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Viruses produce more viruses by manipulating the metabolic and replication systems of their host cells. Many have acquired metabolic genes from ancestral hosts and use the encoded enzymes to subvert host metabolism. The polyamine spermidine is required for bacteriophage and eukaryotic virus replication, and herein, we have identified and functionally characterized diverse phage- and virus-encoded polyamine metabolic enzymes and pathways. These include pyridoxal 5′-phosphate (PLP)-dependent ornithine decarboxylase (ODC), pyruvoyl-dependent ODC and arginine decarboxylase (ADC), arginase, S -adenosylmethionine decarboxylase (AdoMetDC/ speD ), spermidine synthase, homospermidine synthase, spermidine N -acetyltransferase, and N -acetylspermidine amidohydrolase. We identified homologs of the spermidine-modified translation factor eIF5a encoded by giant viruses of the Imitervirales . Although AdoMetDC/ speD is prevalent among marine phages, some homologs have lost AdoMetDC activity and have evolved into pyruvoyl-dependent ADC or ODC. The pelagiphages that encode the pyruvoyl-dependent ADCs infect the abundant ocean bacterium Candidatus Pelagibacter ubique , which we have found encodes a PLP-dependent ODC homolog that has evolved into an ADC, indicating that infected cells would contain both PLP- and pyruvoyl-dependent ADCs. Complete or partial spermidine or homospermidine biosynthetic pathways are found encoded in the giant viruses of the Algavirales and Imitervirales , and in addition, some viruses of the Imitervirales can release spermidine from the inactive N -acetylspermidine. In contrast, diverse phages encode spermidine N -acetyltransferase that can sequester spermidine into its inactive N -acetyl form. Together, the virome-encoded enzymes and pathways for biosynthesis and release or biochemical sequestration of spermidine or its structural analog homospermidine consolidate and expand evidence supporting an important and global role of spermidine in virus biology.

New antimicrobials are needed for the treatment of extensively drug-resistant Acinetobacter baumannii . The de novo pyrimidine biosynthetic enzyme dihydroorotate dehydrogenase (DHODH) is a validated drug target for malaria and human autoimmune diseases. We provide genetic evidence that A. baumannii DHODH ( Ab DHODH) is essential for bacterial survival in rodent infection models. We chemically validate the target by repurposing a unique library of ~450 triazolopyrimidine/imidazopyrimidine analogs developed for our malaria DHODH program to identify 21 compounds with submicromolar activity on Ab DHODH. The most potent (DSM186, DHODH IC 50 28 nM) had a minimal inhibitory concentration of ≤1 µg/ml against geographically diverse A. baumannii strains, including meropenem-resistant isolates. A structurally related analog (DSM161) with a long in vivo half-life conferred significant protection in the neutropenic mouse thigh infection model. Encouragingly, the development of resistance to these compounds was not identified in vitro or in vivo. Lastly, the X-ray structure of Ab DHODH bound to DSM186 was solved to 1.4 Å resolution. These data support the potential of Ab DHODH as a drug target for the development of antimicrobials for the treatment of A. baumannii and potentially other high-risk bacterial infections.

Current malaria treatments are threatened by drug resistance and new drugs are urgently needed. In a phenotypic screen for new antimalarials, we identified (S)-SW228703 ((S)-SW703), a tyrosine amide with asexual blood and liver stage activity and a fast-killing profile. Resistance to (S)-SW703 is associated with mutations inPlasmodium falciparumcyclic amine resistance locus (PfCARL) andP. falciparumacetyl CoA transporter (PfACT), similarly to several other compounds that share features such as fast activity and liver-stage activity. Compounds with these resistance mechanisms are thought to act in the ER, though their target(s) are unknown. The tyramine of (S)-SW703 is shared with some reportedPfCARL-associated compounds; however, we observed that strict S-stereochemistry was required for activity of (S)-SW703, suggesting differences in mechanism of action or binding mode. (S)-SW703 provides a new chemical series with broad activity on multiple life-cycle stages and a fast-killing mechanism of action, available for lead optimization to generate new treatments for malaria.

