KaryoCreate: A CRISPR-based technology to study chromosome-specific aneuploidy by targeting human centromeres.

Aneuploidy, the presence of chromosome gains or losses, is a hallmark of cancer. Here, we describe KaryoCreate (karyotype CRISPR-engineered aneuploidy technology), a system that enables the generation of chromosome-specific aneuploidies by co-expression of an sgRNA targeting chromosome-specific CENPA-binding ɑ-satellite repeats together with dCas9 fused to mutant KNL1. We design unique and highly specific sgRNAs for 19 of the 24 chromosomes. Expression of these constructs leads to missegregation and induction of gains or losses of the targeted chromosome in cellular progeny, with an average efficiency of 8% for gains and 12% for losses (up to 20%) validated across 10 chromosomes. Using KaryoCreate in colon epithelial cells, we show that chromosome 18q loss, frequent in gastrointestinal cancers, promotes resistance to TGF-ß, likely due to synergistic hemizygous deletion of multiple genes. Altogether, we describe an innovative technology to create and study chromosome missegregation and aneuploidy in the context of cancer and beyond.

Immune evasion in HPV- head and neck precancer-cancer transition is driven by an aneuploid switch involving chromosome 9p loss.

An aneuploid-immune paradox encompasses somatic copy-number alterations (SCNAs), unleashing a cytotoxic response in experimental precancer systems, while conversely being associated with immune suppression and cytotoxic-cell depletion in human tumors, especially head and neck cancer (HNSC). We present evidence from patient samples and cell lines that alterations in chromosome dosage contribute to an immune hot-to-cold switch during human papillomavirus-negative (HPV-) head and neck tumorigenesis. Overall SCNA (aneuploidy) level was associated with increased CD3+ and CD8+ T cell microenvironments in precancer (mostly CD3+, linked to trisomy and aneuploidy), but with T cell-deficient tumors. Early lesions with 9p21.3 loss were associated with depletion of cytotoxic T cell infiltration in TP53 mutant tumors; and with aneuploidy were associated with increased NK-cell infiltration. The strongest driver of cytotoxic T cell and Immune Score depletion in oral cancer was 9p-arm level loss, promoting profound decreases of pivotal IFN-γ-related chemokines (e.g., CXCL9) and pathway genes. Chromosome 9p21.3 deletion contributed mainly to cell-intrinsic senescence suppression, but deletion of the entire arm was necessary to diminish levels of cytokine, JAK-STAT, and Hallmark NF-κB pathways. Finally, 9p arm-level loss and JAK2-PD-L1 codeletion (at 9p24) were predictive markers of poor survival in recurrent HPV- HNSC after anti-PD-1 therapy; likely amplified by independent aneuploidy-induced immune-cold microenvironments observed here. We hypothesize that 9p21.3 arm-loss expansion and epistatic interactions allow oral precancer cells to acquire properties to overcome a proimmunogenic aneuploid checkpoint, transform and invade. These findings enable distinct HNSC interception and precision-therapeutic approaches, concepts that may apply to other CN-driven neoplastic, immune or aneuploid diseases, and immunotherapies.

Recent Advances in the Synthesis and Transformation of Carbamoyl Fluorides, Fluoroformates, and Their Analogues.

Carbamoyl fluorides, fluoroformates, and their analogues are a class of important compounds and have been evidenced as versatile building blocks for the preparation of useful molecules in organic chemistry. While major achievements were made in the synthesis of carbamoyl fluorides, fluoroformates, and their analogues in the last half of 20th century, an increasing number of reports have focused on using O/S/Se=CF2 species or their equivalents as the fluorocarbonylation reagents for the direct construction of these compounds from the parent heteroatom-nucleophiles in recent years. This review mainly summarizes the advances in the synthesis and typical application of carbamoyl fluorides, fluoroformates, and their analogues by the halide exchanges and fluorocarbonylation reactions since 1980.

Photoinduced Copper-Promoted Decarboxylative Trifluoromethylselenolation of Aromatic Carboxylic Acids with [Me4N][SeCF3].

Visible-light-induced decarboxylative trifluoromethylselenolation of (hetero)aromatic carboxylic acids with [Me4N][SeCF3], oxidant, and catalysts afforded a variety of (hetero)aryl trifluoromethyl selenoethers in good yields. The reaction might involve a radical process, which generated (hetero)aryl radicals from the stable (hetero)aromatic carboxylic acids via oxidative decarboxylation with NFSI as the oxidant, [di-tBu-Mes-Acr-Ph][BF4] as the photocatalyst, and 1,1'-biphenyl as the cocatalyst. Both catalysts had a decisive influence on the reaction. The trifluoromethylselenolation was further promoted by the copper salts probably via Cu-mediated cross-coupling of the sensitive SeCF3 species with the in situ formed (hetero)aryl radicals. Advantages of the method include visible light irradiation, mild reaction conditions at ambient temperature, good functional group tolerance, no pre-functionalization/activation of the starting carboxylic acids, and applicability to drug molecules. This protocol is promising and synthetically useful, which overcame the limitations of the known trifluoromethylselenolation methods and represented the first decarboxylative trifluoromethylselenolation of (hetero)aromatic carboxylic acids.

Divergent dehydroxyfluorination and carbonation of alcohols with trifluoromethyl trifluoromethanesulfonate.

