The ability of certain antimutagenic agents to prevent development of antibiotic resistance.

Resistance to multiple antimicrobial agents has now become a prominent fact of contemporary life. It is believed that poor patient compliance, e.g. interrupted or premature cessation of therapy; and misuse or abuse of antibiotics, e.g. wrong antibiotic or insufficient dose, play important roles in resistance development. We present evidence that, this form of resistance often stems from spontaneous mutations accompanied by the positive selecting pressure of the doses of antibiotics being between the MIC and MBC levels. A number of antimutagenic agents, e.g. green tea catechins, and other antioxidants, etc. are able to suppress the emergence of resistance. In many cases, these agents are capable of exerting these effects at doses which by themselves produce no visible effect on growth. In a number of cases antimutagenic substances capable of preventing resistance emergence are present in normal food stuffs. These effects are exerted against resistance to tetracyclines, fluoroquinolones, macrolides, beta-lactams, aminoglycosides and the like. The implications of these laboratory findings for practical chemotherapy are discussed.

The 2-pyridone antibacterial agents: bacterial topoisomerase inhibitors.

Many attempts have been made to prepare analogs of 4-quinolone antibacterial agents bearing novel ring systems, which might retain the favorable properties of these widely used antibacterial agents and at the same time increase activity against multidrug-resistant bacteria, streptococci, and anaerobic microorganisms. One such attempt involved bioisosteric exchange of the 1-N atom and 4a-C atom of naphthyridones, quinolones, and benzoxazines to produce a family of highly active pyridopyrimidines, quinolizines, and ofloxacin bioisosteres. These new antibacterial agents have been named collectively as the 2-pyridones. Many hundreds of 2-pyridones have been synthesized and evaluated in vitro and in vivo, and selected members are advancing toward human clinical trials. Preparation of these bioisosteres required the development of enabling chemistry, as previous methods were unsuccessful in producing the needed core structures. This review compares the structure-activity relationships of these agents with known trends among 4-quinolones, from which it is seen that there are many parallels, but also some significant departures as well. Generally, 2-pyridones are more highly active in vitro and in vivo and more water soluble than comparable 4-quinolones. These properties are posited to arise from electronic and conformational alternations in these new substances. Selected members show excellent pharmacodynamic properties, justifying the view that this is a very promising new class of totally synthetic antibacterial agents.

University education of medicinal chemists: comparison of eight countries.

Medicinal chemists are mainly taught in faculties or schools of pharmacy and are available for employment. Yet major pharmaceutical research companies seek organic chemists, rather than medicinal chemists, for new drug discovery. This apparent contradiction led the Medicinal Chemistry Section of IUPAC to send a questionnaire regarding postgraduate academic education for medicinal chemists to the faculties or schools of pharmacy in eight countries, namely, France, Germany, Italy, Japan, Spain, Switzerland, UK and USA. The questionnaire aimed to elicit information about postgraduate medicinal chemistry students, their courses and training, and the occupations taken up after graduation. The replies representing 109 medicinal chemistry departments or sections have been analysed and the results are presented to provide a data base on modern medicinal chemistry curricula for comparative purposes. The information should help guide discussion of the optimum paths to be followed by students in preparation for their careers. The evidence suggests that academic training of medicinal chemists equips them to enter a wide range of occupations, many of which are in industry.

Role of antimutagens/anticarcinogens in cancer prevention.

Recently it has become increasingly clear that chemicals found in our foods and beverages can prevent the genetic damage that leads to cancer initiation. Such substances may also affect subsequent events in the pathways that lead to cancer, and may have the potential to inhibit the mutations that allow tumor cells to become resistant to antitumor agents. We describe here the antimutagenic potential of Glabrene analogs against EMS-induced mutations utilizing modified Ames tests in S. typhimurium TA 100 and E. coli JC 5088. Results of studies of the ability of well-known antioxidants such as EGCG and related compounds to prevent drug resistance mutations in microorganisms are described, and their possible significance in the prevention of chemotherapeutic drug-resistance in tumor cells is discussed.

Multiple drug resistance.

