J . C'hem. Tech. Biotechnol. 1991, 51, 115-140

SCI Biotechnology Group Meeting Analytical Methods in Biotechnology The following are summaries of papers presented at a meeting of' the SCI Biotechnology Group held on 13 December 1989 at the Society of Chemical Indusrry, 14 Belgratie Square, London S W I X 8PS. The papers published here are entirely the responsibilitj~of the authors und do not reflect the views of' the Editorial Board of the Journal of Chemical Technology and Biotechnology.

Analytical Methods in Biotechnology: Introduction

C. R . Hill Celltech Limited, 216 Bath Road, Slough, Berkshire SLI 4EN. U K

A key objective during the development of a protein pharmaceutical is to demonstrate the safety and efficacy of the product. Ultimately this can only be achieved through appropriately designed human clinical studies. However, before the product is introduced to humans it is essential to subject the product to a rigorous programme of characterisation studies. Guidelines for the testing of protein pharmaceutical products derived by DNA recombinant and hybridoma technology are given in documents prepared by the various National Regulatory Agencies.'-4 In comparison with small molecule drugs, protein pharmaceutical compounds are extremely complex molecules. Even small protein drugs such as human growth hormone have a molecular weight of 22 000. The protein molecule may contain a number of disulphide bonds and carbohydrate groups and a wide range of post-translational events may occur. The extent and type of such events can be influenced by the process used to manufacture the product.' The cell type used to express the product, the fermentation and purification process used to recover the product and subsequent formulation and storage prior to use can all influence the quality and structure of the product. Quality control of a protein pharmaceutical product must, therefore, be defined by extensive characterisation of both the product itself and the specific production process used in its manufacture. There is a wide range of potential impurities that may be associated with a protein product. These can be derived from any of the raw materials used in its manufacture or from the product itself. For example, protein impurities may be 115 .1. Chern. Tech. Biorrchnol. (51) (l991)-@'1 SCI. 1991 Printed in Great Britain

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SCI Biofechnoloyy Group Mreiiny

derived from the host organism, from media used in the fermentation process, from reagents used in the purification process or excipients used in the formulation. Impurities derived from the product can include proteolytically cleaved or aggregated protein and deamidated6 or amino-acid substituted forms.7 The presence of such impurities can potentially have severe consequences. They may act, for example, by affecting the stability of the protein drug itself or may themselves exert deleterious immunological or biological effects. A wide range of analytical techniques is required to address the complex technical issues raised above.*q9A number of the assays required form part of the traditional range of methods used to analyse protein purity (e.g. Sodium Dodecyl Sulphate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) and isoelectric focussing). However, more recently there have been a number of technical advances in assay technology. This has been due, to a large extent, to the increasing need to analyse and characterise the new protein pharmaceutical products produced by the Biotechnology Industry. These developments range from improvements in sensitivity of traditional assays such as SDS-PAGE to the application of techniques such as Mass Spectrometry and Nuclear Magnetic Resonance (NMR) to the analysis of protein structure. The objective of this meeting was to discuss a number of the new methods as they apply to the analysis of potential therapeutic proteins. The meeting was opened with a discussion on some of the challenges facing both the regulatory authorities and the Biotechnology Industry in the development and manufacturing of therapeutic peptides and proteins. The methods discussed ranged from electrophoretic and N - and C-terminal sequencing techniques, to the use of immunoassay methods for protein impurity detection and the application of mass spectrometry and NMR techniques to the characterisation of peptide protein and glycoprotein structure and oligosaccharide sequence. For each technique the authors have reviewed the current capabilities of the technique and then considered their future potential for use in characterisation of protein pharmaceuticals.

