Pharmaceutical applications: Editorial overview

Pharmaceutical applications Editorial overview

Tim Harris and George Poste

Glaxo Group Research Ltd, Middlesex, UK and Smith Kline and Beecham, King of Prussia, Philadelphia, USA

Current Opinion in Biotechnology 1991, 2:211-214

Given the difficulty in defining what constitutes biotechnology, the reader is entitled to ask: "What is Pharmaceu- tical Biotechnology?". This editorial attempts to answer the question, and the reviews that comprise this section demonstrate the diversity of the topic and its profound impact on drug discovery and development. Many proteins with therapeutic potential have been produced, following the discovery in the 1970s that genes encoding any protein of interest could be cloned in E~ cherichia coli and expressed in a diverse array of heterol- ogous prokaryotic and eukaryotic cells, including mammalian cells (see Current Opinion in Biotechnology 1990, Vol 1, No 1). The commercial exploitation of heterolo- gous gene expression has been extremely rapid. A num- ber of genetically engineered proteins are already ap- proved for routine clinical use and an impressive reper- toire of recombinant DNA-derived proteins and novel drug delivery systems to improve drug localization are in various stages of clinical testing. In addition to the production of proteins for use as drugs or vaccines, other advances include new gene expression technologies and protein chemistry and engineering techniques. Coupling these with the automated synthesis of natural and syn- thetic genes and proteins, and with immunobiology and cell biology, we have a powerful set of experimental tools for improving our search for traditional drug classes (i.e. low-molecular-weight heterocyclic molecules). The ability to isolate specific proteins selected as 'targets' for therapeutic intervention provides the foundation for increasingly rational strategies for drug discovery, by allowing drug-target interactions to be studied at a molecular level to define the structural requirements for selective drug action. These parallel strategies of exploiting 'proteins as targets' and 'proteins as drugs' have led to a reawakening of interest in biotechnology. Strictly speaking, the use of biotechnology in the pharmaceutical industry is not new. Many antibiotics, starting with penicillin and continuing today, are natural products that either are generated by fermentation technology or require fermentation to make key intermediates. The isolation of natural products from micro-organisms and plants has also enabled important therapeutic advances to be made in the treatment of inflammation, hy- percholesterolemia, cancer and immune rejection.

It is probably a valid generalization, however, to state that the large established pharmaceutical companies were slow to recognize the implications of the advances in cell and molecular biology occuring twenty years ago in the creation of new opportunities for drug discovery. It required the explosive growth of new independent companies between 1975 and 1985 (capitalized by funding sources hitherto uninvolved in sponsoring biomedical research) that concentrated initially on protein therapeutics, to spur the established companies to recognize the potential of biotechnology. Their response was to seek to strengthen their own internal skills in cell and molecular biology and to forge alliances with the new companies via equity and collaboration.

The events of the past 15 years provide a fascinating illustration of the profound impact that a new technology has on industrial strategy and practices, involving the forging of new relationships between industry and academia and the increasing migration of biomedical researchers into industry. The vital role of the investment community in catalyzing the emergence of technologies that might oth- erwise have become stagnated, also provides a dramatic illustration of how to (and how not to) establish new companies to develop products for a particular niche market.

The accomplishments of this period provide a study of stark contrasts. On the one hand, biotechnology has cre- ated a new therapeutic vista and produced a genuine shift in approaches to drug discovery. Cell and molecular biology are now seen as indispensable tools that comple- ment the already substantial array of technical disciplines required for successful pharmaceutical Research and De- velopment. On the other hand, the emergence of the biotechnology industry has been accompanied by an un- precedented level of hyperbole and exaggerated claims. A tidal wave of press releases has trumpeted the immi- nent arrival of miraculous new cures for all major human diseases (the biotechnology panacea) and the pending demise of the established pharmaceutical industry. The legion of start-up companies that were founded in the 1980s with the objective of becoming 'FIPCO's' (fully in- tegrated pharmaceutical companies) face the 1990s with the less attractive reality: that of becoming NON-COs.

Abbreviations CSF~colony-stimulating factor; IL--interleukin; t-PA--tissue-type plasminogen activator.

(~) Current Biology Ltd ISSN 0958-1669 211

212 Pharmaceutical applications

Pharmaceutical biotechnology thus encompasses a diverse set of skills and is relevant to an array of therapeutic applications, all of which are the focus of intense interest in both academia and industry. These are summarized in Table 1.

It is impossible to survey the broad panorama of activ- ities illustrated in Table 1 within a single issue of Cur- rent Opinion in Biotechnology. We have therefore cho- sen to review topics in which significant recent research progress has been made and which provide realistic op- pomanities for translation from research to exploratory therapeutic evaluation over the coming years.

The first therapeutic proteins to be produced by genetic engineering and to reach the market place were insulin, growth hormone and a subunit (surface antigen) vaccine for hepatitis B virus. These successes have provided the paradigm that has been followed for other therapeutic macromolecules produced by recombinant DNA technology.

