An introduction to a novel filtration system

ATF-manufacturing PlatformMicrocarrier Process Unit Operations

VaccinesGene TherapyStem Cells

Microcarrier Screen Filter Module

• All USP Class VI materials (Polysulfone, Polyester, epoxy, 316L SS)

• 70um screen

• Special flow path design to reduce fouling and attachment

• Multiple low shear, rapid, separations, without settling:

– Media exchanges, wash steps

– Seed transfer

– Harvest: cell debris & virus filtration from microcarriers

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Microcarriers: Process Overview

SIP: PBS & ucarriers

Wash ucarriers

Innoculate

Batch/perfusion growth

Trypsinization

Wash

Cell transfer

Reactor seed train 5x volumes

SIP: PBS & ucarriers

Wash ucarriers

Innoculate

Batch/perfusion growth

Trypsinization

Wash

Cell transfer

SIP: PBS & ucarriers

Wash ucarriers

Innoculate

Batch/perfusion growth

Media reduction

Infection

Media addition

Harvest

50+ flasks…

X XMaybe ?

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• Major advantage is liquid and microcarrier

handling: ease and speed to remove or

transfer media or cells– Rapid liquids removal without shear

– Wash steps for carriers

– Low shear cell/seed transfers

– Rapid Harvest

– Microcarriers do not need to settle

• Cell ”health” improved, allowing higher

productivity per cell

• Media removal can be to a low volume

• Simple to operate and scales to manufacturing

• External system which allows easy exchange

of filter if required

Microcarrier Processes: ATF System Advantages

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Microcarriers: ATF2 System Setup

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Microcarriers: ATF10 System Setup

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1. Reactor filled with carriers and PBS

2. Reactor is SIP’d

3. ATF is autoclaved

4. Steamed connection made

1. PBS removed at 1vv/hr for 40-50 mins

2. Addition of media

3. Diafiltrate for 5 minutes

4. Close harvest, media added to working volume

5. Innoculate with cells

ATF rate constant

at 4-5x filtrate rate

Microcarriers: Preparation & Washing

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1. Perfusion carried out at small or large scale

2. Rates at 0.5vv/day to 4vv/day

3. ATF rate at 30+ higher than filtrate rate

Microcarriers: Perfusion

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1. Cell growth to maximum in batch or perfusion mode

2. Media reduction at 1vv/hr for 30-40 mins

3. Trypsinise cells

4. Harvest cells through ATF leaving behind carriers

5. Diafiltrate with new media, to capture maximum number of cells

6. Cells transferred directly to next reactor

7. ATF rate at 3-5x filtrate rate

Microcarriers: Seed Transfer

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1. Remove media at 1vv/hr for 40-50 mins

2. Addition of new media

3. Infect cells with virus

4. Optional: Perfusion

1. Harvest cells and debris at 1 vv/hr for 40-50 mins

2. Diafiltrate for 10-20 minutes

3. Optional: Clarification with ATF and 0.2u filter on holding tank

ATF rate constant

at 4-5x filtrate rate

Microcarriers: Media Exchange, Infection, Harvest

Production reactor

0.2u or 0.5u filtered stream

Clarified Harvest

Adherent Cells: Combined DSP

Vaccine Process – Adherent CellsBuffer, Wash

70u filtered stream

Microcarrier-free Harvest

Ionic Solution(hc-dna precipitation)