Table Of ContentToxicology
VOL. VOL.
26
26
about the book… TARGET ORGAN
TOXICOLOGY T A R G E T O R G A N T O X I C O L O G Y S E R I E S
Despite being regarded as the most common toxicological target in the endocrine system, SERIES
the adrenal gland has often been neglected in regulatory testing. Adrenal Toxicology
addresses the increased interest in adrenocortical toxicology and the need for a resource Series Editors
that makes techniques available to examine adrenal endocrine disruption.
A. Wallace Hayes • John A. Thomas • Donald E. Gardner
Examining current techniques and the latest advancements, Adrenal Toxicology reviews
A
the endocrinology, pharmacology, pathology and toxicology of the adrenal gland. This
text provides information on the range of drugs and chemicals that affect adrenocortical
function and suggests standardized approaches for in vivo and in vitro assessment. This d
volume also presents recent developments in the molecular mechanisms of toxicity to the
r
adrenal cortex and medulla, and considers environmental adrenal endocrine disruption in
sentinel species. e
Adrenal Toxicology: n
• reflects the major developments made over the past decade
• focuses on the latest research techniques, including their uses and limitations A
• provides an integrated strategy for adrenal toxicology evaluation
l
• identifies knowledge and data gaps, providing impetus for regulatory consideration
A
about the editors...
T drenAl
PHILIP W. HARVEY received his honors degree and doctorate (endocrine, reproductive/
developmental, and neurobehavioral toxicology) from the University of Keele, UK, and has o
worked in general industrial/pharmaceutical toxicology for over 20 years. He has particular
x
interests in endocrine toxicology (the adrenal, prolactin and hormonal carcinogenesis, T
estrogenic chemicals and human health) and devised and edited the books The Adrenal in i oxicology
c
Toxicology: Target Organ and Modulator of Toxicity (1996) and Endocrine and Hormonal
Toxicology (1999). He has served on various journal editorial boards and is currently the
o
Editor-in-Chief of Journal of Applied Toxicology.
l
DAVID J. EVERETT received his honors degree in biological sciences from the University
of Sussex, UK, and has been actively involved in toxicological safety assessment of novel o
pharmaceuticals, agrochemicals and industrial chemicals for more than 30 years. His
g
involvement in carcinogenicity has led to a particular interest for him in hormonal
carcinogenesis. He is a Fellow of the Institute of Biology.
y
CHRISTOPHER J. SPRINGALL is a toxicological pathologist who received his original medical
degree from Liverpool University, UK, and has been actively involved in toxicology and
drug development for over 25 years. He has worked with a very wide range of candidate
drugs, including a substantial number which have an impact on endocrine systems.
Edited by
Printed in the United States of America Harvey
Philip W. Harvey
Everett
David J. Everett
Springall
61291
Christopher J. Springall
A
drenAl
T
oxicology
TARGET ORGAN TOXICOLOGY SERIES
SeriesEditors
A.WallaceHayes,JohnA.Thomas,andDonaldE.Gardner
AdrenalToxicology.PhilipW.Harvey,DavidJ.Everett,andChristopherJ.
Springall,editors,336pp.,2008
CardiovascularToxicology,FourthEdition.DanielAcosta,Jr.,editor,712pp.,
2008
ToxicologyoftheGastrointestinalTract.ShayneC.Gad,editor,384pp.,2007
ImmunotoxicologyandImmunopharmacology,ThirdEdition.RobertLuebke,
RobertHouse,andIanKimber,editors,676pp.,2007
ToxicologyoftheLung,FourthEdition.DonaldE.Gardner,editor,696pp.,2006
ToxicologyofthePancreas.ParvizM.Pour,editor,720pp.,2005
ToxicologyoftheKidney,ThirdEdition.JoanB.TarloffandLawrenceH.Lash,
editors,1200pp.,2004
OvarianToxicology.PatriciaB.Hoyer,editor,248pp.,2004
CardiovascularToxicology,ThirdEdition.DanielAcosta,Jr.,editor,616pp.,
2001
NutritionalToxicology,SecondEdition.FrankN.KotsonisandMaureenA.
