Validare i nuovi metodi alternativi in maniera più efficace

[Coleman R, Tsaioun K, Archibald K. A pragmatic comparison of human in vitro with animal in vivo approaches to predicting toxicity in human medicines. AltTox, March 27, 2014.]


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Despite the best efforts of the pharmaceutical industry to weed out potentially toxic drug candidates before they reach patients, the frequency of liver and other toxicities associated with new medicines remains at unacceptable levels (Verma & Kaplowitz, 2009; Chen et al., 2011). The “best efforts” concerned still rely predominantly on studies in experimental animals, mainly rodents, dogs, and nonhuman primates. Unfortunately, drugs that cause organ toxicity in these species do not necessarily cause such effects in humans, and vice versa. These species differences are particularly dramatic in the case of liver, due to large differences in metabolism between species. For this reason it is becoming apparent that something needs to be done about the way that we identify potential human organ toxicity before exposing human subjects to experimental medicines, and there is mounting scientific evidence for the utility of human-, rather than non-human-based test methods (Krewski et al., 2009; FDA, 2012; Firestone et al., 2010; Collins, 2011).

However, despite growing enthusiasm for “humanizing” drug testing, which Safer Medicines Trust believes will reduce the human and financial burden of adverse drug reactions, this approach has been extraordinarily difficult to get established as a viable alternative. The reasons for this difficulty are various, such as the obvious difficulties of modelling the complexity of the whole organism by extrapolating from isolated parts, the fact that the regulatory authorities demand results from animal studies, and the lack of validated alternative human-based methods. While there is a degree of validity in each point, they do not represent the whole case. For example, the objections to human in vitro and in silico approaches ignore the fact that these technologies have become more sophisticated and physiologically relevant in the last decade, yet the regulatory authorities still rely on extrapolation of human safety from animal in vivo studies. Why? Mostly for historical reasons: back in the 1960s animal tests were all that were available. As to the claim that few human-based tests have achieved formal validation, here we have only half the story; the truth is that not only is the validation process, orchestrated by organisations such as ECVAM and ICCVAM, time-consuming (Mak & Perry, 2007; Leist et al., 2012), but the animal in vivo tests that we currently rely on and that (sometimes inappropriately) serve as a “gold standard comparator” for the in vitro approaches, have never themselves been subjected to such scrutiny (ECOPA, 2008). Thus, when a new test method is evaluated, there is no way of knowing what it has to improve on. And finally, we strongly believe that the best model for humans has to be human.

We would like to suggest that rather than subjecting each new test method to several years of in-depth analysis to determine whether it passes all the theoretical criteria, and perfectly matches human outcomes, as is currently the case, a more fruitful approach would be to investigate whether a range of new approaches could actually improve on the patently flawed tests that are our current standards. Safer Medicines Trust proposes that what is needed is an objective independent study comparing human in vitro and in silico approaches with animal in vivo counterparts (Coleman, 2011; Clotworthy & Archibald, 2013), an approach we term “pragmatic validation.”


While to perform such a study prospectively would take many years and be resource-intensive, there is a simpler approach, which recognises that we have a vast wealth of (theoretically) available clinical and pre-clinical information on marketed drugs, whereby we can judge just how reliable the current safety testing paradigm has proven. Safer Medicines Trust’s proposal is based on identifying a number of marketed drugs that have been judged safe and effective for use as human medicines, based on the regulatory panel of animal tests, but have subsequently gone on to cause various toxicities in patients, and pair each of these with a structurally similar marketed drug that lacks such toxicity, acting as test drug and negative control, respectively. These drugs would then be submitted for blind testing using a variety of human-tissue-based technologies to determine whether any of these tests, either alone or in combination, could provide an indication of the toxicity known to be exhibited by the test compound in human subjects. A wide range of different organ toxicities could be studied using this approach, with a variety of high-throughput and high-content technologies selected from those perceived to be among the best available, offering complementary capabilities that should maximise the chances of identifying the majority of toxicities. These would, of course, include 3D and co-culture systems, as well as broad and hypothesis-free screens, in order to test the limits and the synergies of various different types of assays.

Before embarking on such a study, which although obviating the costs of both clinical and animal studies, would still incur the expense of the in vitro studies, we are currently initiating an entirely retrospective proof-of-principle study, whose only costs will be those entailed in data analysis.


Chen, M., Vijay, V., Shi, Q., Liu, Z., Fang, H., & Tong, W. (2011). FDA-approved drug labeling for the study of drug-induced liver injury. Drug Discov Today, 16, 697-703.

Chen, M., Zhang, J., Wang, Y., Liu, Z., Kelly, R., Zhou, G., Fang, H., Borlak, J., & Tong, W. (2013). The liver toxicity knowledge base: a systems approach to a complex end point. Clin Pharmacol Ther, 93, 409-412.

Clotworthy, M., & Archibald, K. (2013). Advances in the development and use of human tissue-based techniques for drug toxicity testing.  Expert Opin Drug Metab Toxicol, 9, 1155-1169.

Coleman, R.A. (2011). Efficacy and safety of new medicines: a human focus. Cell Tissue Bank, 12, 3-5.

Collins, Francis S. (2011). Reengineering Translational Science: The Time Is Right.  Sci Transl Med, 3(90):90cm17.

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European Consensus Platform for Alternatives (ecopa). (2008). 8th Annual Workshop, “Cosmetics Directive, REACH legislation and novel Directive 86/609: realistic 2009/2013 – factual status?,” Brussels, November 29-30, 2008.  Retrieved from

Firestone, M., Kavlock, R., Zenick, H., & Kramer, M. (2010). The U.S. Environmental Protection Agency strategic plan for evaluating the toxicity of chemicals.  J Toxicol Environ Health B Crit Rev, 13, 139-162.

Food and Drug Administration (FDA). (2012, May). S6 Addendum to Preclinical Safety Evaluation of Biotechnology-Derived Pharmaceuticals. Retrieved from

Krewski, D., Andersen, M.E., Mantus, E., & Zeise, L. (2009) Toxicity Testing in the 21st Century: Implications for Human Health Risk Assessment. Risk Anal, 29, 474-479.

Leist, M., Hasiwa, N., Daneshian, M., & Hartung, T. (2012). Validation and quality control of replacement alternatives—current status and future challenges. Toxicol Res, 1, 8–22.

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