Case-Control Tests

When performing case-control-type tests, it is of crucial importance that the control individuals are well matched with those in the case group. If this does not occur, it is possible that markers that appear to be associated with the trait will be identified, even though these markers may be biologically unrelated to the trait. One often-discussed type of mismatching is discussed in the following section. 

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Precipitation and Purification. During the hydrolysis, control tests are made by turbidimetric titration of samples taken intermittently. When the desired degree of hydrolysis is reached, the ester is precipitated from the reaction solution into water. It is important for the precipitate to have the proper texture for subsequent washing to remove acid and salts for thermal stabilization. Before precipitation, the reaction solution is usually diluted with additional aqueous acetic acid to reduce the viscosity. If a flake texture is desired, the solution is poured into a vigorously stirred, 10—15% aqueous acetic acid. To precipitate the acetate in powder form, dilute acetic acid is added to the stirred reaction solution. In both cases, the precipitated ester is suspended in 25—30% aqueous acid solutions and finally washed with deionized water. The dilution, precipitation temperature, agitation, and strength of the acid media must be controlled to ensure uniform texture. 

For drug substances and drug products, applications for enantiomers and racemates should include a stereochemically specific identity test and/or a stereochemically selective assay. The choice of control tests should be based on the method of manufacture and stability characteristics and, in the case of the finished product, its composition. 

Guidance on specifications is divided into universal tests/criteria which are considered generally applicable to all new substances/products and specific tests/criteria which may need to be addressed on a case-by-case basis when they have an impact on the quality for batch control. Tests are expected to follow the ICH guideline on analytical validation (Section 13.5.4). Identification of the drug substance is included in the universal category, and such a test must be able discriminate between compounds of closely related structure which are likely to be present. It is acknowledged here that optically active substances may need specific identification testing or performance of a chiral assay in addition to this requirement. 

If multi-section tires or experimental and control tires are mounted on the same axle, as is often the case for tests with truck tires on commercial fleets, the inverse of the above relation should be applied because the tires are now running under an imposed common slip. For the stiffer compound the slip is now larger and for the softer it is smaller and the stiffer compound becomes poorer and the soft compound becomes better than would be the case if whole tires of the same group had been mounted on the same axle. 

Lohmoller G, Matuschke A et al (1989) [False-positive test of autonomic neuropathy in HIV infection and AIDS Case control study of heart rate variability in 62 HIV positive patients]. Med Klin (Munich) 84(5) 242-245... 

Those aspects critical to the in vivo bioavailability of the product and routine control tests proposed to ensure that the product has consistent bioavailability from batch to batch. Where a product has low in vivo absorption, the evidence should be discussed and a conclusion reached as to whether this is due to intrinsic properties of the active ingredient(s) or whether it is related to the properties of the dosage form concerned. In the case of products intended to have a nonsystemic effect, the potential for systemic absorption may need to be considered. This may involve specific studies to determine the levels of the active ingredient(s) in the blood, plasma, urine, or feces and a discussion of the clinical significance of those results. 

The following procedures have evolved from field testing or from predictive methods using either CAM or PCAM equipment and are in order preferred for accuracy and realism of the determination. However, the nature of the process or circumstances of the test condition controls which of the procedures is used. In many cases, testing over several days is possible and desirable. In other cases, the tests can only last a few minutes, otherwise processing would be interrupted and process conditions would become atypical. 

Acute static bioassays are employed to evaluate sample effects on fish and microinvertebrates. Fathead minnows and Daphnia pulex are the freshwater species employed, while sheepshead minnows and grass shrimp are used for marine assays. In all cases, the test organisms are exposed for 96 hours to prescribed concentrations of the sample introduced into holding tanks maintained at the same environmental conditions as a control population. 

A case control study of four plants where DMF was produced or used showed no statistically significant association between ever having been exposed to DMF and subsequent development of cancers of the buccal cavity and pharynx, liver, prostate, and testes and malignant melanoma. Although prostate cancer was significantly elevated at one plant when examined by plant site, it did not appear to be related to exposure level or duration. 

Transfer each selected embryo to a single well of a multiwell tissue culmre plate (see Note 8) filled with embryo medium containing the substance to be tested (see Note 9). In most cases, the test substance will be evaluated at several concentrations along with a vehicle and/or untreated control, as appropriate. If possible, just prior to transferring embryos pre-fill the multiwell plates with the appropriate medium and test solution... 

In response to the biases inherent in case-control studies, family-based studies utilizing the transmission/ disequilibrium test (TDT) have become the gold standard in detection of association (Thomson, 1995 Spielman and Ewens, 1996). The family-based studies require analysis of alleles of both parents and the affected individual to determine which of the parental alleles are transmitted to the affected individual. Only data from heterozygous parents are informative and, for a two-allele system, each allele has an expected transmission rate of 50%. 

Finally, the statistical methods used to obtain estimates of relative risk, absolute rates of cancer, confidence intervals and significance tests, and to adjust for confounding should have been clearly stated by the authors. The methods used should preferably have been the generally accepted techniques that have been refined since the mid-1970s. These methods have been reviewed for case-control studies (Breslow Day, 1980) and for cohort studies (Breslow Day, 1987). 

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