REGULATORY TOXICOLOGY AND PHARMACOLOGY 6,66-72 ( 1986)

A Critical Appraisal of the Duration of Chronic Animal Toxicity Studies

CYNTHIA E. LUMLEY AND STUART R. WALKER

Centrefor Medicines Research, Woodmansterne Road, Carshalton. Surrey, England

Received December 16. 1985

One method of assessing the contribution of studies of longer than 6 months to a safety evaluation program is to compare retrospectively the findings in toxicity tests carried out for 6 months or less with those observed after 6 months. The Centre for Medicines Research has therefore established a databank comprising animal toxicological data obtained from pharmaceutical companies in Europe. Twenty-one companies have provided data for 124 compounds (2 14 studies), including 88 studies of 65 compounds where comparable short-term (~6 months) and long-term (>6 months) data are available. The results from the 88 studies show that, excluding the possibility of identifying carcinogens, tests of longer than 6 months have not added to the overall safety evaluation of these compounds. @ 1986 Academic press Inc.

INTRODUCTION

The pharmaceutical industry conducts extensive toxicity tests in animals during the development of new medicines. The design of these tests must take into account the guidelines suggested by regulatory authorities in the countries where it is hoped to market the drug. These guidelines include recommendations for the minimum duration of animal exposure, which depends upon the proposed duration of human use. However, there is disagreement regarding the amount and nature of animal testing required before a medicine can be used “ethically and safely” in the clinic (Fig. I) (Alder et al., 198 1; Health Protection Branch 198 1; Kesterson, 1982; Koseisho, 1984).

Regulatory authorities are under public pressure to establish guidelines which might increase the safety of medicines to be marketed. As a result, there has been a growing tendancy for more tests to be required, including those of increasing duration (Gross, 1979). However, greater safety cannot be achieved by simply prolonging animal tests (Gross, 1982) and the value of experiments lasting longer than 6 months has not been established. One method of assessing their contribution to a safety evaluation program is to compare retrospectively the results of tests carried out for 6 months or less with results obtained after 6 months. The Centre for Medicines Research has already es- tablished a databank comprising comprehensive animal toxicological data obtained from pharmaceutical companies in the United Kingdom (Lumley and Walker, 1985a,b)

66

0273-2300186 $3.00 Copyright 0 1986 by Academic Press, Inc. All righu of reproduction in any form resewed.

DURATION OF ANIMAL TOXICITY STUDIES 67

Canada

16. Q Q Q

Japan

0' I

1 3 6

Human Exposure Conthsl

FIG. 1. International guidelines for the minimum duration of animal toxicity studies (studies to be carried out in both a rodent and a nonrodent species).

and these data have been used to conduct such a retrospective analysis (Lumley and Walker, 1985~). This databank has now almost doubled in size by including studies from companies in Switzerland and Germany.

COMPOSITION OF THE EUROPEAN DATABANK

Twenty-one pharmaceutical companies with research and development activities in Europe have provided comprehensive repeated-dose toxicology data for compounds tested and evaluated between 1965 and 1982. Those compounds for which data from more than one time period were available and/or those which had been tested in two or more species were submitted. Because of the differences in design, including dose levels and tissues examined, carcinogenicity tests were excluded unless a combined carcinogenicity/toxicity experiment was carried out. These data comprise the European toxicology databank. The methodology of its establishment and data aquisition have been described in detail elsewhere (Lumley and Walker, 1985a,b).

The European databank contains information for 124 compounds studied in one or more species. There are 2 14 case studies, a case study being defined as the full information for one compound tested in one species for one or more time period(s). One hundred case studies on 77 compounds include tests carried out for longer than 6 months. The therapeutic classes of the compounds in the total databank and of those with long-term data are shown in Fig. 2. Also illustrated are the number marketed (a total of 54), the number still in development (2 l), and those for which research has terminated (48). These 48 compounds will not be marketed for commercial reasons (8), clinical findings ( 15) concern about possible carcinogenicity, mutagenicity, or reproductive effects (5), repeated-dose animal toxicity (1 l), or chemical reasons (2). The reason for not marketing 5 compounds was given as “not animal toxicology,” while no reason was specified for 2 compounds and the market status of 1 compound is not known. The species studied are shown in Fig. 3.

