The re-emerging role of microbial natural products in antibiotic discovery

New classes of antibacterial compounds are urgently needed to respond to the high frequency of occurrence of resistances to all major classes of known antibiotics. Microbial natural products have been for decades one of the most successful sources of drugs to treat infectious diseases but today, the emerging unmet clinical need poses completely new challenges to the discovery of novel candidates with the desired properties to be developed as antibiotics. While natural products discovery programs have been gradually abandoned by the big pharma, smaller biotechnology companies and research organizations are taking over the lead in the discovery of novel antibacterials. Recent years have seen new approaches and technologies being developed and integrated in a multidisciplinary effort to further exploit microbial resources and their biosynthetic potential as an untapped source of novel molecules. New strategies to isolate novel species thought to be uncultivable, and synthetic biology approaches ranging from genome mining of microbial strains for cryptic biosynthetic pathways to their heterologous expression have been emerging in combination with high throughput sequencing platforms, integrated bioinformatic analysis, and on-site analytical detection and dereplication tools for novel compounds. These different innovative approaches are defining a completely new framework that is setting the bases for the future discovery of novel chemical scaffolds that should foster a renewed interest in the identification of novel classes of natural product antibiotics from the microbial world.     

Omics based approach for biodiscovery of microbial natural products in antibiotic resistance era

The need for a new antibiotic pipeline to confront threat imposed by resistant pathogens has become a major global concern for human health. To confront the challenge there is a need for discovery and development of new class of antibiotics. Nature which is considered treasure trove, there is re-emerged interest in exploring untapped microbial to yield novel molecules, due to their wide array of negative effects associated with synthetic drugs. Natural product researchers have developed many new techniques over the past few years for developing diverse compounds of biopotential. Taking edge in the advancement of genomics, genetic engineering, in silico drug design, surface modification, scaffolds, pharmacophores and target-based approach is necessary. These techniques have been economically sustainable and also proven efficient in natural product discovery. This review will focus on recent advances in diverse discipline approach from integrated Bioinformatics predictions, genetic engineering and medicinal chemistry for the synthesis of natural products vital for the discovery of novel antibiotics having potential application.     

Current approaches to exploit actinomycetes as a source of novel natural products

For decades, microbial natural products have been one of the major sources of novel drugs for pharmaceutical companies, and today all evidence suggests that novel molecules with potential therapeutic applications are still waiting to be discovered from these natural sources, especially from actinomycetes. Any appropriate exploitation of the chemical diversity of these microbial sources relies on proper understanding of their biological diversity and other related key factors that maximize the possibility of successful identification of novel molecules. Without doubt, the discovery of platensimycin has shown that microbial natural products can continue to deliver novel scaffolds if appropriate tools are put in place to reveal them in a cost-effective manner. Whereas today innovative technologies involving exploitation of uncultivated environmental diversity, together with chemical biology and in silico approaches, are seeing rapid development in natural products research, maximization of the chances of exploiting chemical diversity from microbial collections is still essential for novel drug discovery. This work provides an overview of the integrated approaches developed at the former Basic Research Center of Merck Sharp and Dohme in Spain to exploit the diversity and biosynthetic potential of actinomycetes, and includes some examples of those that were successfully applied to the discovery of novel antibiotics.     

The role of combichem in antibiotic discovery

Combinatorial chemistry (combichem) has had a significant impact on the discovery of new antibiotics. Most of the successes have come from the use of small libraries to explore a specific pharmacophore. However, large diverse libraries are more appropriate when identifying hits by screening specific bacterial or fungal targets. Combichem has been used to optimize new azole and oxazolidinone leads. An entirely new class of antibiotics, inhibitors of bacterial peptidyl-deformylase, has been discovered by combining mechanism-based drug design and combichem. These compounds are active in vivo. The impact of combichem on discovery projects that aim to develop new antibiotics for the treatment of infectious diseases is discussed.     

In Situ Natural Product Discovery via an Artificial Marine Sponge

There is continuing international interest in exploring and developing the therapeutic potential of marine–derived small molecules. Balancing the strategies for ocean based sampling of source organisms versus the potential to endanger fragile ecosystems poses a substantial challenge. In order to mitigate such environmental impacts, we have developed a deployable artificial sponge. This report provides details on its design followed by evidence that it faithfully recapitulates traditional natural product collection protocols. Retrieving this artificial sponge from a tropical ecosystem after deployment for 320 hours afforded three actin–targeting jasplakinolide depsipeptides that had been discovered two decades earlier using traditional sponge specimen collection and isolation procedures. The successful outcome achieved here could reinvigorate marine natural products research, by producing new environmentally innocuous sources of natural products and providing a means to probe the true biosynthetic origins of complex marine–derived scaffolds.     