The hepatitis B virus surface antigen expressed in yeast and used as a vaccine against serum hepatitis is still the only example of a human subunit recombinant vaccine product. As Brown's review demonstrates (pp 215-219), this may be because a protective immune response against an infectious micro-organism is more compli- cated than the simple maintenance of high levels of neu- tralizing antibodies. Such a response critically depends on both T and B cell function, and the presentation of more than one epitope from one, or many, antigens may be necessary to induce effective protection against dis-

ease. In addition, the role of immune restriction phenom- ena, in which the level of immunity produced by a vaccine is influenced by the histocompatibility phenotype of the recipient, is just beginning to be mapped.

Several new therapies, based on novel proteins whose roles were only revealed by cloning and expressing their genes, have now been introduced. The production of tissue-type plasminogen activator (t-PA) represents a landmark for the biotechnology industry as production shifted from micro-organisms to expression in tissue cul- ture cells. However, as reviewed by Larsen and Bamathan (pp 220-226), knowledge of the molecular biology of t-PA has increased since then. Novel thrombolytic proteins are now being constructed to help gain an understanding of the complex interactions between specific activators and inhibitors in the control of vascular haemostasis.

The importance of the principle that the full function and therapeutic merits of endogenous mediators cannot be revealed until they are available for study, is also well illustrated by the colony-stimulating factors (CSFs), granulocyte CSF, granulocyte macrophage CSF and erythro- poeitin (Watanabe et al. pp 227-237), which are being used clinically to stimulate specific classes of haematopoi- etic cells. It is interesting that the initial ideas were to use other cytokines, such as interleukin (IL)-I, IL-6 and tumour necrosis factor in a similar way, but that the pleiotropic effects of these mediators, demonstrated in animal models and transgenic mice, has revealed severe side effects. The importance of ensuring that the action of such mediators is localized to the tissue affected by

Table 1. The aims of pharmaceutical biotechnology.

1. The production of endogenous mediators for direct use as therapeutics.

2. Chemical and/or genetic modification of endogenous mediators to improve their pharmaceutical and/or therapeutic properties.

3. The genesis of immunogenic proteins, improving antigen-processing and presentation, selective induction of specific immune repertoires and assessment of histocompatibility-mediated restriction of immune responsiveness.

4. The isolation and production of specific cellular proteins as 'targets' for drug action to improve the experimental study of drug-target interactions and design of drugs with even greater selectivity.

5. Selective modulation of gene expression to create new animal models of disease, to evaluate the toxicological risks associated with abnormal patterns of exposure to endogenous mediators and the potential use of genetically modified plants and animals as production sources for scarce therapeutic proteins.

6. Correction of genetic diseases.

7. Directed inactivation/activation of specific genes for therapeutic benefit and other strategies for sequence selective interaction of therapeutic ligands with nucleic acid targets.

8. The design of new drug delivery systems and pharmaceutical dosage forms for non-systemic delivery of peptides and proteins, improved targeting of both conventional and recombinant DNA-derived drugs to specific cell types and new strategies for administration of biological mediators in combination and/or defined sequence to mimic their physiological mode of action.

9. Design of small-molecular-weight drugs that duplicate the functions of biological macromolecules derived from a knowledge of the tertiary structure and mechanism of action of their 'active site' domains.

disease cannot be overstated and remains an unresolved challenge. It is now more likely that inhibitors of the function or synthesis of these proteins will be more useful as immunomodulatory agents than the proteins themselves.

Antibodies represent another major class of proteins with potential therapeutic value. Several companies have been established recently to develop therapeutic and diagnos- tic antibodies. This involves constructing chimeric or hu- manized versions of the proteins, either by removing immunogenic regions and reducing the induction of human anti-mouse antibodies or by augmenting activity (e.g. by joining the antibody to a toxin or another ef- fector molecule). These proteins require specific expression systems as described by Pltickthun (pp 238-246). The same systems are also essential for the expression of recombinant antibodies with enzymatic activity (so called catalytic antibodies or abzymes) for studying organic reaction mechanisms. Recombinant antibodies raised against specific cytokines provide valuable probes for selective immunomodulation. They will also be used to help to understand and treat problems such as autoim- mune disease and to provide insights into the humoral and cellular regulatory pathways involved in immunosup- pression and tolerance (Thomson, pp 247-253).

The delivery of protein pharmaceuticals by routes other than intravenous injection remains a major obstacle to their more widespread use in the treatment of diseases in non-hospitalized patients. Many of the proponents of the use of endogenous mediators for therapy have ignored totally the fundamental requirement of being able to de- liver such mediators in a way that allows their physiological action to be duplicated. Unless this requirement is met, the anticipated therapeutic benefit will not be re- alized and, equally important, new iatrogenic toxicities might arise as cell types within the body that are not or- dinarily exposed to the mediator encounter it.

