Search

The discovery of new antimalarial medicines with novel mechanisms of action is key to combating the problem of increasing resistance to our frontline treatments. The Open Source Malaria (OSM) consortium has been developing compounds ("Series 4") that have potent activity against Plasmodium falciparum in vitro and in vivo and that have been suggested to act through the inhibition of PfATP4, an essential membrane ion pump that regulates the parasite’s intracellular Na+ concentration. The structure of PfATP4 is yet to be determined. In the absence of structural information about this target, a public competition was created to develop a model that would allow the prediction of anti-PfATP4 activity among Series 4 compounds, thereby reducing project costs associated with the unnecessary synthesis of inactive compounds.In the first round, in 2016, six participants used the open data collated by OSM to develop moderately predictive models using diverse methods. Notably, all submitted models were available to all other participants in real time. Since then further bioactivity data have been acquired and machine learning methods have rapidly developed, so a second round of the competition was undertaken, in 2019, again with freely-donated models that other participants could see. The best-performing models from this second round were used to predict novel inhibitory molecules, of which several were synthesised and evaluated against the parasite. One such compound, containing a motif that the human chemists familiar with this series would have dismissed as ill-advised, was active. The project demonstrated the abilities of new machine learning methods in the prediction of active compounds where there is no biological target structure, frequently the central problem in phenotypic drug discovery. Since all data and participant interactions remain in the public domain, this research project “lives” and may be improved by others.

Herein we describe the optimization of a phenotypic hit against Plasmodium falciparum based on an aminoacetamide scaffold. This led to N‐(3‐chloro‐4‐fluorophenyl)‐2‐methyl‐2‐{[4‐methyl‐3‐(morpholinosulfonyl)phenyl]amino}propanamide (compound 28) with low‐nanomolar activity against the intraerythrocytic stages of the malaria parasite, and which was found to be inactive in a mammalian cell counter‐screen up to 25 μm. Inhibition of gametes in the dual gamete activation assay suggests that this family of compounds may also have transmission blocking capabilities. Whilst we were unable to optimize the aqueous solubility and microsomal stability to a point at which the aminoacetamides would be suitable for in vivo pharmacokinetic and efficacy studies, compound 28 displayed excellent antimalarial potency and selectivity; it could therefore serve as a suitable chemical tool for drug target identification., New roads to novel chemotypes: Compound selection from the TCAMS library (GSK), followed by iterative rounds of inhibitor design and synthesis afforded a single‐digit nanomolar inhibitor of P. falciparum (3D7), based on an aminoacetamide core. Compound 28 is a potent antimalarial and displayed excellent selectivity in a mammalian counter‐screen. This compound could be used as a suitable chemical tool for drug target identification or as a lead compound for further optimization.

Phenotypic HTS campaigns with a blood stage malaria assay have been used to discover novel chemotypes for malaria treatment with potential alternative mechanisms of action compared to existing agents. N1-(5-(3-Chloro-4-fluorophenyl)furan-2-yl)-N3,N3-dimethylpropane-1,3-diamine, 1 was identified as a modest inhibitor of P. falciparum NF54 (IC50 = 875 nM) with an apparent long plasma half-life after high dose oral administration to mice, although the compound later showed poor metabolic stability in liver microsomes through ring- and side chain-oxidation and N-dealkylation. We describe here the synthesis of derivatives of 1, exploring the influence of substitution patterns around the aromatic ring, variations on the alkyl chain and modifications in the core heterocycle, in order to probe potency and metabolic stability, where 4k showed a long half-life in rats.

