Tumor-specific T cells Current Strategies to Reduce · Cost of Manufacture Juan Vera. 2 Infusion...

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Tumor-specific T cells

Current Strategies to Reduce

Cost of ManufactureJuan Vera

2

Infusion

Tumor-specific T cells

Antigen

SpecificityP

atient Adoptive T cell

transfer

Blood draw

PBMCs

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T cell therapy for cancer

T cell

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Challenge: Tumor heterogeneity

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Immune escape

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• Simultaneously target multiple TAAs

• Target multiple epitopes (CD4 and CD8) within each antigen

• T cells with native T cell receptor specificity (non-engineered)

Our approach

7MultiTAA T cells

MultiTAA T cell therapy

MAGEA4

PRAME

Survivin

NYESO1

SSX2

8

0

20

40

60

80

100%

Positiv

e c

ells

MultiTAA-T Cell Phenotype

n=39CD4 CD8CD3 DC TCM TEMNK

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MultiTAA-T Cell Phenotype

0.1

1

10

100

1000

0 1 2 3 4 5 6

SF

C/2

x1

05

ce

lls

SSX2 MAGEA4PRAME SurvivinNYESO1

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MultiTAA-T Cell Phenotype

0%

10%

20%

0 0.5 1 1.5 2E:T of 20:1

% S

pecific

Lysis

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0

20

40

60

80

100

120

140

160

180

200

PRE MTH3 MTH9

SurvivinNYESO1MAGEA4SSX2PRAME

SF

C/2

x1

05

Pre Mth3 Mth9

Targeted antigens Non-targeted antigen

0

5

10

15

20

25

30

35

40

45

Pre Mth3 Mth9

MAGEC1

SF

C/2

x1

05

Mth 3Mth 9Pre-Infusion

Clinical response – DLBCL

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Activation

DC

Overlapping pepmixes

PBMCs MultiTAA T cells

Expansion

Marker current PROCESS Current process

7 days 10 days 7 days

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How can we expand

a T cell product in a

practical matter?

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Conventional cultureware limitations

• Limited media volume to allow gas exchange

O2

CO2

T75 flask

75cm2

O2

CO2

1cm2

24 well plate

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Limitations of conventional suspension cell culture methods

•Prolonged culture period

•Extensive manipulation

•Risk of contamination

• Labor intensive

• Highly trained personnel

• Excessive reagent use

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• Gas permeable membrane allows exchange of CO2 and O2

• Supports cell growth with large media volumes

• Reduces frequency of feeding/manipulation

• No rocking or stirring

100 cm2

100

0 m

L

1000U/IL2

1 cm2 = 5x105 cells

Vera JF et al. JIT. 2010

®

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G-Rex vs conventional

cultureware – cell expansion

0

3

6

9

12

15

G-RexPlate

Fold

exp

ansi

on

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Day 1

Day 2

Day 3

Day 4

Day 5

Day 6

CFSE

24-well plate G-Rex

Superior cell numbers

not due to increased

cell division

Vera JF et al. JIT. 2010

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Anexin

PE

24-well plate

G-Rex

7AA-D Prcp

Day 1 Day 3 Day 5 Day 7

24-well plate G-Rex

Improved cell viability in G-Rex

SS

FS

Vera JF et al. JIT. 2010

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Low seeding density results in

greater fold expansion

-5

15

35

55

75

95

115

1.00E+06 5.00E+05 2.50E+05 1.25E+05 6.50E+04

Cells/cm2 (x 106)

Fold

Exp

ansi

on

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Low cell density = Greater cell expansion

Time

Cel

l nu

mb

ers

Max cell density 10E+07

cells/cm2

Op

timal tim

e f

or

cu

lture

harv

est

Optimal seeding density

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10ml of media/cm2 resulted in

maximum cell expansion

Volume of media/cm2

Cel

ls/c

m2

(x 1

06)

0

3

6

9

12

15

0.5 2 5 10 15 20

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10ml of media/cm2 resulted in

maximum cell expansion

Volume of media/cm2

Cel

ls/c

m2

(x 1

06)

0

3

6

9

12

15

0.5 2 5 10 15 2010

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Upfront media addition resulted in shortest culture period

0

2

4

6

8

10

12

14

16

0 3 6 9 13 16 20 24

10mL/cm2 (2.5mL*4)

10mL/cm2 (5mL*2)

10mL/cm2

Cel

ls/c

m2

(x 1

06)

Time in culture

25

0

2

4

6

8

10

12

14

0

50

100

150

200

250

300

Day 0 Day 3 Day 6 Day 9

GlucoseCells

Glu

cose

[mg/

dl] C

ells/cm2

Time in culture

Inverse correlation between cell number and glucose concentration

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Cel

l nu

mb

er (x

10

6)

Days in culture

0

20

40

60

80

100

120

140

0 4 8 12

Hemocytometer

Flow

Formula

Glucose assessment can be used to predict cell output

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Are these observations

reproducible?

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•25E+06 cells•1 L of media

Day 0

G-Rex 100M

•1.4E+09 cells

Day 12

G-Rex 100M

Combination of optimal culture conditions using the G-Rex

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1

10

100

1000

10000

Day 0 Day 12

CelGeneBaylorCity of Hope

Cel

l nu

mb

er (x

10

6)

Days in cultureBajgain et al. Mol Ther Methods Clin Dev. 2014

Multicenter study validates optimal G-Rex culture conditions

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SA: 5 cm2

Vol: 50 ml

G-Rex 5

G-Rex 500

SA: 500 cm2

Vol: 5000 mlSA: 100 cm2

Vol: 1000 ml

G-Rex 100

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G-Rex Culture conditions are

linearly scalable

Cel

l nu

mb

er (x

10

6)

0.1

1

10

100

1000

10000

Day 0 Day 10

G-Rex 5

G-Rex 100

G-Rex 500

Bajgain et al. Mol Ther Methods Clin Dev. 2014

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G-Rex 100

G-Rex 500

Available As A Closed System

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Rapid Cell Harvest

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Cell collection using GatheRex

Video

Cell harvest with the GatheRex

Bajgain et al. Mol Ther Methods Clin Dev. 2014

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Automated closed

manufacture

36BUSINESS CONFIDENTIAL AND PROPRIETARY

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SummaryMultiple AntigensTarget expression of multiple tumor antigens thereby

targeting more tumor cells

Clinical SafetyNo related SAEs or CRS in 60+ patients

No Genetic ModificationNative peptides presented in culture,

natural T cells expand with no mutagenesis risk

Lower CostEfficient process - requires no genetic modification or

extensive product manipulation

Clinical ImpactPotential of increased duration – prevents immune

escape with antigen spread

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Tumor-specific T cells

Current Strategies to Reduce

Cost of ManufactureJuan Vera