Do we need new antibiotics? The search for new targets and new compounds

Resistance to antibiotics and other antimicrobial compounds continues to increase. There are several possibilities for protection against pathogenic microorganisms, for instance, preparation of new vaccines against resistant bacterial strains, use of specific bacteriophages, and searching for new antibiotics. The antibiotic search includes: (1) looking for new antibiotics from nontraditional or less traditional sources, (2) sequencing microbial genomes with the aim of finding genes specifying biosynthesis of antibiotics, (3) analyzing DNA from the environment (metagenomics), (4) reexamining forgotten natural compounds and products of their transformations, and (5) investigating new antibiotic targets in pathogenic bacteria.     

Flavone-based analogues inspired by the natural product simocyclinone D8 as DNA gyrase inhibitors

The increasing occurrence of drug-resistant bacterial infections in the clinic has created a need for new antibacterial agents. Natural products have historically been a rich source of both antibiotics and lead compounds for new antibacterial agents. The natural product simocyclinone D8 (SD8) has been reported to inhibit DNA gyrase, a validated antibacterial drug target, by a unique catalytic inhibition mechanism of action. In this work, we have prepared simplified flavone-based analogues inspired by the complex natural product and evaluated their inhibitory activity and mechanism of action. While two of these compounds do inhibit DNA gyrase, they do so by a different mechanism of action than SD8, namely DNA intercalation.     

Importance of microbial natural products and the need to revitalize their discovery

Microbes are the leading producers of useful natural products. Natural products from microbes and plants make excellent drugs. Significant portions of the microbial genomes are devoted to production of these useful secondary metabolites. A single microbe can make a number of secondary metabolites, as high as 50 compounds. The most useful products include antibiotics, anticancer agents, immunosuppressants, but products for many other applications, e.g., antivirals, anthelmintics, enzyme inhibitors, nutraceuticals, polymers, surfactants, bioherbicides, and vaccines have been commercialized. Unfortunately, due to the decrease in natural product discovery efforts, drug discovery has decreased in the past 20 years. The reasons include excessive costs for clinical trials, too short a window before the products become generics, difficulty in discovery of antibiotics against resistant organisms, and short treatment times by patients for products such as antibiotics. Despite these difficulties, technology to discover new drugs has advanced, e.g., combinatorial chemistry of natural product scaffolds, discoveries in biodiversity, genome mining, and systems biology. Of great help would be government extension of the time before products become generic.     

Using display technologies to identify macrocyclic peptide antibiotics

Antibiotic resistant bacterial infections are now a leading cause of global mortality. While drug resistance continues to spread, the clinical antibiotic pipeline has become bare. This discord has focused attention on developing new strategies for antimicrobial discovery. Natural macrocyclic peptide-based products have provided novel antibiotics and antibiotic scaffolds targeting several essential bacterial cell envelope processes, but discovery of such natural products remains a slow and inefficient process. Synthetic strategies employing peptide display technologies can quickly screen large libraries of macrocyclic sequences for specific target binding and general antibacterial potential providing alternative approaches for new antibiotic discovery. Here we review cell envelope processes that can be targeted with macrocyclic peptide therapeutics, outline important macrocyclic peptide display technologies, and discuss future strategies for both library design and screening.     

Acquisition of a natural resistance gene renders a clinical strain of methicillin-resistant Staphylococcus aureus resistant to the synthetic antibiotic linezolid

Linezolid, which targets the ribosome, is a new synthetic antibiotic that is used for treatment of infections caused by Gram-positive pathogens. Clinical resistance to linezolid, so far, has been developing only slowly and has involved exclusively target site mutations. We have discovered that linezolid resistance in a methicillin-resistant Staphylococcus aureus hospital strain from Colombia is determined by the presence of the cfr gene whose product, Cfr methyltransferase, modifies adenosine at position 2503 in 23S rRNA in the large ribosomal subunit. The molecular model of the linezolid-ribosome complex reveals localization of A2503 within the drug binding site. The natural function of cfr likely involves protection against natural antibiotics whose site of action overlaps that of linezolid. In the chromosome of the clinical strain, cfr is linked to ermB, a gene responsible for dimethylation of A2058 in 23S rRNA. Coexpression of these two genes confers resistance to all the clinically relevant antibiotics that target the large ribosomal subunit. The association of the ermB/cfr operon with transposon and plasmid genetic elements indicates its possible mobile nature. This is the first example of clinical resistance to the synthetic drug linezolid which involves a natural resistance gene with the capability of disseminating among Gram-positive pathogenic strains.     

