Continuous Capture Chromatography · 2017-06-22 · Why use continuous chromatography for capture?...

Continuous Capture Chromatography

A simple, robust approach to move from batch to

continuous capture

LEWA Bioprocess Technologies Group – Devens, MA USA

Contents

– Our approach to multi-column capture chromatography

LEWA-Nikkiso America Inc.

2

– Customer case studies

– Conclusions

– A simple set up

– Who we are and what we do

Why use continuous chromatography for capture?

Under utilization of resin capacity in Protein A step

• Protein A resin costs: $10,000 – $15,000 / L

• Size of Protein A columns used in manufacturing: 100 – 250 L

• One packed column worth $1 – $2.5 million, but utilization is only 40-60%!

© LEWA - Bioprocess Group and 3

Unused resin capacity,

typically 40-60% Feed

(clarified harvest)

Batch single Protein A column Flow-through

4© LEWA - Bioprocess Group

Why continuous chromatography for capture?

The driver for the development of continuous capture processes using counter-current

chromatography was the realization that 40 – 60% of the costly Protein A resin was not being utilized.

Even though people started thinking about the under utilization of Protein A resin back in the mid-

2000s, developing improvements didn’t become a priority until processes went to large scale

production where the cost of one Protein A column could be in excess of several millions of dollars.

The basic principle of continuous capture (2C-PCC*)

5

feed

unused resin

capacity

feed

fully loaded column

elution

*2C-PCC: 2-Column Periodic Countercurrent Chromatography© LEWA - Bioprocess Group and


The basic principle of continuous capture (2C-PCC*)

Different terminology such as, “periodic countercurrent chromatography,” “multi-column continuous

capture chromatography” generally refer to the same technology.

In continuous capture using two columns, the first column is fully loaded and breaks through to the

second column, then the second column is fully loaded so that all resin capacity is utilized. After the

initial loads, while one column is being loaded, the other column is washed, eluted, regenerated, and

re-equilibrated. This cycle is repeated continuously until the entire batch has been processed.

ChromaCon / LEWA Partnership

LEWA is global GMP scale up partner for

ChromaCon two column technology.

Control logic for operating two columns

continuously on a single system.

7

ChromaCon

•Process know-how

•Intellectual property

•Entry-stage equipment

LEWA

•Scale-up Equipment

•Process solutions

•Continuous manufacturing

•Worldwide Footprint

CaptureSMB

MCSGP

• Technical, marketing and sales

collaboration.

• Patented technology

ChromaCon / LEWA Partnership

LEWA is global GMP scale up partner for

ChromaCon two column technology.

Control logic for operating two columns

continuously on a single system.

8

CaptureSMB

MCSGP

• Technical, marketing and sales

collaboration.

• Patented technology

The continuous capture chromatography that we employ in our GMP scale system is based on and

licensed from ChromaCon, a Swiss life science tool company that specializes in developing and

commercializing chromatographic process technology for protein and monoclonal antibodies.

Continuous capture with LEWA EcoPrime® Twin LPLC

© LEWA Bioprocess Group and 9

Huge productivity10 cm ID Protein A columns –

more than 1 kg mAb purified/day

(500 L feed)

SimplicityTwo column design minimizes

complexity and secures

maximum uptime

Easy scale upProcess design on the

Contichrom CUBE, fast scale-up

to EcoPrime Twin

Continuous capture with LEWA EcoPrime® Twin LPLC


There are 3 main advantages for using EcoPrime Twin LPLC for continuous capture.

1. Significant increase in productivity as compared with traditional batch operation. For example,

over 1 kg of mAb can easily be purified in a day on a Twin with two 10-cm ID Protein A columns.

2. The simple 2-column design of the EcoPrime Twin dramatically reduces the complexity when

compared to systems with 3 or more columns. This simplicity minimizes downtime and facilitates

transfer to manufacturing and validation.

3. The EcoPrime Twin was designed to be the scale-up system for the Contichrom CUBE, a bench

scale system. Methods from the CUBE can be easily translated for configuration and operation on

the Twin.