Mackey,editors,480pp.,2001
ToxicologyofSkin.HowardI.Maibach,editor,558pp.,2000
Neurotoxicology,SecondEdition.HughA.TilsonandG.JeanHarry,editors,
386pp.,1999
Toxicant–ReceptorInteractions:ModulationofSignalTransductionsandGene
Expression.MichaelS.DenisonandWilliamG.Helferich,editors,256pp.,1998
ToxicologyoftheLiver,SecondEdition.GabrielL.PlaaandWilliamR.Hewitt,
editors,444pp.,1997
FreeRadicalToxicology.KendallB.Wallace,editor,454pp.,1997
EndocrineToxicology,SecondEdition.RaphaelJ.Witorsch,editor,336pp.,
1995
Carcinogenesis.MichaelP.WaalkesandJerroldM.Ward,editors,496pp.,
1994
DevelopmentalToxicology,SecondEdition.CaroleA.KimmelandJudy
Buelke-Sam,editors,496pp.,1994
NutritionalToxicology.FrankN.Kotsonis,MaureenA.Mackey,andJerryJ.
Hjelle,editors,336pp.,1994
OphthalmicToxicology.GeorgeC.Y.Chiou,editor,352pp.,1992
ToxicologyoftheBloodandBoneMarrow.RichardD.Irons,editor,192pp.,
1985
ToxicologyoftheEye,Ear,andOtherSpecialSenses.A.WallaceHayes,editor,
264pp.,1985
CutaneousToxicity.VictorA.DrillandPaulLazar,editors,288pp.,1984
A
drenAl
T
oxicology
Edited by
Philip W. Harvey
Covance Laboratories UK Ltd.
North Yorkshire, UK
David J. Everett
Covance Laboratories UK Ltd.
North Yorkshire, UK
Christopher J. Springall
Covance Laboratories UK Ltd.
North Yorkshire, UK
InformaHealthcareUSA,Inc.
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Adrenaltoxicology/editedbyPhilipW.Harvey,DavidJ.Everett,
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1. Adrenalglands–Toxicology. 2. Adrenalglands–Effectofdrugson.
I.Harvey,PhilipW. II.Everett,DavidJ.1953- III.Springall,ChristopherJ.
IV.Series.
[DNLM:1. AdrenalGlands–drugeffects. 2.AdrenalGlands–physiology.
3.AdrenalCortexDiseases–chemicallyinduced. 4.AdrenalCortex
Diseases–physiopathology. 5.Toxicology–methods.WK702A24272008]
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childrenJessicaRuth,RebeccaEve,andWilliamJozef.
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Formyfather,John.
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For colleagues at Covance whose support has allowed us the time to undertake
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C.J.S.
Preface
Despite the adrenal gland being the most common target within the endocrine
system (Ribelin, 1984), adrenal dysfunction is poorly recognized in toxicology.
Theadrenalisadefinedvitalorganwithaprimaryroleintheadaptationtostress-
ful circumstances (indeed adrenocortical glucocorticoid production is the single
most important physiological response for survival of an organism post-injury
or infection, e.g., Munck et al., 1984), and toxicological pathology in regula-
torystudiesoftendisregardsadrenocorticalhistologicalfindingsassecondaryto
stress, which can be inappropriate without evidence that the adrenal cortex is
functionally competent. For example, adrenal hypertrophy may certainly be due
tostress-inducedoversecretionofadrenocorticotropichormone(ACTH),butthis
conditionmayalsoarisefrommoreseriousadrenocorticalsteroidogenicenzyme
inhibition,theconsequentlossofglucocorticoid(cortisolorcorticosterone)secre-
tion,andreducedorabolishedfeedbackcontrolofpituitaryACTHsecretion.The
resultant uncontrolled ACTH hypersecretion can then overstimulate the cortex
to produce hypertrophy. Genuine stress-related changes encountered in toxicity
studies are generally considered to be of minimal toxicological consequence,
being physiologically adaptive responses that are reversible upon withdrawal
of treatment. By contrast, pharmacotoxicological suppression of steroidogene-
siscanbeaseriousconditionleadingtoAddisoniancrisis(adrenalinsufficiency
characterizedbylethargy,hemodynamicinstability,andcardiovascularcollapse)
and death. Indeed, there are many examples of drugs and chemicals known to
inhibit critical adrenocortical enzymes, potentially producing adrenal incompe-
tence,insufficiency,orsuppression,andseveralexampleshavebeendiscoveredin
patientsfollowingunexpectedside-effectsandfatalitiesapparentlynotadequately
detected,orindeedignoredasinconsequential,inpreclinicaltoxicology.