Multiple drug resistance to antibacterial agents, antifungals, antivirals, antiprotozoals, and antitumor agents has risen spectacularly in the last decade or so and presently threatens eventually to put an end to successful chemotherapy in all of the above fields. This review summarizes the known origins of the problem, its present dimensions, the means employed to combat the phenomenon and promising avenues for future developments.

Umbelliferone analogues and their potential to inhibit Benzo(a)pyrene- and hydrogen peroxide-induced mutations.

Following the natural product lead, farneciferol-D (kopetdaghin, 8), some ether analogues of umbelliferone were synthesized and assayed for their potential to be antimutagenic/anticarcinogenic against mutations induced by benzo(a)pyrene, a potent mutagen/carcinogen, and hydrogen peroxide, and for their ability to function as free radical scavengers. The "true" antimutagenic effect of these compounds was determined at half the nontoxic concentration in Salmonella typhimurium strains utilizing a modified Ames test protocol, and their free radical-scavenging ability was assayed utilizing a nonenzymatic phenazine methosulfate (PMS)-NADH system. Umbelliferone analogues 4 and 5 demonstrated good potential in preventing mutations induced by benzo(a)pyrene and hydrogen peroxide and also exhibited good superoxide scavenging ability in the PMS-NADH assay, suggesting that the antimutagenic activity of these analogues may be linked to their antioxidative properties.

Structure-antimutagenic activity relationship study of plicatin B.

A systematic structure-activity relationship study of plicatin B (1), an antimutagenic constituent of Psoralea juncea, was undertaken with a view toward elucidating its chemical mode of action and possibly optimizing its antimutagenic activity during the process. Compound 1 and its related analogues were examined for their antimutagenic activity against mutations induced by ethyl methanesulfonate, a direct acting mutagen and alkylating agent, in Salmonella typhimurium strain TA100, utilizing the modified Ames test protocol. The dihydro analogue 3 resulting from saturation of the conjugated alkene double bond of 1 was found to exhibit reduced cytotoxicity and enhanced efficacy relative to the parent compound. This result serves preliminarily to exclude a Michael acceptor role of the alpha,beta-unsaturated carbonyl moiety in connection with its antimutagenic activity.

The Medicinal and Bioorganic Chemistry Foundation--third winter conference. 23-29 January 1999, Steamboat Springs, CO, USA.

This was the third of an ongoing series of biennial conferences held in a relaxed setting, where medicinal chemists congregated to discuss recent enabling developments in organic-medicinal chemistry and biological findings of relevance to the selection of promising drug targets. Approximately 240 academic, industrial and government scientists gathered for this 5-day symposium devoted to optimization in chemical synthesis, methodologies for parallel array medicinal chemistry, applications of structural biology to drug discovery, impact of genomics technologies on pharmaceutical research, and multidrug resistance (MDR). In addition, a keynote address was delivered by Steven Hanessian (University of Montreal, Canada) on "Synthesis... Where do we go from here?" Sixteen oral presentations were delivered and eight papers were presented in a poster session. The plenary lectures are to be published later this year in special issues of Medicinal Research Reviews.

Antimutagenic/antioxidant activity of green tea components and related compounds.

The ability of green tea components and other antioxidant compounds to function as antimutagens/antioxidants has been well established, and their role in cancer prevention is supported by numerous epidemiological studies. We have utilized modified Ames tests, superoxide scavenging assays, and assays for protection against DNA scissions to compare and contrast the protective effects of various teas and commercial and laboratory-isolated tea components to those produced by compounds such as resveratrol, selenium, curcumin, vitamins C and E, quercetin dihydrate, sulforaphane, ellagic acid dihydrate, glutathione reduced, trolox, butylated hydroxanisole (BHA), butylated hydroxytoluene (BHT), and N-acetyl-L-cysteine (NAC). In Ames tests, employing hydrogen peroxide as a mutagen, epigallocatechin gallate (EGCG) produced the highest level of protection of all antioxidants tested. Measurement of protection against DNA scissions produced results that again showed that EGCG produced the strongest protective effects. In scavenging assays using a xanthine-xanthine oxidase (enzymatic system), epicatechin gallate (ECG) showed the highest scavenging potential. In a nonenzymatic (phenazine methosulfate-NADH) oxidizing system, EGCG once again showed the strongest effects. The implications of these and similar results are discussed in relation to cancer prevention and prevention of drug/antibiotic resistance.

