UAB Symposium

Congenital Bone Marrow Failure Path to Cure

Patient’sJourney

• 8yo male• Generallyhealthy• Shortstature(5%ile)• Hypermobilethumbs

withflattenedthenareminences

• Hairandnailsnormal• 1yearhistoryoflowplatelet

counts

Test Result Normal RangeWBC 2.7 x 10*9/L 4.0-11.0ANC 1.8 x 10*9/L 1.5- 8.0Hgb 13.8 g/dL 13.3-17.7MCV 103 fL 78-100Plt 54 x 10*9/L 150-450

ReferraltoHematologistCBCworsenedovertheyear

InterconnectedPathwaysofCongenitalBMF

DiamondBlackfanAnemiaDisorderofribosomalproteins

FanconiAnemiaDisorderofDNArepair

DyskeratosisCongenitaDisorderofproteinsthatmaintaintelomers

DiagnosticMutagenStressTest

ClassicManifestationsofFanconi AnemiaCaneffecteveryorganinthebody

• Skin(caféaulait) 71%• RadialRay 59%• Kidney/urinary 57%• Shortstature 67%• Ears/deafness ?• CNS 1%• Eyes 47%• GI(esoph,duod) 7%• Cardiac(PDA/VSD) 29%

‘Classic’phenotype– missingthumb,missingkidney,shortandbonemarrowfailure

V – VertebralfusionsA – AnalatresiaC – Cardiac(PDAandVSD)TE– Tracheo-EsophagealfistulaR – Renal(hydronephrosis)/radialdefectsL – Limb(nonradial)defects

EffectofIncorrectDiagnosis

DevastatingEffects- NosurveillanceforMDS/leukemia- Incorrectreproductivecounseling(orevaluationofexistingsiblings)

WhentotestforFA• AbsoluteindicationsforDEBtesting

- aplasticanemia(anyage)

• PossibleindicationsforDEBtesting

- youngpatientwithH/Ncancer

-MDSwithcomplexcytogeneticabnormalities

- constellationofmacrocyticanemia,shortstature

- VATER/VACTERL(don’tbeswayedbypriordx)

FAVATERSignatureHigherfrequencyofrenalandlimb(radialand/orthumb)anomalies

93%inFAVATER<30%innonFAVATER

SkeweddistributionwithinFAFANCB(21%)FANCD1/BRCA2(14%)FANCD2(12%)

FANCA(19%)

RecommendationAllVATERpatientswithrenalandlimbanomalies,thentest forFA

General Pattern of Bone Marrow Failure

ISCN:46,XY,dup(1)(q12q32)[9]/46,XY,del(20)(q11.2q13.3)[10]/46,XY[1]

INTERPRETATION:Twoabnormalcloneswereidentified.Clone1,comprises45%ofmetaphases;duplicationwithinthelongarmofchromosome1,extendingfrombandq12toq32.Clone2,comprisestheremaining55%ofmetaphases,hadadeletionwithinthelongarmofchromosome20,extendingfrombandq11.2toq13.3.

ComplexCytogeneticAbnormality Clone1

ImpactofOverridingtheG2/MArrestClonalChromosomal

Abnormalities

0

10

20

30

40

50

60

3qG

7pL/7qL 1q

G 6pL

7pL

7qL11qL 1p

L6qL17pL

Percent

BetsyHirsch,Ph.D.

0%10%20%30%40%50%60%70%80%90%

100%

3qG -7/7qL 1qG

With "other" abnlWith 1qG, 3qG or 7qLSole abnormality

Evolution of Clonal Chromosomal Abnormalities

BetsyHirsch,Ph.D.

Whatdotheymean?Morefrequentmarrowexams

~16%donothaveBMF,MDSorAcuteLeukemiabyage18years

PathophysiologyofBMF,MDSandAcuteLeukemiainFA

Oligoclonalhematopoiesis

- marrowfailure- MDS/leukemia

(evadingG2/Marrest)

DNA damage

Viral suppression [INFg hypersensitivity]

FA-A 60%

FA-C 14.8%

FA-G 9%

AutosomalRecessive(exceptFA-B)

TheFanconi AnemiaPathwayofDNARepairandHumanCancerBy Vaidehi Krishnan, Lavina Sierra Tay and Yoshiaki Ito

Uniqueissues

- impactoncarriers

- genetictestinginchildrenwhoarepotentialdonors

FA/BRCADNARepairPath

BRCA2

Genotype/PhenotypeCorrelationsTimetoMDSorLeukemia

FANCCn=78 FANCA+Gn=253

OtherFANCGroupsn=415

HighriskofothermalignanciesbeforeandafterHSCT

FANCD1/BRCA2n=14

All<7yearsofage

FANCCGenotype-PhenotypeCorrelations

Gillio AP,Verlander PC,Batish SD,Giampietro PF,Auerbach AD.Blood1997;90:105-10.

