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Quality Control and Downstream Processing of Therapeutic Enzymes

  • David GervaisEmail author
Chapter
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 1148)

Abstract

Therapeutic enzymes are a commercially minor but clinically important area of biopharmaceuticals. An array of therapeutic enzymes has been developed for a variety of human diseases, including leukaemia and enzyme-deficiency diseases such as Gaucher’s disease. Production and testing of therapeutic enzymes is strictly governed by regulatory bodies in each country around the world, and batch-to-batch consistency is crucially important. Manufacture of a batch starts with the fermentation or cell culture stage. After expression of the therapeutic enzyme in a cell culture bioreactor, robust and reproducible protein purification, or downstream processing (DSP) of the target product, is critical to ensuring safe delivery of these medicines. Modern processing technology, including the use of disposable processing equipment, has greatly improved the DSP development pathway in terms of robustness and speed to clinic. Once purified, the drug substance undergoes rigorous quality control (QC) testing according to current regulatory guidance, to enable release to the clinic and patient. QC testing is conducted to ensure the safety, purity, identity, potency and strength of the medicinal product, requiring multiple analytical methods that are rigorously validated and monitored for robust performance. Several case studies, including L-asparaginase and asfotase alfa, are discussed to illustrate the methods described herein.

Keywords

Therapeutic enzymes Downstream processing Quality control Enzyme characterisation Enzyme manufacturing 

Abbreviations

2D-GE

Two-dimensional gel electrophoresis

ABG

Acid β-glucosidase

ADA

Anti-drug antibodies

ADC

Antibody-drug conjugates

AUC

Analytical ultracentrifugation

CFU

Colony-forming units

CGE

Capillary gel electrophoresis

CHO

Chinese hamster ovary

cIEF

Capillary isoelectric focussing

CM

Carboxymethyl

CNS

Central nervous system

DEAE

Diethyl amino ethyl

DNA

Deoxyribonucleic acid

DOE

Design of experiments

DP

Drug product

DS

Drug substance

DSP

Downstream processing

ELISA

Enzyme-linked immunosorbent assay

EMA

European medicines agency

ERT

Enzyme replacement therapy

EU

Endotoxin units

Fc

Constant domain (antibody)

FDA

United States Food and Drug Administration

GD

Gaucher’s disease

GMP

Good manufacturing practice

HCP

Host cell proteins

HER2

Human epidermal growth factor receptor 2

HIC

Hydrophobic interaction chromatography

HPLC

High-pressure liquid chromatography

HPP

Hypophosphatasia

HVAC

Heating, ventilation and air conditioning

ICH

International Council on Harmonisation

IEX

Ion-exchange

IEX-HPLC

Ion-exchange high-pressure liquid chromatography

IgG

Immunoglobulin

IMAC

Immobilised-metal affinity chromatography

ITC

Isothermal calorimetry

IU

International unit (of enzymatic activity)

kcat

Enzyme catalytic constant

KM

Michaelis constant

LAL

Limulus amoebocyte lysate

LAL-D

Lysosomal acid lipase deficiency

LC-MS

Liquid chromatography coupled mass spectrometry

LOD

Limit of detection

mAbs

Monoclonal antibodies

MALDI

Matrix-assisted laser desorption/ionisation

MF

Microfiltration

MS

Mass spectrometry

MS/MS

Tandem mass spectrometry

NAD

Nicotinamide adenine dinucleotide

NADP

Nicotinamide adenine dinucleotide phosphate

NCPPB

National Collection of Plant Pathogenic Bacteria (UK)

PCR

Polymerase chain reaction

PEG

Poly(ethylene glycol)

pI

Isoelectric point

PMDA

Japanese Pharmaceuticals and Medical Devices Agency

Q

Quaternary ammonium

QA

Quality assurance

QbD

Quality by design

QC

Quality control

QMS

Quality management system

qPCR

Quantitative polymerase chain reaction

R&D

Research and development

RP-HPLC

Reversed-phase high-pressure liquid chromatography

RT-PCR

Real-time polymerase chain reaction

S

Sulphopropyl

SDS-PAGE

Sodium dodecyl sulphate polyacrylamide gel electrophoresis

SEC

Size-exclusion chromatography

SOP

Standard operating procedures

SPR

Surface plasmon resonance

TFF

Tangential-flow filtration

TFF-MF

Tangential-flow filtration – microfiltration

TNALP

Tissue-nonspecific alkaline phosphatase

U

Units (of enzyme activity)

US

United States

UV

Ultraviolet

Vmax

Maximum enzymatic reaction velocity

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© Springer Nature Singapore Pte Ltd. 2019

Authors and Affiliations

  1. 1.Porton Biopharma LimitedSalisburyUK

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