Malaria control programs continue to be threatened by drug resistance. To identify new antimalarials, we conducted a phenotypic screen and identified a novel tetrazole-based series that shows fast-kill kinetics and a relatively low propensity to develop high-level resistance. Preliminary structure-activity relationships were established including identification of a subseries of related amides with antiplasmodial activity. Assaying parasites with resistance to antimalarials led us to test whether the series had a similar mechanism of action to chloroquine (CQ). Treatment of synchronized Plasmodium falciparum parasites with active analogues revealed a pattern of intracellular inhibition of hemozoin (Hz) formation reminiscent of CQ's action. Drug selections yielded only modest resistance that was associated with amplification of the multidrug resistance gene 1 (pfmdr1). Thus, we have identified a novel chemical series that targets the historically druggable heme polymerization pathway and that can form the basis of future optimization efforts to develop a new malaria treatment.

The spiroindimicins are a unique class of chlorinated indole alkaloids characterized by three heteroaromatic rings structured around a congested spirocyclic stereocenter. Here, we report the first total synthesis of (+)-spiroindimicin A, which bears a challenging C-3′/C-5′′-linked spiroindolenine. We detail our initial efforts to effect a biomimetic oxidative spirocyclization from its proposed natural precursor, lynamicin D, and describe how these studies shaped our final abiotic 9-step solution to this complex alkaloid built around a key Pd-catalyzed asymmetric spirocyclization. Scalable access to spiroindimicins A, H, and their congeners has enabled discovery of their activity against several parasites relevant to human health, providing potential starting points for new therapeutics for the neglected tropical diseases leishmaniasis and African sleeping sickness., Spiroindimicins A and H have been synthesized for the first time via a key palladium-catalyzed spirocyclization. Access to these alkaloids and several congeners has allowed the discovery of their antiparasitic properties.

Malaria puts at risk nearly half the world's population and causes high mortality in sub-Saharan Africa, while drug resistance threatens current therapies. The pyrimidine biosynthetic enzyme dihydroorotate dehydrogenase (DHODH) is a validated target for malaria treatment based on our finding that triazolopyrimidine DSM265 (1) showed efficacy in clinical studies. Herein, we describe optimization of a pyrrole-based series identified using a target-based DHODH screen. Compounds with nanomolar potency versus Plasmodium DHODH and Plasmodium parasites were identified with good pharmacological properties. X-ray studies showed that the pyrroles bind an alternative enzyme conformation from 1 leading to improved species selectivity versus mammalian enzymes and equivalent activity on Plasmodium falciparum and Plasmodium vivax DHODH. The best lead DSM502 (37) showed in vivo efficacy at similar levels of blood exposure to 1, although metabolic stability was reduced. Overall, the pyrrole-based DHODH inhibitors provide an attractive alternative scaffold for the development of new antimalarial compounds.

Dihydroorotate dehydrogenase (DHODH) catalyzes a key step in pyrimidine biosynthesis and has recently been validated as a therapeutic target for malaria through clinical studies on the triazolopyrimidine-based Plasmodium DHODH inhibitor DSM265. Selective toxicity towards Plasmodium species could be achieved because malaria parasites lack pyrimidine salvage pathways, and DSM265 selectively inhibits Plasmodium DHODH over the human enzyme. However, while DSM265 does not inhibit human DHODH, it inhibits DHODH from several preclinical species, including mice, suggesting that toxicity could result from on-target DHODH inhibition in those species. We describe here the use of dihydroorotate (DHO) as a biomarker of DHODH inhibition. Treatment of mammalian cells with DSM265 or the mammalian DHODH inhibitor teriflunomide led to increases in DHO where the extent of biomarker buildup correlated with both dose and inhibitor potency on DHODH. Treatment of mice with leflunomide (teriflunomide prodrug) caused a large dose-dependent buildup of DHO in blood (up to 16-fold) and urine (up to 5,400-fold) that was not observed for mice treated with DSM265. Unbound plasma teriflunomide levels reached 20-85-fold above the mouse DHODH ICsub50/sub, while free DSM265 levels were only 1.6-4.2-fold above, barely achieving ∼ ICsub90/subconcentrations, suggesting that unbound DSM265 plasma levels are not sufficient to block the pathway in vivo. Thus, any toxicity associated with DSM265 treatment in mice is likely caused by off-target mechanisms. The identification of a robust biomarker for mammalian DHODH inhibition represents an important advance to generally monitor for on-target effects in preclinical and clinical applications of DHODH inhibitors used to treat human disease.

