Spotlight pubs.acs.org/crt
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USING INTRACELLULAR DRUG BIOAVAILABILITY TO PREDICT CELLULAR POTENCY It is important to assess the amount of compound available at intracellular drug targets. A team led by Per Artursson developed a high-throughput, label-free, cell-based method to directly measure available drug concentrations inside the cell ((2017) PNAS, 114, E6231−E6239). This technique quantifies the fraction of intracellular unbound drug which is bioavailable for target interactions. The authors explored the potency of 35 mitogen-activated protein kinase 14 (MAPK14) inhibitors. They found that the potency of the compounds in cellular assays declined, on average, by 1 order of magnitude from their potency in biochemical assays, where the molecules were able to directly access and interact with their targets. To determine if drug exposure to the targets was limited in cellular assays, the researchers calculated the intracellular bioavailability (Fic) of the inhibitors by measuring cellular accumulation of the compounds and the intracellular fraction of unbound compounds. Most of the MAPK14 compounds displayed low Fic values, in keeping with the observed decrease in cellular potency. The scientists then used the Fic along with the biochemical pIC50 to predict the cellular pIC50, and their forecasts correlated well with the measured cellular potencies. While this method gives information on whole cell exposure, it can be used in conjunction with mathematical modeling to assess compound accumulation in subcellular compartments. Abigail Druck Shudofsky
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ANTIBIOTIC RADIOTRACER TO IMAGE AND DIAGNOSE LIVE BACTERIAL INFECTIONS
in the presence of bacteria, clearly differentiating bacterial infection from cancer and inflammation. Nonhuman primate experiments were performed for biodistribution assessment, and forthcoming clinical trials will assess the success of [18F]FPTMP in identifying bacterial infection in patients. Abigail Druck Shudofsky
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NITRIC OXIDE-RELEASING FUROXANS INDUCE PSEUDOMONAS AERUGINOSA BIOFILM DISPERSAL
Courtesy of Dr. Mark A. Sellmyer.
Clinical exams and imaging techniques are often not able to distinguish bacterial infections from other pathologies with overlapping presentations, such as cancer and inflammation. A broad spectrum bacterial-specific imaging agent would improve diagnosis. A team led by Mark A. Sellmyer and Robert H. Mach created a small molecule positron emission tomography (PET) probe based on the synthetic antibiotic trimethoprim (TMP). The fluorine-18-labeled derivative combines bacterial specificity with PET sensitivity and spatial resolution ((2017) PNAS, 114, 8372−8377). TMP inhibits dihydrofolate reductase (dhfr), an enzyme conserved across Gram-positive and Gram-negative bacteria, along with some parasitic and mycobacterial species. It is active in nanomolar concentrations, with broad tissue distribution and a short blood half-life in humans. The authors synthesized a TMP-based fluoropropyl derivative radiotracer, [18F]fluoropropyltrimethoprim ([18F]FPTMP), and assessed it in vitro in different bacterial species for dhfr affinity and cell uptake. The probe showed rapid uptake which resulted in >100-fold signal increases in three different bacterial species. When tested in a rodent model, uptake of [18F]FPTMP in live bacteria was increased nearly 3-fold compared to that in heat-killed bacteria. The scientists then tested the imaging specificity of [18F]FPTMP by implanting mouse mammary carcinoma cells, turpentine (which induces chemical inflammation), or live E. coli in different parts of mice. While a standard PET radiotracer showed uptake in all pathologies, the only significant [18F]FPTMP accumulation occurred © 2017 American Chemical Society
Courtesy of Dr. Scott Rice.
