Subchronic and Chronic Toxicity Studies

This chapter seeks to provide a firm grasp of the objectives for repeat-dose studies, the regulatory requirements governing them, the key factors in their design and conduct, and the interpretation of their results. There are a number of review chapters which deal with the general case of such studies (Wilson et al., 2001 being the most recent) that provide useful background. 

As with any scientific study or experiment (but especially for those in safety assessment), the essential first step is to define and understand the reason(s) for the conduct of the study, its objectives. There are three major (scientific) reasons for conducting subchronic and chronic studies, but a basic characteristic of all but a few 

The objectives of the typical subchronic study fall into three categories. The first is to broadly define the toxicity (and, if one is wise, the pharmacology and hyperpharmacology) of repeated doses of a potential therapeutic agent in an animal model (Traina, 1983). This definition is both qualitative (what are the target organs and the nature of the effects seen ) and quantitative (at what dose levels, or, more important, at what plasma and tissue levels, are effects definitely seen and not seen ). 

TABLE 7.1. Duration of Treatment Supported by Preclinical Studies 

Animal Study Length Generally Allowed Human Dosing 

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The traditional acute, subchronic, and chronic toxicity studies performed in rodents and other species also can be considered to constitute multiple endpoint screens. Although the numerically measured continuous variables (body weight, food consumption, hematology values) generally can be statistically evaluated individually by traditional means, the same concerns of loss of information present in the interrelationship of such variables apply. Generally, traditional multivariate methods are not available, efficient, sensitive, or practical (Young, 1985). 

Pharmacokinetics. All subchronic and chronic toxicity studies now incorporate (either in the study itself or in a parallel study) evaluation of the basic pharmacokinetics of a compound. This is discussed in detail in Chapter 18. 

Metabolism and pharmacokinetic studies have greater relevance when conducted in both sexes of young adult animals of the same species and strain used for other toxicity tests with the test substance. The number of animals used in metabolism and pharmacokinetic studies would be sufficient to reliably estimate population variability. This usually means a separate (but parallel) set of groups of animals in rodent studies. A single set of intravenous and oral dosing results from adult animals, when combined with some in vitro kinetic results, may provide an adequate data set for the design and interpretation of short-term, subchronic and chronic toxicity studies. 

Lee CC, Hong CB, Ellis El ( et al Subchronic and chronic toxicity studies of 2,4-dinitrotoluene. Part II. CD rats. J Am Coll TmVo/4 243-256, 1985... 

Parental Hazards (Effects). For assessment of reproductive and developmental risk, parental hazards, both paternal and maternal, must be identified and evaluated. Parental hazards can be expressed as altered nutritional state, functional impairment, and systemic toxicity. Because of possible indirect affects, knowledge and evaluation of non-reproductive/non-developmental toxicity studies are useful. This information is available by examination of subchronic and chronic toxicity studies. 

No review of subacute, subchronic, or chronic toxicity of chemical warfare nerve agents would be complete without discussion of the significant paper by Munro et al. that reviewed both animal and human studies of the nerve agents tabun (GA), sarin (GB), and VX. These studies included subacute, subchronic, and chronic toxicity studies in animals. Special attention was paid to the phenomenon of Organophosphorus-Induced Delayed Neuropathy (OPIDN). Reproductive toxicity and carcinogenicity tests were reviewed as well as in vitro studies of mutagenicity. Munro et al. s findings can be summarized as follows ... 

A review of reproductive and developmental toxicity studies of methyl salicylate indicated that, under conditions of sufficient exposure, there is a pattern of embryotoxicity and teratogenesis that is similar to that caused by comparable doses of salicylic acid. The abnormalities included neural tube defects and malformations of the skeleton and viscera. Studies of orally administered methyl salicylate indicated that the no-observed-adverse-effect level (NOAEL) for reproductive toxicity is 75-100 mg/kg daily, which is a level consistent with data from subchronic and chronic toxicity studies and is also consistent with the reproductive NOAEL for salicylic acid, which has been reported as 80 mg/kg daily (Belsito et al. 2007). 

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