Recent Risk Assessments Of Dioxins

Comparing the WHO, COT, SCF and JECFA evaluations

J.C. Larsen(using many slides prepared by A.G. Renwick)Danish Institute of Food and Veterinary Research, Division of Toxicology and Risk Assessment, Mørkhøj Bygade 19, DK-2860 Søborg, Denmark

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Polychlorinated dibenzodioxins

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ClPolychlorinated dibenzofurans

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Co-planar polychlorinated biphenyls

HAZARD IDENTIFICATION

Human studies

Epidemiology

Local accidental high exposures

Sevesoe.g. Bertazzi et al., 2001

Differences in background low level exposures

Rotterdam and Groningen cohortse.g. Huisman et al., 1995Koopman-Esseboom et al., 1996

Exposed workers

e.g.Fingerhut et al., 1991Ott and Zober, 1996Manz et al., 1991Flesch-Janys et al., 1995Becher et al., 1996Coggon et al., 1991

Environmental

HAZARD IDENTIFICATION (TCDD)

Genetic Studies in bacteria and mammalian cells were generally negative, does not bind to DNA, not genotoxic

Acute Oral LD50 is 1 - 5000g/kg (guinea pig – hamster)

Short-term Studies in mice rats, guinea pigs and monkeys; metabolic effects and changes in liver, thymus and haematology

Long-term andcarcinogenicity

Decreased weight gain, increased mortality (F), splenic and thymic atrophy, hepatic degeneration plus increase in thyroid follicular and hepatic carcinomas in rats

Reproductive Multi-generation studies – deceased fertility (rats)Single-generation studies – decreased sperm count (rats)Developmental studies – cleft palate and hydronephrosis (mice), effects on testes, prostate and sperm count (rats)

In vitro and animal studies

HAZARD CHARACTERISATION

1. Selection of effects that are of relevance to humans

2. Identification of effect(s) produced at the lowest doses

3. Dose-response assessment of effects and consideration of mode of action (threshold or non-threshold)

4. Derivation of a health-based advice related to the risk associated with the critical effect(s) (threshold or non-threshold)

For Dioxins the effects are considered to be mediated via the AhR [or other threshold mechanisms] so that the usual NOAEL + uncertainty factor approach is suitable

Recent studies with knockout mice show that the AhR is important in:-

Effects on liver, thymus(?), epididymis, testes – Lin et al., 2001Effects on prostate - Lin et al., 2002Oxidative stress and vascular inflammation – Hennig et al., 2002T-lymphocyte suppression – Kerkvliet et al., 2002Keratinocyte differentiation – Loertscher et al., 2002

Are the extensive epidemiological data adequate for risk assessment?Are exposure data reliable - both quantitatively and in relation to confounding exposures?Does hazard characterisation in human studies include all hazards identified in animal studies?

HAZARD CHARACTERISATION

Human studies

Chloracne – no clear dose-response reported

Liver – increases in liver enzymes

Cardiovascular disease – inconsistent evidence

Blood lipids – inconsistent changes

Reproductive hormones – inconsistent changes

Reproductive outcomes – change in sex ratio (Serveso)

Thyroid function – small and inconsistent changes

Neurological effects – inconsistent findings

Respiratory system – inconsistent evidence

Urinary system – no major changes reported

Immunological effects – inconsistent findings

Neurodevelopmental – some differences in ongoing Dutch studies

Cancer – largely based on occupational exposures

Based on COT, 2001

Mably et al (1992a,b,c) – reported decreased weights of epididymis and sperm numbers in rats after a single oral dose of 64ng/kg on gestation day 15 (GD15).

Animal studies

HAZARD CHARACTERISATION

Gray et al (1995, 1997a, 1997b) – reported greater effects on male offspring following single oral dosage on GD15 compared with GD8, with significant decreases in sperm count after 50ng/kg

Faqi et al (1998) – reported effects on sperm production in offspring following 25ng/kg as a subcutaneous loading dose 2 weeks prior to mating followed by weekly maintenance doses of 5ng/kg

Ohsako et al (2001) – reported decreased anogenital distance in offspring following 50ng/kg by gavage on GD15. NOAEL was 12.5 ng/kg bw.