There is a substantial need to develop new medicines against parasitic diseases via public-private partnerships. Based on high throughput phenotypic screens of largely protozoal pathogens and bacteria, the Medicines for Malaria Venture (MMV) has recently assembled an open-access ‘Pathogen Box’ containing 400 well-curated chemical compounds. In the present study, we tested these compounds for activity against parasitic stages of the nematode Haemonchus contortus (barber's pole worm). In an optimised, whole-organism screening assay, using exsheathed third-stage (xL3) and fourth-stage (L4) larvae, we measured the inhibition of larval motility, growth and development of H. contortus. We also studied the effect of the ‘hit’ compound on mitochondrial function by measuring oxygen consumption. Among the 400 Pathogen Box compounds, we identified one chemical, called tolfenpyrad (compound identification code: MMV688934) that reproducibly inhibits xL3 motility as well as L4 motility, growth and development, with IC50 values ranging between 0.02 and 3 μM. An assessment of mitochondrial function showed that xL3s treated with tolfenpyrad consumed significantly less oxygen than untreated xL3s, which was consistent with specific inhibition of complex I of the respiratory electron transport chain in arthropods. Given that tolfenpyrad was developed as a pesticide and has already been tested for absorption, distribution, excretion, biotransformation, toxicity and metabolism, it shows considerable promise for hit-to-lead optimisation and/or repurposing for use against H. contortus and other parasitic nematodes. Future work should assess its activity against hookworms and other pathogens that cause neglected tropical diseases., Graphical abstract, Highlights • We screened compounds in the ‘Pathogen Box’ for activity against the parasitic nematode Haemonchus contortus. • Tolfenpyrad, a pyrazole-5-carboxamide-based insecticide, has significant anthelmintic activity. • Tolfenpyrad inhibits oxygen consumption in parasitic larvae of H. contortus.

Microbial resistance to chemotherapy has caused countless deaths where malaria is endemic. Chemotherapy may fail either due to pre-existing resistance or evolution of drug-resistant parasites. Here we use a diverse set of antimalarial compounds to investigate the acquisition of drug resistance and the degree of cross-resistance against common resistance alleles. We assess cross-resistance using a set of 15 parasite lines carrying resistance-conferring alleles in pfatp4, cytochrome bc1, pfcarl, pfdhod, pfcrt, pfmdr, pfdhfr, cytoplasmic prolyl t-RNA synthetase or hsp90. Subsequently, we assess whether resistant parasites can be obtained after several rounds of drug selection. Twenty-three of the 48 in vitro selections result in resistant parasites, with time to resistance onset ranging from 15 to 300 days. Our data indicate that pre-existing resistance may not be a major hurdle for novel-target antimalarial candidates, and focusing our attention on fast-killing compounds may result in a slower onset of clinical resistance., It is unclear whether new antimalarial compounds may rapidly lose effectiveness in the field because of parasite resistance. Here, Corey et al. investigate the acquisition of drug resistance and the extent to which common resistance mechanisms decrease susceptibility to a diverse set of 50 antimalarial compounds.

The discovery and optimisation of a novel series of potent and selective p38α inhibitors is described. Evaluating the structure-activity relationship of an aminoalkyl substituent at the 3 position of the 2(1H)-pyrazinone core, p38α potency was increased 20000-fold. The most advanced compound (25) demonstrated excellent in vivo properties suitable for an inhaled route of administration.

Dissecting Plasmodium drug resistance Malaria is a deadly disease with no effective vaccine. Physicians thus depend on antimalarial drugs to save lives, but such compounds are often rendered ineffective when parasites evolve resistance. Cowell et al. systematically studied patterns of Plasmodium falciparum genome evolution by analyzing the sequences of clones that were resistant to diverse antimalarial compounds across the P. falciparum life cycle (see the Perspective by Carlton). The findings identify hitherto unrecognized drug targets and drug-resistance genes, as well as additional alleles in known drug-resistance genes. Science , this issue p. 191 ; see also p. 159

Chemogenetic characterization throughin vitroevolution combined with whole genome analysis is a powerful tool to discover novel antimalarial drug targets and identify drug resistance genes. Our comprehensive genome analysis of 262Plasmodium falciparumparasites treated with 37 diverse compounds reveals how the parasite evolves to evade the action of small molecule growth inhibitors. This detailed data set revealed 159 gene amplifications and 148 nonsynonymous changes in 83 genes which developed during resistance acquisition. Using a new algorithm, we show that gene amplifications contribute to 1/3 of drug resistance acquisition events. In addition to confirming known multidrug resistance mechanisms, we discovered novel multidrug resistance genes. Furthermore, we identified promising new drug target-inhibitor pairs to advance the malaria elimination campaign, including: thymidylate synthase and a benzoquinazolinone, farnesyltransferase and a pyrimidinedione, and a dipeptidylpeptidase and an arylurea. This deep exploration of theP. falciparumresistome and drug-able genome will guide future drug discovery and structural biology efforts, while also advancing our understanding of resistance mechanisms of the deadliest malaria parasite.One Sentence SummaryWhole genome sequencing reveals howPlasmodium falciparumevolves resistance to diverse compounds and identifies new antimalarial drug targets.

