Recent reports of increased tolerance to artemisinin derivatives-the last widely effective class of antimalarials -bolster the medical need for new treatments. The spirotetrahydro-β-carbolines, or spiroindolones, are a new class of fast-acting and potent schizonticidal drugs displaying low nanomolar potency against Plasmodium falciparum and Plasmodium vivax clinical isolates. Spiroindolones rapidly diminish protein synthesis in P. falciparum, an effect that is ablated in parasites bearing non-synonymous mutations in the gene encoding the P-type cation-transporter ATPase4 (PfATP4). The optimized spiroindolone NITD609 shows an acceptable safety profile and pharmacokinetic properties compatible with once-daily oral dosing; and demonstrates singledose efficacy in a rodent malaria model. Collectively, these data demonstrate that NITD609 possesses a pharmacological profile suitable for a new drug candidate for the treatment of malaria.Globally, 3.3 billion people are exposed to malaria, a devastating disease that causes over 800,000 deaths each year and kills more under five-year-olds than any other infectious agent (1). Fifty years ago, malaria had been eliminated from many areas of the world through effective antimalarial drug treatments, vector control interventions and disease prevention # Corresponding authors (Winzeler@scripps.edu and Thierry.diagana@novartis.com). * These authors equally contributed to this work One-sentence summary We describe the pharmacological profile of a new antimalarial drug candidate-the spiroindolone NITD609-which through a novel mechanism of action rapidly clears a Plasmodium infection upon administration of a single oral dose in a malaria mouse model. NIH Public Access Author ManuscriptScience. Author manuscript; available in PMC 2011 September 3. (2). However, the global spread of drug resistance resulted, by the 1980s, in a substantial increase in disease incidence and mortality. Today, some encouraging epidemiological data suggest that the introduction of new drugs (notably the artemisinin-based combination therapies or ACTs) may have reversed that trend (3). Derivatives of the endoperoxide artemisinin constitute the only antimalarial drugs that remain effective in all malariaendemic regions, but recent reports suggest that decades of continuous use as monotherapies might have fostered the emergence of resistance (4-6). This realization has triggered a concerted search for new drugs that could be deployed if artemisinin resistance were to spread.Many of the therapies currently in development utilize known antimalarial pharmacophores (e.g. aminoquinolines and/or peroxides) chemically modified to overcome the liabilities of their predecessors (7). While these compounds may prove to be important in the treatment of malaria, it would be preferable to discover novel chemotypes with a distinct mechanism of action (8). However, despite significant advances in our understanding of Plasmodium genome biology, the identification and validation of new drug targets has proven challengi...

Dengue virus (DENV), a mosquito-borne flavivirus, is a major public health threat. The virus poses risk to 2.5 billion people worldwide and causes 50 to 100 million human infections each year. Neither a vaccine nor an antiviral therapy is currently available for prevention and treatment of DENV infection. Here, we report a previously undescribed adenosine analog, NITD008, that potently inhibits DENV both in vitro and in vivo. In addition to the 4 serotypes of DENV, NITD008 inhibits other flaviviruses, including West Nile virus, yellow fever virus, and Powassan virus. The compound also suppresses hepatitis C virus, but it does not inhibit nonflaviviruses, such as Western equine encephalitis virus and vesicular stomatitis virus. A triphosphate form of NITD008 directly inhibits the RNA-dependent RNA polymerase activity of DENV, indicating that the compound functions as a chain terminator during viral RNA synthesis. NITD008 has good in vivo pharmacokinetic properties and is biologically available through oral administration. Treatment of DENV-infected mice with NITD008 suppressed peak viremia, reduced cytokine elevation, and completely prevented the infected mice from death. No observed adverse effect level (NOAEL) was achieved when rats were orally dosed with NITD008 at 50 mg/kg daily for 1 week. However, NOAEL could not be accomplished when rats and dogs were dosed daily for 2 weeks. Nevertheless, our results have proved the concept that a nucleoside inhibitor could be developed for potential treatment of flavivirus infections.

Candidate antibacterials are usually identified on the basis of their in vitro activity. However, the apparent inhibitory activity of new leads can be misleading because most culture media do not reproduce an environment relevant to infection in vivo. In this study, while screening for novel anti-tuberculars, we uncovered how carbon metabolism can affect antimicrobial activity. Novel pyrimidine–imidazoles (PIs) were identified in a whole-cell screen against Mycobacterium tuberculosis. Lead optimization generated in vitro potent derivatives with desirable pharmacokinetic properties, yet without in vivo efficacy. Mechanism of action studies linked the PI activity to glycerol metabolism, which is not relevant for M. tuberculosis during infection. PIs induced self-poisoning of M. tuberculosis by promoting the accumulation of glycerol phosphate and rapid ATP depletion. This study underlines the importance of understanding central bacterial metabolism in vivo and of developing predictive in vitro culture conditions as a prerequisite for the rational discovery of new antibiotics.

The antiplasmodial activity of a series of spirotetrahydro beta-carbolines is described. Racemic spiroazepineindole (1) was identified from a phenotypic screen on wild type Plasmodium falciparum with an in vitro IC(50) of 90 nM. Structure-activity relationships for the optimization of 1 to compound 20a (IC(50) = 0.2 nM) including the identification of the active 1R,3S enantiomer and elimination of metabolic liabilities is presented. Improvement of the pharmacokinetic profile of the series translated to exceptional oral efficacy in the P. berghei infected malaria mouse model where full cure was achieved in four of five mice with three daily doses of 30 mg/kg.

