Abstract
The idealized supercell simulations in a previous study by Roberts et al. are further analyzed to clarify the physical mechanisms leading to differences in mesocyclone intensification ...between an experiment with surface friction applied to the full wind (FWFRIC) and an experiment with friction applied to the environmental wind only (EnvFRIC). The low-level mesocyclone intensifies rapidly during the 3 min preceding tornadogenesis in FWFRIC, while the intensification during the same period is much weaker in EnvFRIC, which fails to produce a tornado. To quantify the mechanisms responsible for this discrepancy in mesocyclone evolution, material circuits enclosing the low-level mesocyclone are initialized and traced back in time, and circulation budgets for these circuits are analyzed. The results show that in FWFRIC, surface drag directly generates a substantial proportion of the final circulation around the mesocyclone, especially below 1 km AGL; in EnvFRIC, circulation budgets indicate the mesocyclone circulation is overwhelmingly barotropic. It is proposed that the import of near-ground, frictionally generated vorticity into the low-level mesocyclone in FWFRIC is a key factor causing the intensification and lowering of the mesocyclone toward the ground, creating a large upward vertical pressure gradient force that leads to tornadogenesis. Similar circulation analyses are also performed for circuits enclosing the tornado at its genesis stage. The frictionally generated circulation component is found to contribute more than half of the final circulation for circuits enclosing the tornado vortex below 400 m AGL, and the frictional contribution decreases monotonically with the height of the final circuit.
Glutarimides such as thalidomide, pomalidomide, and lenalidomide are the most frequently used ligands to recruit E3 ubiquitin ligase cereblon (CRBN) for the development of proteolysis-targeting ...chimeras (PROTACs). Due to the rapid and spontaneous racemization of glutarimides, most CRBN-recruiting PROTACs are synthesized as a mixture of racemates or diastereomers. Since the (S)-enantiomer is primarily responsible for binding to CRBN, the existence of the largely inactive (R)-enantiomer complicates the drug development process. Herein, we report that substituted achiral phenyl dihydrouracil (PDHU) can be used as a novel class of CRBN ligands for the development of PROTACs. Although the parent PDHU has a minimal binding affinity to CRBN, we found that some substituted PDHUs had a comparable binding affinity to lenalidomide. Structural modeling provided a further understanding of the molecular interactions between PDHU ligands and CRBN. PDHUs also have greater stability than lenalidomide. Finally, potent BRD4 degraders were developed by employing trisubstituted PDHUs.
RNA plays a myriad of roles in the body including the coding, decoding, regulation, and expression of genes. RNA oligonucleotides have garnered significant interest as therapeutics via antisense ...oligonucleotides or small interfering RNA strategies for the treatment of diseases ranging from hyperlipidemia, HCV, and others. Additionally, the recently developed CRISPR-Cas9 mediated gene editing strategy also relies on Cas9-associated RNA strands. However, RNA presents numerous challenges as both a synthetic target and a potential therapeutic. RNA is inherently unstable, difficult to deliver into cells, and potentially immunogenic by itself or upon modification. Despite these challenges, with the help of chemically modified oligonucleotides, multiple RNA-based drugs have been approved by the FDA. The progress is made possible due to the nature of chemically modified oligonucleotides bearing advantages of nuclease stability, stronger binding affinity, and some other unique properties. This review will focus on the chemical synthesis of RNA and its modified versions. How chemical modifications of the ribose units and of the phosphatediester backbone address the inherent issues with using native RNA for biological applications will be discussed along the way.
Proteolysis targeting chimeras (PROTACs) have emerged as useful chemical probes and potential therapeutics by taking advantage of the ubiquitin-proteasome system to degrade intracellular ...disease-associated proteins. PROTACs are heterobifunctional molecules composed of a target protein ligand, E3 ubiquitin ligase ligand, and a linker between them. The generation of efficient PROTACs requires screening of many parameters, especially the lengths and types of the linkers. We report our proof-of-concept study using a two-stage strategy to facilitate the development of PROTACs against the estrogen receptor (ER). In stage one, a library of close to 100 PROTACs was synthesized by simply mixing a library of ERα ligands containing a hydrazide functional group at different positions with a preassembled library of E3 ligase ligands bearing different types and lengths of linkers with a terminal aldehyde group in a 1:1 ratio. Cell-based screening occurred without further purification, because the formation of the acylhydrazone linkage is highly efficient and produces water as the only byproduct. Compound A3 was the most potent ER degrader in two ER+ cell lines (DC
= ∼ 10 nM,
= ≥ 95%). Stage two involved transformation to a more stable amide linker to generate a more drug-like molecule. The new compound, AM-A3, showed comparable biological activity (DC
= 1.1 nM,
= 98%) and induced potent antiproliferation (IC
= 13.2 nM,
= 69%) in MCF-7. This proof-of -concept study demonstrates that the two-stage strategy can significantly facilitate the development of PROTACs against ER without the tedious process of making large numbers of PROTACs one by one. It has the potential to be expanded to many other targets.
