Antimicrobial Indole-3-Carboxamido-Polyamine Conjugates Target Bacterial Membranes and Are Antibiotic Potentiators.

Small molecules that can restore the action of legacy antibiotics toward drug-resistant bacteria represent an area of ongoing research interest. We have previously reported indole-3-glyoxylamido and indole-3-acetamido-polyamine conjugates that exhibit intrinsic activity toward bacterial and fungal species, and the ability to enhance the action of doxycycline toward the Gram-negative bacteria Pseudomonas aeruginosa; however, these desirable activities were commonly associated with unfavorable cytotoxicity and/or red blood cell hemolytic properties. In this paper, we report the synthesis and biological investigation of a new class of α,ω-di(indole-3-carboxamido)polyamine derivatives, leading to the identification of several analogues that exhibit antimicrobial- and antibiotic-potentiating activities without detectable cytotoxic or hemolytic properties. 5-Bromo-substituted indole analogues 3 and 12-18 were generally more broad-spectrum in their activity than others in the set, with 13b (polyamine PA-3-6-3) being particularly notable for its anti-Staphylococcus aureus, Acinetobacter baumannii, and Cryptococcus neoformans activities (MIC ≤ 0.28 µM). The same analogue also restored the action of doxycycline toward P. aeruginosa with a 21-fold enhancement, while the corresponding 5-bromo-indole-3-carboxamide-PA3-7-3 analogue was able to enhance the action of both doxycycline and erythromycin toward P. aeruginosa and Escherichia coli, respectively. The analogue 13b was capable of disrupting the bacterial membrane of both S. aureus and methicillin-resistant S. aureus (MRSA) and the outer membrane of P. aeruginosa, suggesting that membrane perturbation could be a mechanism of action of both intrinsic antimicrobial activities and antibiotic potentiation.

Preliminary SAR of Novel Pleuromutilin-Polyamine Conjugates.

While pleuromutilin (1) and its clinically available derivatives (2-6) are highly effective against Gram-positive bacteria, they remain inactive against many pathogenic Gram-negative bacteria due to the efflux pump AcrAB-TolC. In an effort to broaden the spectrum of activity of pleuromutilin (1), we developed a series of novel pleuromutilin-polyamine conjugates (9a-f) which exhibited promising intrinsic antimicrobial properties, targeting both Gram-positive and Gram-negative bacteria, including Staphylococcus aureus, methicillin-resistant S. aureus (MRSA), and Escherichia coli, along with the fungal strain Cryptococcus neoformans, and were devoid of cytotoxic and hemolytic properties with the exception of one conjugate. Furthermore, this series displayed moderate to low antibiotic potentiation of legacy antibiotics doxycycline and erythromycin, with three conjugates enhancing the activity four-fold in combination with doxycycline. In comparison to pleuromutilin (1) and tiamulin (2), one of the conjugates exhibited an expanded spectrum of activity, including Gram-negative bacteria and fungi, making it a promising option for combating microbial infections.

Indole-3-Acetamido-Polyamines as Antimicrobial Agents and Antibiotic Adjuvants.

The widespread incidence of antimicrobial resistance necessitates the discovery of new classes of antimicrobials as well as adjuvant molecules that can restore the action of ineffective antibiotics. Herein, we report the synthesis of a new class of indole-3-acetamido-polyamine conjugates that were evaluated for antimicrobial activities against a panel of bacteria and two fungi, and for the ability to enhance the action of doxycycline against Pseudomonas aeruginosa and erythromycin against Escherichia coli. Compounds 14b, 15b, 17c, 18a, 18b, 18d, 19b, 19e, 20c and 20d exhibited strong growth inhibition of methicillin-resistant Staphylococcus aureus (MRSA) and Cryptococcus neoformans, with minimum inhibitory concentrations (MIC) typically less than 0.2 µM. Four analogues, including a 5-bromo 15c and three 5-methoxyls 16d-f, also exhibited intrinsic activity towards E. coli. Antibiotic kill curve analysis of 15c identified it to be a bactericide. While only one derivative was found to (weakly) enhance the action of erythromycin against E. coli, three examples, including 15c, were found to be strong enhancers of the antibiotic action of doxycycline against P. aeruginosa. Collectively, these results highlight the promising potential of α,ω-disubstituted indole-3-acetamido polyamine conjugates as antimicrobials and antibiotic adjuvants.

Structure-Activity Relationship Studies of Indolglyoxyl-Polyamine Conjugates as Antimicrobials and Antibiotic Potentiators.