Tuberculosis: a search for novel therapy starting with natural products.

The reemergence of tuberculosis and closely related diseases as significant public health problems is briefly reviewed with particular emphasis on the exacerbating role of AIDS and multiple drug resistance. Screening methods available for discovering new chemical entities active against resistant strains are discussed and their use in screening extracts and compounds for activity is illustrated with a number of newly discovered structures being presented. In particular, the properties of the potent and structurally novel indoloquinazolinone alkaloid, tryptanthrin, is described. Many analogs of this lead structure were synthesized by combinatorial and multiple parallel synthetic techniques and evaluated in vitro and in vivo for their potential in the chemotherapy of human infections.

Antitubercular natural products: berberine from the roots of commercial Hydrastis canadensis powder. Isolation of inactive 8-oxotetrahydrothalifendine, canadine, beta-hydrastine, and two new quinic acid esters, hycandinic acid esters-1 and -2.

Berberine (4) is responsible for the activity of an extract of a commercial root sample of Hydrastis canadensis against multiply drug resistant Mycobacterium tuberculosis. Two new quinic acid feruloyl esters, compounds 2 and 3, have been isolated from the same source along with canadine (1c), 8-oxotetrahydrothalifendine (1), and beta-hydrastine (5). These were found to be inactive. The structures of the new compounds were elucidated from spectral (1H, 13C, HMQC, HMBC, and H-H COSY) and chemical evidences.

The search for orally active medications through combinatorial chemistry.

The literature of combinatorial chemistry is reviewed with particular attention paid to considerations of absorption, distribution, metabolism and excretion in the design and evaluation of libraries containing drug-like molecules. Published libraries are evaluated in particular for the likelihood that the products would possess oral bioavailability.

Education of medicinal chemists in Departments of Medicinal Chemistry (U.S.A.).

The present state of faculties, student bodies, and curricula in departments of medicinal chemistry have been surveyed by questionnaire and analyzed in the context of perceptions of quality and content. The results reveal a healthy diversity of educational objectives and a broader range of educational objectives than those uncovered in previous surveys of the perceived needs of industrial departments of medicinal chemistry in their search for drug discovery personnel.

A simple, inexpensive apparatus for performance of preparative scale solution phase multiple parallel synthesis of drug analogs. I. Preparation of a retrospective library of quinolone antiinfective agents.

A simple inexpensive apparatus is described consisting of conveniently commercially available components which is suitable for the solution phase multiple parallel synthesis of 24-72 analogs of drug-like molecules. The use of the apparatus is illustrated by preparation of a retrospective library of over 100 analogs of antimicrobial fluoroquinolones prepared in 0% to quantitative yields. Each analog was prepared in up to 150 mg quantity and each was analyzed by NMR and mass spectrometric techniques to verify its purity and identity.

A simple, inexpensive apparatus for performance of preparative scale solution phase multiple parallel synthesis of drug analogs. II. Biological evaluation of a retrospective library of quinolone antiinfective agents.

A series of pure fluoroquinolone antiinfective agents was prepared by multiple parallel synthesis using a simple new apparatus. These compounds were evaluated biologically against Gram-positive and Gram-negative microorganisms and against a BCG strain transfected with luciferase in a fluorescence-based antitubercular assay. Activity against relatively fast growing, acid-fast Mycobacterium smegmatis was determined in part by agar-dilution streak assays. Data obtained against Escherichia coli-derived DNA gyrase does not correlate well with whole cell assays against E. coli. These compounds were assayed by a convenient glass-fiber filter binding method modified for high throughput screening. In these analogs, the results with a N-1 cyclopropyl substituent were often inferior to those obtained with a N-1 2',4'-difluorophenyl substituent. None of the new compounds prepared was superior in its antimycobacterial potency to ciprofloxacin or temafloxacin.