Huge productivity

System/ModelColumn ID Range

(cm)

Feed per day

(L/day)

Product recovered*

(kg/day)

EP Twin 1000.004 – 0.60 L/min

2.5 10 23 360 0.11 1.7

EP Twin 2500.02 – 2.4 L/min

5 20 92 1,460 0.44 7.0

EP Twin 5000.06 – 9.0 L/min

8 45 206 7,420 1.0 35.0


10 cm ID columns

More than 300 L feed processed/day

More than 1 kg mAb purified/day

*Process dependent

Huge productivity

System/ModelColumn ID Range

(cm)

Feed per day

(L/day)

Product recovered*

(kg/day)

EP Twin 1000.004 – 0.60 L/min

2.5 10 23 360 0.11 1.7

EP Twin 2500.02 – 2.4 L/min

5 20 92 1,460 0.44 7.0

EP Twin 5000.06 – 9.0 L/min

8 45 206 7,420 1.0 35.0


10 cm ID columns

More than 300 L feed processed/day

More than 1 kg mAb purified/day

*Process dependent

Modeling parameters:

Linear velocity feed (cm/h) – 250

Linear velocity process(cm/h) – 300

Step yield (%) – 95

Titer (g/L) – 5

Column length – 10

DBC (g/L resin) – 60

Productivity calculated – 1.11 kg/L resin/ day

Buffer consumption calculated – 0.41 L/g product

The numbers in the table are estimates of the productivity that can be achieved with the EcoPrime

Twin. The EcoPrime Twin is available in 3 sizes based on the flow rate capability. Even with the

smallest Twin and 10-cm ID columns, over 300L of feed can be processed in a day generated 1.7 kg

of product.

Simplicity

The 2-column approach reduces risk

• Accelerates validation

• Simplifies maintenance

• Lessens costly downtime

• Minimizes operating expenses

• Streamlines implementation, easy-to-

understand and use


8 column multicolumn process

~240 Valves

~40 Valves

Same productivity as systems with 3 or

more columns, in a simple 2-column design.

CUBE from ChromaCon®

Simplicity

The 2-column approach reduces risk

• Accelerates validation

• Simplifies maintenance

• Lessens costly downtime

• Minimizes operating expenses

• Streamlines implementation, easy-to-

understand and use


8 column multicolumn process

~240 Valves

~40 Valves

CUBE from ChromaCon®

A key element of the simplicity inherent in the 2-column design of the EcoPrime Twin and the CUBE is

the significant reduction in valves required. Fewer mechanical components not only means less

downtime for repair and maintenance, but also accelerates validation and streamlines implementation

in a manufacturing setting because the equipment and it’s operation is so much easier to understand.

The real beauty of the simple, 2-column design is that we can achieve the same productivity as

systems with 3 or more columns.

Easy scale-up, scale-down


Contichrom CUBE EcoPrime Twin 100 EcoPrime Twin 500

CUBE is a simple tool for scaling-up and scaling down.• Develop new processes

• Troubleshoot existing processes

EcoPrime Twin mirrors CUBE operation with continuous capture, batch, and

sequential batch functionality.

Easy scale-up, scale-down


Contichrom CUBE EcoPrime Twin 100 EcoPrime Twin 500

CUBE is a simple tool for scaling-up and scaling down.• Develop new processes

• Troubleshoot existing processes

Whether you are developing new processes or troubleshooting an existing one, scaling up to the Twin

or down to CUBE is relatively easy because the design of the Twin mirrors the operation of the CUBE.

There is nothing worse for a schedule than having to redo your process development when scaling

up. Or, when things go wrong, not have a scaled-down system to do quick troubleshooting on.

UV signals of four cycles (column 1 and column 2)

Seamless scale-up of a continuous capture process

17

Contichrom

CUBE

© LEWA - Bioprocess Group and

EcoPrime

Twin

UV signals of four cycles (column 1 and column 2)

Seamless scale-up of a continuous capture process

18

Contichrom

CUBE

© LEWA - Bioprocess Group and

EcoPrime

Twin

Shown on this slide is an example of what you can expect when scaling up to the Twin from the

CUBE. The transfer of the method operation is seamless and reproducible.