Tricyclic GyrB/ParE (TriBE) Inhibitors: A New Class of Broad-Spectrum Dual-Targeting Antibacterial Agents

Increasing resistance to every major class of antibiotics and a dearth of novel classes of antibacterial agents in development pipelines has created a dwindling reservoir of treatment options for serious bacterial infections. The bacterial type IIA topoisomerases, DNA gyrase and topoisomerase IV, are validated antibacterial drug targets with multiple prospective drug binding sites, including the catalytic site targeted by the fluoroquinolone antibiotics. However, growing resistance to fluoroquinolones, frequently mediated by mutations in the drug-binding site, is increasingly limiting the utility of this antibiotic class, prompting the search for other inhibitor classes that target different sites on the topoisomerase complexes. The highly conserved ATP-binding subunits of DNA gyrase (GyrB) and topoisomerase IV (ParE) have long been recognized as excellent candidates for the development of dual-targeting antibacterial agents with broad-spectrum potential. However, to date, no natural product or small molecule inhibitors targeting these sites have succeeded in the clinic, and no inhibitors of these enzymes have yet been reported with broad-spectrum antibacterial activity encompassing the majority of Gram-negative pathogens. Using structure-based drug design (SBDD), we have created a novel dual-targeting pyrimidoindole inhibitor series with exquisite potency against GyrB and ParE enzymes from a broad range of clinically important pathogens. Inhibitors from this series demonstrate potent, broad-spectrum antibacterial activity against Gram-positive and Gram-negative pathogens of clinical importance, including fluoroquinolone resistant and multidrug resistant strains. Lead compounds have been discovered with clinical potential; they are well tolerated in animals, and efficacious in Gram-negative infection models.     

Drug discovery from natural products

Natural product compounds are the source of numerous therapeutic agents. Recent progress to discover drugs from natural product sources has resulted in compounds that are being developed to treat cancer, resistant bacteria and viruses and immunosuppressive disorders. Many of these compounds were discovered by applying recent advances in understanding the genetics of secondary metabolism in actinomycetes, exploring the marine environment and applying new screening technologies. In many instances, the discovery of a novel natural product serves as a tool to better understand targets and pathways in the disease process. This review describes recent progress in drug discovery from natural sources including several examples of compounds that inhibit novel drug targets.     

Prospects for novel inhibitors of peptidoglycan transglycosylases

The lack of novel antimicrobial drugs under development coupled with the increasing occurrence of resistance to existing antibiotics by community and hospital acquired infections is of grave concern. The targeting of biosynthesis of the peptidoglycan component of the bacterial cell wall has proven to be clinically valuable but relatively little therapeutic development has been directed towards the transglycosylase step of this process. Advances towards the isolation of new antimicrobials that target transglycosylase activity will rely on the development of the enzymological tools required to identify and characterise novel inhibitors of these enzymes. Therefore, in this article, we review the assay methods developed for transglycosylases and review recent novel chemical inhibitors discovered in relation to both the lipidic substrates and natural product inhibitors of the transglycosylase step.     

Drug Discovery from Marine Microbes

The marine environment has been a source of more than 20,000 inspirational natural products discovered over the past 50 years. From these efforts, 9 approved drugs and 12 current clinical trial agents have been discovered, either as natural products or as molecules inspired from the natural product structure. To a significant degree, these have come from collections of marine invertebrates largely obtained from shallow-water tropical ecosystems. However, there is a growing recognition that marine invertebrates are oftentimes populated with enormous quantities of “associated” or symbiotic microorganisms and that microorganisms are the true metabolic sources of these most valuable of marine natural products. Also, because of the inherently multidisciplinary nature of this field, a high degree of innovation is characteristic of marine natural product drug discovery efforts.     

