A metagenomic analysis for combination therapy of multiple classes of antibiotics on the prevention of the spread of antibiotic-resistant genes.

Antibiotics used systemically to treat infections may have off-target effects on the gut microbiome, potentially resulting in the emergence of drug-resistant bacteria or selection of pathogenic species. These organisms may present a risk to the host and spread to the environment with a risk of transmission in the community. To investigate the risk of emergent antibiotic resistance in the gut microbiome following systemic treatment with antibiotics, this metagenomic analysis project used next-generation sequencing, a custom-built metagenomics pipeline, and differential abundance analysis to study the effect of antibiotics (ampicillin, ciprofloxacin, and fosfomycin) in monotherapy and different combinations at high and low doses, to determine the effect on resistome and taxonomic composition in the gut of Balb/c mice. The results showed that low-dose monotherapy treatments showed little change in microbiome composition but did show an increase in expression of many antibiotic-resistant genes (ARGs) posttreatment. Dual combination treatments allowed the emergence of some conditionally pathogenic bacteria and some increase in the abundance of ARGs despite a general decrease in microbiota diversity. Triple combination treatment was the most successful in inhibiting emergence of relevant opportunistic pathogens and completely suppressed all ARGs after 72 h of treatment. The relative abundances of mobile genetic elements that can enhance transmission of antibiotic resistance either decreased or remained the same for combination therapy while increasing for low-dose monotherapy. Combination therapy prevented the emergence of ARGs and decreased bacterial diversity, while low-dose monotherapy treatment increased ARGs and did not greatly change bacterial diversity.

Scientific and Regulatory Policy Committee Points-to-consider Paper*: Drug-induced Vascular Injury Associated with Nonsmall Molecule Therapeutics in Preclinical Development: Part 2. Antisense Oligonucleotides.

Drug-induced vascular injury (DIVI) is a recurrent challenge in the development of novel pharmaceutical agents. In recent years, DIVI has been occasionally observed in nonhuman primates given RNA-targeting therapeutics such as antisense oligonucleotide therapies (ASOs) during chronic toxicity studies. While DIVI in laboratory animal species has been well characterized for vasoactive small molecules, and immune-mediated responses against large molecule biotherapeutics have been well described, there is little published information regarding DIVI induced by ASOs to date. Preclinical DIVI findings in monkeys have caused considerable delays in development of promising new ASO therapies, because of the uncertainty about whether DIVI in preclinical studies is predictive of effects in humans, and the lack of robust biomarkers of DIVI. This review of DIVI discusses clinical and microscopic features of vasculitis in monkeys, their pathogenic mechanisms, and points to consider for the toxicologist and pathologist when confronted with ASO-related DIVI. Relevant examples of regulatory feedback are included to provide insight into risk assessment of ASO therapies.

Scientific and Regulatory Policy Committee Points-to-consider Paper*: Drug-induced Vascular Injury Associated with Nonsmall Molecule Therapeutics in Preclinical Development: Part I. Biotherapeutics.

Drug-induced vascular injury (DIVI) is a recurrent challenge in the development of novel pharmaceutical agents. Although DIVI in laboratory animal species has been well characterized for vasoactive small molecules, there is little available information regarding DIVI associated with biotherapeutics such as peptides/proteins or antibodies. Because of the uncertainty about whether DIVI in preclinical studies is predictive of effects in humans and the lack of robust biomarkers of DIVI, preclinical DIVI findings can cause considerable delays in or even halt development of promising new drugs. This review discusses standard terminology, characteristics, and mechanisms of DIVI associated with biotherapeutics. Guidance and points to consider for the toxicologist and pathologist facing preclinical cases of biotherapeutic-related DIVI are outlined, and examples of regulatory feedback for each of the mechanistic types of DIVI are included to provide insight into risk assessment.

Nonclinical safety biomarkers of drug-induced vascular injury: current status and blueprint for the future.

Better biomarkers are needed to identify, characterize, and/or monitor drug-induced vascular injury (DIVI) in nonclinical species and patients. The Predictive Safety Testing Consortium (PSTC), a precompetitive collaboration of pharmaceutical companies and the U.S. Food and Drug Administration (FDA), formed the Vascular Injury Working Group (VIWG) to develop and qualify translatable biomarkers of DIVI. The VIWG focused its research on acute DIVI because early detection for clinical and nonclinical safety monitoring is desirable. The VIWG developed a strategy based on the premise that biomarkers of DIVI in rat would be translatable to humans due to the morphologic similarity of vascular injury between species regardless of mechanism. The histomorphologic lexicon for DIVI in rat defines degenerative and adaptive findings of the vascular endothelium and smooth muscles, and characterizes inflammatory components. We describe the mechanisms of these changes and their associations with candidate biomarkers for which advanced analytical method validation was completed. Further development is recommended for circulating microRNAs, endothelial microparticles, and imaging techniques. Recommendations for sample collection and processing, analytical methods, and confirmation of target localization using immunohistochemistry and in situ hybridization are described. The methods described are anticipated to aid in the identification and qualification of translational biomarkers for DIVI.

High-resolution imaging spectrometer for recording absolutely calibrated far ultraviolet spectra from laser-produced plasmas.

An imaging spectrometer was designed and fabricated for recording far ultraviolet spectra from laser-produced plasmas with wavelengths as short as 155 nm. The spectrometer implements a Cassegrain telescope and two gratings in a tandem Wadsworth optical configuration that provides diffraction limited resolution. Spectral images were recorded from plasmas produced by the irradiation of various target materials by intense KrF laser radiation with 248 nm wavelength. Two pairs of high-resolution gratings can be selected for the coverage of two wavebands, one grating pair with 1800 grooves/mm and covering approximately 155-175 nm and another grating pair with 1200 grooves/mm covering 230-260 nm. The latter waveband includes the 248 nm KrF laser wavelength, and the former waveband includes the wavelength of the two-plasmon decay instability at 23 the KrF laser wavelength (165 nm). The detection media consist of a complementary metal oxide semiconductor imager, photostimulable phosphor image plates, and a linear array of 1 mm(2) square silicon photodiodes with 0.4 ns rise time. The telescope mirrors, spectrometer gratings, and 1 mm(2) photodiode were calibrated using synchrotron radiation, and this enables the measurement of the absolute emission from the laser-produced plasmas with temporal, spatial, and spectral resolutions. The spectrometer is capable of measuring absolute spectral emissions at 165 nm wavelength as small as 5x10(-7) J/nm from a plasma source area of 0.37 mm(2) and with 0.4 ns time resolution.