Continuous BioManufacturing of Mabs by QbD

Continuous BioManufacturing of Mabs by QbD

Maurizio Cattaneo, PhD, CPIP President

BioVolutions Inc. Woburn, Massachusetts, USA

Continuous Biomanufacturing

of Therapeutic Mabs Smaller equipment and facilities �  More flexible operation �  Reduced inventory �  Lower capital costs, less work-in-progress materials �  Smaller ecological footprint Integrated processing with fewer steps. �  No manual handling, increased safety �  Shorter processing times �  Increased efficiency

On-line monitoring and control for increased product quality assurance in real-time (PAT) �  Amenable to Real Time Release Testing approaches �  Consistent quality

Continuous Manufacturing of Mabs Upstream and Protein A Downstream

(50L and 1000L Scale Up)

(0.5L and 20L Scale Up)

CONFIDENTIAL 4

Our Continuous Biomanufacturing Pilot Plant

CQAs (eg. Glycosylation, HCP, DNA, Aggregation)

�  Glycosylation (eg. Galactosylation, Fucosylation)

IgG1   % HighMan   % G0   % G1   % G2  % Sialic acid  

% Gal Alpha  

% Core Fucose  

A-Mab   7±5   71±10   19±10   3±10   1±5   0±5   91±20  

B-Mab   1±5   78±10   17±10   4±10   2±5   0±5   94±20  

R-Mab   2±5   46±10   40±10   12±10   3±5   0±5   95±20  

CQA Acceptable Range

HCP 0-100ng/mg

DNA <10-3 ng/dose

Aggregates 0-5%

Risk Assessment Mitigation Matrix (RAMM) for selecting CPPs

Rela&ve  Importance  of  Output  on  CQA   9   1   1   9   3   3   3   3   3   3  

pH  

CO2  

VCD  

Viability  

Biob

urde

n  

Endo

toxin  

Titer  

Mycop

lasm

In  Vitro  viral  

MMV  PC

R  

Process  Parameters   Tot   Impacts   Proc  Perfusion  Rate   1   1   9   9   1   1   1   1   1   1   127  Quality  &  Growth   WC-­‐CPP  Sparge  Oxygen  Flowrate     1   9   1   1   1   1   1   1   1   1   55          Agita&on  110-­‐125  RPM   3   9   3   3   1   1   3   1   1   1   99  Gas  Evolu&on  Rate   WC-­‐CPP  pH   0   3   3   3   1   1   3   1   1   1   138  Quality  &  Growth   CPP  DO  (>40%)   1   1   3   3   1   1   3   1   1   1   73  Titer      Seed  Density  (0.5  x  10E6)   1   1   3   9   1   1   3   1   1   1   127  Cell  Growth   WC-­‐CPP  N-­‐1  Inoculum  Volume   1   1   1   1   1   1   1   1   1   1   47          Temperature     1   1   3   9   1   1   9   1   1   1   163  Quality  &Growth   CPP  Dura&on  of  Perfusion     1   1   1   3   1   1   1   1   1   1   83          Glucose  Control   1   1   9   9   1   1   9   1   1   1   151  Quality  &  Growth   CPP  Lactate  Control   1   1   9   9   1   1   9   1   1   1   151  Quality  &  Growth   CPP  An&-­‐Foam  Control     3   1   1   1   1   1   1   1   1   1   65          Protein  Load  Protein  A   1   1   1   1   3   3   9   3   1   1   161  Quality  &  Titer   CPP    Elu&on  pH  Protein  A     1   1   1   1   3   3   9   3   1   1   161  Quality  &  Titer   CPP  Totals   17   32   48   62   18   18   62   18   14   14              

Design of Experiments (DoE) (Taguchi L9)

Settings Temperature (°C)

pH Glucose

1 36.5 7.2 0

2 36.5 7 1

3 36.5 6.8 3

4 36 7.2 1

5 36 7 3

6 36 6.8 0

7 35.5 7.2 3

8 35.5 7 0

9 35.5 6.8 1

Continuous Culture Overview

•  It can produce high titers and good quality product

•  Equipment is utilized more efficiently and the equipment footprint is much smaller

•  The Continous System pushes productivity to

the next level, ~10x higher cell densities

Upstream Continuous Cell Culture

1.E+05

1.E+07

2.E+07

3.E+07

4.E+07

5.E+07

6.E+07

7.E+07

8.E+07

0 5 10 15 20 25 30 35

Via

ble

Cel

l Den

sity

(vc/

mL

)

Run Time (Days)

-0.5

0

0.5

1

1.5

2

2.5

3

3.5

0 10 20 30

Tite

r (g

/L)

Run Time (Days)

Run 1

Run 2

Run 3

Key

Glucose and Lactate Control

-­‐1  

0  

1  

2  

3  

4  

5  

6  

7  

0.00   5.00   10.00   15.00   20.00   25.00   30.00   35.00   40.00  

Analyte  (g/L)  

Time  (Days)  