Contents



19


– Conclusions

– A simple set up



Batch Capture Total CV 7.85 L

Continuous Capture Total CV 1.6 L

Simple transfer from batch to continuous capture

Customer A - process development run comparison with 1g/L titer mAb

Essentially same process steps = simple process transfer

Higher load volume 40 vs 49 CVs = higher resin utilization & reduce buffer

consumption

Shorter residence times = faster loading & shorter processing time

Linear

[cm/h]

Column Equilibration 3 250

Load 40 150

Wash 1 2 150

Wash 2 2 250

Wash 3 3 250

Elution 3 250

post wash 1 3 250

post wash 2* 2 250

Regen* 3 250

CV

Process time: 9 h Cycle time: 4 h at linear velocity of 250 cm/h

CV

Load Start-Up 15

Load Connected 27

Load Parallel 22

Wash 1 connected 1

Wash 1 Parallel 1

Wash 2 2

Wash 3 3

Elution 3

Post-Wash1 2

Post Wash 2 1

Regeneration 3

Re-Equilibration 5


Batch Capture Total CV 7.85 L

Continuous Capture Total CV 1.6 L

Simple transfer from batch to continuous capture

Customer A - process development run comparison with 1g/L titer mAb

Process time: 9 h Cycle time: 4 h at linear velocity of 250 cm/h

In the first case study, the customer transferred their traditional mAb purification platform batch

capture step to a continuous capture step on the LEWA EcoPrime Twin for a 1 g/L titer mAb.

Essentially the same steps are employed when transferring from a batch process to a continuous

capture process so the method or recipe transfer is easy and straightforward. You will notice that with

the continuous capture method, higher load volumes are possible (49 CV and compared to 40 CV)

since the breakthrough from one column on the second column. The result is higher resin utilization

and reduced buffer consumption. In addition, because the breakthrough can be captured on the

second column, less residence time is needed when loading the column, so faster loading results in

shorter processing time.


EcoPrime Twin continuous capture reproducibility

Customer A –1g/L titer mAb: 50 g/L load

• At steady state, variability of peak area is <1 %.

• Steady state can be reached with first elution.

Column 1 Column 2

1 0.250 0.294

2 0.263 0.297

3 0.265 0.297

4 0.265 0.297

5 0.265 0.297

Average 0.264 0.297

error %* 0.639 0.025

SD 0.0064 0.0014

AreaPeak

5 peaks overlaid

Column 1

Column 2


EcoPrime Twin continuous capture reproducibility

Customer A –1g/L titer mAb: 50 g/L load 5 peaks overlaid

Column 1

Column 2

The data on this slide demonstrates the reproducibility of five cycles of a continuous capture run with 1

g/L mAb. A load of 50 g/L was processed with peak area variability of less than 1% for both columns.

Some notes about this work:

• No process development work was done. The first elution peak is slightly under loaded because the first load of

column was based on an assumed titer. A steady state was reached at the second cycle, but with proper

process development, steady state is easily reached at the first cycle.

• In comparing the elution profiles from columns 1 and 2, the differences in the intensity of the two UV signals is

due to a calibration issue.


Continuous capture delivers higher productivity

with less buffer consumption

Customer A - process development run: 1g/L titer mAb

Triple productivity or 67% less resin and ~ 30% buffer reduction


Continuous capture delivers higher productivity

with less buffer consumption

Customer A - process development run: 1g/L titer mAb

When comparing the platform batch process to the continuous capture process where the load was

increased from 40 to 50 g/L resin, either productivity was tripled or 67% less resin is used while using

30% less buffer. This is a significant increase in productivity or resin utilization as compared with the

traditional batch process.


EcoPrime Twin scale-up estimate

Customer A – 1 g/L titer estimate for 100-kg process scale-up

Savings up to $900K in resin or 22 days in processing time.



