2/8/2019

1

T cell therapy for viruses

Ann Leen

Viral infections post-transplant

• 40% deaths after alternative donor transplant due to viral infections

• Antiviral drugs –Costly–Significant side effects –Often ineffective

• Alternative - Adoptive T cell transfer

2/8/2019

2

InfusionCell

expansion

Virus-specific T cellsSC

T r

ecip

ient

Antigen Specificity

Adoptive T cell

transfer

Blood draw

Donor lymphocytes

SC

T d

onor

Immunotherapy for viral infections

• Virus-specific T cells as prophylaxis and treatment–EBV–Adv/EBV (bivirus)–Adv/EBV/CMV (trivirus)

2/8/2019

3

Generation of trivirus-specific T cell lines using Ad5f35 vectors

6 wk

PBMCs EBV LCL

B95-8 EBV virus Ad5f35pp65 vector

Ad5f35pp65transduced EBV LCLs

PBMCs

Ad5f35pp65 vector

Restimulation

6 wk

+IL2

• In vitro expanded donor-derived virus-specific T cells targeting Adv, EBV, CMV

–Safe–Reconstituted antiviral immunity for

EBV, CMV and Adv–Effective in clearing disease–Considerable expansion in vivo

Clinical Outcome Summary –Donor-specific setting

Leen et al, Nat Med. 2006Leen et al, Blood. 2009

2/8/2019

4

InfusionCell

expansion

Virus specific T-cellsSC

T r

ecip

ient

Antigen Specificity

Adoptive T-cell

transfer

Blood draw

Donor lymphocytes

SC

T d

onor

Limitations

- Cost

- Complexity

- Limited viral range

2/8/2019

5

Cost

6 wk

PBMCs EBV LCL

B95-8 EBV virus Ad5f35pp65 vector

Ad5f35pp65transduced EBV LCLs

PBMCs

Ad5f35pp65 vector

Restimulation

6 wk

+IL2

EBV LCL

Ad5f35pp65

15mer pepmixes

Target antigens- EBV- EBNA1, LMP2, BZLF1- CMV- IE1, pp65- Adv- Hexon, Penton

Reduce cost:Replace virus/vector with peptides

2/8/2019

6

1

10

100

1000

10000

HEXON PENTON IE1 PP65 EBNA1 LMP2 BZLF1 NO PEP

AdV CMV EBV

SpecificityS

FC

/2x1

05

Reduce cost – peptides

Complexityday 0 – VST initiation 2.4 x 107

day 12 – IL2 feed

day 20 – split + IL2 feed

Day 30 - harvest/freeze

day 27 – split + IL2 feed

3.6 x 108

day 9 - restim 2 x 107centrifuge

day 16-harvest and reseed 6 x 107

centrifuge

day 23 – harvest and reseed 1.2 x 108

centrifuge

2/8/2019

7

Reduce complexity:Simplifying Production using G-Rex devices

• Gas permeable membrane allows CO2/O2 exchange• Supports cell growth with large volumes of media• Reduces feeding frequency and manipulation• No rocking or stirring

G-Rex 10

CO2

O2

Manufacturing limitations

Reduce cost – peptides

Reduce complexity – G-Rex

Extend target viruses - HHV6+ BKGerdemann et al, Blood, 2013

Gerdemann et al, Mol Ther, 2012

Vera et al, JIT, 2010

Clinical trial – PACT application

2/8/2019

8

AdV– Hexon, PentonEBV– EBNA1, LMP2, BZLF1CMV– IE1, pp65BKV– LT, VP1HHV6– U11, U14, U90

ARMS - Rapidly generated T cell lines targeting 5 viruses

Papadopoulou et al, Sci Trans Med, 2014

mVSTs(multivirus VSTs)

+IL4/7

T cell stimulation/ expansion

10 days

PACT - 0053

• 11 pts infused on study: 4 pts: 5x106/m2 (DL1) 4 pts: 1x107/m2 (DL2) 3 pts: 2x107/m2 (DL3)

• No dose limiting toxicity• 1 Grade II skin GvHD (improved with topical steroids)

