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

Nitric oxide is an inorganic free radical gas, of formula *N=O (abbreviated as NO). The discovery in 1987/88 that vascular endothelial cells are able to synthesize NO from L-arginine as a transcellular signal [1-4] was initially received by most biologists with considerable scepticism. By now, however, the existence of the L-arginine:NO pathway has been thoroughly documented and its relevance in biology is slowly being unravelled. All of this has led to the appearance of a new and vigorous field of research, as evidenced by the increasing number of publications relating to NO and NO synthases (Figure 1). This review will describe the known biochemical mechanisms involved in the synthesis of NO from L-arginine by the NO synthases and will also describe the nature of these enzymes, their inhibition and their molecular characteristics. For more extensive reviews about the biological roles of NO, see [5-7].

/pdf/nitric-oxide-synthases-in-mammals-1etyzijagx.pdf

Nitric oxide synthases in mammals.

■ Abstract Apoptosis is a genetically programmed, morphologically distinct form of cell death that can be triggered by a variety of physiological and pathological stimuli. Studies performed over the past 10 years have demonstrated that proteases play critical roles in initiation and execution of this process. The caspases, a family of cysteine-dependent aspartate-directed proteases, are prominent among the death proteases. Caspases are synthesized as relatively inactive zymogens that become activated by scaffold-mediated transactivation or by cleavage via upstream proteases in an intracellular cascade. Regulation of caspase activation and activity occurs at several different levels: ( a) Zymogen gene transcription is regulated; ( b) antiapoptotic members of the Bcl-2 family and other cellular polypeptides block proximity-induced activation of certain procaspases; and ( c) certain cellular inhibitor of apoptosis proteins (cIAPs) can bind to and inhibit active caspases. Once activated, caspases cleave a variety of intracellular polypeptides, including major structural elements of the cytoplasm and nucleus, components of the DNA repair machinery, and a number of protein kinases. Collectively, these scissions disrupt survival pathways and disassemble important architectural components of the cell, contributing to the stereotypic morphological and biochemical changes that characterize apoptotic cell death.

Mammalian caspases: structure ,a ctivation ,s ubstrates, and functions during apoptosis

▪ Abstract Apoptosis is a genetically programmed, morphologically distinct form of cell death that can be triggered by a variety of physiological and pathological stimuli. Studies performed over the past 10 years have demonstrated that proteases play critical roles in initiation and execution of this process. The caspases, a family of cysteine-dependent aspartate-directed proteases, are prominent among the death proteases. Caspases are synthesized as relatively inactive zymogens that become activated by scaffold-mediated transactivation or by cleavage via upstream proteases in an intracellular cascade. Regulation of caspase activation and activity occurs at several different levels: (a) Zymogen gene transcription is regulated; (b) antiapoptotic members of the Bcl-2 family and other cellular polypeptides block proximity-induced activation of certain procaspases; and (c) certain cellular inhibitor of apoptosis proteins (cIAPs) can bind to and inhibit active caspases. Once activated, caspases cleave a variet...

Mammalian Caspases: Structure, Activation, Substrates, and Functions During Apoptosis

Years of cumulative research can result in the development of a clinically useful drug, providing either a cure for a particular disease or symptomatic relief from a physiological disorder. A lead compound with a desired pharmacological activity may have associated with it undesirable side effects, characteristics that limit its bioavailability, or structural features which adversely influence its metabolism and excretion from the body. Bioisosterism represents one approach used by the medicinal chemist for the rational modification of lead compounds into safer and more clinically effective agents. The concept of bioisosterism is often considered to be qualitative and intuitive.1 The prevalence of the use of bioisosteric replacements in drug design need not be emphasized. This topic has been reviewed in previous years.2-5 The objective of this review is to provide an overview of bioisosteres that incorporates sufficient detail to enable the reader to understand the concepts being delineated. While a few popular examples of the successful use of bioisosteres have been included, the George Patani graduated with a B.Pharm. in 1992 from the College of Pharmaceutical Sciences, Mangalore University at Manipal, India. In 1996, he received his M.S. in Pharmaceutical Science at Rutgers University under the direction of Professor Edmond J. LaVoie. He is presently pursuing graduate studies in pharmaceutics. His current research interests are focused on drug design and controlled drug delivery.

https://www.mch.estranky.sk/file/228/1996chemrev96-3147-76_bioisosterism.pdf

Bioisosterism: A Rational Approach in Drug Design.

Biosynthesis and metabolism of tetrahydrobiopterin and molybdopterin.