Polyamines are fundamental molecules of life, and their deep evolutionary history is reflected in extensive biosynthetic diversification. The polyamines putrescine, agmatine, and cadaverine are produced by pyridoxal 5′-phosphate-dependent L-ornithine, L-arginine, and L-lysine decarboxylases (ODC, ADC, LDC), respectively, from both the alanine racemase (AR) and aspartate aminotransferase (AAT) folds. Two homologous forms of AAT-fold decarboxylase are present in bacteria: an ancestral form and a derived, acid-inducible extended form containing an N-terminal fusion to the receiver-like domain of a bacterial response regulator. Only ADC was known from the ancestral form and limited to the Firmicutes phylum, whereas extended forms of ADC, ODC, and LDC are present in Proteobacteria and Firmicutes. Here, we report the discovery of ancestral form ODC, LDC, and bifunctional O/LDC and extend the phylogenetic diversity of functionally characterized ancestral ADC, ODC, and LDC to include phyla Fusobacteria, Caldiserica, Nitrospirae, and Euryarchaeota. Using purified recombinant enzymes, we show that these ancestral forms have a nascent ability to decarboxylate kinetically less preferred amino acid substrates with low efficiency, and that product inhibition primarily affects preferred substrates. We also note a correlation between the presence of ancestral ODC and ornithine/arginine auxotrophy and link this with a known symbiotic dependence on exogenous ornithine produced by species using the arginine deiminase system. Finally, we show that ADC, ODC, and LDC activities emerged independently, in parallel, in the homologous AAT-fold ancestral and extended forms. The emergence of the same ODC, ADC, and LDC activities in the nonhomologous AR-fold suggests that polyamine biosynthesis may be inevitable.

Purpose Evaluate visual outcomes in relation to time from injury to intervention in patients who undergo lateral canthotomy with cantholysis (LCC) for retrobulbar hemorrhage (RBH). Methods Retrospective study of patients with orbital compartment syndrome (OCS) secondary to RBH who underwent LCC. OCS due to RBH was defined by a combination of decreased vision, proptosis, resistance to retropulsion, increased intraocular pressure, and relative afferent pupillary defect. Time from injury to intervention and change in visual acuity were calculated, with regression analysis identifying predictors of vision recovery. Results Fifteen participants were included. Three (20%) participants presented with no light perception, 7 (47%) with count fingers (CF) to light perception, and 5 (33%) with better than count fingers vision. All 5 participants who had LCC within 3 hours (twice the standard 90 minutes) gained some vision, and 6 of 10 participants who had LCC after 3 hours recovered some vision. The latest intervention with visual acuity improvement was performed 9 hours postinjury. Of 3 participants who presented with no light perception vision, 1 regained vision to 20/40 (intervention 1.7 hours postinjury), and 2 did not regain any vision (interventions at 5 and 8.7 hours postinjury). Duration from injury to intervention was associated with decreased amount of vision recovery (P = 0.03). Conclusions Increased time to intervention with LCC was associated with less vision recovery after OCS from RBH. However, over half of participants with intervention more than 90 minutes after injury still showed visual acuity improvement. The authors recommend LCC in all patients who present with OCS regardless of the time since injury.Patients with orbital compartment syndrome may see visual recovery after lateral canthotomy and cantholysis, even if performed outside of the previously accepted 3-hour window.