With ∼429,000 deaths in 2016, malaria remains a major infectious disease where the need to treat the fever symptoms, but also to provide relevant post-treatment prophylaxis, is of major importance. An azepanylcarbazole amino alcohol is disclosed with a long- and fast-acting in vivo antiplasmodial efficacy and meets numerous attributes of a desired post-treatment chemoprophylactic antimalarial agent. The synthesis, the parasitological characterization, and the animal pharmacokinetics and pharmacodynamics of this compound are presented along with a proposed target.

SUMMARYOpen science is a new concept for the practice of experimental laboratory-based research, such as drug discovery. The authors have recently gained experience in how to run such projects and here describe some straightforward steps others may wish to take towards more openness in their own research programmes. Existing and inexpensive online tools can solve many challenges, while some psychological barriers to the free sharing of all data and ideas are more substantial.

SUMMARYMalaria is a disease that still affects a significant proportion of the global human population. Whilst advances have been made in lowering the numbers of cases and deaths, it is clear that a strategy based solely on disease control year on year, without reducing transmission and ultimately eradicating the parasite, is unsustainable. This article highlights the current mainstay treatments alongside a selection of emerging new clinical molecules from the portfolio of Medicines for Malaria Venture (MMV) and our partners. In each case, the key highlights from each research phase are described to demonstrate how these new potential medicines were discovered. Given the increased focus of the community on eradicating the disease, the strategy for next generation combination medicines that will provide such potential is explained.

A major cause of the paucity of new starting points for drug discovery is the lack of interaction between academia and industry. Much of the global resource in biology is present in universities, whereas the focus of medicinal chemistry is still largely within industry. Open source drug discovery, with sharing of information, is clearly a first step towards overcoming this gap. But the interface could especially be bridged through a scale-up of open sharing of physical compounds, which would accelerate the finding of new starting points for drug discovery. The Medicines for Malaria Venture Malaria Box is a collection of over 400 compounds representing families of structures identified in phenotypic screens of pharmaceutical and academic libraries against the Plasmodium falciparum malaria parasite. The set has now been distributed to almost 200 research groups globally in the last two years, with the only stipulation that information from the screens is deposited in the public domain. This paper reports for the first time on 236 screens that have been carried out against the Malaria Box and compares these results with 55 assays that were previously published, in a format that allows a meta-analysis of the combined dataset. The combined biochemical and cellular assays presented here suggest mechanisms of action for 135 (34%) of the compounds active in killing multiple life-cycle stages of the malaria parasite, including asexual blood, liver, gametocyte, gametes and insect ookinete stages. In addition, many compounds demonstrated activity against other pathogens, showing hits in assays with 16 protozoa, 7 helminths, 9 bacterial and mycobacterial species, the dengue fever mosquito vector, and the NCI60 human cancer cell line panel of 60 human tumor cell lines. Toxicological, pharmacokinetic and metabolic properties were collected on all the compounds, assisting in the selection of the most promising candidates for murine proof-of-concept experiments and medicinal chemistry programs. The data for all of these assays are presented and analyzed to show how outstanding leads for many indications can be selected. These results reveal the immense potential for translating the dispersed expertise in biological assays involving human pathogens into drug discovery starting points, by providing open access to new families of molecules, and emphasize how a small additional investment made to help acquire and distribute compounds, and sharing the data, can catalyze drug discovery for dozens of different indications. Another lesson is that when multiple screens from different groups are run on the same library, results can be integrated quickly to select the most valuable starting points for subsequent medicinal chemistry efforts., Author Summary Malaria leads to the loss of over 440,000 lives annually; accelerating research to discover new candidate drugs is a priority. Medicines for Malaria Venture (MMV) has distilled over 25,000 compounds that kill malaria parasites in vitro into a group of 400 representative compounds, called the "Malaria Box". These Malaria Box sets were distributed free-of-charge to research laboratories in 30 different countries that work on a wide variety of pathogens. Fifty-five groups compiled >290 assay results for this paper describing the many activities of the Malaria Box compounds. The collective results suggest a potential mechanism of action for over 130 compounds against malaria and illuminate the most promising compounds for further malaria drug development research. Excitingly some of these compounds also showed outstanding activity against other disease agents including fungi, bacteria, other single-cellular parasites, worms, and even human cancer cells. The results have ignited over 30 drug development programs for a variety of diseases. This open access effort was so successful that MMV has begun to distribute another set of compounds with initial activity against a wider range of infectious agents that are of public health concern, called the Pathogen Box, available now to scientific labs all over the world (www.PathogenBox.org).