Comparable short-term (~6 months) and long-term (~6 months) data are available

68 LUMLEY AND WALKER

FIG. 2. Therapeutic class of pharmaceutical compounds in the toxicology databank. GI, gastrointestinal: CVS, cardiovascular system: CNS, central nervous system; A-lnf, anti-infectives; Resp, respiratory system; Endo, endocrine; A-All, anti-allergics; Mist, miscellaneous. Total databank: q , marketed; RI, under devel-

research terminated. Long-term studies: 8, marketed, R!, under development; minated.

in 88 case studies (65 compounds). In 48 of these, the short-term data include pathology examination at, or before, 3 months, while in 40 studies 6 month data are available. The data have been analyzed to determine whether any new information became apparent after 6 months.

RESULTS

Analysis shows that in 70 of 88 cases there were no novel, significant toxicological findings after 3 or 6 months (Table 1). In 14 of the remaining 18 studies there were new findings after 3 months but, as there was no pathology examination at 6 months,

Prinate

FIG. 3. Species used in studies of pharmace 2 14 case studies; long-term: 100 case studies).

compounds in the toxicology databank (total databank: ong-term studies; %, short-term studies.

DURATION OF ANIMAL TOXICITY STUDIES 69

TABLE I

RESULTS OF THE EUROPEAN STUDY

No. of case studies

Conclusion Pathology,

1 or 3 months Pathology, 6 months Total

All toxicologically significant effects seen in short-term study

New findings were seen in an alternate species within 6 months

The new findings did not influence the progression of the compound to market or termination of research

Total

34 36 70

3 0 3

11 4 15

48 40 88

no conclusion can be drawn as to whether all effects would have been apparent by then. In neither of the two studies in dogs where 18 month data are available were new findings observed after 12 months.

The 40 case studies with pathology examinations at 6 months are summarized in Table 2 by therapeutic class and species. In 4 of these there were new significant findings after 6 months. Details of these are shown in Table 3. It is apparent that experimental design is an important factor, as in all of these cases the dose levels used in the long-term test were the same or higher than those used in the short-term test. In one case (No. 26) the number of animals examined in the short-term test may have been too small to reveal the organ weight changes observed in the long-term test, when a larger number was included. With more appropriate choice of dose levels and number of animals studied, some of the effects first seen after 6 months may well have become apparent earlier. In no case study did the findings after 6 months influence the progression of the compound to market or termination of research.

CONCLUSIONS

The toxicology databank established on the basis of U. K. case studies has doubled in size by the inclusion of data from pharmaceutical companies in Switzerland and Germany. The number of studies with comparable short- and long-term data has increased from 45 (United Kingdom alone) to 88 (European databank). The results presented here reinforce the conclusion from the U. K. analysis (Lumley and Walker, 1985c), namely, that, excluding those to investigate carcinogenicity, tests of longer than 6 months have not added to the overall safety evaluation of these compounds.

Many regulatory authorities still suggest that tests of longer than 6 months be com- pleted before applying for marketing authorization (Fig. I), presumably in the belief that this might guarantee the increased clinical safety of new medicines. In particular, the Canadian guidelines suggest l&month repeated-dose toxicity studies in both a rodent and a nonrodent species for oral or parenteral drugs to be administered clinically for 1 month or longer (Health Protection Branch, 198 1). At a workshop held in October 1984 (Walker and Dayan, in press), a short list of effects that had apparently taken

70 LUMLEY AND WALKER

TABLE 2

TOXICITY DATA FOR PHARMACEUTICAL COMPOUNDS

Time (months)

No. Species Therapeutic class 1-6 12 18 24

1 Rat 2 Rat 3 Rat 4 Rat 5 Rat 6 Rat 7 Rat 8 Rat 9 Rat

10 Dog II Dog 12 Primate 13 Primate 14 Primate 15 Primate 16 Primate 17 Primate

18 Rat 19 Rat 20 Rat 21 Rat 22 Rat 23 Rat 24 Dog 25 Dog 26 Primate 27 Primate 28 Primate 29 Primate

30 Rat 31 Rat 32 Rat 33 Rat 34 Rat 35 Dog 36 Dog 37 Dog 38 Dog 39 Dog 40 Dog

Drugs acting on the central nervous system

NSAI xxx X NSAI xxx X NSAI xxx X NSAI xxx X Tranquillizer xxx X Anticonvulsant xxx X Psychotropic xxx X Analgesic xxx X Antidepressant xxx X Tranquillizer xxx X NSAI xxx X NSAI xxx xxxa NSAI xxx X NSAI xxx X NSAI xxx X NSAI xxx X NSAI xxx X