Free radical nitric oxide (NO) can induce bacterial biofilms to disperse into single cells, increasing their susceptibility to conventional antibiotics. Heterocyclic furoxans are soluble NO donors that contain a 1,2,5-oxadiazole 2-oxide ring with two substituent groups at positions 3 and 4. These substituent groups determine compound solubility and modulate the kinetics of NO production and release. A group of collaborators led by Scott A. Rice studied the effect of three furoxans (LL4212, LL4216, and LL4254) on Pseudomonas aeruginosa biofilms ((2017) ACS Chem. Biol., 12, 2097−2106). After determining that LL4212, LL4216, and LL4254 spontaneously release nitrogen oxide species under physiologically relevant conditions, the authors investigated whether the compounds induce biofilm dispersal. They found that treating P. aeruginosa biofilms for 1 h with 200 μM LL4254 reduced biofilm biomass by >70%. Cotreatment of biofilms with LL4254 and the NO scavenger cPTIO reduced the dispersal level by Published: September 18, 2017 1653
DOI: 10.1021/acs.chemrestox.7b00236 Chem. Res. Toxicol. 2017, 30, 1653−1654
Chemical Research in Toxicology
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∼55%, suggesting that the effect of LL4254 is NO-dependent. As LL4212 and LL4216 have slower rates of NO release compared to LL4254, the authors treated biofilms with compound concentrations that correspond to the amount of nitrogen oxide species released in 1 h from ∼200−400 μM LL4254. 3.6 mM of LL4212 and LL4216 dispersed 48 and 70% of the biofilm biomass, respectively. Additionally, LL4212 and LL4216 prevented free-floating bacteria from forming biofilms; when the furoxans were added to medium at the time of P. aeruginosa inoculation, there were significant concentration-dependent reductions in biofilm biomass after 6 h compared to that in untreated controls. cPTIO did not inhibit the effects of LL4212 and LL4216 on biofilm dispersal, suggesting that their functional activities were NO-independent. Finally, a marker of biofilm dispersal is decreased production of the iron-chelating siderophore pyoverdine. LL4212, LL4216, and LL4254 reduced the level of P. aeruginosa pyoverdine synthesis, further supporting the idea that furoxans can be used to promote biofilm dispersal by NO-dependent or NO-independent mechanisms. Abigail Druck Shudofsky
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Spotlight
ORGANOMETALLIC OXA DERIVATIVES HAVE POTENT CROSS-SPECIES ACTIVITY AGAINST SCHISTOSOMA MANSONI AND S. HAEMATOBIUM
Courtesy of Dr. Jeannine Hess.
The tropical disease schistosomiasis is caused by Schistosoma helminths and affects more than 250 million people annually. The drug oxamniquine (OXA) is only effective against one of six Schistosoma species, namely, S. mansoni. Researchers led by Jennifer Keiser and Gilles Gasser attached organometallic moieties onto OXA to create derivatives effective against different Schistosoma species ((2017) ACS Infect. Dis., DOI: 10.1021/acsinfecdis.7b00054). The authors N-alkylated the exocyclic amine of OXA with a ferrocenyl unit (1) which they then tested in vitro on larval and adult stages of S. mansoni. S. mansoni were incubated with 100 μM of either OXA or 1 for 72 h. The compounds reduced larval worm viability by 67% and 100%, respectively. Furthermore, while OXA had no significant effect on adult worm viability, 1 killed all adult worms after only 24 h. When used in an S. mansoni−mouse infection model, both OXA and 1 (200 mg/kg, orally) eliminated all worms without causing any toxicity. The scientists synthesized two analogues of 1, a ruthenocenyl (5) and a benzyl derivative (6). All three compounds displayed similar IC50 values against adult S. mansoni worms in vitro. When tested in vivo (100 mg/kg, orally), 1, 5, and 6 displayed no toxicity and exhibited worm burden reductions of 81%, 93.7%, and 76.1%, respectively, compared to OXA at 99%. Lastly, the researchers treated adult S. haematobium worms in vitro with 100 μM of OXA or a derivative. While OXA treatment had no effect, 1, 5, and 6 showed excellent in vitro activity against S. haematobium. As S. haematobium is the most prevalent Schistosoma species, the antischistosomal potency of these derivatives against that species is promising and suggests their potential for application against all Schistosoma species. Abigail Druck Shudofsky
CLIMATE VARIABILITY INFLUENCES RODENT POPULATION DYNAMICS AND HANTAVIRUS TRANSMISSION
Hantavirus is a rodent-borne zoonotic pathogen which causes ∼200,000 disease cases annually. Human infection occurs through exposure to aerosolized virus-containing rodent waste. As the reasons for interannual variability in hantavirus transmission are unclear, Nils Chr. Stenseth and Huaiyu Tian led a group which used model fitting and simulation to determine the role of climate in hantavirus disease dynamics ((2017) PNAS, 114, 8041−8046). The authors used rodent field surveillance data gathered between 1960−2013 from the Weihe Plain in Central China in combination with contemporaneous epidemiological records of hantavirus disease in that area and local daily climate reports. They found that rodent population density and human disease incidence fluctuated with climate variables, specifically temperature and rainfall. The team found that hantavirus outbreaks followed an 8 y interannual cycle, likely due to the El Niñ o−Southern Oscillation. Climate also provided a seasonal effect; modeling results indicated that winter temperatures and summer rainfall levels could positively or negatively affect rodent abundance, directly impacting hantavirus disease epidemics. The researchers developed a climate-driven model for hantaviruses transmission dynamics which considers direct and indirect factors affecting the animal host reservoir and quantifies their role in disease transmission. Using this model, they were able to causally predict hantavirus outbreaks. Abigail Druck Shudofsky 1654
DOI: 10.1021/acs.chemrestox.7b00236 Chem. Res. Toxicol. 2017, 30, 1653−1654
