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Low-Dose Hypothesis Again Not Replicated
Reproducibility is Key Factor in Assessment of Potential Endocrine Disruptors

October 11, 2002

The Low-Dose Hypothesis: A Case Study in Irreproducibility

The low-dose hypothesis claims that exposure to extremely low levels of certain substances could cause adverse health effects in humans by disruption of normal hormonal functions. Health effects are purported to occur at doses well below levels that have been demonstrated to cause no health effects using well-established toxicological methods. Of even more scientific significance, the hypothesis is based on the concept of a “non-monotonic” dose-response relationship, meaning that health effects may occur at low doses while much higher doses result in no effects. This aspect of the low-dose hypothesis conflicts with a long-standing and fundamental principle of toxicology – “the dose makes the poison.”

Since it’s inception, the low-dose hypothesis has suffered from a lack of reproducibility. For example, in one of the earliest reports claiming low-dose effects, researchers at the University of Missouri reported an increase in prostate weight in the male offspring of mice that were exposed to low levels of Bisphenol A (BPA) during pregnancy.¹ However, in two independent larger-scale studies conducted using the University of Missouri test conditions, the reported low-dose effects could not be replicated.^2,3

In a similar sequence of events, researchers in Scotland reported reduced testis size and sperm production in the male offspring of rats exposed to very low concentrations of BPA in drinking water.⁴ Again, an independent larger-scale study was unable to replicate the reported effects.⁵ Similar results on other chemicals also could not be replicated by either the original researchers or in an independent laboratory.⁶,⁷

Low-Dose Hypothesis Not Replicated - Again

Most recently, independent researchers have found that another prominent report claiming low-dose effects from exposure to BPA cannot be replicated.⁸ In a series of abstracts^{9,10,11,12,13} and one paper¹⁴ , a group of researchers at the Freie Universität Berlin reported a wide range of effects on the sexual development of both male and female offspring of rats exposed to BPA during pregnancy, including very low and high doses. Doubt had previously been cast on the validity of these results when it was determined that the control animals were from a timeframe not concurrent with the dosed animals.¹⁵

As reported in the peer-reviewed journal Toxicological Sciences, a group of researchers in the United Kingdom attempted to replicate the original results in both Sprague-Dawley rats, which were used by the Berlin researchers, as well as another strain of laboratory rat. With the exception of several expected effects at the highest dose, “the general absence of effects for BPA in the present study is in clear contrast to the results reported by Chahoud and his colleagues.”

The authors further note “pending the resolution of this problem of data reproducibility, adequate intra- and interlaboratory confirmation of endocrine toxicities is indicated, especially in cases of new and subtle effects being observed.”

Reproducibility is Key Factor in Assessment of Potential Endocrine Disruptors

Reproducibility of results in independent laboratories is a critical step in the scientific process of testing the validity of a hypothesis. The lack of reproducibility for the low-dose hypothesis indicates that it is not valid, however, the conflicting results can create a dilemma for risk assessors, who are obligated to consider all relevant information.

A pragmatic and sound scientific solution to this dilemma has been proposed in a recent paper from the National Centre for Environmental Toxicology (NCET) in the United Kingdom.¹⁶ The authors propose that the most appropriate procedure for assessing evidence for endocrine disruption is a tiered approach that incorporates a weight-of-evidence evaluation of all available data. The four tiers are based on a hierarchy of data in which the first three tiers estimate the potential and possible mechanisms for endocrine disruption, while the highest tier provides the most robust and definitive evidence of endocrine disruption.

1. Structure-activity relationships;
2. Specific in vitro screening studies;
3. Targeted short-term and mechanism-specific in vivo studies; and
4. Repeat-dose, regulatory guideline, GLP studies focused on carcinogenicity, reproduction and fertility, and prenatal development.

In the overall weight-of-evidence assessment, Tier 4 studies, when available, “are taken to provide the strongest evidence of endocrine effects, or their absence, and should accordingly be weighted above less-comprehensive data.”

Specifically in regard to the issue of reproducibility of low-dose effects, the authors state “because there has been a failure to replicate low-dose effects to date, caution should be exercised in a weight of evidence approach – there is clearly a need for substantiation of genuine reproducible findings (i.e. data must be robust) before data can be considered meaningfully.” In addition, the authors note “findings occurring only at the low dose level, where there is no dose response information at higher doses tested, should be considered, as with any other toxicity data, on a case basis in relation to historical control and any other relevant data.”

Weight-of-Evidence Reaffirms Safety of Bisphenol A

The weight-of-evidence approach proposed by the NCET authors has already been implemented for BPA. In particular, two definitive Tier 4 reproductive toxicity studies have been completed and both confirm the absence of low-dose effects from BPA.^17,18 These studies are integral components of several overall weight-of-evidence assessments that have recently been conducted on BPA.