PURPOSE Define incidence of severe ocular trauma in orbital fracture patients and determine if ocular signs and symptoms are useful predictors of severe ocular injuries. METHODS Retrospective chart review was performed on all patients with orbital fractures between April 1, 2013, and December 31, 2014. Patients were included if they had radiographic evidence of acute fracture of at least one orbital wall and were evaluated by the Ophthalmology service. Demographics, concurrent injury data, and symptoms and signs of ocular trauma were collected. Concurrent ocular injuries were grouped by severity. Predictive signs or symptoms for severe ocular trauma were identified by stepwise logistic regression analysis. The threshold point for predictive signs and symptoms was detected by a receiver operating characteristic (ROC). RESULTS Five-hundred-twelve patients were included. The most common mechanisms of injury were assault (39%), fall (25%), and motor vehicle accident (21%). The incidence of any concurrent ocular trauma was 75% (383/512), with 14% (70/512) being severe. Four signs and symptoms were predictors of severity: blurred vision (P < 0.0001), pain with eye movements (P < 0.0001), visual acuity worse than 20/40 in the ipsilateral eye (P < 0.001), and restricted motility (P < 0.001). The presence of 2 or more of these signs or symptoms was predictive of severe ocular trauma with high sensitivity (91%) and specificity (86%). CONCLUSIONS In cooperative patients with acute orbital wall fractures, the presence of 2 or more signs or symptoms is predictive of severe ocular trauma and necessitates the need for urgent ophthalmic consultation.Severe ocular injury associated with orbital wall fracture is more likely in patients with 2 or more ophthalmic signs or symptoms.

Background Pain is a common stressor for ICU patients, necessitating routine assessment. For patients who are unable to communicate, self-report tools are unsuitable, and the use of an observational tool is required to assess pain appropriately. The Critical Care Pain Observation Tool (CPOT) is the most reliable tool currently available to assess pain in these patients. We investigated whether the implementation of the CPOT in one Australian ICU could increase frequency of appropriate pain assessments, and if this would affect the administration of analgesia and sedation. Methods In this before and after study, we first performed a retrospective chart audit on 441 adult ICU patient charts, over 49 days. Data collected included frequency and type of pain assessments, sedation and analgesia administered, communication and CAM-ICU status, and bedside nurse-perceived pain. During the implementation phase, new policy and guideline documents were released, and ICU charts were redesigned to incorporate the CPOT. All nursing staff attended an education session on pain assessment and correct use of the CPOT. The chart audit was repeated, capturing 344 charts over 43 days. Results Mean total assessments in 24 hours increased from 7.2 to 7.9 for communicative, 3.0 to 8.9 for non-communicative, and 5.1 to 9.1 for transitioning patients. For non-communicative patients there was a significant increase in observational assessments including the CPOT (1.7 to 8.3), and a decrease in inappropriate use of self-report tools (1.3 to 0.2). We also observed significant increases in administration of paracetamol, opiates, propofol, patient-controlled analgesia, modified-release opiates, and neuropathic pain agents. Conclusions Implementation of the CPOT using standardised education and resources led to increased frequency of pain assessment, particularly for non-communicative patients. Appropriate observational assessments were also more frequently used for these patients. Analgesic administration generally increased, as did the use of propofol.

The spiroindimicins are a unique class of chlorinated indole alkaloids characterized by three heteroaromatic rings structured around a congested spirocyclic stereocenter. Here, we report the first total synthesis of (+)-spiroindimicin A, which bears a challenging C-3’/C-5’’-linked spiroindolenine. We detail our initial efforts to effect a biomimetic oxidative spirocyclization from its proposed natural precursor, lynamicin D, and describe how these studies shaped our final abiotic 9-step solution to this complex alkaloid built around a key asymmetric Pd-catalyzed spirocyclization. Scalable access to spiroindimicins A, H, and their congeners has enabled discovery of their activity against several parasites relevant to human health, providing potential starting points for new therapeutics for the neglected tropical diseases leishmaniasis and African sleeping sickness.

Malaria control programs continue to be threatened by drug resistance. To identify new antimalarials we conducted a phenotypic screen and identified a novel tetrazole-based series that shows fast-kill kinetics and a relatively low propensity to develop high-level resistance. Preliminary structure activity relationships (SAR) were established including identification of a sub-series of related amides with antiplasmodial activity. Assaying parasites with resistance to antimalarials led us to test whether the series had a similar mechanism of action to chloroquine (CQ). Treatment of synchronized P. falciparum parasites with active analogs revealed a pattern of intracellular inhibition of hemozoin (Hz) formation reminiscent of CQ’s action. Drug selections yielded only modest resistance that was associated with amplification of the multidrug resistance gene 1 (pfmdr1). Thus, we have identified a novel chemical series that targets the historically druggable heme polymerization pathway, and that can form the basis of future optimization efforts to develop a new malaria treatment.

