Gene therapy and therapeutic gene editing: what are the predictors of success?

Bobby Gaspar

Professor of Paediatrics and Immunology

Centre for Immunodeficiency

UCL Institute of Child Health/Great Ormond Street NHS Trust

Gene therapy and paediatric conditions

Cystic fibrosis

Duchenne’s MD

Inborn errorsOCT, HurlersX-ALD

Retinalabnormalities

Haemophilia Severeimmunodeficiencies

Skin conditionsEB

Fanconi’s anaemiaHaemoglobinopathiesAcute leukaemias

Primary Immunodeficiency

red blood cell

platelets

white blood cells neutrophils /monocytes

white blood cells / Immune cells

stem cells

Virus

Bacteria

Fungi

Pneumonia, diarrhoea, repeated infections

Severe combined immunodeficiency (SCID)

T B NK

HSC-multi

SCID/CID

c, IL7Ra, JAK3, ZAP-70RAG1/2, artemis, ligase IV,CernunnosADA, PNPMHC I/II, CD3//, CD45,ORAI1

Molecular defects in SCID

How do we get genes into cells?

Disabled viruses

Adenovirus

Adeno-associated virus

Non-viral vectors

Retrovirus

X

Stem cells

lymphocytes

B

BloodBone marrow

T

NK

ThymusXIn X-SCID growth of lymphocytes is blocked

a bg

Pathology of SCID-X1

MoLV U5U5 Y

RSD SA

RQ

IL2RG

LTR-driven gammaretroviral vector: MFG gC

MoLV

Criteria for entry:No matched sibling donorMolecularly confirmed diagnosis

Common gamma chain vector:PG 13 producer cells (GALV envelope)titre approximately 1x10e6 transducing units per ml

SCID-X1 gene therapy protocol

Lymphocyte recovery CD3

0 50 100 150 200 250 300

0

1000

2000

3000

4000

5000

P1

P2

P3

P4

P5

P6P7

P8

P10T L

ym

ph

ocyte

s/µ

l

Weeks

P9

 

Patient details

 

Age at therapy

(months)

Maternal graft

Mutation Gamma chain expression

Total cells infused(x10e6)

 

P1 10 ++ R289X ++ 180 A+W

P2 10 ++ S238N - 180 A+W

P3 4 - Y125C +/- 78 A+W

P4 3y - R289X ++ 115 A+W

P5 10 - R222C ++ 200 A+W

P6 10 - PolyA - 200 A+W

P7 6 - M1i - 84 A+W

P8 13 - C182Y + 207 A+W

P9 7 - S108P + 160 A+W

P10 12 - del - 60 A+W

Currentstatus

 

Biochemical defect in ADA deficiency

DNA

d-adenosine d-adenosine d-inosine

d-adenosine

d-ATPincrease is toxic to lymphocyte function

dCydK

ADA

T cell recovery post ADA gene therapy

Pre-diagnosis Pre-GT Post-GT0

10

20

30

40

50

AD

A A

cti

vit

y (n

mo

l/m

gH

b/h

)

Pre-diagnosis Pre-GT Post-GT0

20

40

60

80

800

1000

1200

1400

1600

dA

TP (

mm

ol/

L)

Metabolic correction post gene therapy

Summary of ADA-Deficient SCID Patients Retroviral Vectors, Myeloreductive Conditioning–Milan/London/CHLA-NHGRI, NIH-UCLA

Center # Pts F/U (yrs)1 Off Enzyme Survival DFS2

Milan 18 0.8 – 11.5 15/18 100% 83.3%

London 8 0.5 – 7.5 4/8 100% 50%

CHLA-NHGRIUCLA-NHGRI

68

2– 50.1-2

3/67/8

100%100%

50%87.5%

TOTAL 40 0.1 – 11.5 29/40 100% 72.5%

1 As of April 20122DFS ≡ Alive without BMT or PEG-ADA re-start

Data: Courtesy HB Gaspar (London) and Alessandro Aiuti (Milan)

 

Patient details

 

Age at therapy(months)

Maternal graft

Mutation Gamma chain expression

Total cells infused(x10e6)

 P1 10 ++ R289X ++ 180 A+W

P2 10 ++ S238N - 180 A+W

P3 4 - Y125C +/- 78 A+W

P4 3y - R289X ++ 115 A+W

P5 10 - R222C ++ 200 A+W

P6 10 - PolyA - 200 A+W

P7 6 - M1i - 84 A+W

P8 13 - C182Y + 207 A+W

P9 7 - S108P + 160 A+W

P10 12 - del - 60 A+W

Currentstatus

 

