Antimicrobial activity of the DNA minor groove binders furamidine and analogs

[摘要] Pentamidine(1) and berenil(2) are aromatic diamidines that are known to bind to the DNA minor groove at AT tracts1-4 and to have a long history as antimicrobial agents.5 These molecules may be viewed as benzamidines connected by acyclic linkers. In the 1930's these type diamidine molecules were studied for their use against African trypanosomes.6,7 A large number of synthetic aromatic diamidines which may be viewed as derived from 1 and 2 have demonstrated broad spectrum antimicrobial activity against several protozoan and fungal infections.8 At present, for this diamidine class of compounds only pentamidine has been found to have significant clinical use. Currently, pentamidine is used against primary stage African trypanosomiasis, antimony-resistant leishmaniasis and AIDs associated Pneumocystis carinii.9-11 Pentamidine is not effective when given orally and several toxic effects including hypotension, dysglycemia, and renal and hepatic toxicity have been reported.12-14 Some of the toxicity of pentamidine has been attributed to its metabolism involving cleavage of the linker.15    In an effort to develop aromatic diamidines with improved properties we have studied analogs in which the acyclic linkers of 1 and 2 have been replaced by heterocyclic rings. Sometime ago it was reported that 2,5-bis[4-amidinophenyl]furan (3,furamidine) was effective against trypanosomes in both murine and simian models.16,17 More recently furamidine was found to be highly active in animal models for Pneumocystis carinii and Cryptosporidium parvum.18,19 Extensive biophysical studies including NMR, X-ray crystallographic and footprinting investigations have shown that the primary mode of DNA binding for furamidine is at AT-rich sites of the minor groove.20-27 Furthermore, it has been recently clearly shown that furamidine and its analogs enter cells and bind to the nucleus, however their mode of cell penetration has not yet been determined.28 A detailed mechanism of antimicrobial action of furamidine and related diamidines has not been elucidated but it remains under investigation. Generally, antimicrobially active molecules of this type show strong affinity for the DNA minor groove, whereas close structural analogs of furamidine (monoamidines) do not significantly bind and are biologically inactive. These results have lead to our working hypothesis that active compounds first form a complex with the minor groove of DNA which leads to inhibition of one or more DNA dependent enzymes or control proteins. Different enzymes in different organisms are thought to be inhibited.22,29-32 Another possible mode of action for these type molecules may be direct inhibition of transcription.33, 34 It seems plausible that these molecules may act by multiple modes. Both the potent and the broad spectrum of activities showed by furamidine and related analogs coupled with their selective minor groove binding has led us to design other groove binders in an effort to discover improved antimicrobial agents. Despite the fact that both pentamidine and furamidine are highly effective on intravenous administration both have been show to be ineffective on oral administration in animal models.17,19,35,36 Development of potent dicationic antimicrobial agents alone is not sufficient to be of public health benefit. Consequently, another major part of our investigation focuses on prodrug approaches for amidines as a means to effectively deliver these potent antimicrobial agents.    2. Chemistry Results 2.1. Synthesis of Furamidine and Analogs The synthetic approaches to furamidine and analogs have evolved over the years and they are summarized here. Our original synthesis of furamidine is outlined in Scheme 116 and is essentially the approach described by Dann.37 The first step involves a Friedel-Crafts reaction between bromobenzene and furmaroyl chloride to form the corresponding dibenzoylethene. The reduction of 1,2-di-[4-bromobenzoyl]ethene can be performed by the action of Zn/HOAc or stannous chloride; the later is now the preferred reagent.38 Furan ring formation is achieved using acetic anhydride/H2SO4 which accomplishes the cyclodehydration reaction. 2,5-Bis [4-bromophenyl] furan is converted into 2,5-bis [4-cyanophenyl] furan by the action of copper (I) cyanide in refluxing DMF. The bis-nitrile is converted to furamidine by Pinner methodology.16 This synthetic route works well on the laboratory scale yet it involves several steps and the first step requires the use of either carbon disulfide or bromobenzene in excess. All of these factors are disadvantageous for large-scale synthesis. The key intermediate for preparation of furamidine and analogs is 2,5-bis [4-cyanophenyl] furan and therefore we have explored other synthetic approaches for this compound. Setter methodology was used to make 2,5-bis [4-cyanophenyl] furan as outlined in Scheme 2. In this approach a thiazolium catalyzed reaction of divinyl sulfone and 4-cyanobenzaldehyde gave the saturated 1,4-diketone in one step.39 Nevertheless, the yield for this reaction was consistently only about 40-50%. Conversion of the saturated 1,4-diketone to the furan was achieved in good yields under similar conditions as noted previously in Scheme 1. Stille coupling of 2,5-bis[tri-n-butylstannyl]furan with 4-bromobenzonitrile provided a short route to 2,5-bis [4-cyanophenyl] furan in an approximately 70% yield as outlined in Scheme 3.40 This route has been used for the scale-up GMP synthesis of a compound which is currently in clinical trials.41      Pinner methodology was used for the synthesis of 2,5-bis[4(N-alkylamidino)] furans and the structurally related bis-amidoximes directly from 2,5-bis [4-cyanophenyl] furan (Scheme 4). Arguably, the Pinner approach is the most widely used method for preparation of amidines. Although the method generally works quite well, it does require the rigorous exclusion of water. 16,19,42 A recently described approach for nitrile to amidine conversion involves formation of and subsequent reduction of an amidoxime.43 This method does not require rigorous water exclusion and therefore is extremely attractive. Unfortunately, this method is not easily applicable to the preparation of N-alkylamidines. Scheme 5 outlines our approach for the synthesis of the 2,4-bis[4-amidinophenyl]furans.42 The first step involves bromine addition to the 4,4'-bis-cyanochalcone to yield the corresponding dibromo compound. Reaction of excess sodium methoxide with the dibromo analog converts it into an enol ether. Reaction of dimethyl sulfonium ylide (formed in dimethylsulfoxide) with the enol ether yielded 2,4-bis[4-cyanophenyl]furan. An alternative route to the 2,4-bis[phenyl]furan system has been developed more recently.44 2,4-Bis[4-cyanophenyl]furan was converted into the desired diamidines using Pinner methodology. The syntheses of the extended analogs of furamidine 4-6 which incorporate benzimidazole ring systems are outlined in Scheme 6.45 In each

[学科分类] 化学（综合）

[关键词] furamidines;prodrugs;DNA minor groove binders