The spread of plasmids as a function of bacterial adaptability

Abstract The horizontal spread of plasmids among natural bacterial populations serves as an evolutionary function for adaptation to the ups and downs in nature. Recent evolutionary challenges are the introduction of antibiotics and the creation of new environmental conditions in agriculture and medical care. As a consequence, surviving bacterial populations have acquired new genetic determinants which enable the colonisation and maintenance in distinct ecological niches. The acquisition of new genetic determinants can take place rather rapidly because of the plasmids' biology: their self-transferability and their ability to pick up genes. As an example of horizontal gene transfer, from an ecological and evolutionary viewpoint, the emergence of resistance to streptothricins ( sat genes) is described.     

Hybrid isoprenoid secondary metabolite production in terrestrial and marine actinomycetes

Terpenoids are among the most ubiquitous and diverse secondary metabolites observed in nature. Although actinomycete bacteria are one of the primary sources of microbially derived secondary metabolites, they rarely produce compounds in this biosynthetic class. The terpenoid secondary metabolites that have been discovered from actinomycetes are often in the form of biosynthetic hybrids called hybrid isoprenoids (HIs). HIs include significant structural diversity and biological activity and thus are important targets for natural product discovery. Recent screening of marine actinomycetes has led to the discovery of a new lineage that is enriched in the production of biologically active HI secondary metabolites. These strains represent a promising resource for natural product discovery and provide unique opportunities to study the evolutionary history and ecological functions of an unusual group of secondary metabolites.     

Advancing cell wall inhibitors towards clinical applications

Natural products represent a major source of approved drugs and still play an important role in supplying chemical diversity. Consistently, 2014 has seen new, natural product-derived antibiotics approved for human use by the US Food and Drug Administration. One of the recently approved second-generation glycopeptides is dalbavancin, a semi-synthetic derivative of the natural product A40,926. This compound inhibits bacterial growth by binding to lipid intermediate II (Lipid II), a key intermediate in peptidoglycan biosynthesis. Like other recently approved antibiotics, dalbavancin has a complex history of preclinical and clinical development, with several companies contributing to different steps in different years. While our work on dalbavancin development stopped at the previous company, intriguingly our current pipeline includes two more Lipid II-binding natural products or derivatives thereof. In particular, we will focus on the properties of NAI-107 and related lantibiotics, which originated from recent screening and characterization efforts.     

Antimalarials from nature

Malaria is a major public health problem mainly due to the development of resistance by the most lethal causative parasitic species, Plasmodium falciparum to the mainstay drugs like chloroquine. New drugs with unique structures and mechanism of action are urgently required to treat sensitive and drug-resistant strains of malaria. Historically, compounds containing novel structure from natural origin represent a major source for the discovery and development of new drugs for several diseases. This review presents recent advances in antimalarial drug discovery from natural sources, including plant extracts, and compounds isolated from plants, bacteria, fungi and marine organisms. These compounds offer new and novel scaffolds for development as antimalarials. The literature from 1998 to October 2008 is reviewed. The review present literature compilation from plant and marine extracts, alkaloids (naphthylisoquinolines, bisbenzylisoquinolines, protoberberines and aporphines, indoles, manzamines, and miscellaneous alkaloids) terpenes (sesquiterpenes, triterpenes, diterpenes, and miscellaneous terpenes) quassinoids, flavonoids, limonoids, chalcones, peptides, xanthones, quinones and coumarines, and miscellaneous antimalarials from nature. The review also provides an outlook to recent semisynthetic approaches to antimalarial drugs discovered from natural sources.     

Renewed interests in the discovery of bioactive actinomycete metabolites driven by emerging technologies

Actinomycetes are prolific sources of bioactive molecules. Traditional workflows including bacterial isolation, fermentation, metabolite identification and structure elucidation have resulted in high rates of natural product rediscovery in recent years. Recent advancements in multi-omics techniques have uncovered cryptic gene clusters within the genomes of actinomycetes, potentially introducing vast resources for the investigation of bioactive molecules. While developments in culture techniques have allowed for the fermentation of difficult-to-culture actinomycetes, high-throughput metabolite screening has offered plenary tools to accelerate hits discovery. A variety of new bioactive molecules have been isolated from actinomycetes of unique environmental origins, such as endophytic and symbiotic actinomycetes. Synthetic biology and genome mining have also emerged as new frontiers for the discovery of bioactive molecules. This review covers the highlights of recent developments in actinomycete-derived natural product drug discovery.