OnsetofMarrowFailure Survival

FANCCexon1

FANCCexon14FANCCIVS4

NonFANCCFANCCexon1

FANCCexon14

FANCCIVS4

NonFANCC

Indications for HSC Transplant

• Severe cytopeniao Hgb <8 g/dLo ANC <500/mm3

o PLT <20,000/mm3

• MDS/Leukemia

Eligibility Requirements

• Adequate organ functiono Renal (GFR >30)o Liver (ALT <3 x normal)o Cardic (EF >45)

• Available donoro Sibling BM (without FA)o Unrelated BMo Unrelated UCBo Haplo BM

-7-6-5-4-3-2-10123456

ATG

TBI450ratherthan1200cGyCyclophosphamide40ratherthan200mg/kg

CSA

TcelldepletedHSC

DestroyLeukemiaandtheHost’sImmuneSystem

PreventionofHVGandGVHreactions

Myeloablation ImmunosuppressionPrinciples

ConditioningDilemmainFA

Matched Sibling Donor BMT for FATrial 1 - CY, ATG, TLI 450Trial 2 - CY, ATG, FLU, TCD

State of the Art Sibling Donor Transplants after 2000

Years0 1 2 3 4

0.0

0.2

0.4

0.6

0.8

1.0

Complications are rare except in those with MDS/Leukemia or adult age

Unrelated Donor BMT for FA

ThymusblockPriortoHCT,CTisperformedtolocatethethymus

ThymusblocksattachedtoTBIstandbracketssecuringthelungcompensators

ThymusShielding

Perserving FertilityTransientTranspositionoftheOvary

Years

Cum

ulat

ive

Prop

ortio

n

0.0

0.2

0.4

0.6

0.8

1.0

0 1 2 3 4 5

I I I IIIIIIIIIII

I I I I I II I I

I

I I IIIII I I I I IIIII

FLU CY TBI (300) + TS 89%

FLU CY TBI (450) + TS 75%

FLU CY TBI (450) 67%

CY TBI (450) 23%

IIIII I I I I IIIII

I I I I I II I I

Survival in FA Patients Unrelated Donors

Androgens – negative effect in univariate analysis (trend in multivariate)

Alternative Donor BMT for FA Survival Risk Factors

Effect of Transfusions Effect of Infection History

HighExpectationsforSurvivalafterBMT

AlternativestoTransplant

KidneyLiverCardiac

Verypoororganfunction

Preexistinginfections

OlderPatientAge

LungBlood

~45(siblingdonor)~40(unrelateddonor)

Alternatives§ Androgens

§ Intermittentuseofbloodgrowthfactorandtransfusions– othersupportivecare

§ Experimentaltherapy(genetherapyornoveltransplantprotocol)

Nodonor

CancerRiskIncidence30%byage45years

ObservedtoExpectedRatio

• Leukemia 785

• Head/Neck 706

• Esophagus 2362

• Liver 386

• Total 50

RosenbergPS,GreeneMH,AlterBP.CancerincidenceinpersonswithFanconi’sanemia.Blood2003;101:822–826.

DoesTransplantAltertheRisk

Conclusions• Transplantisassociatedwithhighrateofsolidtumors,particularlySCCs

• AssociationbetweenGVHD[oritstreatment]andSCCs- 33-foldhigherriskwithcGVHD

Rosenberg,Blood2005;105:67-73

• Eliminate Bu and TBI– Antibody based conditioning– Higher stem cell doses– PGD/IVF for HLA match and absence of FA

• Prevent GVHD– T cell depletion of the graft– Use of UCB– PGD/IVF for HLA match and absence of FA– Gene therapy

• Enhance immune reconstitution– More HSCs– Thymic progenitors Opportunistic infection– Thymic shielding– PGD/IVF for HLA match and absence of FA– Gene therapy

How do we reduce cancer risk?

Low risk of Grade II-IV AGvHDwith T Cell Depletion

Days Post-TX

Cum

ulat

ive

Inci

denc

e

Alternative donor (std. risk)

Matched sibling

P = 0.47

0.0

0.2

0.4

0.6

0.8

1.0

0 10 20 30 40 50 60 70 80 90 100

Months Post-TX

Cum

ulat

ive

Inci

denc

e

Alternative donor (std. risk)Matched sibling

P = 0.24

0.0

0.2

0.4

0.6

0.8

1.0

0 3 6 9 12 15 18 21 24

Low risk of Chronic GvHD with T Cell Depletion

19%(7-31%)byage45years

DirectionstoReduceCancerRiskinFanconi Anemia

RiskbyPatientAge

YearsofAge

0.0

0.2

0.4

0.6

0.8

1.0

0 5 10 15 20 25 30 35 40 45

Cum

ulat

ive

Inci

denc

e

UniversityofMinnesota

CD34+ Cell

Growth factor

receptors

Differentiation

AhR

AhRL

XRE

Nucleus

AhRL

CYP1B1/AHRR

Cytosol

• SR-1directlybindsandinhibitstheAhR,blockinginductionofCYP1B1andimpedingstemcelldifferentiation

