A series of bis(amidinobenzimidazoles) and bis(amidinoindoles) with varied linking chains connecting the aromatic groups and various modifications to the basic amidino groups have been prepared. The calf thymus (CT) DNA and nucleic acid homopolymer [poly(dA).poly(dT),poly(dA-dT).poly-(dA-dT), and poly(dG-dC).poly(dG-dC)] binding properties of these compounds have been studied by thermal denaturation (delta Tm) and viscosity. The compounds show a greater affinity for poly(dA).poly(dT) and poly(dA-dT).poly(dA-dT) than for poly(dG-dC).poly(dG-dC). Viscometric titrations indicate that the compounds do not bind by intercalation. Molecular modeling studies and the biophysical data suggest that the molecules bind to the minor groove of CT DNA and homopolymers. Analysis of the shape of the molecules is consistent with this mode of nucleic acid binding. Compounds with an even number of methylenes connecting the benzimidazole rings have a higher affinity for DNA than those with an odd number of methylenes. Molecular modeling calculations that determine the radius of curvature of four defined groups in the molecule show that the shape of the molecule, as a function of chain length, affects the strength of nucleic acid binding. Electronic effects from cationic substituents as well as hydrogen bonding from the imidazole nitrogens also contribute to the nucleic acid affinity. The bis(amidinoindoles) show no structurally associated differential in nucleic acid base pair specificity or affinity.

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On the basis of a previously observed correlation between the antimicrobial activity and DNA binding strength of dicationic molecules, a series of 10 dicationically substituted bis-benzimidazoles were tested for activity in the rat model of Pneumocystis carinii pneumonia. One of the compounds, 1,4-bis[5-(2-imidazolinyl)-2-benzimidazolyl]butane, was found to be more potent and less toxic than pentamidine.

Nine dicationically substituted bis-benzimidazoles were examined for their in vitro activities against Giardia lamblia WB (ATCC 30957). The potential mechanisms of action of these compounds were evaluated by investigating the relationship among in vitro antigiardial activity and the affinity of the molecules for DNA and their ability to inhibit the activity of giardial topoisomerase II. Each compound demonstrated antigiardial activity, as measured by assessing the incorporation of [methyl-3H]thymidine by giardial trophozoites exposed to the test agents. Three compounds exhibited excellent in vitro antigiardial activities, with 50%o inhibitory concentrations which compared very favorably with those of two currently used drugs, quinacrine HCI and metronidazole. Putative mechanisms of action for these compounds were suggested by the strong correlation observed among in vitro antigiardial activity and the affinity of the molecules for natural and synthetic DNA and their ability to inhibit the relaxation activity of giardial topoisomerase II. A strong correlation between the DNA binding affinity of these compounds and their inhibition of giardial topoisomerase II activity was also observed.Giardia lamblia is a common cause of endemic and epidemic diarrheal disease throughout the world. Some individuals harbor asymptomatic infections, while others may exhibit acute or chronic gastrointestinal disease. Four agents are presently used to treat giardiasis: the nitroimidazoles metronidazole and tinidazole, the nitrofuran furazolidone, and quinacrine HCl, an acridine. Many problems are associated with the currently used chemotherapeutic agents, including treatment failures, unpleasant side effects, activity against normal intestinal flora, and possible carcinogenicity. While treatment of symptomatic individuals is recommended, there is controversy as to whether asymptomatic cyst passers should be treated, especially in light of the problems associated with the antigiardial agents presently available. More-effective and less-toxic agents are therefore needed for the treatment of giardiasis.The search for new antigiardial agents has been aided by improvements in axenic culturing of the organism (13) and drug susceptibility testing (3,12). Among the classes of compounds recently examined for antigiardial activity are anthelminthic benzimidazoles (1,8,14,15) Washington, DC 20307. which are used at present to treat the infection (2). In addition, there was a strong correlation between the antigiardial activities of the pentamidine analogs and their affinity for calf thymus DNA and poly(dA) poly(dT). The ability of pentamidine to bind to DNA has been proposed as a mechanism of action for this and related compounds. Pentamidine and related molecules have also been identified as potential inhibitors of the activity of type II topoisomerases (7,17).A number of dicationically substituted bis-benzimidazoles, originally developed as protease inhibitors and DNA binding agents, were available in our laboratory for in vitro antimi...