Female Rhesus monkeys given 0.15 ng/kg bw/day in the diet for 3.5 years Schantz & Bowman (1989) - reported subtle, non-persistent neurobehavioural changes (object learning) in offspring (16.2 and 36.3 months).Rier et al (1993) – reported increased incidence of endometriosis after 10 years

Hazard Characterisation

Rodents require higher doses (100-200-fold) to reach the same equivalent body burdens as seen in humans at background exposures.

Body burden estimations are considered the most appropriate dosimetric parameter for interspecies comparison.

What intake in humans would give a maternal body burden of TCDD of for example 30ng/kg?

TCDD would show slow accumulation on repeated daily intake, because the half-life of TCDD in humans is very long (7.5 years)

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4-5 half-lives

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Body burden = daily intake x bioavailability x half-life0.693

Daily intake = body burden x 0.693bioavailability x half-life

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Daily intake = body burden x 0.693bioavailability x half-life

Daily intake = 30 ng/kg x 0.693 0.5 x 7.5 years

Daily intake = 15 pg/kg/day

ANIMAL BODY BURDENS OF TCDD AND ASSOCIATED HUMAN ESTIMATED DAILY INTAKES (EDI) (WHO 1998)

STUDY RESPONSE

(LOAELs)

MATERNAL BODY BURDEN

(ng/kg)

ASSOCIATED

HUMAN EDI

(pg/kg bw/day)

Gray et al., 1997a RATS: decreased sperm count in offspring

28 14

Gehrs et al., 1997b; Gehrs & Smailowicz 1998

Immune suppression in offspring

50 25

Gray et al., 1997b Increased genital malformations in offspring

73 37

Schantz & Bowman, 1989

MONKEYS: Neurobehavioural (object learning) effects in offspring

42 21

Rier et al., 1993 Endometriosis 42 21

Estimation of a TDI for dioxins (WHO

1998)

The LOAELs for the most sensitive adverse responses in experimental animals could be transformed into a range of estimated long-term human daily intakes of 14-37 pg/kg bw/ day.

Composite uncertainty factor of 10:

TDI 1 - 4 TEQ pg/kg bw (rounded figures)

Limitations of the design of the rat studies used by WHO

The bioavailability of 2,3,7,8-TCDD to the foetus is likely to be higher following a single oral bolus dose on days 15 - 16 of gestation (the sensitive window) than after low-level chronic dietary exposure at steady state.

The key to the use of these data for risk assessment came with the studies of Hurst et al (2000a,b) who reported the tissue distribution of 3H in maternal and fetal tissue on GD16 after administration of [3H]-TCDD as a single dose on GD15 and following subchronic exposure (5 days per week for 13 weeks)

Animal studies

HAZARD CHARACTERISATION

Maternal Fetal

Dose on GD 15Dose on GD 15

Maternal Fetal

Daily doseDaily dose

Animal studies

HAZARD CHARACTERISATION

Single dose on GD15

Doseng/kg

Maternalng/kg

Fetalng/kg

Mat/Fet ratio

1000 585 55.7 10.5

50 30 5.3 5.7

200 97 13.2 7.4

800 523 39.1 13.4

Subchronic exposure

Daily doseng/kg

Maternalng/kg

Fetalng/kg

Mat/Fet ratio

0.71 20 1.4 14.3

7.1 120 7.5 16.0

21.3 300 15.2 19.7

The key to the use of these data for risk assessment came with the studies of Hurst et al (2000a,b) who reported the tissue distribution of 3H in maternal and fetal tissue on GD16 after administration of [3H]-TCDD as a single dose on GD15 and following subchronic exposure (5 days per week for 13 weeks)

