Notes

Stereotactic body ablative radiotherapy for reirradiation involving small quantity head and neck types of cancer is owned by extended success: A large, single-institution, contemporary cohort research.
CD19 is among the most relevant targets in cancer immunotherapy. Trametinib molecular weight However, its extracellular domain (ECD) is prone to aggregation and misfolding, representing a major obstacle for the development and analysis of CD19-targeted therapeutics. Here, we engineered stabilized CD19-ECD (termed SuperFolder) variants, which also showed improved expression rates and, in contrast to the wild type protein, they could be efficiently purified in their monomeric forms. Despite being considerably more stable, these engineered mutants largely preserved the wild type sequence (>98.8%). We demonstrate that the variant SF05 enabled the determination of the monovalent affinity between CD19 and a clinically approved FMC63-based CAR, as well as monitoring and phenotypic characterization of CD19-directed CAR-T cells in the blood of lymphoma patients. We anticipate that the SuperFolder mutants generated in this study will be highly valuable tools for a range of applications in basic immunology and CD19-targeted cancer immunotherapy.Optogenetic protein dimerization systems are powerful tools to investigate the biochemical networks that cells use to make decisions and coordinate their activities. These tools, including the improved Light-Inducible Dimer (iLID) system, offer the ability to selectively recruit components to subcellular locations, such as micron-scale regions of the plasma membrane. In this way, the role of individual proteins within signaling networks can be examined with high spatiotemporal resolution. Currently, consistent recruitment is limited by heterogeneous optogenetic component expression, and spatial precision is diminished by protein diffusion, especially over long time scales. Here, we address these challenges within the iLID system with alternative membrane anchoring domains and fusion configurations. Using live cell imaging and mathematical modeling, we demonstrate that the anchoring strategy affects both component expression and diffusion, which in turn impact recruitment strength, kinetics, and spatial dynamics. Compared to the commonly used C-terminal iLID fusion, fusion proteins with large N-terminal anchors show stronger local recruitment, slower diffusion of recruited components, efficient recruitment over wider gene expression ranges, and improved spatial control over signaling outputs. We also define guidelines for component expression regimes for optimal recruitment for both cell-wide and subcellular recruitment strategies. Our findings highlight key sources of imprecision within light-inducible dimer systems and provide tools that allow greater control of subcellular protein localization across diverse cell biological applications.We report a spectrometer employing optically switched dual-wavelength cavity ring-down spectroscopy (OSDW-CRDS) for high-precision measurements of methane isotope ratios. A waveguide optical switch rapidly alternated between two wavelengths to detect absorption by two isotopologues using near-infrared CRDS. This approach alleviated common-mode noise that originated primarily from temperature and frequency fluctuations. We demonstrated the measurement of δD in natural abundance methane to a precision of 2.3 ‰, despite the lack of active temperature or frequency stabilization of the cavity. The ability of alternating OSDW-CRDS to improve the isotope precision in the absence of cavity stabilization were measured by comparing the Allan deviation with that obtained when frequency-stabilizing the cavity length. The system can be extended to a wide variety of applications such as isotope analysis of other species, kinetic isotope effects, ortho-para ratio measurements, and isomer abundance measurements. Furthermore, our technique can be extended to multiple isotope analysis or two species involved in kinetics studies through the use of multiport or high-speed optical switches, respectively.The contribution of the gut microbiome to human health has long been established, with normal gut microbiota conferring protection against invasive pathogens. Antibiotics can disrupt the microbial balance of the gut, resulting in disease and the development of antimicrobial resistance. The effect of antibiotic administration route on gut dysbiosis remains under-studied to date, with conflicting evidence on the differential effects of oral and parenteral delivery. We have profiled the rat gut microbiome following treatment with commonly prescribed antibiotics (amoxicillin and levofloxacin), via either oral or intravenous administration. Fecal pellets were collected over a 13-day period and bacterial populations were analyzed by 16S rRNA gene sequencing. Significant dysbiosis was observed in all treatment groups, regardless of administration route. More profound dysbiotic effects were observed following amoxicillin treatment than those with levofloxacin, with population richness and diversity significantly reduced, regardless of delivery route. The effect on specific taxonomic groups was assessed, revealing significant disruption following treatment with both antibiotics. Enrichment of a number of groups containing known gut pathogens was observed, in particular, with amoxicillin, such as the family Enterobacteriaceae. Depletion of other commensal groups was also observed. The degree of dysbiosis was significantly reduced toward the end of the sampling period, as bacterial populations began to return to pretreatment composition. Richness and diversity levels appeared to return to pretreatment levels more quickly in intravenous groups, suggesting convenient parenteral delivery systems may have a role to play in reducing longer term gut dysbiosis in the treatment of infection.Saccharomyces boulardii is a probiotic yeast that exhibits rapid growth at 37 °C, is easy to transform, and can produce therapeutic proteins in the gut. To establish its ability to produce small molecules encoded by multigene pathways, we measured the amount and variance in protein expression enabled by promoters, terminators, selective markers, and copy number control elements. We next demonstrated efficient (>95%) CRISPR-mediated genome editing in this strain, allowing us to probe engineered gene expression across different genomic sites. We leveraged these strategies to assemble pathways enabling a wide range of vitamin precursor (β-carotene) and drug (violacein) titers. We found that S. boulardii colonizes germ-free mice stably for over 30 days and competes for niche space with commensal microbes, exhibiting short (1-2 day) gut residence times in conventional and antibiotic-treated mice. Using these tools, we enabled β-carotene synthesis (194 μg total) in the germ-free mouse gut over 14 days, estimating that the total mass of additional β-carotene recovered in feces was 56-fold higher than the β-carotene present in the initial probiotic dose.
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