Dance training may be considered as the interweaving of two critical factors knowledge and experience. Consequently, while dance training in Australia is marked by lineages handed down from European and later, North American forebears, its pattern of forms and forces have been shaped by the experiences of the teachers, choreographers and performers who pursued dance within this continent. It is a story of cultural displacement, exploitation and adaptation where the most stereotypical characteristics distance and its associated state of isolation provide a promising point of departure to examine specific influences of place on the evolution of Australian dance training. Consequently, in a current day mapping of dance training in Australia, the transition from a private to a public system needs to be acknowledged, with private dance schools continuing to maintain a vital role in the pre-tertiary network. Indeed, composed movement is a language, different, but equally expressive as the language of words.

Malaria is one of the most challenging human infectious diseases and both prevention and control have been hindered by the development of Plasmodium falciparum resistance to existing therapies. Several new compounds with novel mechanisms are in clinical development for the treatment of malaria including DSM265, an inhibitor of Plasmodium dihydroorotate dehydrogenase. In order to explore the mechanisms by which resistance might develop to DSM265 in the field, we selected for DSM265-resistant P. falciparum parasites in vitro. Any of five different amino acid changes led to reduced efficacy on the parasite and to decreased DSM265 binding to P. falciparum DHODH. The DSM265-resistant parasites retained full sensitivity to atovaquone. All but one of the observed mutations were in the DSM265 binding site, and the remaining C276F was in the adjacent flavin cofactor site. The C276F mutation was previously identified in a recrudescent parasite during a Phase IIa clinical study. We confirmed that this mutation (and the related C276Y) accounted for the full level of observed DSM265 resistance by re-generating the mutation using CRISPR/Cas9 genome editing. X-ray structure analysis of the C276F mutant enzyme showed that conformational changes of nearby residues were required to accommodate the larger F276 residue, which in turn led to a restriction in the size of the DSM265 binding pocket. These findings underscore the importance of developing DSM265 as part of a combination therapy with other agents for successful use against malaria.

Malaria kills nearly 0.5 million people yearly and impacts the lives of those living in over 90 countries where it is endemic. The current treatment programs are threatened by increasing drug resistance. Dihydroorotate dehydrogenase (DHODH) is now clinically validated as a target for antimalarial drug discovery as a triazolopyrimidine class inhibitor (DSM265) is currently undergoing clinical development. We discovered a related isoxazolopyrimidine series in a phenotypic screen, later determining that it targeted DHODH. To determine if the isoxazolopyrimidines could yield a drug candidate, we initiated hit-to-lead medicinal chemistry. Several potent analogues were identified, including a compound that showed in vivo antimalarial activity. The isoxazolopyrimidines were more rapidly metabolized than their triazolopyrimidine counterparts, and the pharmacokinetic data were not consistent with the goal of a single-dose treatment for malaria.

Objectives Workers who fabricate stone countertops using hand tools are at risk of silicosis from overexposure to respirable crystalline silica. This study explored the efficacy of simple engineering controls that can be used for dust suppression during use of hand tools by stone countertop fabricators. Methods Controlled experiments were conducted to measure whether wet methods and on-tool local exhaust ventilation (LEV) reduced respirable dust (RD) exposures during use of various powered hand tools on quartz-rich engineered stone. RD samples collected during edge grinding with a diamond cup wheel and a silicon carbide abrasive wheel were analyzed gravimetrically as well as by X-ray diffraction to determine silica content. A personal optical aerosol monitor was used simultaneously with the RD samples and also for rapid assessment of controls for polishing, blade cutting, and core drilling. Results On-tool LEV and sheet-flow-wetting were effective in reducing exposures, especially when used in combination. Sheet-flow-wetting with LEV reduced geometric mean exposures by as much as 95%. However, typical water-spray-wetting on a grinding cup was less effective when combined with LEV than without LEV. Mean silica content of RD samples from grinding operations was 53%, and respirable mass and silica mass were very highly correlated (r = 0.980). Optical concentration measures were moderately well correlated with gravimetric measures (r = 0.817), but on average the optical measures during a single trial using the factory calibration were only one-fifth the simultaneous gravimetric measures. Conclusions Sheet-flow-wetting combined with on-tool LEV is an effective engineering control for reducing RD exposures during engineered stone edge grinding and blade cutting. On the other hand, addition of LEV to some water-spray-wetted tools may reduce the effectiveness of the wet method.

