Pharmacokinetic, mass balance studies

Mass Balance Studies. Pharmacokinetic mass balance studies apply unlabeled, stable isotopes or radiolabeled compounds to study the extent of absorption and first-pass metabolism, distribution, and excretion of a given compound. In the microdosing approach, a C-labeled compound is administered to human volunteers at doses from as low as one microgram blood, urine, and fecal samples are collected over time and analyzed for C content by accelerator mass spectroscopy to determine half-life, plasma AUC, and maximal concentration (Cmax)- However, these methods are not very popular even when very low doses of radioactivity are involved. Highly sensitive, and more readily available, tech-niques for separation and analysis (e.g., LC-MS, LC-MS/MS) are frequently used alternatives that enable pharmacokinetic investigations and metabolite profiling of nonradiolabeled compounds. 

The permeability of the drug substance can be determined by different approaches such as pharmacokinetic studies in humans (fraction absorbed or mass balance studies) or intestinal permeability studies (in vivo intestinal perfusion studies in humans or suitable animal models or in vitro permeation studies using excised intestinal tissue or epithelial cell culture monolayers like CaCo-2 cell line). In order to avoid misclassification of a drug subject to efflux transporters such as P-glycoprotein, functional expression of such proteins should be investigated. Low- and high-permeability model... 

Permeability can be assessed by pharmacokinetic studies (for example, mass balance studies), or intestinal permeability methods, e.g. intestinal perfusion in humans, animal models, Caco 2 cell lines or other suitable, validated cell lines. In vivo or in situ animal models or in vitro models (cell lines) are only considered appropriate by HHS-FDA for passively transported drugs. It should be noted that all of these measurements assess the fraction absorbed (as opposed to the bioavailability, which can be reduced substantially by first-pass metabolism). 

The two most common drug metabolism studies are mass balance and tissue distribution. Mass balance studies are usually conducted in both the rodent and the nonrodent species used for toxicology evaluations, whereas tissue distribution is performed only in the rodent. For mass balance, a radio-labeled compound is administered to the test species and urine, feces, and, if necessary, expired air are collected at intervals and counted for total radioactivity. Commonly used intervals are 0-4, 4-8, 8-12, 12-24, and then daily, up to 168 hours or until more than 95% of the administered dose has been excreted. Depending on the pharmacokinetic profile of the candidate, other collection intervals can be selected to give a better picture of the excretion profile. For tissue distribution, a radiolabeled compound is administered to the test species, and after predefined times, usually 2, 4, 8, 24, and 48 hours, the test species... 

Beumer, J.H., Beijnen, J.H., and Schellens, J.H., Mass balance studies, with a focus on anticancer drugs, Clin. Pharmacokinet., 45, 33, 2006. 

Both of these approaches allow for assessment of systemic absorption by not conducting complete mass balance studies (e.g., expired air to catch absorbed compound metabolized to COj or HjO expired end products). In vivo dermal absorption studies not taking into account other routes of excretion must be interpreted with caution. One extension of this mass balance excretory analysis is to assess dermal absorption by only monitoring the primary excretory route for the compound studied. Dermal bioavailability has been assessed in exhaled breath using real-time ion trap mass spectrometry to track the uptake and ehmination of compounds (e.g., trichloroethylene) from dermal exposure in humans and rats (Poet et al., 2000). A physiologically based pharmacokinetic model can be used to estimate the total bioavailability of compoimds. The same approach was extended to determine the dermal uptake of volatile chemicals imder non-steady-state conditions using real-time breath analysis in rats, monkeys, and humans (Thrall et al., 2000). 

McDowell, J.A. Chittick, G.E. Ravitch, J.R. Polk, R.E. Kerkering, T.M. Stein, D.S. Pharmacokinetics of abacavir, a human immunodeficiency virus type 1 (HIV-1) reverse transcriptase inhibitor, administered in a single oral dose to HIV-1-infected adults a mass balance study, AntimicrobAgents Chemother., 1999, 43, 2855-2861. 

Exposure Assessment. Single and multiple dose pharmacokinetics, toxicokinetics and tissue distribution studies in relevant species are useful. Proteins are not given orally demonstrating absorption and mass balance is not typically a primary consideration. Rather, this segment of the test should be designed to determine... 

Gamer, R. C., Goris, I., Laenen, A. A., Vanhoutte, E., Meuldermans, W., Gregory, S., Gamer, J. V., Leong, D., Whattam, M., Calam, A., and Snel, C. A. (2002). Evaluation of accelerator mass spectrometry in a human mass balance and pharmacokinetic study—Experience with 14C-labeled (R)-6-[amino(4-chlorophenyl)(l-methyl-l //-imidazol-5-yl)methyl -4-(3-chlorophenyl)- l-methyl-2(17/ i-quinolinone (R115777), a famesyl transferase inhibitor. Drug Metab. Dispos. 30 823-830. 

The pharmacokinetic evaluation of biopharmaceuticals is generally simplified by the usual metabolism of products to small peptides and to amino acids, and thus classical biotransformation and metabolism studies are rarely necessary. Routine studies to assess mass balance are not useful. However, both single- and multiple-dose toxicokinetic data are essential in safety pharmacology asessments, and these can be complicated by two factors (1) biphasic clearance with a saturable, initial, receptor-dependent clearance phase, which may cause nonlinearity in dose-exposure relationships and doseresponses [14] and (2) antibody production against an antigenic biopharmaceutical that can alter clearance or activity in more chronic repeat-dose safety studies in the preclinical model. 

Accelerator mass spectrometry (AMS) is an ultrasensitive analytical method for radioactivity analysis. AMS offers 10 -10 -fold increases in sensitivity over LSC or other decay counting methods so that levels as low as 0.0001 DPM can be detected (Brown et al., 2005, 2006). AMS has been applied to mass balance determination, pharmacokinetic studies of total radioactivity, and measurement of chemically modified DNA and proteins in humans after the administration of a low radioisotope dose (approximately lOnCi/person for mass balance and drug metabolism studies) (Buchholz et al., 1999 Garner, 2000 Garner et al., 2002 Liberman et al., 2004 White and Brown, 2004). In addition, off-line HPLC-AMS has been explored for metabolite profiling after... 

It is difficult to establish uniform guidelines for pharmacokinetic studies for biotechnology-derived pharmaceuticals. Single and multiple dose pharmacokinetics, toxicokinetics, and tissue distribution studies in relevant species are useful however, routine studies that attempt to assess mass balance are not useful. Differences in pharmacokinetics among animal species may have a significant impact on the predictiveness of animal studies or on the assessment of dose response relationships in toxicity studies. Alterations in the pharmacokinetic profile due to immune-mediated clearance mechanisms may affect the kinetic profiles and the interpretation of the toxicity data. For some products there may also be inherent, significant delays in the expression of pharmacodynamic effects relative to the pharmacokinetic profile (e.g., cytokines) or there may be prolonged expression of pharmacodynamic effects relative to plasma levels. 

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