Rapid screening of natural products for antimycobacterial activity by using luciferase-expressing strains of Mycobacterium bovis BCG and Mycobacterium intracellulare.

The object of this study was to investigate the ability of a rapid luciferase assay to detect antimycobacterial activity in plant extracts. Recombinant strains of Mycobacterium bovis BCG (rBCG) and Mycobacterium intracellulare expressing firefly luciferase were used as the test organisms. Assays were conducted in a 96-well minitube format under biosafety level 2 conditions. Control and test wells were sampled immediately after inoculation and after 3 (recombinant M. intracellulare) and 5 (rBCG) days of incubation to measure luminescence with a microplate luminometer, and the relative change in luminescence was calculated as a percentage of control values. As an alternative test method, Alamar blue was added after 12 days of incubation, and changes in color were read visually. A total of 480 extracts were tested. Sixteen extracts were active against rBCG, and of those, seven were also active against recombinant M. intracellulare. With activity defined as a relative change in luminescence of < or = 1% (i.e., > or = 99% inhibition) and a persistence of blue color after addition of Alamar blue, there was 99.0% agreement between the two methods. Our results suggest that the luciferase assay is rapid and accurate and has the potential to greatly accelerate the evaluation of antimycobacterial activity in plant extracts in vitro. With this method, it is possible to screen a large number of samples in a short period of time.

Butitaxel analogues: synthesis and structure-activity relationships.

N-Acyl analogues 8, 9, and 12-26 of butitaxel (3) were prepared in one or two steps from amines 5 and 6 through Schotten-Baumann acylation. Seventeen novel analogues, consisting of aliphatic carbamates, alicyclic amides, and heteroaromatic amides, were synthesized. They were evaluated for their in vitro ability to stimulate the formation of microtubules, their cytotoxicity toward B16 melanoma cells, and their solubility in water. The most potent analogue found in this study was N-debenzoyl-N-(2-thenoyl)butitaxel (20), possessing ca. 2-fold better tubulin assembly properties and cytotoxic activity against B16 melanoma cells than paclitaxel. Compound 20 was ca. 25 times more water soluble than paclitaxel.

Natural antimutagenic agents.

Following a brief review of recent discoveries in the field of natural antimutagenic and tumor chemopreventive agents, contemporary findings in the author's laboratories employing the direct acting mutagen, ethyl methanesulfonate, in modified Ames tests and eukaryotic murine FM3A mammary tumor cells modified to be subject to thymidine-less death are described to illustrate the underlying principles. The EMS studies are illustrated with the isolation of the novel antimutagen, plicatin B, from the medicinal plants, Psoralea juncaea and P. plicata. The FM3A studies are carried out with extracts of Styrax asiatica, a plant previously studied extensively with the EMS system. The FM3A findings closely parallel the earlier work with EMS showing that the responsible agents, cinnamic acid, cinnamoyl ricinoleate and cinnamoyl cinnamate are effective both in prokaryotic and eukaryotic tests and that the new FM3A assay system has useful properties for screening and assay of novel antimutagenic agents.

Novel cytotoxic 3'-(tert-butyl) 3'-dephenyl analogs of paclitaxel and docetaxel.

3'-(tert-Butyl) 3'-dephenyl analogs of paclitaxel were synthesized from 10-deacetylbaccatin III and oxazolidinecarboxylic acid 7 followed by acylation of intermediate amines 10 and 11. Oxazolidinecarboxylic acid 7 was prepared in five steps and in good overall yield from L-tert-leucine. Twelve analogs were synthesized and evaluated for their in vitro ability to stimulate the formation of microtubules and for their cytotoxicity against B16 melanoma cells. Amide, carbamate, urea, and thiourea congeners were prepared. The most potent derivatives found in this study are the docetaxel analog 13, the N-[(tert-amyloxy)carbonyl] analog 17, and the 3'-phenylurea and 3'-tert-butylurea derivatives 20 and 23. Six of these analogs were shown to be ca. 90 times more soluble in water than paclitaxel and ca. 4-5 times more water-soluble than docetaxel.