Continuous and batch microwave reactors were constructed for efficient, “green” synthesis with low-boiling solvents at high temperature in closed vessels. Capabilities for rapid heating and cooling, ...concurrent heating and cooling, and differential heating facilitated novel chemical reactions and processes. Commercial microwave systems based on these developments are available. Times required for conventional reactions typically are decreased by 2−3 orders of magnitude. Green processes also have resulted through use of less or no catalyst, readily recyclable solvents, or media and yields that are often higher than normal. Complementary interactive software for calculating optimal conditions was developed.
To investigate the effect of surface drag on tornadogenesis, a pair of idealized simulations is conducted with 50-m horizontal grid spacing. In the first experiment (full-wind drag case), surface ...drag is applied to the full wind; in the second experiment (environmental drag case), drag is applied only to the background environmental wind, with storm-induced perturbations unaffected. The simulations are initialized using a thermal bubble within a horizontally homogeneous background environment that has reached a balance between the pressure gradient, Coriolis, and frictional forces. The environmental sounding is derived from a prior simulation of the 3 May 1999 Oklahoma tornado outbreak but modified to account for near-ground frictional effects. In the full-wind drag experiment, a tornado develops around 25 min into the simulation and persists for more than 10 min; in the environmental-only drag experiment, no tornado occurs. Three distinct mechanisms are identified by which surface drag influences tornadogenesis. The first mechanism is the creation by drag of near-ground vertical wind shear (and associated horizontal vorticity) in the background environment. The second mechanism is generation of near-ground crosswise horizontal vorticity by drag on the storm scale as air accelerates into the low-level mesocyclone; this vorticity is subsequently exchanged into the streamwise direction and eventually tilted into the vertical. The third mechanism is frictional enhancement of horizontal convergence, which strengthens the low-level updraft and stretching of vertical vorticity. The second and third mechanisms are found to work together to produce a tornado, while baroclinic vorticity plays a negligible role.
Breast cancer is the second most common cancer diagnosed in women in the US with almost 280,000 new cases anticipated in 2023. Currently, on-site pathology for location guidance is not available ...during the collection of breast biopsies or during surgical intervention procedures. This shortcoming contributes to repeat biopsy and re-excision procedures, increasing the cost and patient discomfort during the cancer management process. Both procedures could benefit from on-site feedback, but current clinical on-site evaluation techniques are not commonly used on breast tissue because they are destructive and inaccurate. Ex-vivo microscopy is an emerging field aimed at creating histology-analogous images from non- or minimally-processed tissues, and is a promising tool for addressing this pain point in clinical cancer management. We investigated the ability structured illumination microscopy (SIM) to generate images from freshly-obtained breast tissues for structure identification and cancer identification at a speed compatible with potential on-site clinical implementation. We imaged 47 biopsies from patients undergoing a guided breast biopsy procedure using a customized SIM system and a dual-color fluorescent hematoxylin & eosin (H&E) analog. These biopsies had an average size of 0.92 cm2 (minimum 0.1, maximum 4.2) and had an average imaging time of 7:29 (minimum 0:22, maximum 37:44). After imaging, breast biopsies were submitted for standard histopathological processing and review. A board-certified pathologist returned a binary diagnostic accuracy of 96% when compared to diagnoses from gold-standard histology slides, and key tissue features including stroma, vessels, ducts, and lobules were identified from the resulting images.
Desmosomes are prominent cell-cell adhesive junctions in stratified squamous epithelia and disruption of desmosomal adhesion has been shown to have dramatic effects on the function and integrity of ...these tissues. During normal physiologic processes, such as tissue development and wound healing, intercellular adhesion must be modified locally to allow coordinated cell movements. The mechanisms that control junction integrity and adhesive strength under these conditions are poorly understood. We utilized a proteomics approach to identify plakophilin-3 associated proteins and identified the 14-3-3 family member stratifin. Stratifin interacts specifically with plakophilin-3 and not with other plakophilin isoforms and mutation analysis demonstrated the binding site includes serine 285 in the amino terminal head domain of plakophilin-3. Stratifin interacts with a cytoplasmic pool of plakophilin-3 and is not associated with the desmosome in cultured cells. FRAP analysis revealed that decreased stratifin expression leads to an increase in the exchange rate of cytoplasmic plakophilin-3/GFP with the pool of plakophilin-3/GFP in the desmosome resulting in decreased desmosomal adhesion and increased cell migration. We propose a model by which stratifin plays a role in regulating plakophilin-3 incorporation into the desmosomal plaque by forming a plakophilin-3 stratifin complex in the cytosol and thereby affecting desmosome dynamics in squamous epithelial cells.
