The median-effect equation derived from the mass-action law principle at equilibrium-steady state via mathematical induction and deduction for different reaction sequences and mechanisms and different types of inhibition has been shown to be the unified theory for the Michaelis-Menten equation, Hill equation, Henderson-Hasselbalch equation, and Scatchard equation. It is shown that dose and effect are interchangeable via defined parameters. This general equation for the single drug effect has been extended to the multiple drug effect equation for n drugs. These equations provide the theoretical basis for the combination index (CI)-isobologram equation that allows quantitative determination of drug interactions, where CI 1 indicate synergism, additive effect, and antagonism, respectively. Based on these algorithms, computer software has been developed to allow automated simulation of synergism and antagonism at all dose or effect levels. It displays the dose-effect curve, median-effect plot, combination index plot, isobologram, dose-reduction index plot, and polygonogram for in vitro or in vivo studies. This theoretical development, experimental design, and computerized data analysis have facilitated dose-effect analysis for single drug evaluation or carcinogen and radiation risk assessment, as well as for drug or other entity combinations in a vast field of disciplines of biomedical sciences. In this review, selected examples of applications are given, and step-by-step examples of experimental designs and real data analysis are also illustrated. The merging of the mass-action law principle with mathematical induction-deduction has been proven to be a unique and effective scientific method for general theory development. The median-effect principle and its mass-action law based computer software are gaining increased applications in biomedical sciences, from how to effectively evaluate a single compound or entity to how to beneficially use multiple drugs or modalities in combination therapies.

http://www.protocol-online.org/forums/uploads/monthly_09_2010/post-8165-008503000%201284677034.ipb

Theoretical Basis, Experimental Design, and Computerized Simulation of Synergism and Antagonism in Drug Combination Studies

Oxygenated derivatives of cholesterol (oxysterols) present a remarkably diverse profile of biological activities, including effects on sphingolipid metabolism, platelet aggregation, apoptosis, and ...

Oxysterols: modulators of cholesterol metabolism and other processes.

Sigma-ligands comprise several chemically unrelated drugs such as haloperidol, pentazocine, and ditolylguanidine, which bind to a family of low molecular mass proteins in the endoplasmic reticulum. These so-called sigma-receptors are believed to mediate various pharmacological effects of sigma-ligands by as yet unknown mechanisms. Based on their opposite enantioselectivity for benzomorphans and different molecular masses, two subtypes are differentiated. We purified the sigma1-binding site as a single 30-kDa protein from guinea pig liver employing the benzomorphan(+)[3H]pentazocine and the arylazide (-)[3H]azidopamil as specific probes. The purified (+)[3H]pentazocine-binding protein retained its high affinity for haloperidol, pentazocine, and ditolylguanidine. Partial amino acid sequence obtained after trypsinolysis revealed no homology to known proteins. Radiation inactivation of the pentazocine-labeled sigma1-binding site yielded a molecular mass of 24 +/- 2 kDa. The corresponding cDNA was cloned using degenerate oligonucleotides and cDNA library screening. Its open reading frame encoded a 25.3-kDa protein with at least one putative transmembrane segment. The protein expressed in yeast cells transformed with the cDNA showed the pharmacological characteristics of the brain and liver sigma1-binding site. The deduced amino acid sequence was structurally unrelated to known mammalian proteins but it shared homology with fungal proteins involved in sterol synthesis. Northern blots showed high densities of the sigma1-binding site mRNA in sterol-producing tissues. This is also in agreement with the known ability of sigma1-binding sites to interact with steroids, such as progesterone.

https://www.pnas.org/content/pnas/93/15/8072.full.pdf

Purification, molecular cloning, and expression of the mammalian sigma1-binding site

Properties of electrophoretically homogeneous phenobarbital-inducible and beta-naphthoflavone-inducible forms of liver microsomal cytochrome P-450.

Studies involving the metabolism of chemical carcinogens were important in the discovery and initial characterization of the cytochrome P-450 enzyme system. The Millers and their associates identified NADPH-dependent microsomal enzymes involved in the biotransformation of azo dyes (1, 2). These reactions, reduction and mixed-function oxidation, were also found to be important in the processing of many other carcin ogens, as well as drugs and steroids. Subsequent work led to the identification of the hemoprotein P-4503 as the terminal oxidase involved in such microsomal mixed-function oxidations (3,4). Other early studies revealed that administration of many different chemicals to animals could alter the metabolism of carcinogens (5-7). As we know now, many forms of P-450 exist and the expression of these enzymes is influenced by the com pounds which are given to the animals. Administration of the carcinogenic polycyclic hydrocarbon 3-methylcholanthrene to rats and analysis of the liver microsomes provided some of the early evidence that multiple forms of P-450 exist (8, 9). Since that time, a great deal of effort has been concentrated upon understanding the biochemistry of these P-450 enzymes, in terms of how they catalyze reactions and how their expression is regulated. Although much of the focus regarding P-450 proteins has involved their contributions to the metabolism of steroids and drugs, considerable effort is still directed towards understanding the roles of P-450s in carcinogen metabolism. The characteristics of P-450 enzymes will not be reviewed at length here.4 The microsomal enzymes are found in most tissues but are concentrated in liver. All have characteristic ferrouscarbon monoxide complex Soret peaks near 450 nm, have montimene molecular weights of about 50,000, and accept electrons from the flavoprotein NADPH-P-450 reducÃ-ase. Nearly 20 different P-450 proteins have been isolated from rat liver and all appear to be distinct gene products. Levels of the individual proteins are altered by administration of or exposure to a wide variety of chemicals, many of which are carcinogens themselves (including tumor initiators such as the polycyclic hydrocarbons and tumor promoters such as phÃ©nobarbital). Mitochondrial P-450s accept electrons from ferridoxins and seem to be primarily involved in anabolic steroid metabolism, although some evidence for roles in carcinogen oxidation has been presented (14). Several reviews and monographs deal with various aspects of the catalytic mechanisms and regulation of the P-450 enzymes

https://cancerres.aacrjournals.org/content/canres/48/11/2946.full.pdf

James L. Gaylor

Papers

Ligand interactions with hemoprotein P-450. II. Influence of phenobarbital and methylcholanthrene induction processes on P-450 spectra.

Cytochrome P-450-dependent oxidation of lanosterol in cholesterol biosynthesis. Microsomal electron transport and C-32 demethylation.

Microsomal enzymes of cholesterol biosynthesis. Purification of lanosterol 14 alpha-methyl demethylase cytochrome P-450 from hepatic microsomes.

Membrane-bound enzymes of cholesterol synthesis from lanosterol.

Microsomal enzymes of cholesterol biosynthesis from lanosterol. Solubilization and purification of steroid 8-isomerase.