Application of Quantitative NMR in Pharmacological Evaluation of Biologically Generated Metabolites: Implications in Drug Discovery

Abstract

ABSTRACT:It is important to gain an understanding of the pharmacological activities of metabolite(s) of compounds in development, especially if they are found in systemic circulation in humans. Pharmacological evaluation of metabolites is normally conducted with synthetic standards, which become available during various stages of drug development. However, the synthesis of metabolite standards may be protracted, taking anywhere from several weeks to months to be completed. This often slows down early pharmacological evaluation of metabolites. Once a metabolite(s) is found to possess comparable (or greater) pharmacological activity than the parent compound, additional studies are performed to better understand the implications of circulating pharmacologically active metabolite(s). To conduct some of these studies as early as possible without slowing the progression of a compound in development is important, especially if critical go or no-go decisions impinge on the outcomes from these studies. Early pharmacological evaluation of significant metabolites is hereby proposed to be conducted in the drug discovery stage so that all pertinent studies and information can be gathered in a timely manner for decisionmaking. It is suggested that these major metabolites be isolated, either from biological or chemical sources, and quantified appropriately. For biologically generated metabolites, NMR is proposed as the tool of choice to quantitate these metabolites before their evaluation in pharmacological assays. For metabolites that have the same UV characteristics as the parent compound, quantitation can be conducted using UV spectroscopy instead of NMR. In this article, we propose a strategy that could be used to determine the pharmacological activities of metabolites isolated in submilligram quantities.

Small Molecule Quantification by Liquid Chromatography-Mass Spectrometry for Metabolites of Drugs and Drug Candidates

“…For example, a number of investigators have used nanospray LC-MS to decrease the differences between the signal from parent drug and metabolites (Benetton et al, 2003;Hop et al, 2005;Valaskovic et al, 2006;Ramanathan et al, 2007). Furthermore, a number of other methods to estimate exposure to a metabolite using corrected responses in LC-MS/MS (Vishwanathan et al, 2009;Gao et al, 2010;Ma et al, 2010;Tong et al, 2010;Mutlib et al, 2011;Walker et al, 2011) or NMR based methods (Vishwanathan et al, 2009;Mutlib et al, 2011;Walker et al, 2011) have been developed.…”

Chromatographic resolution of closely related species: Drug metabolites and analogs

“…3) including testosterone (13), 1-dehydrotestosterone (14), 6␤-hydroxytestosterone (15), and 17␤-estradiol (16). These steroids have an essential role in mammals and other vertebrates, where they exhibit a wide-spectrum bioactivities [35,36].…”

Metabolism of F18, a Derivative of Calanolide A, in Human Liver Microsomes and Cytosol

“…Detecting and characterizing reactive metabolites in both experimental animals and humans are highly important initial steps in evaluating the underlying risk and can, in turn, guide the rational design of compounds that do not lead to the formation of reactive metabolites (Evans and Baillie, 2005). Furthermore, an early awareness of the pharmacological activities of major metabolites could facilitate the design of appropriate studies to better investigate the implications of such metabolites, particularly if they have biological activities similar to that of the parent drug (Mutlib et al, 2011). …”