Although simple arylpiperazines are commonly considered to be moderately selective for 5-HT1B serotonin binding sites, N4-substitution of such compounds can enhance their affinity for 5-HT1A sites and/or decrease their affinity for 5-HT1B sites. A small series of 4-substituted 1-arylpiperazines was prepared in an attempt to develop agents with high affinity for 5-HT1A sites. Derivatives where the aryl portion is phenyl, 2-methoxyphenyl, or 1-naphthyl, and the 4-substituent is either a phthalimido or benzamido group at a distance of four methylene units away from the piperazine 4-position, display high affinity for these sites. One of these compounds, 1-(2-methoxyphenyl)-4-[4-(2-phthalimido)butyl]piperazine (18), possesses a higher affinity than 5-HT and represents the highest affinity (Ki = 0.6 nM) agent yet reported for 5-HT1A sites.

The antiprotozoal drug pentamidine [1,5-bis(4'-amidinophenoxy)pentaneI has been previously shown to be metabolized by rat liver microsomes, and five of the seven putative primary metabolites have been identified. With the synthesis and identification of 5-(4'-amidinophenoxy)pentanoic acid and 5-(4'-amidinophenoxy)-1-pentanol as the remaining two metabolites, the primary metabolism of pentamidine in rats appears fully characterized. Use of [14C] Pentamidine [1,5-bis(4'-amidinophenoxy)pentane] has been used for decades in the prophylaxis and treatment of African trypanosomiasis and treatment of antimony-resistant leishmaniasis and Pneumocystis carinii pneumonia (11,20). The increased incidence of P. carini pneumonia associated with the AIDS epidemic has brought about an increase in the clinical use of pentamidine in North America (15). Despite several early studies of the pharmacological properties of the drug (14,16,17,22,23), little was known about the compound. With the development of sensitive and accurate highperformance liquid chromatography (HPLC) assays (1, 9, 18), more detailed studies on the distribution and pharmacokinetics of pentamidine have been conducted (2,7,8). However, these studies have been performed with the misconception that pentamidine is metabolically inert, a conclusion primarily based on the work of Launoy et al. (16,17).We have previously demonstrated that pentamidine is readily converted to seven putative metabolites by rat liver microsomes (3, 4). Five of the seven primary metabolites have been identified as hydroxylated derivatives of the parent compound (see Fig. 1). The cytochrome P-450-dependent mixed-function oxidases have been identified as the enzyme system responsible for this activity (3). In this paper, we describe the characterization of the two remaining primary metabolites of pentamidine. In addition, isolated, perfused rat livers were used to determine the secondary metabolic pathways and to determine the extent of pentamidine metabolism in a model in which both primary and secondary metabolic systems were active. MATERIALS AND METHODSCompounds. Pentamidine (Darco, Durham, N.C.). The structures of all pentamidine metabolites are shown in Fig. 1.Synthesis of 5-(4'-amidinophenoxy)pentanoic acid. A solution of 5-bromovaleric acid (Aldrich Chemical Co., Milwaukee, Wis.) (6.0 g, 0.03 mol) in absolute ethanol (150 ml) and a few drops of H2SO4 was heated under reflux for 24 h. The ethanol was removed under a vacuum to afford 6.7 g (97%) of ethyl-5-bromo-pentanoate as an oil. Sodium (0.63 g, 27.5 mmol) was dissolved in absolute ethanol (20 ml), and 4-cyanophenol (3.0 g, 25 mmol) was added. The solution was heated at reflux for 30 min before ethyl-5-bromo-pentanoate (6.67 g, 32 mmol) in absolute ethanol (10 ml) was added and the solution was heated under reflux for 60 h. The ethanol was removed under reduced pressure, and the resulting ethyl-5-(4'-cyanophenoxy)pentanoate was recrystallized from ethanol-H20 to afford 4.5 g (73%) of the desired product as a white solid.A suspen...

Several classes of agents are known to bind at central 5-HT1A serotonin sites In order to challenge the hypothesis that these agents bind in a relatively similar manner (i.e., share common aryl and terminal amine sites), we prepared N-(phthalimidobutyl) derivatives of examples of several such agents. With regard to arylpiperazines, we had previously shown that introduction of this functionality at the terminal amine is tolerated by the receptor and normally results in a significant (greater than 10-fold) enhancement in affinity. The results of the present study show that this bulky functionality is also tolerated by the receptor when incorporated into examples of all other major classes of 5-HT1A agents (e.g., 2-aminotetralin, phenylalklamine, indolylalkylamine, and (aryloxy)alkylamine derivatives). The length of the alkyl chain that separates the terminal amine from the phthalimido group is of major importance, and a four-carbon chain appears optimal. Alteration of the length of this chain can have a significant influence on affinity; decreasing the chain length from four to three carbon atoms can reduce affinity by an order of magnitude, and further shortening can have an even more pronounced effect.