The adrenal medulla and cortex, respectively have acute and prolonged
adaptive functions in the stress response, but it is the cortex that has additional
important roles in regulating water and electrolyte balance, metabolism, inflam-
mation,immunefunction,andvariousreproductiveanddevelopmentalprocesses
depending on life stage and species. It is therefore surprising that the adrenal
has been neglected in endocrine toxicology and this has been pointed out as a
criticalomissioninaregulatorytoxicologycontext (HarveyandJohnson,2002;
Harvey and Everett, 2003; Harvey and Everett, 2006; Hinson and Raven, 2006;
Harvey, Everett, and Springall, 2007). The United States Environmental Pro-
tection Agency (USEPA), Endocrine Disrupter Screening and Testing Advisory
v
vi Preface
Committee (EDSTAC), in not incorporating adrenal evaluation studies in its
endocrine disruption strategy, failed to recognize the adrenal as an important
endocrine gland influencing health, development, and survival fitness, or the
unique vulnerability of the adrenal to toxic insult (see chap. 1) compared with
other endocrine organs. Further, the adrenal cortex is also unique in possessing
almost universal steroidogenic capability, and this was also overlooked in the
recommendations for the development of models/assays to examine the effects
of chemicals on steroidogenesis. The human adrenocortical carcinoma derived
H295Rcelllinehasbeensuggestedasausefulsystemtoaddressbothissues(e.g.,
HarveyandEverett,2003;Harvey,Everett,andSpringall,2007).Otherregulatory
bodieshavesincepartiallyrectifiedthissituationandrecognizedtheimportance
oftheadrenalintoxicology,oratleasttheutilityofadrenocorticalcellsasamodel
forsteroidogenesisasawhole,andtheOECDhasrecentlyundertakenaprogram
tovalidatetheH295Rcelllineasauniversalmodeltoevaluatesteroidogenictox-
icity(e.g.,Heckeretal.,2007).Oskarssonetal.(2006)reportthatsteroidogenic
geneexpressionfollowingchemicalchallengeiscomparablebetweentheH295R
celllineandthenormalhumanadrenaldemonstratingthevalidityofthiscellline.
The consensus is that the H295R cell line is currently the best available model,
and even though ACTH receptors are not well expressed, effective downstream
challengeaugmentationcanbeachievedbydrugs,suchasforskolin.
The number of literature studies on adrenocortical toxicity has risen
markedlyoverthepastdecade,whichindicatesthegrowingscientificandregula-
toryinterestintheadrenalasatargetfor“endocrinedisruption,”withthemajority
of studies using in vitro techniques. Numerous laboratories are now using the
H295Rcelllinetoinvestigatethediverseeffectsofchemicalsonmolecularmech-
anisms of adrenocortical toxicity, and/or the general process of steroidogenesis,
using steroid production, gene regulation, and enzyme expression as endpoints
seeSandersonetal.,2001;Muller-Vieiraetal.,2005;Graciaetal.,2006;Hecker
etal.,2006;Oskarssonetal.,2006;Furutaetal.,2008andStiglianoetal.,2008
for recent examples of the range of endpoints that can be assessed in this cell
line,andchapters7and8forthoroughreviews).Whilethisisanimportantstep
forward, in vitro systems will not detect drugs or chemicals that affect adrenal
function higher in the hypothalamo-pituitary axis, or effects on carrier proteins,
andthereforethereisalsoaneedtovalidateinvivomodels.Astrategyforevalu-
ationofdrugandchemicaleffectsonadrenocorticalfunctionhasbeenproposed
(Harvey,Everett,andSpringall,2007,andalsoseechap.1forfurtherdetails)and
involvesashort-terminvivoadrenalchallengetestinrodents(orotherlaboratory
speciesifindicated—seeColagiovannietal.,2006,andinthisvolume,forinfor-
mation on adrenal function and mechanism evaluation in the dog) coupled with
the H295R in vitro assay to shed light on molecular sites of toxicity/functional
inhibition.