Correction Factor Applied

SCF – fitted power functions to the data for the GD15 and subchronic data

and derived a correction factor of 2.5 at maternal body burdens of

<30ng/kg and 2.6 from 30-100ng/kg. The SCF established corresponding values of fetal, acute maternal and subchronic steady state maternal body burdens for 2,3,7,8-TCDD

COT - used ratio of 2.5 based on the 2 lowest doses

Single dose on GD15

Doseng/kg

Maternalng/kg

Fetalng/kg

Mat/Fet ratio

50 30 5.3 5.7

Subchronic exposure

Daily doseng/kg

Maternalng/kg

Fetalng/kg

Mat/Fet ratio

0.71 20 1.4 14.3

Ratio

2.5

JECFA – fitted power functions to the data for the GD15 and subchronic

data and derived a correction factor of 2.6; also fitted a linear model and estimated possible TDI values using each correction method

Selection of a NOAEL or LOAEL on which to base the TDI

Faqi et al (1998) appear to have detected the most sensitive endpoints for which the LOAEL is 25ng/kg loading dose plus 5ng/kg every 7 days

Table 33. Studies providing the body burdens at which no effect and the lowest observed effect were seen for the most sensitive adverse effects of TCDD on developmental end-points in rats (From: JECFA, 2002)

Reference; rat strain End-point Dosing regimen No-effect body burden (ng/kg bw)

Lowest effective body burden (ng/kg bw)

Ohsako et al. (2001); Holtzman

Ventral prostate weight; decreased anogenital distance in male offspring

Single oral bolus dose by gavage on day 15 of gestation

13 51

Faqi et al. (1998); Wistar Decreased sperm production and altered sexual behaviour in male offspring

Loading dose; maintenance dose by subcutaneous injection

25

Gray et al. (1997a,b); Long-Evans

Accelerated eye opening and decreased sperm count in offspring

Single oral bolus dose by gavage on day 15 of gestation

28

Mably et al. (1992c); Holtzman

Decreased sperm count in offspring

Single oral bolus dose by gavage on day 15 of gestation

28

Gehrs et al (1997); Gehrs & Smialowicz (1998);

Immune suppression in offspring Single oral bolus dose by gavage on day 14 of gestation

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Loading dose

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Final dose

Estimation of the maternal subchronic steady state body burden (BB) associated with the Faqi et al. (1998) LOAEL

At GD 16 the maternal BB consisted of a (“pseudo”) steady state BB of 20 ng/kg bw and an acute BB of 5 ng/kg bw from the injected loading dose. Using the tabulated corresponding values of fetal, acute maternal and subchronic steady state maternal BBs the SCF estimated a fetal BB of 1.8 + 1.2 = 3.0 ng/kg bw. The 3.0 ng/kg bw fetal BB would correspond to a maternal subchronic steady state BB of 39 ng/kg bw.

Estimation of the daily intake in humans that would give the same maternal body burden

Daily intake = 39 ng/kg x 0.693

0.5 x 7.5 years

Daily intake = 20 pg/kg bw

Uncertainty factors

Database deficiencies Usually 3 or 10

No chronic bioassay

No developmental/repro’ toxicity data

No NOAEL only a LOAEL in critical study

Usually 100

KINETICS DYNAMICSKINETICS DYNAMICS

SPECIESDIFFERENCES

HUMANVARIABILITY

10 10

Uncertainty factors

Inter-species extrapolation

Inter-individual variability

Chemical specific data can be used to replace a default uncertainty factor (UF) by an adjustment factor (AF) - based on IPCS 1999, 2001