The management of blind, painful eyes in Sturge–Weber syndrome patients poses unique challenges to the oculoplastic surgeon. Intraocular and orbital vascular malformations and calcification may theoretically lead to unexpected hemorrhage and difficulty placing an implant in a calcified scleral shell. We present two cases of patients with Sturge–Weber syndrome with blind, painful eyes who underwent evisceration with silicone implant and discuss the relevant current literature. Both of our patients had uncomplicated surgeries and post-operative courses. Our literature review reveals that both evisceration and enucleation are viable surgical options for globe removal in Sturge–Weber syndrome, yet careful preoperative planning must be undertaken to minimize risk.

Summary Background DSM265 is a novel antimalarial that inhibits plasmodial dihydroorotate dehydrogenase, an enzyme essential for pyrimidine biosynthesis. We investigated the safety, tolerability, and pharmacokinetics of DSM265, and tested its antimalarial activity. Methods Healthy participants aged 18–55 years were enrolled in a two-part study: part 1, a single ascending dose (25–1200 mg), double-blind, randomised, placebo-controlled study, and part 2, an open-label, randomised, active-comparator controlled study, in which participants were inoculated with Plasmodium falciparum induced blood-stage malaria (IBSM) and treated with DSM265 (150 mg) or mefloquine (10 mg/kg). Primary endpoints were DSM265 safety, tolerability, and pharmacokinetics. Randomisation lists were created using a validated, automated system. Both parts were registered with the Australian New Zealand Clinical Trials Registry, number ACTRN12613000522718 (part 1) and number ACTRN12613000527763 (part 2). Findings In part 1, 73 participants were enrolled between April 12, 2013, and July 14, 2015 (DSM265, n=55; placebo, n=18). In part 2, nine participants were enrolled between Sept 30 and Nov 25, 2013 (150 mg DSM265, n=7; 10 mg/kg mefloquine, n=2). In part 1, 117 adverse events were reported; no drug-related serious or severe events were reported. The most common drug-related adverse event was headache. The mean DSM265 peak plasma concentration (Cmax) ranged between 1310 ng/mL and 34 800 ng/mL and was reached in a median time (tmax) between 1·5 h and 4 h, with a mean elimination half-life between 86 h and 118 h. In part 2, the log10 parasite reduction ratio at 48 h in the DSM265 (150 mg) group was 1·55 (95% CI 1·42–1·67) and in the mefloquine (10 mg/kg) group was 2·34 (2·17–2·52), corresponding to a parasite clearance half-life of 9·4 h (8·7–10·2) and 6·2 h (5·7–6·7), respectively. The median minimum inhibitory concentration of DSM265 in blood was estimated as 1040 ng/mL (range 552–1500), resulting in a predicted single efficacious dose of 340 mg. Parasite clearance was significantly faster in participants who received mefloquine than in participants who received DSM265 (p

Purpose: To describe a case of orbital apex syndrome as a result of isolated bacterial sinusitis. Observations: A 63-year-old woman presented with an orbital apex syndrome from isolated bacterial sinusitis with rapidly declining visual acuity to no light perception. We compared our case with 6 similar cases of severe vision loss from isolated bacterial sinusitis. In contrast to previously published cases, our patient presented with good vision yet deteriorated to no light perception despite appropriate treatment. Conclusions and importance: Orbital apex syndrome can present as a constellation of cranial neuropathies including optic neuropathy from conditions affecting the orbital apex. Although vision loss remained permanent, prompt initiation of broad-spectrum antibiotics and antifungals and surgical intervention prevented further extension of infection into intracranial structures. Keywords: Orbital apex syndrome, Optic neuropathy