Malaria is estimated to have caused 584,000 deaths and 198 million cases of the disease globally in 2013. Though mortality rates are down 47% globally since 2000 and significant progress has been made in the quest for eradication, reported occurrences of resistance against current therapeutics threaten to reverse that progress. Recently, antimalarials which were once considered unsuitable therapeutic agents have been revisited to improve physicochemical properties and efficacy required for selection as a drug candidate. One such compound is 4(1H)-quinolone ICI 56,780, which is known to be a causal prophylactic that also displays blood schizontocidal activity against P. berghei. Rapid induction of parasite resistance in rodent malaria models however, stalled its further development. We have completed a full structure-activity relationship study on ICI 56,780 focusing on the reduction of cross-resistance with atovaquone for activity against the clinical isolates W2 and TM90-C2B. The optimization work focusing primarily on the 3-, 6-, and 7-positions of the 4(1H)-quinolone core. The best compound 16c, a 3-bromo-substituted 4(1H)-quinolone, was found to possess low nanomolar EC(50) values against the blood stages of the P. falciparum strains W2 and TM90-C2B, as well as low digit nanomolar inhibitory constant against P. berghei liver stages. Furthermore, this bromo compound 16c was also shown in an in vivo efficacy study to reduce the parasitemia by 61 % on day 6 post-exposure, whereas the original 4(1H)-quinolone displayed no inhibition.

Malaria causes a huge humanitarian and economic burden. Parasite resistance to established and recently launched anti-malarials is a major issue which, when combined with a malaria eradication agenda, means there is a considerable need for new small molecule anti-malarials. Catalyzed by a recent surge in funding for malaria drug discovery and development, there is an increasing number of compounds in the malaria pipeline.This review covers patents published in English between January 2010 and June 2011, which feature small molecules for the treatment of malaria. Approximately 50 series of compounds are described. Patents covering clinical applications, diagnosis kits or vaccines are not included, nor patents where the principle disease focus is not malaria.There is considerable activity in the field of small molecules for malaria which is likely to continue. The ultimate goal is to identify novel drugs to support the malaria eradication agenda. This requires safe and efficacious compounds, from novel chemotypes, which rapidly kill parasites and which are readily synthesized from cheap starting materials. In addition, compounds which have activity in the liver stages or in transmission blocking may be prioritized for development over analogs related to established anti-malarial series targeting the asexual blood stages of the parasite.

The identification of a new series of P. falciparum growth inhibitors is described. Starting from a series of known human class I HDAC inhibitors a SAR exploration based on growth inhibitory activity in parasite and human cells-based assays led to the identification of compounds with submicromolar inhibition of P. falciparum growth (EC50500 nM) and good selectivity over the activity of human HDAC in cells (up to50-fold). Inhibition of parasital HDACs as the mechanism of action of this new class of selective growth inhibitors is supported by hyperacetylation studies.