Oxysterol-binding Protein (OSBP) is a human lipid-transport protein required for the cellular replication of many types of viruses, including several human pathogens. The structurally-diverse small ...molecule compounds OSW-1, itraconazole (ITZ), T-00127-HEV2 (THEV) and TTP-8307 (TTP) inhibit viral replication through interaction with the OSBP protein. The OSW-1 compound reduces intracellular OSBP, and the reduction of OSBP protein levels persists multiple days after the OSW-1-compound treatment is stopped. The OSW-1-induced reduction of OSBP levels inhibited Enterovirus replication prophylactically in cells. In this report, the OSBP-interacting compounds ITZ, THEV, and TTP are shown not to reduce OSBP levels in cells, unlike the OSW-1-compound, and the OSW-1 compound is determined to be the only compound capable of providing prophylactic antiviral activity in cells. Furthermore, OSW-1 and THEV inhibit the binding of 25-hydroxycholesterol (25-OHC) to OSBP indicating that these compounds bind at the conserved sterol ligand binding site. The ITZ and TTP compounds do not inhibit 25-hydroxycholesterol binding to OSBP, and therefore ITZ and TTP interact with OSBP through other, unidentified binding sites. Co-administration of the THEV compound partially blocks the cellular activity of OSW-1, including the reduction of cellular OSBP protein levels; co-administration of the ITZ and TTP compounds have minimal effect on OSW-1 cellular activity further supporting different modes of interaction with these compounds to OSBP. OSW-1, ITZ, THEV, and TTP treatment alter OSBP cellular localization and levels, but in four distinct ways. Co-administration of OSW-1 and ITZ induced OSBP cellular localization patterns with features similar to the effects of ITZ and OSW-1 treatment alone. Based on these results, OSBP is capable of interacting with multiple structural classes of antiviral small molecule compounds at different binding sites, and the different compounds have distinct effects on OSBP cellular activity.
•Multiple antiviral compounds target OSBP, but only the OSW-1 compound induces a reduction of OSBP levels in cells.•The OSW-1 compound is the only OSBP-targeting compound with prophylactic antiviral activity in cells.•The ITZ and TTP compounds do not inhibit 25-hydroxycholesterol binding to OSBP, suggesting alternative binding sites.•At high concentrations, THEV and ITZ can significantly block OSW-1 activity in cells.•OSW-1, ITZ, THEV, and TTP compound alter OSBP cellular localization but with differing effects.
Oxysterol-binding protein (OSBP) is a lipid transport and regulatory protein required for the replication of Enterovirus genus viruses, which includes many significant human pathogens. Short-term ...exposure (i.e., 1–6 h) to a low dose (i.e., 1 nM) of the natural product compound OSW-1 induces a reduction of cellular OSBP levels by ∼90% in multiple different cell lines with no measurable cytotoxicity, defect in cellular proliferation, or global proteome reduction. Interestingly, the reduction of OSBP levels persists multiple days after the low-dose, transient OSW-1 compound treatment is ended and the intracellular OSW-1 compound levels drop to undetectable levels. The reduction in OSBP levels is inherited in multiple generations of cells that are propagated after the OSW-1 compound treatment is stopped. The enduring multiday, multigenerational reduction of OSBP levels triggered by the OSW-1 compound is not due to proteasome degradation of OSBP or due to a reduction in OSBP mRNA levels. OSW-1 compound treatment induces transient autophagy in cells, but blocking autophagy does not rescue OSBP levels. Although the specific cellular mechanism of long-term OSBP repression is not yet identified, these results clearly show the existence of an OSBP specific cellular regulation process that is triggered upon treatment with an OSBP-binding compound. The stable reduction of OSBP levels upon short-term, transient OSW-1 compound treatment will be a powerful tool to understand OSBP regulation and cellular function. Additionally, the persistent reduction in OSBP levels triggered by the transient OSW-1 compound treatment substantially reduces viral replication in treated cells. Therefore, the long-term, compound-induced reduction of OSBP in cells presents a new route to broad spectrum anti-Enterovirus activity, including as a novel route to antiviral prophylactic treatment through small molecule targeting a human host protein.
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