Supplemental Digital Content is available in the text., Summary: The purpose of this article is to present a modified approach to the reconstruction of the upper and lower eyelids in floppy eyelid syndrome. A retrospective chart review was performed on all floppy eyelid patients who underwent simultaneous tightening of the upper and lower eyelid with a lateral tarsal strip, using a V-shaped incision in the lateral canthus, at University of Tennessee Hamilton Eye Institute from 2011 to 2012. Preoperative symptoms, surgical outcomes, complication rates, and postoperative symptoms were recorded. Nine eyes of 7 patients who underwent surgical correction for symptomatic floppy eyelids were included. All patients noted improvement in symptoms postoperatively, after reduction in the laxity of the upper and lower eyelid. Postoperative complications included buried lashes in the lateral canthus in 1 eye and a pyogenic granuloma in the lateral canthus of 1 eye. An excellent cosmetic outcome was noted in 78% (7/9) of eyes. No patients reported dissatisfaction nor required secondary surgical correction. The lateral canthal “V” incision provides an additional approach in the successful management of floppy eyelid syndrome involving the upper and lower eyelids. The design of the incision allows for excellent exposure of the lateral canthus for shortening of the eyelids with tarsal strip fixation, and it preserves the lateral canthal skin and canthus architecture. Further, the “V” incision is easily continued into the eyelid crease for blepharoplasty and ptosis repair when necessary.

The siderophore rhizoferrin (N1,N4-dicitrylputrescine) is produced in fungi and bacteria to scavenge iron. Putrescine-producing bacterium Ralstonia pickettii synthesizes rhizoferrin and encodes a single nonribosomal peptide synthetase-independent siderophore (NIS) synthetase. From biosynthetic logic, we hypothesized that this single enzyme is sufficient for rhizoferrin biosynthesis. We confirmed this by expression of R. pickettii NIS synthetase in Escherichia coli, resulting in rhizoferrin production. This was further confirmed in vitro using the recombinant NIS synthetase, synthesizing rhizoferrin from putrescine and citrate. Heterologous expression of homologous lbtA from Legionella pneumophila, required for rhizoferrin biosynthesis in that species, produced siderophore activity in E. coli. Rhizoferrin is also synthesized by Francisella tularensis and Francisella novicida, but unlike R. pickettii or L. pneumophila, Francisella species lack putrescine biosynthetic pathways because of genomic decay. Francisella encodes a NIS synthetase FslA/FigA and an ornithine decarboxylase homolog FslC/FigC, required for rhizoferrin biosynthesis. Ornithine decarboxylase produces putrescine from ornithine, but we show here in vitro that FigA synthesizes N-citrylornithine, and FigC is an N-citrylornithine decarboxylase that together synthesize rhizoferrin without using putrescine. We co-expressed F. novicida figA and figC in E. coli and produced rhizoferrin. A 2.1 Å X-ray crystal structure of the FigC N-citrylornithine decarboxylase reveals how the larger substrate is accommodated and how active site residues have changed to recognize N-citrylornithine. FigC belongs to a new subfamily of alanine racemase-fold PLP-dependent decarboxylases that are not involved in polyamine biosynthesis. These data reveal a natural product biosynthetic workaround that evolved to bypass a missing precursor and re-establish it in the final structure.

The emergence of drug-resistant malaria parasites continues to hamper efforts to control this lethal disease. Dihydroorotate dehydrogenase has recently been validated as a new target for the treatment of malaria, and a selective inhibitor (DSM265) of the Plasmodium enzyme is currently in clinical development. With the goal of identifying a backup compound to DSM265, we explored replacement of the SF5-aniline moiety of DSM265 with a series of CF3-pyridinyls while maintaining the core triazolopyrimidine scaffold. This effort led to the identification of DSM421, which has improved solubility, lower intrinsic clearance, and increased plasma exposure after oral dosing compared to DSM265, while maintaining a long predicted human half-life. Its improved physical and chemical properties will allow it to be formulated more readily than DSM265. DSM421 showed excellent efficacy in the SCID mouse model of P. falciparum malaria that supports the prediction of a low human dose (