• 3 infused prophylactically• All remained infection-free for at least 3 months

• 8 pts had active infections

Patients infused

2/8/2019

9

Patient with AdV CMV EBV BKV HHV6

1 virusx

x

2 viruses

x x

x x

x x

3 virusesx x x

x x x

4 viruses x x x x

8 pts treated for infections

0

50

100

150

200

250

1

10

100

1000

10000

100000

1000000

wk-3 wk-2 wk-1 Infusion wk1 wk2 wk3 wk4 wk6 wk8 wk12

SF

C/5

x105

Adv

cop

ies/

ml

Viral load AdV T cells

Clinical response - Pt with AdV

mVSTs

2/8/2019

10

0

100

200

300

400

500

600

1

10

100

1000

10000

100000

1000000

wk-2 wk-1 Infusion wk1 wk2 wk3 wk4 wk6 wk8 wk12

SF

C/5

x105

EBV T cells

Clinical response - Pt with EBV-PTLD

mVSTs

Viral load

Pre mVSTs Post mVSTsE

BV

cop

ies/μ

g D

NA

0

200

400

600

800

0

10

20

30

40

50

60

Pre wk3 wk5 wk6 wk8 wk12

CMV T cells

Viral load

Clinical response - Pt with CMV

mVSTs

Ant

igen

emia

(C

MV

)

SF

C/5

x105

2/8/2019

11

0

20

40

60

80

1

10

100

1000

10000

wk-3 wk-2 wk-1 Infusion wk1 wk2 wk3 wk4 wk6 wk8 wk10 wk12

1.E+00

1.E+02

1.E+04

1.E+06

1.E+08

1.E+10

wk-3 wk-2 wk-1 Infusion wk1 wk2 wk3 wk4 wk6 wk8 wk10 wk12

BKV T cells

Clinical response - Pt with BKV

Viral load

Viral load

SF

C/5

x105

BK

V c

opie

s/m

lB

KV

cop

ies/

ml

Blood

Urine

mVSTs

mVSTs

0

5

10

15

20

25

1

10

100

1000

10000

Infusion wk1 wk2 wk3 wk4 wk6 w8 w12

Viral load

HHV6 T cells

Clinical response - Pt with HHV6

mVSTs

HH

V6

copi

es/m

l

SF

C/5

x105

2/8/2019

12

Outcomes

• Complete response: (√)

viral load to the normal range

resolution of clinical signs/symptoms

• Partial response: (PR)

>50% reduction in viral load

Patient with AdV CMV EBV BKV HHV6

1 virusx

2 viruses

PR

3 viruses

x 4 viruses PR

8 pts treated for infections

2/8/2019

13

Conventional Rapid

SpecificityAdV, EBV, CMV AdV, EBV, CMV,

BK, HHV6

Blood vol. 60ml 20ml

Cells LCL, VSTs VSTs

Time 56-84 days 10 days

Cell # 200x106 390x106

Old vs New

Papadopoulou et al, Sci Trans Med, 2014

Summary• Feasible to generate broad spectrum

VST products

• Manufacturing simple and robust

• VST therapy safe

• VST therapy effective

2/8/2019

14

Thanks

CAGTLisa RollinsShanna Fulton

CAGT PIsJuan VeraHelen HeslopCliona RooneyMalcolm Brenner

Leen-Vera LaboratoryAyumi WatanabeManik KuvalekarPremal LullaShivani MukhiSpyridoula VasileiouTsung-Yen ChangNorihiro WatanabePradip BajgainSujita SukumaranAndrew JacksonAlejandro TorresYovana VelazquezElizabeth Laval

Ulrike GerdemannAnastasia PapadopoulouIfigeneia Tzannou

Clinical ResearchBambi GrilleyAmy Reyna

QA/QC Laboratory

Adrian GeeSara RichmanNatasha LaptevaMygiao LeDebbie LyonApril DurettCrystal Silva-Lentz

Clinical PIsBilal OmerRobert KranceCarlos RamosPremal LullaSwati NaikStephen Gottschalk

GMP LaboratoryHuimin ZhangBirju MehtaGouqing GeSilvana PercontiJuntima Sritabal-Ramirez