Customer A – 1 g/L titer estimate for 100-kg process scale-up

This graph demonstrates scale up estimates for both traditional batch and continuous capture

processes that produce 100-kg of purified product based on Customer A’s mAb where operation can

be optimized for either productivity or resin utilization.

Scenario 1 – When using the same amount of resin, the 100-kg of product can be produced in 10

days with a continuous capture process as compared to 32 days with the traditional batch capture

step saving 22 days for processing time.

Scenario 2 – When using the same processing time or throughput (10 days), $900,000 of saving in

resin cost is realized when running the continuous capture process.


Batch captureLoad 50 g/L resin

Total CV 17.3 L

Continuous capture Load 80 g/L resin

Total CV 1.6 L

Seamless transfer from batch to continuous capture

Customer B – pilot plant process comparison; 5 g/L titer mAb

CVlinear

[cm/h]

Pre‐Sanitization 3 400

Equilibration 3 400

Load 4.6 100

Load interconnected 11.4 150

Wash 1 2 400

Wash 2 5 400

Wash 3 3 400

Elution 3.5 400

CIP 3 400

Re-equilibration 3 400

Sanitization 3 400

Neutralization 2 3 400

Storage 3 400

CVlinear

[cm/h]

Pre‐Sanitization 3 300

Equilibration 3 300

Load 10 150

Wash 1 2 300

Wash 2 5 300

Wash 3 3 300

Elution 5 300

CIP 3 300


Sanitization 3 300


Storage 3 300

Same process steps = simple process transfer

Faster linear velocity = shorter processing time

Process time: 4.5 h Cycle time: 2.5 h


Batch captureLoad 50 g/L resin

Total CV 17.3 L

Continuous capture Load 80 g/L resin

Total CV 1.6 L

Seamless transfer from batch to continuous capture

Customer B – pilot plant process comparison; 5 g/L titer mAb

Process time: 4.5 h Cycle time: 2.5 h

In the second case study, Customer B used the Contichrom CUBE to obtain initial process

parameters. The process was then scaled-up and optimized on the EcoPrime Twin. To verify the

ability to scale-down, the process from the Twin was re-run on the CUBE.

Shown here is the pilot plant process (5 g/L titer mAb) that was transferred from batch to continuous

capture on the EcoPrime Twin. The process optimization included an increase in load from 50 g/L

resin to 80 g/L resin when going from the batch to the continuous capture process while also

increasing the linear velocity for faster processing.


Reproducible scale up of 5 g/L mAb capture

EcoPrime Twin run with 14-h process time

peak area peak area

1 0.239 0.276

2 0.255 0.283

3 0.259 0.285

4 0.260 0.286

average 0.2583 0.2849

error %* 1.1 0.7

Column 1 Column 2peak #

Variability of peak

area is ~1 % at

steady state

Contichrom Cube run with 12-h process time

Steady state can be

reached with first

elution.Similar chromatograms when scaling up


Reproducible scale up of 5 g/L mAb capture

EcoPrime Twin run with 14-h process time

Contichrom Cube run with 12-h process time

Similar chromatograms when scaling up

Shown here is evidence of reproducible scale from the CUBE to the EcoPrime Twin where steady

state continuous operation is reached during the first elution cycle on the Twin. Peak area

reproducibility is ~1% with the pilot scale Twin run.


Comparison batch to continuous capture

Customer B - process development run: 5 g/L titer mAb

> 2X productivity or 50 % less resin and ~ 50 % buffer reduction

0

5

10

15

20

25

Batch EcoPrime Twin Contichrom Cube

8.4

20.622.2

Pro

ductivity [

g/L

resin

/h]

Productivity - Customer B

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8


0.73

0.36 0.34

Buff

er

Co

nsum

ption [

L/g

]

Buffer Consumption - Customer B


Comparison batch to continuous capture

Customer B - process development run: 5 g/L titer mAb

0

5

10

15

20

25


8.4

20.622.2

Pro

ductivity [

g/L

resin

/h]

Productivity - Customer B

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8


0.73

0.36 0.34

Buff

er

Co

nsum

ption [

L/g

]

Buffer Consumption - Customer B

When comparing the platform batch process to the continuous capture process on both the EcoPrime

Twin and CUBE where the load was increased from 50 to 80 g/L resin, either 2x productivity is

realized or 50% less resin is used while using approximately 50% less buffer.