References 1 . Office of Biologics Research and Review, Points to Consider in the Mutzufirc.rurc, tind Testing of' Monoclonul Antibody Products ,/Or Huwiun Use. Food and Drug Administration, Bethesda, Maryland, 20892, USA, 1987. 2. Australian Department of Community Services and Health, Guiclrlirws f i j r /lie Prepurution and Presentation of' Applicutions ,for General Murketing qf' Monochmrl Antibodies Intended ,/br Use in Humans, ADCSH, G P O Box 9848, Canberra, ACT 2601, Australia, 1987. 3. Committee for Proprietary Medicinal Products, Guidelines on the production and quality control of Medicinal products derived by recombinant DNA technology. Triwds Biotechno/., 5 (1988) Gl-G4. 4. Committee for Proprietary Medicinal Products, Guidelines on the production and quality control of monoclonal antibodies of murine origin intended for use in humans. Triwds Biotidinol., 6 (1988) G5-G8. 5 . Utsumi, J., Mizono, Y., Hosoi, K . , Okana, K.. Sawada, R., Kajitani, M., Sakai, J., Naruto, M. & Shimizu, H . , Characterisation of four different mammalian-cell-derived recombinant human interferon-Beta's: identical polypeptides and non-identical

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carbohydrate moieties compared to natural ones. Eur. J . Biochm~.,259 (1989) 679@7. 6. Hancock, W. S., Canova-Davis, E., Chloupek, R. C.. Therapeutic Peptides and Proteins: Assessing the New Technologies, In Bunbury Report 29, 1988, Cold Spring Harbour, pp. 95-107. 7. Bogosian, G., Violand, B . N., Dornward-King, E. J., Workman, W. E., Jung, P. E. & Kane, J. F., Biosynthesis and incorporation into protein of norleucine by Eschrrichia coli. J . Biol. Chrm., 264 (1989) 531-9. 8. Anicetti, V. R., Keyt, B. A. & Hancock, W. S., Purity analysis of protein pharmaceuticals produced by recombinant DNA technology. Trends Biotechnd., 7 (1989) 342-9. 9. Garnick, R. L., Solli, N. J. & Papa, P. A., The role of quality control in biotechnology: an analytical perspective. Anal. Chern.. 60 (1988) 2456-557.

Therapeutic Peptides and Proteins-Challenges for the Regulatory Authorities

S. L. Jeffcoate National Institute for Biological Standards and Control. Potters Bar. UK

This presentation discusses how the recent developments in the manufacturing processes, purification procedures and analytical test methods for the products of biotechnology have created particular challenges for the authorities that control and regulate these products in order to protect public health. Particular reference is made to hormones which currently dominate the ‘Biotech Market’. The National Institute for Biological Standards and Control (NIBSC) (with its new facility which came into operation some two years ago) is responsible to the UK Department of Health for advising on licensing applications and for the on-going control of biological medicinal products. The UK is almost alone in the world in including hormones as ‘biologicals’ and many countries, including the USA and most European countries, make the mistake of classifying them as ‘chemical drugs’. One can now see that in the new era of biotechnology, substances over a wide range of types, from hormones through to vaccines, are made by essentially the same sorts of biotechnological processes and this has revealed the difficulties that can arise from outdated methods of classification. The work of NIBSC places the UK in a strong position in Europe, both for advising on licensing and for the on-going control of medicinal products made by the new biotechnology procedures. Peptide drugs are a challenge for the regulatory authorities. Regulatory authorities around the world are facing major challenges in the 1990s. These challenges are a result of a number of changes. For example, the rate at which the field is changing puts particular pressure on regulatory authorities to respond to changes and to ensure that products of appropriate purity and efficacy reach the market without undue delay. The watchword is ‘to make haste slowly’. The pace of change, the pathway from ‘clone to the clinic’, or from ‘notion to potion’, gets increasingly rapid and short. This is well exemplified by erythropoietin which is discussed below.

Analytical methods in biotechnology: introduction.

J . C'hem. Tech. Biotechnol. 1991, 51, 115-140 SCI Biotechnology Group Meeting Analytical Methods in Biotechnology The following are summaries of pap...
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