Glucose   Lactate  

Upstream Design Space (DS) based on

Glycosylation CQA

IgG1   % HighMan   % G0   % G1   % G2  % Sialic acid  

% Gal Alpha  

% Core Fucose  

A-­‐Mab  (Test  Ar7cle)   0 54 39 7 2 0 97

A-Mab (Reference)   7±5 71±10 19±10 3±10 1±5 0±5 91±20

B-­‐Mab  (Test  Ar7cle)   0 85 15 1 0 0 96

B-Mab (Reference)   1±5 78±10 17±10 4±10 2±5 0±5 94±20

Overview of Downstream Manufacturing Steps

Downstream Continuous Purification Strategy

�  Synchronize the protein load (g/mL of resin) with the upstream rate of perfusion

�  Replace interim holding tanks with peristaltic pumps to perform continuous downstream purification

�  Reduce the amount of Protein A resin by ~ 20-fold

Continuous Purification of A-Mab

Continuous Purification of A-Mab

0.00

5.00

10.00

15.00

20.00

25.00

30.00

0

400

800

1200

1600

0.00   5.00   10.00   15.00   20.00   25.00   30.00   35.00   40.00   45.00   50.00  

Con

duc

tivi

ty (

mS

/cm

)

Abso

rban

ce (

mA

u)

Time (mins)

Antibody production Perfusion Per Day * 30-Day

Production 12 Runs/Yr

50L 25g 0.75Kg 9Kg

1000L 500g 15Kg 180Kg

Fed-Batch Per Run * 20 Runs/Yr

50L 25g 0.5Kg

1000L 500g 10Kg

* 50% Purfication Yield

Assumptions Parameter   FB   CP  

Seed Train   20L, 80L, 400L, 2,000L   200mL (bag), 50L

(50L run in CFB mode

10 days; 6 reactor vols)  

Production Bioreactor   10,000L   1,000L  

Product Titre   2 g/L   2 g/L  

Production Phase   3 days, 3 days, 13 days   30 days  

Media Consumption   10,000L   1vvd  

Harvest Volume   10,000L per batch   12,000L Continuous

12 days  

Media Cost   $20/L   $5/L  

Protein A Resin 400L  ($4.5M)   20L ($224K)

Direct Comparison between the Fed Batch (FB) and Continuous Perfusion (CP) processes.

The FB process has a higher upfront capital burden ($53M) due to a longer seed train and larger fermentation volume. The CP option benefits from increased throughput and reduced bioreactor sizes.

The CP technology offers substantial capital charge reductions of 32% relative to the FB process.

The CP process produces the most product (265 kg/year) and FB producing the least amount of product (130 kg/year).

Effect of Vessel Size on Cost of Goods (COG)

The CoG of the CP process decreases as the bioreactor size is increased, from $133/g at 500L scale to $87/g at the 1000L scale.

At 500L, the CP process generates about the same annual output (132 kg) as the 10kL FB process (130 kg) and is still more economical than the FB process.

It should be noted that the simplification of material and people flow provided by reduced bioreactor capacity in the CP facility cannot be calculated easily but may provide the key driver to such technology being increasingly adopted.

Capital (US$)

106.7M 33.1M Capital (US$)

103.9M 44.1M

Floor Area (m2)

4,012 2,447 Floor Area (m2)

5,096 3,152

Results at 500Kg / year

•  A comparison of fed-batch to perfusion with the ATF System at “large” scale:

–  Perfusion utilizes a smaller footprint and

–  Lower capital investment costs

–  And has lower operating costs

•  The advantage of disposable bioreactors reduces with increasing bioreactor size,

as required for large scale fed-batch operation for example

21

Summary for Cost of Goods Model

•  A ~$30m capital budget could give you a facility that annually produces:

–  50Kg of protein in FB mode in stainless steel tanks

–  50Kg of protein in FB mode in disposables, and have $10m left over

–  500Kg of protein in perfusion mode in disposables.

•  If you plan to expand an existing facility with small bioreactors (50L-1000L), it is possibly more cost effective to move to a concentrated process than to expand fed-batch capabilities

–  50Kg of protein in FB mode is made using 2x 1000L tanks

22

Summary for Cost of Goods Models - continued

•  The best case scenario if you need 500Kg per year of a single product is a perfusion facility, which provides cost savings of:

–  68% reduction of your capital budget ($33.1m vs $103.9m)

–  41% reduction in cost of goods ($57.2/g vs $97.9/g)

•  If you had 10 products to manufacture, and need 50Kg of each per year:

–  $30m capital is required for the perfusion facility, with 20% spare capacity

–  $100m capital is required for a fed-batch facility, with no spare capacity

•  This model is limited to new build only and focused on the cost of goods:

–  A more sophisticated analysis in future will look at retrofit options and costs for different facilities

–  Operating risks, such as having fewer bioreactors, are not yet

captured

KEY POINTS

•  Making therapeutic antibodies can benefit from new advances in continuous biomanufacturing such as

Continuous Perfusion (CP) and Continuous

Chromatography (CC).

•  The Capex for a new facility is significantly lower than

conventional batch facilities

CONFIDENTIAL 23