LMO2 CAPRIN1 NAT10CD59 FBXO3

Forward Strand

Chromosome 11p13

33.65Mb 33.85Mb 34.05Mb

ABTB2C11orf41

X

VIRUS INTEGRATION

Fold difference in gene expression

0.1

1

10

100

C11orf41 CD59 FBXO3 LMO2 CAPRIN1 Nat10 ABTB2

Gene

Difference

Leukemia

DP1 T cells

DP2 T cells

Fold difference in gene expression

0.1

1

10

100

C11orf41 CD59 FBXO3 LMO2 CAPRIN1 Nat10 ABTB2

Gene

Difference

Leukemia

DP1 T cells

DP2 T cells

Fold difference in gene expression

0.1

1

10

100

C11orf41 CD59 FBXO3 LMO2 CAPRIN1 Nat10 ABTB2

Gene

Differenc

e

Leukemia

DP1 T cells

DP2 T cells

Fold difference in gene expression

0.1

1

10

100

C11orf41 CD59 FBXO3 LMO2 CAPRIN1 Nat10 ABTB2

Gene

Difference

Leukemia

DP1 T cells

DP2 T cells

Fold difference in gene expression

0.1

1

10

100

C11orf41 CD59 FBXO3 LMO2 CAPRIN1 Nat10 ABTB2

Gene

Differenc

e

Leukemia

DP1 T cells

DP2 T cells

Fold difference in gene expression

0.1

1

10

100

C11orf41 CD59 FBXO3 LMO2 CAPRIN1 Nat10 ABTB2

Gene

Difference

Leukemia

DP1 T cells

DP2 T cells

Fold difference in gene expression

0.1

1

10

100

C11orf41 CD59 FBXO3 LMO2 CAPRIN1 Nat10 ABTB2

Gene

Difference

Leukemia

DP1 T cells

DP2 T cells

Fold difference in gene expression

0.1

1

10

100

C11orf41 CD59 FBXO3 LMO2 CAPRIN1 Nat10 ABTB2

Gene

Difference

Leukemia

DP1 T cells

DP2 T cells

Fold

Diff

ere

nce

Fold difference in gene expression

0.1

1

10

100

C11orf41 CD59 FBXO3 LMO2 CAPRIN1 Nat10 ABTB2

Gene

Difference

Leukemia

DP1 T cells

DP2 T cells

Relative to:Leukaemia panel

DP1 T cellsDP2 T cells

Activation of LMO2

MoLV U5U5 Y

RSD SA

RQ

IL2RG

LTR-driven gammaretroviral vector: MFG gC

MoLV

U5U5 Y

R RQ

MP ΔSD

PREProm. IL2RG

EF1(S)SF

RSV

New gammaretroviral SIN vectors: Sin11 / SRS11

Development of a new vector for SCID-X1

SF EFS SF.HS

P < 0.001

detection limit

Control Cohort Survival (n=14)

0 100 200 300 4000

20

40

60

80

100

Days

Per

cen

t su

rviv

al

23/24 mice long term survival

Reduced mutagenesis with SIN configuration

Gene transfer for SCID-X1 using a self-inactivating (SIN)

gammaretroviral vectorA multi-institutional phase I/II trial evaluating

the treatment of SCID-X1 patients with retrovirus-mediated gene transfer

Sites:Great Ormond Street Hospital, UK (1)

Hôpital Necker Enfants Malades, France (4)Children’s Hospital Boston, US (2)

Cincinnati Children’s Hospital Medical Center, USMattel Children’s Hospital, Los Angeles, US (1)

Lentiviral vector gene therapy for ADA-SCID

Phase I/II, non-controlled, open-label, nonrandomised, trial to assess the

safety and efficacy of EF1αS-ADA lentiviralvector mediated gene modification of

autologousCD34+ cells from ADA-deficient individuals

Sites:Great Ormond Street Hospital, UK (2)

Mattel Children’s Hospital, Los Angeles, US

Vector production:Indiana University Vector Production Facility

Testing of efficacy and safety

Phase I/II study of 10 patients

Development of vector

January 2007 July 2009 December 2012 January 2018

MRC Research Grant MRC DPFS MRC DCS

Lentiviral vector mediated Gene Therapy for ADA SCID

Collaborations:UCLAIndiana University Vector Production Facility

1st patient treated Feb 20123 patients treated in total

LV Phase I/II trial for Wiskott-Aldrich syndrome

Lentiviral Vector for CGD

Prov irus pC C L c him GP91 W PR E45 4 8 7 b p

G P 91cP P T cts

HIV 1 p si W P RE * (P RE 4)

d el taU3

d el taU3

HIV 1 R HIV 1-p o lyA

HIV 1 P BS

Ch im eric p ro mo ter

HIV 1 RRE

HIV U5 HIV 1 R

HIV U5

Therapeutic gene editing

Gene therapy and paediatric conditions

Cystic fibrosis

Duchenne’s MD

Inborn errorsOCT, HurlersX-ALD

Retinalabnormalities

Haemophilia Severeimmunodeficiencies

Skin conditionsEB

Fanconi’s anaemiaHaemoglobinopathiesAcute leukaemias

Cures now available for specific SCID conditions

First ever cures with Gene Therapy

Morbidity and mortality still low compared to other treatments and prev innovations

Need new safer vectors

Each disease should be considered individually (gene regulation, expression, tissue specificity)

Gene therapy in PID

Great Ormond Street Hospital

Paul VeysPersis AmroliaKanchan RaoGraham DaviesAlison JonesCathy CaleLesley HendersonJane GasparJin Hua Xu-BayfordLucie BrownNursing and support staff

Jude CopeJodi New

Institute of Child HealthEmma BjorkerenKate ParsleyKimberly GilmourSam CoorayElena de FalcoJo SinclairDoug KingSteve HoweStuart AdamsSuzy ThornhillMichelle QuayeDaleen Lopez-BeggSue SwiftFang ZhangLin ZhangClaudia Montiel EquihaMaria Alonso-FerreroMarlene CarmoChristine RivatClaire BoothKaren BucklandSue SwiftNourredine HimoudiAnne-Marie McNicol

Purine Res Lab, Guys Hospital Lynette Fairbanks

ManchesterBrian BiggerFiona Wilkinson

Collaborators

Chris BaumAxel SchambachChristof von KalleManfred Schmidt

Mik AntoniouMarina Cavazzana-CalvoAlain FischerDavid WilliamsManuel GrezVicky BordonTheoni Petropolou

Many thanks to

Adrian ThrasherBobby Gaspar