Drugs acting on the cardiovascular system

Beta-blocker xxx X Antihypertensive xxx XXX” Antihypertensive xxx XXX” Antihypertensive xxx X Hypolipidemic xxx X Not specified xxx X Beta-blocker xxx X Calcium antagonist xxx X Antihypertensive xxx XXX” Hypolipidemic xxx X Hypohpidemic xxx X Hypolipidemic xxx X

Miscellaneous drugs

Endocrine xxx X Cytotoxic xxx X GI agent xxx X GI agent xxx X GI agent xxx X Endocrine xxx X Endocrine xxx X Endocrine xxx X GI agent xxx X GI agent xxx X Respiratory xxx X

X X X X X X X X X X X X X X

X

X

XXX” X X X X X

X

X X X X

Nofe. 0, no toxicity; X, toxic findings already noted seen again; XXX, new significant toxic findings; NSAI, nonsteroidal anti-inflammatory.

0 Did not influence the progression of the compound.

Num

ber

Tim

e (m

onth

s)”

Ther

apeu

tic

clas

s M

ax

dose

s (m

g/kg

) sp

ecie

s Nu

mbe

r of

ani

mal

s

TABL

E 3

CASE

ST

UDIE

S W

ITH

NEW

SI

GNIF

ICAN

T FI

NDIN

GS

AFTE

R 6

MON

THS

Find

ings

wi

thin

Ne

w fin

ding

s af

ter

6 m

onth

s 6

mon

ths

-

12

3 6

12

NSA

I 32

0 32

0 45

0 Pr

imat

e 4

12

14

19

1 6

12

Antih

yper

tens

ive

2000

16

00

1600

R

at

20

24

36

20

3 6

9 18

An

tihyp

erte

nsiv

e 65

2 10

0 10

0 10

0 R

at

30

30

20

36

26

Antih

yper

tens

ive

Prim

ate

6 12

80

0 80

0 6

14

Com

men

ts

Dea

th;

body

wei

ght

and

food

inta

ke

decr

ease

d: e

dem

a; d

eter

iora

tion

in c

ondi

tion;

re

d ce

ll pa

ram

eter

s,

prot

ein,

al

bum

in

and

seru

m

enzy

mes

dec

reas

ed; b

rown

ur

ine,

os

mol

ality

de

crea

sed;

ga

stro

inte

stin

al

hem

orrh

age/

er

osio

n/ed

ema

Body

wei

ght

gain

and

foo

d in

take

in

crea

sed;

ser

um e

nzym

e ch

ange

s: c

olor

atio

n of

som

e tis

sues

Dea

th;

food

inta

ke d

ecre

ased

; re

spira

tory

dep

ress

ion;

he

mat

olog

ical

and

clini

cal

chem

istry

cha

nges

; liv

er w

eigh

t in

crea

sed;

gas

tric

ulce

rs; t

hym

us

smal

l: ad

rena

l we

ight

in

crea

sed

Atax

ia;

beha

viora

l ch

ange

s; h

eart

rate

dec

reas

ed; v

omiti

ng;

colo

ratio

n of

som

e tis

sues

; liv

er

weig

ht i

ncre

ased

Kidn

ey:

inte

rstit

ial

neph

ritis

(3/7

hi

gh

dose

ani

mal

s; f

irst n

oted

35

week

s)

Hea

rt: w

eigh

t in

crea

sed

(hig

h an

d in

term

edia

te

dose

s)

Hea

rt: r

elat

ive

weig

ht

incr

ease

d;

scle

rotic

int

erst

itial

m

yoca

rdia

l st

reak

s (9

mon

ths)

Se

min

ifero

us

tubu

les:

deg

ener

ative

ch

ange

s ( 1

8 m

onth

s)

Adre

nals

and

pitu

itary

: he

mat

ic

suffu

sion

s (1

8 m

onth

s)

Hea

rt: w

eigh

t de

crea

sed

(all

dose

s)