Theprimarypurposeofthistextistoreviewthescopeof,anddevelopments
in, adrenal toxicology. It is specifically designed to be both complementary to,
and an update of, the first text in this area The Adrenal in Toxicology: Target
Preface vii
OrganandModulatorofToxicity(Harvey,1996a).Thepresenttexthasminimal
overlap with the former, but reviews the major developments in the field over
the last decade, and identifies research requirements including the validation of
standardizedmodelsandmethods.Thefocusofthisnewtextisontheadrenalas
a target organ, both in the main mammalian species used in pharmaceutical and
chemical regulatory toxicology and environmental sentinel species, and also on
theadvancesinidentifyingmolecularmechanismsofaction.Mosttoxicological
researchhasfocusedonthecortexratherthanthemedulla,whichreflectsitswider
and more complex role in physiological processes, and this is also by necessity
reflectedinthistext,althoughpathologyofthemedullaisexaminedintherodent
andhuman.
Following an introduction and overview of adrenal endocrine control and
toxicology covering the range of toxicants, targets, mechanisms, interactions,
models, and species differences (see chap. 1), there is a section covering the
endocrinologyandpharmacologyoftheadrenalinhealthanddisease.Anexem-
plary chapter on human adrenal dysfunction (see chap. 2) reinforces the critical
importanceofthecorrectfunctionofthisgland,bothmedullaandcortex,andfrom
this,thepotentialconsequencesofdrugorchemical-induceddysfunctionmaybe
inferred.Forexample,manyadrenaldiseasesandsyndromesaregenerallydueto
faults in gene, enzyme, or receptor expression, and the knowledge derived from
human medicine of the molecular basis of dysfunction can assist in identifying
the significance and consequences of pharmacotoxicologically induced effects
on these targets. Further, the fact that many of the enzymes involved in human
adrenocorticaldisorderscanbepharmacologicallymanipulated(HakkiandBern-
hardt, 2006) raises the real possibility that these adrenal conditions could occur
asaconsequenceofoff-targettoxicityfromdrugsandchemicals,bothinnormal
subjects,butespeciallyinpatientspredisposedtodevelopthenaturaletiological
factorsofthedisease,whereadditionalinsultmayaccelerateorprecipitateonset.
Inaddition,thissectionalsoprovidesahighlyauthoritativereview(seechap.3)
of the endocrinology, pharmacology and pathophysiology of the hypothalamo-
pituitary-adrenal(HPA)axis,includingnewemergingknowledgeofdevelopmen-
taleffectsofHPAhormones.Theearlylifeinfluenceofglucocorticoidhormones
isanimportantpharmacologicalfieldwithimmensepotentialapplication;adrenal
glucocorticoidanalogueshavewelldocumentedusestoacceleratematurationof
premature/fetal lung, but have also long been known to have adverse effects on
generalgrowthanddevelopmentintermsofteratogenicity(Hawkins,1983).New
evidencesuggeststheymayalsobeassociatedwithadultsusceptibilitytomajor
diseasessuchashypertension,type2diabetes,coronaryheartdisease,hyperlipi-
demia and nervous system disturbance following early life exposure. The phar-
macologyofglucocorticoidsandeffectsonimmuneandinflammatoryresponses
isintroduced(propertieswhichhavebeenexploitedinoneofthemostimportant
classes of medicines in the past 50 years, for example, in asthma and allergy—
see also Harvey, 1996a, and chapters therein for accounts of the well-known
classicalpharmacologyofadrenalglucocorticoidsandtheirsyntheticanalogues).