New chemotherapeutic compounds against multidrug-resistant Mycobacterium tuberculosis (Mtb) are urgently needed to combat drug resistance in tuberculosis (TB). We have identified and characterized the indolcarboxamides as a new class of antitubercular bactericidal agent. Genetic and lipid profiling studies identified the likely molecular target of indolcarboxamides as MmpL3, a transporter of trehalose monomycolate that is essential for mycobacterial cell wall biosynthesis. Two lead candidates, NITD-304 and NITD-349, showed potent activity against both drug-sensitive and multidrug-resistant clinical isolates of Mtb. Promising pharmacokinetic profiles of both compounds after oral dosing in several species enabled further evaluation for efficacy and safety. NITD-304 and NITD-349 were efficacious in treating both acute and chronic Mtb infections in mouse efficacy models. Furthermore, dosing of NITD-304 and NITD-349 for 2 weeks in exploratory rat toxicology studies revealed a promising safety margin. Finally, neither compound inhibited the activity of major cytochrome P-450 enzymes or the hERG (human ether-a-go-go related gene) channel. These results suggest that NITD-304 and NITD-349 should undergo further development as a potential treatment for multidrug-resistant TB.

However, most attempts to detect in vitro-in vivo correlations were unsuccessful, emphasizing the challenges of anti-TB drug discovery. The objective of this work is to provide a reference dataset for the TB drug discovery community with a focus on comparative in vitro potency and pharmacokinetics.

Diarrheal disease is responsible for 8.6% of global child mortality. Recent epidemiological studies found the protozoan parasite Cryptosporidium to be a leading cause of pediatric diarrhea with particularly grave impact on infants and immunocompromised individuals. There is neither a vaccine nor effective treatment. We establish a drug discovery process built on scalable phenotypic assays and mouse models that takes advantage of transgenic parasites. Screening a library of compounds with anti-parasitic activity we identified pyrazolopyridines as inhibitors of Cryptosporidium parvum and C. hominis. Oral treatment with the pyrazolopyridine KDU731 results in potent reduction in intestinal infection of immunocompromised mice. Treatment also leads to rapid resolution of diarrhea and dehydration in neonatal calves, a clinical model of cryptosporidiosis that closely resembles human infection. Our results suggest the Cryptosporidium lipid kinase PI(4)K as a target for pyrazolopyridines and warrant further preclinical evaluation of KDU731 as a drug candidate for the treatment of cryptosporidiosis.

Alleviating the burden of tuberculosis (TB) requires an understanding of the genetic basis that determines the emergence of drug-resistant mutants. PA-824 (pretomanid) is a bicyclic nitroimidazole class compound presently undergoing the phase III STAND clinical trial, despite lacking identifiable genetic markers for drug-specific resistant Mycobacterium tuberculosis. In the present study, we aimed to characterize the genetic polymorphisms of spontaneously generated PA-824-resistant mutant strains by surveying drug metabolism genes for potential mutations. Of the 183 independently selected PA-824-resistant M. tuberculosis mutants, 83% harbored a single mutation in one of five nonessential genes associated with either PA-824 prodrug activation (ddn, 29%; fgd1, 7%) or the tangential F 420 biosynthetic pathway (fbiA, 19%; fbiB, 2%; fbiC, 26%). Crystal structure analysis indicated that identified mutations were specifically located within the protein catalytic domain that would hinder the activity of the enzymes required for prodrug activation. This systematic analysis conducted of genotypes resistant to PA-824 may contribute to future efforts in monitoring clinical strain susceptibility with this new drug therapy.T uberculosis (TB) remains a major global health concern, with Ͼ8 million new cases and 1.8 million deaths occurring annually (WHO). This pandemic is exacerbated by the pervasive spread of multidrug-resistant (MDR)-TB that challenges clinicians to fight a disease with a limited arsenal of resources. The bicyclic 4-nitroimidazole chemotype has yielded two promising candidates, delamanid (OPC67683) and pretomanid (PA-824), which actively inhibit both nonreplicating and rapidly growing bacilli under aerobic and anaerobic conditions (1). Both drugs are undergoing clinical evaluation and FDA approval is pending for the treatment of MDR-TB. In 2013, delamanid received conditional marketing authorization by the European Medicines Agency (EMA) for use in adult patients deprived of other treatment options (2). PA-824 is in the phase III STAND clinical trial, and at this stage of the development pipeline, it would be beneficial to monitor the genetic basis of resistant clinical strains as they emerge in the wake of future implementation into a treatment protocol.Bicyclic 4-nitroimidazoles are prodrugs that require metabolic activation by a deazaflavin (cofactor F 420 )-dependent nitroreductase (Ddn) (3). Ddn (Rv3547) converts the prodrugs into three primary metabolites, a des-nitroimidazole and two unstable byproducts (4). Ddn is likely a membrane-bound protein (5) that is involved in a protective mechanism under oxidative stress (6). The major mechanism of action of nitroimidazole in active disease under aerobic conditions is to hinder the formation of mycolic acids, and under anaerobic conditions, the mechanism involves the induction of respiratory poisoning (4, 7). By inhibiting the formation of ketomycolates, a class of mycolic acids, nitroimidazole interferes with Mycobacterium tuberculosis cell wall formation, ...