These weight-of-evidence assessments have consistently reached the same conclusion that the low-dose hypothesis for BPA is unproven. The two definitive studies and the multiple weight-of-evidence assessments provide strong reassurance that there is no basis for human health concerns from exposure to low doses of BPA.

More information on the definitive Tier 4 studies and the weight-of-evidence assessments is available at the following links:

• Definitive Peer-Reviewed Study Reports No Low-Dose Effects
   
• US and Japanese Regulatory Agencies View the Low-Dose Hypothesis as Unproven
   
• European Commission’s CSTEE Confirms No Low-Dose Effects from BPA
   
• European Commission's Scientific Committee on Food Confirms That Products Made with Bisphenol A Are Safe

References

¹Nagel, S.C., F.S. vom Saal, K.A. Thayer, M.G. Dhar, M. Boechler and W.V. Welshons, 1997, Environmental Health Perspectives, vol. 105, pages 70-76.

²Ashby, J., H. Tinwell and J. Haseman, 1999, Regulatory Toxicology and Pharmacology, vol. 30, pages 156-166.

³Cagen, S.Z., J.M. Waechter, Jr., S.S. Dimond, W.J. Breslin, J.H. Butala, F.W. Jekat, R.L. Joiner, R.N. Shiotsuka, G.E. Veenstra and L.R. Harris, 1999a, Toxicological Sciences, vol. 50, pages 36-44.

⁴Sharpe, R.M., G. Majdic, J. Fisher, P. Parte, M.R. Millar and P.T.K. Saunders, 1996, Abstract #S23-4, 10^th International Congress of Endocrinology, San Fancisco, June 1996 (The Endocrine Society, Washington, D.C.).

⁵Cagen, S.Z., J.M. Waechter, Jr., S.S. Dimond, W.J. Breslin, J.H. Butala, F.W. Jekat, R.L. Joiner, R.N. Shiotsuka, G.E. Veenstra and L.R. Harris, 1999b, Regulatory Toxicology and Pharmacology, October 1999.

⁶Ashby, J., H. Tinwell, P.A. Lefevre, J. Odum, D. Paton, S.W. Millward, S. Tittensor and A.N. Brooks, 1997, Regulatory Toxicology and Pharmacology, vol. 28, pages 102-118.

⁷Sharpe, R.M., K.J. Turner and J.P. Sumpter, 1998, Environmental Health Perspectives, vol. 106, pages A220-A221.

⁸Tinwell, H., J. Haseman, P. A. Lefevre, N. Wallis and J. Ashby, 2002, Toxicological Sciences, vol. 68, pages 339-348.

⁹Chahoud, I., O. Fialkowski and C. E. Talsness, 2001, Reproductive Toxicology, vol. 15, page 589.

¹⁰Fialkowski, O., H.-J. Merker, C. E. Talsness and I. Chahoud, 2000, Abs. 55 in Hormones and Endocrine Disrupters in Food and Water: Possible Impact on Human Health, Copenhagen, May 2000.

¹¹Schönfelder, G., B. Flick, L. Mayr, C. Talsness, M. Paul and I. Chahoud, 2001, Reproductive Toxicology, vol. 15, page 594.

¹²Talsness, C. E., H.-J. Merker and I. Chahoud, 2000, Abs. 56 in Hormones and Endocrine Disrupters in Food and Water: Possible Impact on Human Health, Copenhagen, May 2000.

¹³Talsness, C. E., X. Wu, W. Witthoft and I. Chahoud, 2001, Reproductive Toxicology, vol. 15, page 596.

¹⁴Talsness, C. E., O. Fialkowski, C. Gericke, H.-J. Merker and I. Chahoud, 2000, Congenital Anom., vol. 40, pages S94-S107.

¹⁵“The National Toxicology Program Endocrine Disruptors Low-Dose Peer Review Final Report, August, 2001,” available on the Internet at http://ntp.niehs.nih.gov/ntp/htdocs/liason/LowDosePeerFinalRpt.pdf.

¹⁶ Harvey, P. W. and I. Johnson, 2002, Journal of Applied Toxicology, vol. 22, pages 241-247.

¹⁷Tyl, R. W., C. B. Myers, M. C. Marr, B. F. Thomas, A. R. Keimowitz, D. R. Brine, M. M. Veselica, P. A. Fail, T. Y. Chang, J. C. Seely, R. L. Joiner, J. H. Butala, S. S. Dimond, S. Z. Cagen, R. N. Shiotsuka, G. D. Stropp, and J. M. Waechter, 2002, Toxicological Sciences, vol. 68, pages 121-146. For a summary, see http://www.bisphenol-a.org/new/july2news.html.

¹⁸Ema, M., S. Fujii, M. Furukawa, M. Kiguchi, T. Ikka, and A. Harazono, 2001, Reproductive Toxicology, vol. 15, pages 505-523.