With the extensive focus on lean conformation in the finished hog over the past 25 years, there is some indication that pork quality has suffered and taste has been bred out of today’s pork. Similar to the Certified Angus Beef program (a breed noted for intramuscular fat), small-scale farmers can promote a different ‘upscale’ pork by using breeds that will focus on pork taste exclusively, and feeding diets (possibly apart from corn and soybeans) to enhance flavor. Two experiments were devised to examine the influence of breed, rearing environment and diet on fresh pork quality and flavor. In Trial 1, three sow breed groups (Tamworth, Tamworth · Landrace, or Hampshire · Landrace) were mated to Duroc boars. Littermates (91 pigs total) were assigned randomly at weaning to one of three treatments: (1) confinement, (2) dry-lot and (3) pasture. All pigs were full fed a 16% crude protein (CP) grow-finish ration. Pasture pigs were allowed access to plots consisting of predominately white and crimson clovers with warm-season grasses (Bermuda grass and crab grass). Hampshire crosses had higher Minolta L* scores, indicating a paler, less desirable loin. Pork quality was similar across rearing environments except for lower initial pH levels observed in the pasture system and higher drip-loss percentage recorded in both outdoor systems. In Trial 2, 42 Tamworth · Duroc littermates were randomly assigned to one of two rearing environments (confinement or pasture) at 55 kg and full fed a 14% CP diet. Pigs finishing on pasture had access to standing, mature barley. Pork from the pasture system was darker than that from pigs reared in confinement. No differences were observed in sensory evaluation of the pork for the rearing environments examined. For both trials, intramuscular fat levels (< 2%) and visual color scores were too low to be considered for ‘upscale’ markets. Alternative diets to produce niche-market pork are unlikely to influence flavor without adequate levels of marbling.

The discovery of new chemotypes to feed the pipeline of antimalarial drugs remains a constant challenge, particularly in light of emerging resistance to current therapies. Recently, phenotypic screenings have been successfully used for antimalarial hit generation where the biological target(s) may often not be clearly identified. To catalyse malaria research by both filling the pipeline and having a better understanding of ligand-target relationships, a unique screening tool has been elaborated: the Malaria Box. The Malaria Box is a set composed of 400 commercially available chemical entities derived from a selection of more than 20,000 hits from the screening of corporate and academic libraries [1-3]. The originality of the Malaria Box relies in its composition of 200 lead-like and 200 probe-like compounds that have confirmed activity on blood-staged Plasmodium falciparum and that have been assessed for cytotoxicity. Lead-like compounds commensurate with oral absorption and the presence of known toxicophores has been reviewed. Conversely probe-like compounds are intended to represent the broadest cross-section of structural diversity. Significantly, the scope of the Malaria Box goes beyond the Malaria field as active compounds may have utility in other parasitic or neglected diseases. It is well-documented that artemisinin was initially discovered from helminth research and is currently a gold standard drug against Malaria. Also, the presence of orthologues of various molecular targets may lead to new therapeutic applications in orphan diseases or for example oncology. Ultimately, the data collection resulting from the Malaria Box would enable the community to better understand similarities and differences between parasite diseases or orphan diseases by mining data sets that were previously considered separately [4]. Herein we disclose the selection process applied to assemble the Malaria Box as well a preliminary results.

The Global Enteric Multicenter Study (GEMS), a study of infectious diarrhea involving over 20,000 children at seven sites in sub-Saharan Africa and South Asia, recently reported that the little-studied protozoan parasite Cryptosporidium ranks second to rotavirus as a cause of life-threatening diarrhea in infants and was also associated with growth stunting and excess mortality [1]. Cryptosporidium species, predominantly Cryptosporidium hominis and Cryptosporidium parvum, were previously well known for causing chronic diarrhea in AIDS patients, as well as for their chlorine resistance and their association with waterborne outbreaks in the developed world [2–4]. Numerous smaller studies had also demonstrated the importance of cryptosporidiosis in young children and its association with malnutrition (reviewed in [5]) [5–19], but Cryptosporidium had not previously garnered significant attention from the pharmaceutical industry or major funding organizations such as the Bill and Melinda Gates Foundation. The GEMS put cryptosporidiosis into context amongst more studied diarrheal pathogens and brought it to the attention of these organizations. No vaccine for cryptosporidiosis exists, and the available treatments for those most at risk are inadequate. The only licensed drug, nitazoxanide, is unreliable in severely malnourished children (~56% improvement in diarrhea at 7 days versus 26% in controls [20]), and shows no benefit relative to placebo in HIV-infected patients [20–22]. More reliable, efficacious, and faster-acting treatments are needed for these populations. Interest in Cryptosporidium drug development has correspondingly increased, and despite a limited experimental system, several new molecules with in vitro and, in some cases, validated in vivo activity against Cryptosporidium have been reported [23–35]. However, data regarding these compounds are difficult to compare due to use of differing in vitro and in vivo models and lack of assay standardization (see [36] for a thorough review of this and other barriers). A major current challenge then is to develop a common framework to help Cryptosporidium researchers and funding agencies to prioritize screening hits and leads for further development and to define essential developmental milestones. In this article, we propose a target product profile (TPP) (Table 1) and testing cascade (Fig 1) that can be used to guide this process. This TPP will undoubtedly require revision as progress is made, but can provide clear initial goals and a framework for go/no-go decision making in order to facilitate the appropriate distribution of effort and limited resources. Fig 1 Proposed development scheme and anticipated costs for Cryptosporidium drug development. Table 1 Proposed target product profile for treatments for diarrhea due to cryptosporidiosis.