Subm

itted

to

mar

ket;

dose

s in

12-m

onth

st

udy

were

hig

her

than

tho

se in

3- a

nd

6-m

onth

st

udie

s; n

ew f

indi

ng

seen

in

high

-dos

e gr

oup

only

Not

mar

kete

d be

caus

e of

find

ings

in

a sp

ecia

l rat

stu

dy; d

oses

at 6

and

12

mon

ths

were

the

sam

e

Not

mar

kete

d bu

t st

ill un

der

deve

lopm

ent;

all

findi

ngs

afte

r 6

mon

ths

were

see

n in

bot

h tre

ated

and

co

ntro

l gr

oups

and

wer

e no

t dos

e re

late

d; h

owev

er,

beca

use

they

wer

e m

ild

in c

ontro

ls

and

occa

sion

ally

m

oder

ate

and

mor

e fre

quen

t in

tre

ated

th

e po

ssib

ility

that

the

y we

re

treat

men

t-rel

ated

ca

nnot

be

tota

lly

exclu

ded

Not

mar

kete

d be

caus

e of

the

resu

lts o

f a

spec

ial r

at s

tudy

; the

sam

e do

ses w

ere

used

in b

oth

stud

ies;

the

num

ber

of

anim

als

exam

ined

at

6 m

onth

s m

ay

have

bee

n to

o sm

all

to d

etec

t wei

ght

chan

ge s

een

at 1

2 m

onth

s.

a Ti

me

of p

atho

logi

cal

exam

inat

ion.

72 LUMLEY AND WALKER

longer than 12 months to develop in 15 separate rat studies was presented by the Health Protection Branch and discussed. However, the relevant experimental details were not provided and there were no data from tests longer than 12 months in a nonrodent species on which to base the requirement for 18-month studies in the dog. The conclusion of the group attending the meeting (with one exception) was that, on the basis of the data currently available, there was no justification for conducting 18-month tests in two species.

To date there is no published evidence to support 18-month tests. The results of the present investigation suggest that erroneous conclusions may be drawn regarding the value of long-term tests if careful attention is not given to experimental design. Rather than prolonging investigations to identify toxicity in animals, the emphasis should be on postmarketing surveillance to determine safety in the target species-man.

ACKNOWLEDGMENTS

We are grateful to the following companies for supporting this study and particularly for providing data: Bayer AG, Beecham Pharmaceuticals; Boehringer Mannheim GmbH; The Boots Company PLC; Ciba- Geigy AG and Ciba-Geigy Pharmaceuticals; Fisons Pharmaceuticals; Glaxo Group Research Ltd.; Hoechst AG; Knoll AG, May & Baker Ltd.; Pfizer; Reckitt & Colman; Roche Products Ltd. and F. Hoffman- LaRoche & Co.; Sandoz Ltd; Schering AG; Smith Kline & French Research Ltd.; Sterling-Winthrop Group Ltd.: Thiemann Arzneimittel GmbH; Thomae GmbH; The Wellcome Research Laboratories: Wyeth Research (U. K.).

REFERENCES

ALDER, S., DANTON, C., AND ZBINDEN, G. (198 1). Pre-clinical Safety Requirements in 1980. Swiss Federal Institute of Technology and University of Zurich.

GROSS, F. H. (1979). Constraints of drug regulation on the development of new drugs. Arch. Toxicol. 43, 9-17.

GROSS, F. H. (1982). The scientific basis of drug safety regulations. In Drug Safety: Progress and Contro- versies-proceedings, IVth International Congress of Pharmaceutical Physicians, Paris, April 1981. (M. Auriche, J. Burke, and J. Duchier, eds.), pp. 9-18. Pergamon Press, Oxford.

Health Protection Branch (198 I). Pre-clinical Toxicologic Guidelines. Bureau of Human Prescription Drugs, Health Protection Branch, Health and Welfare, Canada.

KESTERSON, J. W. (1982). Drug safety evaluation: Animal toxicology studies and their interpretations. Drug Info. J., January/June, 22-34.

KOSEISHO (1984). Toxicity Test Guidelines 1984. Yakugyo Jiho Co. Ltd, Japan. LUMLEY, C. E., AND WALKER, S. R. (1985a). The establishment of a computer-based toxicology databank.

Med. Info. 10(2), 173-174. LUMLEY, C. E., AND WALKER, S. R. (1985b). A toxicology databank based on animal safety evaluation

studies of pharmaceutical compounds. Hum. Toxicol. 4,447-460. LUMLEY, C. E., AND WALKER, S. R. (1985~). The value of chronic animal toxicology studies of pharmaceutical

compounds: A retrospective analysis. Fundam. Appl. Toxicol. 5, 1007- 1024. WALKER, S. R., AND DAYAN, A. D. (in press). Long-Term Animal Studies-Their Predictive Valuefor Man:

Proceedings, Centre for Medicines Research Workshop, held at the Ciba Foundation, London, October 2, 1984. MTP Press Ltd, Lancaster.