• SR-1mediatedmassiveUCBstemcellexpansion

Boitano et al. Science 2010; 1345-1348

StemRegenin-1 Aryl Hydrocarbon Receptor (AhR) Pathway

SR-1

Massive Expansion of Cord Blood Stem Cells327-fold (n=37)

C D 3 4 + C e l l N u m b e r s

1 e 6

1 e 7

1 e 8

1 e 9

1 e 1 0

B e f o r e

C u l t u r e

A f t e r

C u l t u r e

1 0

1 0 0

1 0 0 0

1 0 0 0 0

M a n u f a c t u r i n g o v e r v i e w

Fo

ld E

xp

an

sio

n

T o t a l c e l l s C D 3 4 + c e l l s

4 . 3

1 , 3 7 2

8 3 2

3 2 7

million

millionExpansion

MGTA-456 after TBI 1320 cGyFaster Neutrophil Recovery, 100% engraftment

Days

Inci

denc

e

0.0

0.2

0.4

0.6

0.8

1.0

0 7 14 21 28 35 42

Historical 83% (CI, 74-91)N=121 median 26.5 d (13-42)

MGTA-456 (single) 100% N=9/9

median 15 days

p < 0.001

Absolute CD4 200 at 2-3 months

Accelerated Immune Reconstitution after UCBT

0 .0

0 .2

0 .4

0 .6

0 .8

1 .0

M A C p ro to c o l

M o n th s p o s t T x

x1

03

/µl

C D 19

C D 4

C D 8

NK

0 1 2 3 6 12

Myeloablative Conditioning

T Progenitor Dose Escalation

• 0.1 x 106 /kg

• 0.3 x 106 /kg

• 1.0 x 106 /kg

• 3.0 x 106 /kg

• 10.0 x 106 /kgTprog ex vivo culture

Repeat CD34+ Selection

T cell tracking

Days

TBIFLU

CY

FLU

CY

FLU

RES

T

-2 -1-3-4-7 -6 -5 0 3012 100 180…6 60-8

CD34-

Re-Cryopreserved

CD34+ Selection

Ex vivo culture – HSC835

UCB 1

20% or maximum available

Future of SR-1 Expanded HSPCsManufacture of Thymic Progenitors

Alcohol/FoodExposuresandCancerRisk

Use of assisted reproductive techniques

IVF

Preimplantation GeneticDiagnosis

Savior Sibling IVF / PGD

PGD to prevent disease and save the life of an existing

childPGD for mutation PGD

for HLA

To eliminate risk of genetic

disease

To guarantee an HLA

identical HSC donor

Savior Sibling

The Candidates 1995

Molly Henry

FANCC IVS4 A>T MutationPredicting the Future

Verlinsky et al. JAMA 2004; 285,3130

Pregnancy

Headlines The Creation of

Adam

BackLash

Thepathofthe‘slipperyslope’

SlipperySlope

DiseasePrevention HLAMatch

GenderPhysicalFeatures

Longevity

Parental Motivation

Risks to the Donor

Future

GeneTherapy

Willitwork?

CreatingSomaticMosaicism

NOSelectiveGrowthAdvantage

SelectiveGrowthAdvantage

Chemotherapy?

HSC

LessonsofNature

HSCSomaticMosaicism‘NaturalGeneTherapy’

43yearoldfemale- Healthy- MMCresistant- Clonalhematopoiesis- Doublecrossoverevent

Selectivegrowthadvantage

mitosis

x

x

x

x

x

x

ControlPatient’s Blood

FANCAgenemutations

RiskofMDS/ALinresidualFAcellsLessonsforFAGeneTherapy

MMC resistant colonies

GregoryJJetalPNAS2001;98:2532

1) Maintenance of adequate (not normal) blood counts as an adult

2) Expected competitive growth advantage of revertant HSC not clear

BetsyHirsch,Ph.D.

FISH with MLL probe exclusively found in colonies with FANCA mutations

Rare Disease Transforms the Practice of Medicine

NMDP / Be the MatchUCB Banking and TransplantationDNA Repair GenesFA Cures

UniversityofMinnesotaBMTProgram

PediatricBMT

MuktaAurora,M.D.VeronicaBachanova,M.D.NelliBejanyanClaudioBrunstein,M.D.SarahCooley,M.D.DanKaufman,M.D.,Ph.D.BrianMcClune,D.O.PhilipMcGlave,M.D.JeffreyMiller,M.D.ArneSlungaard,M.D.CelalettinUstun,M.D.GregVercelotti,M.D.EricaWarlick,M.D.DanWeisdorf,M.D.

BruceBlazar,M.D.ChristenEbens,M.D.KeliHippen,Ph.D.MargaretMacMillan,M.D.WesMiller,M.D.AngelaMortari,Ph.D.PaulOrchard,M.D.JakubTolar,M.D.,Ph.D.AngelaSmith,M.D.HeatherStefanski,MD,Ph.D.MichaelVerneris,M.D.ToddDefor,M.S.(StatisticalSection)

AdultBMT

MagentaTeamTonyBoitano,Ph.D.MichaelCooke,Ph.D.BastianoSanna,Ph.D.JasonGardner,Ph.D.