Customer B – 5 g/L titer estimate for 100-kg process scale-up

Savings up to $320K in resin or 6.1 days in processing times



Customer B – 5 g/L titer estimate for 100-kg process scale-up

This graph shows the scale up estimates for both traditional batch and continuous capture processes

that produce 100 kg of purified product based on Customer B’s 5 g/L titer mAb where operation can

be optimized for either productivity or for resin utilization.

Scenario 1 – When using the same amount of resin, the 100-kg of product can be produced in 5 days

with continuous capture as compared to 11.5 days with the traditional batch capture process.

Scenario 2 – When using the same processing time or throughput (5 days), $320,000 of saving in

resin cost is realized when running the continuous capture process.

Contents– Our approach to multi-column capture chromatography


36


– Conclusions

– A simple set up


Simple design, simple to set up & run

© LEWA Nikkiso America Inc. 37

Video Link Here

Set up / run system <1 day

../../Videos/EP Twin Commission 8 Hours Simple V3.2.mov


https://www.youtube.com/watch?v=6v80YZYw7zs&t=5s

Simple design, simple to set up & run

© LEWA Nikkiso America Inc. 38

Video Link Here

Set up / run system <1 day

Because of the simple, straightforward design, the EcoPrime Twin is simple to setup and run. Just

attach to the utilities and columns and you are set to go.

The following is a link to video demonstrating the simple setup. Just copy/paste the link into your

browser.





Contents



39


– Conclusions

– A simple set up


Customer experience on mAb shows

Twin column capture delivers:

40

• Significant productivity gains

Better Protein A utilization

• Substantial cost or time savings

Faster loading

Shorter processing time

Reduced buffer consumption

• Seamless scale up

Same process steps

Proven reproducibility

• Simple 2-column approach and system

Accelerate implementation

Minimize risk

© LEWA Nikkiso America Inc.

Customer experience on mAb shows

Twin column capture delivers:

41

• Significant productivity gains

Better Protein A utilization

• Substantial cost or time savings

Faster loading

Shorter processing time

Reduced buffer consumption

• Seamless scale up

Same process steps

Proven reproducibility

• Simple 2-column approach and system

Accelerate implementation

Minimize risk

© LEWA Nikkiso America Inc.

The two customer case studies that were presented demonstrate that for a typical mAb capture step

significant productivity gains and better resin utilization are realized when compared to traditional

batch operation.

Substantial cost or time savings are achieved with faster loading, shorter processing times and

reduced buffer consumption.

Scale up (or down) from the Contichrom CUBE is easy and reproducible because the same process

steps are used with directly scalable column sizes.

All of this is achieved on the EcoPrime Twin LPLC, a simple 2-column system where the less complex

design minimizes downtime, lowers operating cost, and accelerates validation and implementation in

manufacturing.

Contents



42

– Two customer case studies

– Conclusions

– A simple set up


LEWA Bioprocess Group

– Engineering

– Fabrication

LEWA-Nikkiso

America, Inc.

– Sales

– Engineering

– Fabrication

LEWA GmbH

Headquarters

Industrial Division

– Sales

– Engineering

– Pump production

– Fabrication

Nikkiso Co., Ltd.

Group

Headquarters

LEWA Bioprocess Group is part of the global family of companies

under the $1.3B US Nikkiso Corporation (1953)

Business sites in >80 countries, >6,000 employees

LEWA Bioprocess Group

– Engineering

– Fabrication

LEWA-Nikkiso

America, Inc.

– Sales

– Engineering

– Fabrication

LEWA GmbH

Headquarters

Industrial Division

– Sales

– Engineering

– Pump production

– Fabrication

Nikkiso Co., Ltd.