Historically, one of the key problems in neglected disease drug discovery has been identifying new and interesting chemotypes. Phenotypic screening of the malaria parasite, Plasmodium falciparum has yielded almost 30,000 submicromolar hits in recent years. To make this collection more accessible, a collection of 400 chemotypes has been assembled, termed the Malaria Box. Half of these compounds were selected based on their drug-like properties and the others as molecular probes. These can now be requested as a pharmacological test set by malaria biologists, but importantly by groups working on related parasites, as part of a program to make both data and compounds readily available. In this paper, the analysis and selection methodology and characteristics of the compounds are described.

The development of new antimalarial compounds remains a pivotal part of the strategy for malaria elimination. Recent large-scale phenotypic screens have provided a wealth of potential starting points for hit-to-lead campaigns. One such public set is explored, employing an open source research mechanism in which all data and ideas were shared in real time, anyone was able to participate, and patents were not sought. One chemical subseries was found to exhibit oral activity but contained a labile ester that could not be replaced without loss of activity, and the original hit exhibited remarkable sensitivity to minor structural change. A second subseries displayed high potency, including activity within gametocyte and liver stage assays, but at the cost of low solubility. As an open source research project, unexplored avenues are clearly identified and may be explored further by the community; new findings may be cumulatively added to the present work., Starting from hits identified by a pharmaceutical company, new antimalarials have been discovered using an “open source” approach that discloses data in real time and eschews secrecy.

Background In view of the need to continuously feed the pipeline with new anti-malarial agents adapted to differentiated and more stringent target product profiles (e.g., new modes of action, transmission-blocking activity or long-duration chemo-protection), a chemical library consisting of more than 250,000 compounds has been evaluated in a blood-stage Plasmodium falciparum growth inhibition assay and further assessed for chemical diversity and novelty. Methods The selection cascade used for the triaging of hits from the chemical library started with a robust three-step in vitro assay followed by an in silico analysis of the resulting confirmed hits. Upon reaching the predefined requirements for selectivity and potency, the set of hits was subjected to computational analysis to assess chemical properties and diversity. Furthermore, known marketed anti-malarial drugs were co-clustered acting as ‘signposts’ in the chemical space defined by the hits. Then, in cerebro evaluation of the chemical structures was performed to identify scaffolds that currently are or have been the focus of anti-malarial medicinal chemistry programmes. Next, prioritization according to relaxed physicochemical parameters took place, along with the search for structural analogues. Ultimately, synthesis of novel chemotypes with desired properties was performed and the resulting compounds were subsequently retested in a P. falciparum growth inhibition assay. Results This screening campaign led to a 1.25% primary hit rate, which decreased to 0.77% upon confirmatory repeat screening. With the predefined potency (EC50 10) criteria, 178 compounds progressed to the next steps where chemical diversity, physicochemical properties and novelty assessment were taken into account. This resulted in the selection of 15 distinct chemical series. Conclusion A selection cascade was applied to prioritize hits resulting from the screening of a medium-sized chemical library against blood-stage P. falciparum. Emphasis was placed on chemical novelty whereby computational clustering, data mining of known anti-malarial chemotypes and the application of relaxed physicochemical filters, were key to the process. This led to the selection of 15 chemical series from which ten confirmed their activity when newly synthesized sample were tested.