteosteomyelitis,index ulcer,requiredextensivework- upincludingabonebiopsy andNPWT • Pneumonia,requiredextensive testing/treatment • Osteomyelitis,righthallux, requiredamputation. FIGURE4PhotosDepictingPRBMTreatedWoundsDuringTheirStudyParticipation.Representativecasesdepictingthetime progressionofwoundhealingfollowingtreatmentwithPRBM.Patient1:Type2DiabeticFemalewithUlcerx15weeks,HbA1c:6.9%, SerumCreatinine1.11mg/dLandWoundAreaof7.5cm 2 .After3PRBMtreatmentsthewoundareadecreasedover90%,and,followingthe fifthtreatment,theulcerwasconfirmedtobefullyhealed.Patient2:Type2DiabeticFemalewithUlcerx5weeks,HbA1c:8.4%,Serum Creatinine1.26mg/dLandWouldAreaof12.25cm 2 .Afterthe3PRBMtreatmentthewoundareahaddecreasedbyover35%andafter 12treatmentswasconfirmedtobefullyhealed.Patient3:Type2DiabeticFemalewithulcerx4weeks,HbA1c:7.0%,SerumCreatinine 1.3mg/dLandaWoundAreaof1.5cm 2 .After5PRBMtreatmentsthewoundareahaddecreasedbyover50%andafterninetreatments wasconfirmedtobefullyhealed. 10of13ARMSTRONG ETAL . 1742481x, 2024, 4, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/iwj.14882, Wiley Online Library on [23/04/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons LicenseNext >