Group

Headquarters

LEWA Bioprocess Group is part of the global family of companies

under the $1.3B US Nikkiso Corporation (1953)

Business sites in >80 countries, >6,000 employees

The LEWA Bioprocess group located in Devens, MA is part of the global family of companies of

Nikkiso. With facilities and subsidiaries in 80 countries, we are able to provide support and service for

our products across the globe.

6/21/2017

© LEWA Process Technologies, Inc.45

6/21/201745

GMP systems engineering, manufacturing & automation;• Chromatography• Buffer In-line Dilution• TFF• Downstream process skidsMarkets:• Biopharmaceutical• Pharmaceutical• OEM (bio/pharma)

Devens, MA USA

2000

Acquired in 2014, we are now agroup within LEWA-Nikkiso America, Inc.

The LEWA group ($280M US) is a manufacturer of metering pump headquartered in Leonberg, DE.LEWA was acquired by Nikkiso in 2008. Both Nikkiso & LEWA founded ~65 years ago

LEWA Bioprocess Technologies Group

A group of ~ 60+ professionals dedicated to GMP scale bioprocess systems’ design and manufacture.

Part of the ~ 6,000 employees of Nikkiso Corp. – a diversified company with $1.3B in revenue.

Location

Date of Establishment

Ownership

About LEWA

Video Link Here

Who we are, what we do

https://www.youtube.com/watch?v=6IS04Iso-_s&t=39s

6/21/2017

© LEWA Process Technologies, Inc.46

6/21/201746

GMP systems engineering, manufacturing & automation;• Chromatography• Buffer In-line Dilution• TFF• Downstream process skidsMarkets:• Biopharmaceutical• Pharmaceutical• OEM (bio/pharma)

Devens, MA USA

2000

Acquired in 2014, we are now agroup within LEWA-Nikkiso America, Inc.

The LEWA group ($280M US) is a manufacturer of metering pump headquartered in Leonberg, DE.LEWA was acquired by Nikkiso in 2008. Both Nikkiso & LEWA founded ~65 years ago

LEWA Bioprocess Technologies Group

A group of ~ 60+ professionals dedicated to GMP scale bioprocess systems’ design and manufacture.

Part of the ~ 6,000 employees of Nikkiso Corp. – a diversified company with $1.3B in revenue.

Location

Date of Establishment

Ownership

About LEWA

Video Link Here

Who we are, what we do

The LEWA Bioprocess group is a center of excellence dedicated to developing and manufacturing

automated systems for use in GMP scale bioprocessing.

Link to video. Copy/paste the link into your browser.



A sampling of systems we produce at LEWA Bioprocess

A sampling of systems we produce at LEWA Bioprocess

In addition to process LPLC systems for both traditional and multi-column chromatography, we

manufacture a variety of other systems, including process HPLC, buffer and formulation preparation

systems, TFF, and cell harvest systems all with our control systems and automation software.

LEWA, Creating Fluid Solutions, the LEWA logo, LEWA Intellidrive and LEWA EcoPrime are trademarks of LEWA GmbH.

ChromaCon, Contichrom and CaptureSMB are registered marks of ChromaCon AG.

All goods and services are sold subject to the terms and conditions of sale of the company within LEWA which supplies them. A copy of these terms and conditions is available on request. Contact your local LEWA representative for the most current information.

© 2017 LEWA – Nikkiso America, Inc. – All rights reserved.

For more information: www.lewapt.com

Headquarters LEWA GmbHUlmer Str.1071229 Leonberg GERMANYTel: +49 7152 14-0Fax: +49 7152 [email protected]

LEWA Bioprocess Technologies, Inc. 8 Charlestown RoadDevens, MA 01434 USATel: +978 487 [email protected]

NIKKISO CO. LTD.Yebisu Garden Place Tower 22nd Floor20-3, Ebisu 4-Chome, Shibuya-ku150-6022 TokyoTel: +81 3 3443-3711Fax: +81 3 3473-4963

LEWA-Nikkiso America, Inc.132 Hopping Brook RoadMA 01746 HollistonTel: +1 508 429-7403Fax: +1 508 [email protected]

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