Antibiotic resistance is a growing global health threat, requiring urgent attention. One approach to overcome antibiotic resistance is to discover and develop new antibiotic enhancers, molecules that work with legacy antibiotics to enhance their efficacy against resistant bacteria. Our previous screening of a library of purified marine natural products and their synthetic analogues led to the discovery of an indolglyoxyl-spermine derivative that exhibited intrinsic antimicrobial properties and was also able to potentiate the action of doxycycline towards the difficult to treat, Gram-negative bacterium Pseudomonas aeruginosa. A set of analogues have now been prepared, exploring the influence of indole substitution at the 5- and 7- positions and length of the polyamine chain on biological activity. While limiting cytotoxicity and/or hemolytic activities were observed for many analogues, two 7-methyl substituted analogues (23b and 23c) were found to exhibit strong activity towards Gram-positive bacteria with no detectable cytotoxicity or hemolytic properties. Different molecular attributes were required for antibiotic enhancing properties, with one example identified, a 5-methoxy-substitiuted analogue (19a), as being a non-toxic, non-hemolytic enhancer of the action of two tetracycline antibiotics, doxycycline and minocycline, towards P. aeruginosa. These results provide further stimulation for the search for novel antimicrobials and antibiotic enhancers amongst marine natural products and related synthetic analogues.

Investigation of α,ω-Disubstituted Polyamine-Cholic Acid Conjugates Identifies Hyodeoxycholic and Chenodeoxycholic Scaffolds as Non-Toxic, Potent Antimicrobials.

With the increased incidence of antibiotic resistance, the discovery and development of new antibacterials is of increasing importance and urgency. The report of the natural product antibiotic squalamine in 1993 has stimulated a lot of interest in the study of structurally simplified cholic acid-polyamine derivatives. We report the synthesis of a focused set of deoxycholic acid-polyamine conjugates and the identification of hyodeoxycholic acid derivatives as being potently active towards S. aureus MRSA and some fungal strains, but with no attendant cytotoxicity or hemolytic properties. Analogue 7e exhibited bactericidal activity towards a range of Gram-positive bacteria, while preliminary investigation of its mechanism of action ruled out the bacterial membrane as being a primary cellular target as determined using an ATP-release bioluminescence assay.

Discovery and Preliminary Structure-Activity Investigation of 3-Substituted-1H-imidazol-5-yl-1H-indoles with In Vitro Activity towards Methicillin-Resistant Staphylococcus aureus.

Antibiotics have been the cornerstone of modern medicine saving lives by virtue of being able to cure infectious diseases and to prevent infections in those who are immune compromised. Their intense use has led to a surging increase in the incidence of antibiotic-resistant bacteria resulting in a desperate need for antibiotics with new mechanisms of action. As part of our search for new antimicrobials we have screened an in-house library of compounds and identified two 3-substituted-1H-imidazol-5-yl-1H-indoles as weak growth inhibitors (MIC 16 µg/mL) against methicillin-resistant Staphylococcus aureus (MRSA). An extensive library of analogues was prepared using the Van Leusen three-component reaction, biological evaluation of which led to the identification of two analogues (26 and 32) with favorable anti-MRSA activity (MIC ≤ 0.25 µg/mL) which also lacked cytotoxic or hemolytic properties. The screening campaign also identified two derivatives, a phenethyl-indole-imidazole 57 and a 5-phenyl-1H-imidazole 111 that were non-toxic selective antifungals towards Cryptococcus neoformans. These results have identified 3-substituted-1H-imidazol-5-yl-1H-indoles and 5-phenyl-1H-imidazoles as new structural scaffolds for further investigation as anti-MRSA and anti-C. neoformans agents, respectively.

6-Bromoindolglyoxylamido derivatives as antimicrobial agents and antibiotic enhancers.

The combination of increased incidence of drug-resistant strains of bacteria and a lack of novel drugs in development creates an urgency for the search for new antimicrobials. Initial screening of compounds from an in-house library identified two 6-bromoindolglyoxylamide polyamine derivatives (3 and 4) that exhibited intrinsic antimicrobial activity towards Gram-positive bacteria, Staphylococcus aureus and S. intermedius with polyamine 3 also displaying in vitro antibiotic enhancing properties against the resistant Gram-negative bacterium Pseudomonas aeruginosa. A series of 6-bromo derivatives (5-15) were prepared and biologically evaluated, identifying analogues with enhanced antibacterial activity towards Escherichia coli and with moderate to excellent antifungal properties. Polyamine 3, which includes a spermine chain, was the most potent of the series - its mechanism of action was attributed to rapid membrane permeabilization and depolarization in both Gram-positive and Gram-negative bacteria.