100 - FOLD UNCERTAINTY FACTOR

INTER-SPECIES

DIFFERENCES

10 - FOLD

INTER-INDIVIDUAL

DIFFERENCES

10 - FOLD

TOXICO-DYNAMIC

ADUF

10 0.4

2.5

TOXICO-KINETIC

AKUF

10 0.6

4.0

TOXICO-DYNAMIC

HDUF

10 0.5

3.2

TOXICO-KINETIC

HKUF

10 0.5

3.2

TOXICO-DYNAMIC

ADUF

10 0.4

2.5

TOXICO-KINETIC

AKUF

10 0.6

4.0

TOXICO-DYNAMIC

HDUF

10 0.5

3.2

TOXICO-KINETIC

HKUF

10 0.5

3.2

Taken into account by calculation of maternal body burden

TOXICO-DYNAMIC

ADUF

10 0.4

2.5

TOXICO-DYNAMIC

HDUF

10 0.5

3.2

TOXICO-KINETIC

HKUF

10 0.5

3.2

Taken into account by calculation of maternal body burden

TOXICO-KINETIC

AKAF

TCDD1.0

Rats are more sensitive than humans and as sensitive as the most sensitive human

Taken into account by calculation of maternal body burden

TOXICO-DYNAMIC

ADAF

TCDD1.0

TOXICO-KINETIC

AKAF

TCDD1.0

TOXICO-DYNAMIC

HDAF

TCDD1.0

TOXICO-KINETIC

HKUF

10 0.5

3.2

Humans are generally less sensitive than rats, but the most sensitive human might be as sensitive as rats

No data available

9.6

Uncertainty factor for TCDD

Database deficiencies 3

Inter-individual variability 3.2

Inter-species extrapolation 1

20 pg/kg/day gives a maternal body burden of 39 ng/kg

Dividing by the 9.6-fold uncertainty factor gives a daily intake of 2.1 pg/kgThis was rounded to give a tolerable intake of

2pg/kg/day

Table 34. Summary of four calculations of provisional tolerable monthly intake intakes (PTMIs) for PCDDs, PCDFs and coplanar PCBs (From JECFA, 2002)

Linear model  Power model 

Ohsako et al. (2001)

Faqi et al. (1998)

Ohsako et al. (2001)

Faqi et al. (1998)

Administered dose (ng/kg bw)a 12.5 12.5

Maternal body burden (ng/kg bw)b 7.6 25b 7.6 25b

Equivalent maternal body burden with repeated dosing (ng/kg bw)

13c 25c 19d 39d

Body burden from feed (ng/kg bw) 3 3 3 3

Total body burden (ng/kg bw) 16 28 22 42

EHMI (pg/kg bw per month) 237 423 330 630

Safety factor 3.2 9.6 3.2 9.6

PTMI (pg/kg bw per month) 74 44 103 66

EHMI, equivalent human monthly intake

a Bolus dose (NOEL)b Target maternal body burden after repeated dosing (LOEL)c Linear relationship between fetal and maternal body burden assumed from data in Table 30d Non-linear relationship between fetal and maternal body burden assumed from data in Table 30e For humans, 7.6 year half-life and 50% uptake from food assumed (Eq. 1)

WHO (1998) – gave a range of 1- 4 pg/kg/day – because intake data are usually expressed on a daily basis

SCF (2001) – gave a value of 14pg/kg/week – to reflect the long half-life of TCDD

Guidance values expressed as 2,3,7,8-TCDD equivalents

JECFA (2002) – gave a value of 70pg/kg/month – to reflect the long half-life of TCDD

Note – one day = 0.04% of the half-life – one week = 0.25% of the half-life – one month = 1.11% of the half-life

Therefore – the main rationale of longer time intervals is to emphasise that short-term peaks of exposure will not greatly affect body burden - rather than indicate a duration of exposure that could affect body burden

Conclusions

Recent evaluations by the COT, SCF and JECFA have been based on changes in the male rat reproductive system following in utero exposure

Conversion of the animal data into a tolerable intake for humans has allowed for

the fetal to maternal ratio after bolus dose the long half-life and potential for accumulation the relative sensitivity of rats and humans potential human variability in kinetics and dynamics

The COT, SCF and JECFA differ in the time-base of the guidance value but not in the data or approach used

The intake by a significant proportion of the population exceeds the guidance value