C Pharmacology & Toxicology 2000, 86, Supplement I, 6–7. Copyright CPrinted in Denmark . All rights reserved

ISSN 0901-9928

Preclinical Safety Assessment: In vitro – in vivo TestingFlemming Højelse

Non-Clinical Safety Research, H. Lundbeck A/S, Ottiliavej 9, DK-2500 Valby, Denmark

Abstract: In vitro – and in vivo preclinical safety tests on drug candidates needed before first dose in man and beforemarketing authorisation are as follows: The acute and repeated dose toxicity studies, the reproductive toxicity studies, thegenotoxicity studies, the carcinogenicity studies and finally the safety pharmacology studies. The Safety Assessment ofthe results with respect to predictability for humans is discussed, as well as new tests under validation. Suggestions forchanges in the future of Non-Clinical Safety tests are mentioned.

Preclinical safety testing of a drug candidate starts 1–2years before first dose in man and cast 5–6 years. The resultsof the last study are available a few months before com-pletion of the clinical testing.

Preclinical safety testing is constantly being developed inorder to improve the safety assessment of a drug candidatebefore clinical testing and marketing authorisation.

The LD50 test in rodents has now been replaced by anacute toxicity test, where the endpoint, instead of lethality,is clinical signs and determination of target organs/tissueafter administration of very high doses. The acute toxicitytesting should also focus on a recommendation, how medi-cal doctors should treat cases of overdosing and perhaps re-commend an antidote. In the future the acute toxicity testmight be replaced by the safety pharmacology tests men-tioned below. This however, requires a validation of thesetests with respect to predictability for human safety.

For many years the repeated dose toxicity testing in ro-dents and non-rodents have been carried out and used forsafety assessment of drug candidates. These tests are pivotalto highlight the target organs/tissues and to establish a ‘‘NoObserved Effect Level’’ (NOEL) for calculation of thesafety margin between exposure to patients and the NOELin experimental animals. These tests should not and prob-ably will not be replaced in the future, but new endpointswill be introduced in the studies, as new techniques and newsafety diagnostic assays are being developed. The mechan-ism behind changes in the target organ needs to be eluci-dated before the relevance to man can be assessed.

The effects of drug candidates on reproduction such asfertility, embryo-foetal development and pre- and postnataldevelopment, including effects on the mother have been infocus since the thalidomide disastrous event forty years ago.The design of these studies has been changed to optimizeand to harmonize. ICH (International Conference on Har-monisation) plays a very important part, and today, thesafety assessment of a drug candidate of effects on repro-duction is valid and nothing like the thalidomide event has

Author for correspondence: Flemming Højelse, Non-clinical SafetyResearch, H. Lundbeck A/S, Ottiliavej 9, DK-2500 Valby, Denmark.

occurred since then (CPMP/SWP/386/95). These tests last1–2 years and much effort has been invested in findingshort-term in vitro tests. ECVAM (European Centre of theValidation of Alternative Methods) conducts an interlab-oratory prevalidation trial, testing 3 in vitro systems for em-bryonic development (Brown 1995). However, the develop-ment of a foetus is so complex that a simple in vitro modelis difficult or perhaps impossible.

ICH also plays a major part with respect to genotoxicitytests, which has resulted in a global strategy for safety geno-toxicity testing of drug candicates before first dose in manand marketing authorisation (CPMP/ICH/141/98). Manynew genotoxicity tests are being developed and validationof these tests is a continuous process. A combination ofrepeated dose toxicity tests and chromosome-damaging testlooks promising, and so do techniques involving transgenicanimals.

In the course of the last few years, the predictability ofand thus the necessity of two rodents (rat and mouse)studies for the carcinogen potential testing of a drug candi-date has been questioned. ICH recommends an alternativeapproach: one rodent (rat) carcinogenicity test with life-long dosing (standard test of today) and a short-term ormedium-term rodent (mouse) study, which might revealmechanisms of carcinogenicity. Several of these short-termmouse tests and the use of transgenic animals are being vali-dated now, and in a few years they will be the standard tests.

Safety pharmacology or secondary pharmacology is adiscipline, where safety assessment on vital major bodysystems/functions is carried out. The dosing may be singledose or dosing escalating over a few hours until a maxi-mum tolerated dose is reached. Before, during and afterdosing, vital functions are monitored by optimal methods,often developed for use in man (CPMP/SWP/872/98). Thepredictability of several of these tests on respiratory andcardiovascular functions is high and due to a lot of inter-est from the pharmaceutical industry, the authorities andscientists will increase the knowledge. ICH and many au-thorities now discuss the design and which test resultsshould be used before first dose in man.

The side effects or the toxicity of a drug candidate ob-

7PRECLINICAL SAFETY ASSESSMENT: IN VITRO – IN VIVO TESTING

served in preclinical/non-clinical safety testing is never buta surprise, it is always a chock, even when the structure andthe pharmacological mode of action have been carefullyconsidered. We definitely need preclinical safety testing be-fore first dose in humans and marketing authorisation, thusnew tests will be developed for safety screening purposes.When validation and predictability for human safety hasbeen demonstrated, these tests will be part of the standardnon-clinical safety battery, accepted by regulatory authorit-ies. In the future some of these tests will replace some ofthe tests of today, but authorities need to be conservative,since their role is the protection of patients. New tests willcontinuously be added to the battery and before too long,we will no doubt see some replacement.

References

Brown, N. A., H. Spielman, R. Bechter et al.: Screening chemicalsfor reproductive toxicity, the current alternatives. The report andrecommendations of an ECVAM/ETS workshop (ECVAM work-shop 12). ATLA 1995, 23, 868–882.

ICH Harmonised Tripartite Guideline, CPMP Guideline for– Carcinogenicity studies of pharmaceuticals (CPMP/ICH(141/95)– Genotoxicity tests for pharmaceuticals (CPMP/ICH/141/95)– Detection of toxicity to reproduction for medicinal products

(CPMP/SWP/386/95)– Note for guidance on safety pharmacology studies in medicinal

product development (draft from 13 May, 1998) (CPMP/SWP/872/98).