Clearly, systemic administration can be used effectively for treatments in which the target resides within the bloodstream, or when the purpose is merely to duplicate the response of endocrine hormones or other blood- borne mediators. The problems imposed by the con- straint of systemic administration are far more challeng- ing, however, when the goal is to duplicate the action of autocrine and paracrine mediators that operate in ex- travascular tissue environments. First, the large size of the macromolecules is a serious handicap to the extravasa- tion necessary to yield therapeutic concentrations in the tissues. Second, the complexity of the dose response patterns and the fact that many of these agents act only in concert with other mediators, pose major challenges that cannot yet be duplicated reliably by current approaches to macromolecular drug delivery. Third, many autocrine and paracrine mediators are highly pleiotropic and evoke multiple biological responses in different cell types throughout the body. There are a variety of control systems in the body to ensure that the action of such mediators is restricted to the immediate vicinity in which they are required. These include the extremely short half- lives and rapid breakdown of mediators, down-regulation of receptors, and Induction of inhibitors to block further

Editorial overview Harris and Poste 213

synthesis of the mediator. Until these formidable problems in drug delivery are resolved, many of the claimed therapeutic applications of endogenous mediators will remain in the realm of unrealized promise.

The applicability of pharmaceutical proteins would be increased considerably if drug delivery systems could be developed to promote oral absorption. There has been much work in this area, but not many success stories (Ilium and Davies, pp 254-259). Coupled with this is the targeting concept (the 'magic biotechnological bul- let'). Drug delivery and targeting represent major challenges and the complexity of antibody targeting of toxic molecules illustrates the problems involved.

The reviews on growth factors and their receptors (Har- ris, pp 260-268) and small molecule receptors (Strange, pp 269-277) exemplify the other side of the biotechnology coin-- the use of proteins as targets for drug action and as enabling technologies for improved discovery of conventional drugs. The provision of cloned genes for receptors and ligands and their expression in many sur- rogate systems has provided important new insights into disease pathogenesis and drug action, teaching us new strategies for therapy. For example, measurement of the levels of various growth factors or their ligands using cloned probes in various cancers has provided the oncol- ogist with new ways of classifying tumours. Expression of particular genes can be correlated to malignant phenotype, disease progression and prognosis. Catecholamine receptors can now be classified by amino acid sequence homology, as well as by pharmacological profile. Cell lines expressing independent receptor subtypes can be used to study signalling systems and can be put to prac- tical use in identifying new antagonists and agonists. It is even possible to express mammalian receptors in yeast and to obtain signalling via the mating type response (Strange, pp 269-277).

Large amounts of purified growth factors can now be provided for those interested in studying their structure, although a major challenge exists in understanding the structure-function relationships in those receptors that are integral membrane proteins. There are many technical problems concerning expression and purification that remain to be solved before the structure of any therapeu- tically important seven-transmembrane-domain receptor protein will be determined at high resolution. The deter- mination of the structure of protein domains (expressed in yeast) may be an interim solution.

Another class of transmembrane protein affecting membrane transport processes has also come into the lime- light. The relationship between the multidrug resistance protein and bacterial transport proteins has been known for sometime. The review by Higgins (pp 278-281) puts recent studies into perspective and there is every possi- bility that new approaches to the therapy of multidrug resistance tumours will be designed as a result.

In addition to its valuable role in improving current approaches to the treatment of disease, biotechnology is also creating entirely novel opportunities for disease modulation. The ultimate form of disease therapy will

214 Pharmaceutical applications

come in the ability to control gene expression selec- tively and to practice therapeutic intervention by switch- ing genes on and off in a predictable fashion. Although this 'holy grail' remains far distant, the first steps of the journey have begun with the construction of antisense and triplex oligonucleotides, ribozymes and intercalating agents that recognize specific nucleotide sequences in nucleic acid species located in both the nucleus and the cytoplasm. The excitement of this new research area is reflected in the review by Crooke (pp 282-287) and rapid progress is being made in the development of oligonu- cleotide chemistry for therapeutic inhibition of gene expression. As emphasized earlier, the topics included in this issue provide only a sample of the myriad examples of the cre- ative application of biotechnology to human and veteri- nary therapeutics. All of these efforts are linked, however, by the fundamental recognition that molecular biology has forever changed the way in which disease is studied

and the search for new medicines is practiced. The description of disease in terms of specific alterations in gene structure and function has heralded the era of'reverse ge- netics'. Knowledge of the alterations in genomic function now provide a logical foundation for understanding disease pathogenesis in terms of the underlying molecular pathology, and the identification of increasingly rational therapeutic strategies directed against defined biochemi- cal pathways and molecular targets. We look forward to the description of further examples of these important new opportunities for pharmaceutical b/otechnology in future issues.

T Harris, Director of Biotechnology, Glaxo Group Research Ltd, Green- ford Road, Greenford, Middlesex UB60HE, UK.

G Poste, President of Research and Development, Smith Kline and Beecham, 709 Swedeland Road, King of Prussia, Philadelphia 19406- 2799, USA.

Pharmaceutical applications: Editorial overview

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