Longitudinal Studies of Cardiac Troponin-I Concentrations in Serum from Male Sprague Dawley Rats: Baseline Reference Ranges and Effects of Handling and Placebo Dosing on Biological Variability

Abstract

ABSTRSCTSerum cardiac troponin-I has been validated as a biomarker for cardiotoxicity in numerous animal models; however, baseline reference ranges for cTnI concentration in a healthy population of laboratory rats, as well as an investigation of biological cTnI variability in rats with respect to time, handling, and placebo dosing methods, have not been reported. In this study, we used an ultrasensitive cTnI immunoassay to quantify hourly concentrations of cTnI in live rats handled under standard laboratory conditions using 15 mL of serum per determination. The baseline reference range (mean 4.94 pg/mL, range 1-15 pg/mL, 99% confidence interval [CI]) of cTnI concentration in rats was consistent with previously reported reference ranges for cTnI in humans (1-12 pg/mL) and with preliminary studies in dogs (1-4 pg/mL) and monkeys (4-5 pg/mL) using the same cTnI assay method. In addition, cTnI concentrations in individual rat serum samples show minimal biological variability over a twenty-four-hour interval when compared to a meaningful reference change value of 193% to 206%. Furthermore, measurements of cTnI concentration were consistent within the reference limits in individual rats over long periods and under three different standard laboratory handling conditions. Thus, using this new method, rats can be followed longitudinally at hourly intervals, and a doubling of cTnI concentration would be significant above biological variability. This is a new paradigm for preclinical testing, which allows transient changes in cTnI concentration to be accurately quantified. This understanding of baseline and biological variability in rats will be fundamental for designing and analyzing future studies that assess potential cardiotoxicity in drug development.

Longitudinal studies of cardiac troponin I concentrations in serum from male cynomolgus monkeys: resting values and effects of oral and intravenous dosing on biologic variability

“…In resting animals, the cTnI concentrations were relatively low and constant and ranged from 0.2 to 9.6 pg/mL (Mean = 2.5 pg/mL) with minimal variability between animals (Figures A, B and ). Values for cTnI concentration from the catheterized animals used in this study were similar to measurements of cTnI concentrations in noncatheterized male Cynomolgus monkeys determined previously using the Erenna immunoassay system …”

“…However, 2 monkeys given oral sham dosing had relatively small, transient increases of ~60 or 215 pg/mL that resolved within the 24‐hour period (Figure ). These results follow a pattern similar to previous studies of cTnI concentrations in Sprague–Dawley rats given oral sham dosing in which the majority of animals showed little or no change in cTnI concentration, while a few outliers had increased cTnI concentrations …”

“…There are numerous assays to measure cTn in laboratory animals, including many commercially available assays for cTnI quantitation but only a few for measurement of cTnT . We selected an ultrasensitive assay for measurement of cTnI concentration in our studies because it can quantify cTnI concentrations in very small sample volumes of serum or plasma and has an analytic threshold of < 1 ng/L in people and laboratory animals, which is below the lower limit of quantitation for many automated systems used in preclinical studies …”

“…The ultrasensitive method for determination of cTnI concentration employed has been used previously to determine the cTnI concentrations and biologic variability in healthy people and those with cardiac injury secondary to myocardial infarctions . Our group has used these technologies to evaluate resting cTnI concentrations in healthy rats and to determine the response of rats to oral dosing and simulated transport …”

Experimental determination of diagnostic window of cardiac troponins in the development of chronic anthracycline cardiotoxicity and estimation of its predictive value

“…In addition, it remains unknown whether the cTn concentration-time profile is the same at different stages of cardiotoxicity development. Furthermore, there is also no agreement on the predictive value of the very early cTn rise (after the first standard ANT dose) for the development of chronic ANT cardiotoxicity in the future [24][25][26][27][28][29]. Indeed, optimal timing a type of the assay may be crucial, particularly in the new era of highly sensitive troponin assays.…”