The synthesis of 2,4-diamino-6-(2,5-dimethoxybenzyl)-5-methylpyrido[2,3-d]pyrimidine (BW301U, 7) by a route that has general applicability to the preparation of many 6-(substituted benzyl)-5-methylpyrido[2,3-d]pyrimidines is described. The key intermediate, 2,4-diamino-7,8-dihydro-6-(2,5-dimethoxybenzyl)-5-methyl-7-oxopyrido[2,3-d]pyrimidine (4), is converted to the 7-chloro compound 5 by treatment with a 1:1 complex of N,N-dimethylformamide--thionyl chloride, and 5 is hydrogenolyzed with palladium on charcoal in the presence of potassium hydroxide to yield 7. BW301U is a potent lipid-soluble inhibitor of mammalian dihydrofolate reductase and has significant activity against the Walker 256 carcinosarcoma in rats.

Synthesis and antitumor activity of 2,4-diamino-6-(2,5-dimethoxybenzyl)-5-methylpyrido[2,3-d]pyrimidine

The antiproliferative effects of interferon alpha (IFN-alpha) and interferon gamma (IFN-gamma) were found to be cell-dependent. Among the human cell lines examined, IFN-gamma had a greater antiproliferative effect against cell lines that exhibited induction of indoleamine 2,3-dioxygenase, such as the KB oral carcinoma or WiDr colon adenocarcinoma, than against those that lacked the enzyme activity, such as the SW480 colon adenocarcinoma or NCI-H128 small-cell lung carcinoma. Induction of this dioxygenase showed a clear temporal relationship with increased metabolism of L-tryptophan and the depletion of this amino acid in the culture medium. While 70-80% of L-tryptophan remained in the medium of IFN-alpha- or vehicle-treated cells, virtually all of this amino acid was depleted in the medium of the IFN-gamma-treated group following 2-3 days of culture. Supplementing the growth medium with additional L-tryptophan reversed the antiproliferative effect of IFN-gamma against KB cells in a dose- and time-dependent manner. The antiproliferative effects of IFN-alpha and IFN-gamma on SW480 and NCI-H128 cells, which are independent of the dioxygenase activity, and the inability of added L-tryptophan to reverse the effects of IFN-gamma in WiDr cells suggest multiple mechanisms of action of the IFNs. The data show that the antiproliferative effect of IFN-gamma through induction of indoleamine 2,3-dioxygenase, with a consequent L-tryptophan deprivation, is an effective means of regulating cell growth.

https://www.pnas.org/content/pnas/85/4/1242.full.pdf

Induction of indoleamine 2,3-dioxygenase: a mechanism of the antitumor activity of interferon gamma

Mammalian cells and tissues were found to have two pathways for the biosynthesis of tetrahydrobiopterin (BH4): (i) the conversion of GTP to BH4 by a methotrexate-insensitive de novo pathway, and (ii) the conversion of sepiapterin to BH4 by a pterin salvage pathway dependent on dihydrofolate reductase (5,6,7,8-tetrahydrofolate: NADP+ oxidoreductase, EC 1.5.1.3) activity. In a Chinese hamster ovary cell mutant lacking dihydrofolate reductase (DUKX-B11), endogenous formation of BH4 proceeds normally but, unlike the parent cells, these cells or extracts of them do not convert sepiapterin or 7,8-dihydrobiopterin to BH4. KB cells, which do not contain detectable levels of GTP cyclohydrolase or BH4 but do contain dihydrofolate reductase, readily convert sepiapterin to BH4 and this conversion is completely prevented by methotrexate. In supernatant fractions of bovine adrenal medulla, the conversion of sepiapterin to BH4 is completely inhibited by methotrexate. Similarly, this conversion in rat brain in vivo is methotrexate-sensitive. Sepiapterin and 7,8-dihydrobiopterin apparently do not enter the de novo pathway of BH4 biosynthesis and may be derived from labile intermediates which have not yet been characterized.

David S. Duch

Papers

Biosynthesis and metabolism of tetrahydrobiopterin and molybdopterin.

Synthesis and antitumor activity of 2,4-diamino-6-(2,5-dimethoxybenzyl)-5-methylpyrido[2,3-d]pyrimidine

Induction of indoleamine 2,3-dioxygenase: a mechanism of the antitumor activity of interferon gamma

Biosynthesis of tetrahydrobiopterin by de novo and salvage pathways in adrenal medulla extracts, mammalian cell cultures, and rat brain in vivo.

Biopterin cofactor biosynthesis: GTP cyclohydrolase, neopterin and biopterin in tissues and body fluids of mammalian species