Preparation of alkyl fluorides and carbonates via divergent dehydroxyfluorination and carbonation of alcohols with trifluoromethyl trifluoromethanesulfonate (CF3SO2OCF3) is described. The reactions performed with BTMG in THF provided alkyl fluorides in good yields, whereas those of two different alcohols with Et3N in DCM formed asymmetric carbonates in moderate to excellent yields. CF3SO2OCF3 was demonstrated to be either a "F" or a "CO" reagent in the reactions by changing the base, allowing the selective construction of alkyl fluorides and carbonates from the corresponding alcohols with high efficiency. Notably, the fluorine-containing asymmetric carbonates that are difficult to synthesize by other methods were comprehensively prepared by this method, which would have great application potential in both academic and industrial fields.

An EVOH nanofibrous sterile membrane with a robust and antifouling surface for high-performance sterile filtration via glutaraldehyde crosslinking and a plasma-assisted process.

Constructing a sterile membrane with a robust and antifouling surface is a powerful means to improve the sterile filtration efficiency of sterile membranes. In this work, a robust EVOH nanofibrous sterile membrane was facilely fabricated by the method of in situ crosslinking with glutaraldehyde and surface plasma treatment. The resultant EVOH nanofibrous sterile membrane possessed a carboxylated-crosslinked surface, with high hydrophilicity, which generated high chemical stability, high-temperature steam resistance, and an ultrahigh antifouling performance against bovine serum albumin, ribonucleic acid and nanoparticle pollutants. Moreover, the membrane also exhibited a reasonably high primary water permeance (4522.2 LMH bar-1 at 0.2 MPa), as well as an absolute interception rate (100%) of Escherichia coli, Staphylococcus aureus cells and Brevundimonas diminuta superior to the state-of-the-art sterile membrane. Moreover, the modified membrane packed syringe-driven filter presented 100% interception (LRV ≥ 7) to Brevundimonas diminuta and high permeation flux (from 10.8 to 41.8 L·h-1) in a wide operating pressure range of 0.1 MPa to 0.6 MPa, indicating its potential in real bio-separation applications. This work provides a facile strategy for the preparation of a high-performance sterile membrane for biological drug product sterilization.

Poor ventilation worsens short-range airborne transmission of respiratory infection.

To explain the observed phenomenon that most SARS-CoV-2 transmission occurs indoors whereas its outdoor transmission is rare, a simple macroscopic aerosol balance model is developed to link short- and long-range airborne transmission. The model considers the involvement of exhaled droplets with initial diameter ≤50 µm in the short-range airborne route, whereas only a fraction of these droplets with an initial diameter within 15 µm or equivalently a final diameter within 5 µm considered in the long-range airborne route. One surprising finding is that the room ventilation rate significantly affects the short-range airborne route, in contrast to traditional belief. When the ventilation rate in a room is insufficient, the airborne infection risks due to both short- and long-range transmission are high. A ventilation rate of 10 L/s per person provides a similar concentration vs distance decay profile to that in outdoor settings, which provides additional justification for the widely adopted ventilation standard of 10 L/s per person. The newly obtained data do not support the basic assumption in the existing ventilation standard ASHRAE 62.1 (2019) that the required people outdoor air rate is constant if the standard is used directly for respiratory infection control. Instead, it is necessary to increase the ventilation rate when the physical distance between people is less than approximately 2 m.

The value of color Doppler sonography in diagnosis and hemodynamic analysis of innominate artery occlusion.

The occlusion of the innominate artery caused a significant decrease in the distal end of the right subclavian artery and the right common carotid artery, internal carotid artery, and external carotid artery (ECA). Due to the different pressure and the abundant communicating arteries between the ECA and the bilateral vertebral artery (VA), different paths of blood steal in the anterior and posterior circulation occurred.

Exposure and respiratory infection risk via the short-range airborne route.

Leading health authorities have suggested short-range airborne transmission as a major route of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). However, there is no simple method to assess the short-range airborne infection risk or identify its governing parameters. We proposed a short-range airborne infection risk assessment model based on the continuum model and two-stage jet model. The effects of ventilation, physical distance and activity intensity on the short-range airborne exposure were studied systematically. The results suggested that increasing physical distance and ventilation reduced short-range airborne exposure and infection risk. However, a diminishing return phenomenon was observed when the ventilation rate or physical distance was beyond a certain threshold. When the infectious quantum concentration was less than 1 quantum/L at the mouth, our newly defined threshold distance and threshold ventilation rate were independent of quantum concentration. We estimated threshold distances of 0.59, 1.1, 1.7 and 2.6 m for sedentary/passive, light, moderate and intense activities, respectively. At these distances, the threshold ventilation was estimated to be 8, 20, 43, and 83 L/s per person, respectively. The findings show that both physical distancing and adequate ventilation are essential for minimising infection risk, especially in high-intensity activity or densely populated spaces.

Probable cross-corridor transmission of SARS-CoV-2 due to cross airflows and its control.

A COVID-19 outbreak occurred in May 2020 in a public housing building in Hong Kong - Luk Chuen House, located in Lek Yuen Estate. The horizontal cluster linked to the index case' flat (flat 812) remains to be explained. Computational fluid dynamics simulations were conducted to obtain the wind-pressure coefficients of each external opening on the eighth floor of the building. The data were then used in a multi-zone airflow model to estimate the airflow rate and aerosol concentration in the flats and corridors on that floor. Apart from flat 812 and corridors, the virus-laden aerosol concentrations in flats 811, 813, 815, 817 and 819 (opposite to flat 812, across the corridor) were the highest on the eighth floor. When the doors of flats 813 and 817 were opened by 20%, the hourly-averaged aerosol concentrations in these two flats were at least four times as high as those in flats 811, 815 and 819 during the index case's home hours or the suspected exposure period of secondary cases. Thus, the flats across the corridor that were immediately downstream from flat 812 were at the highest exposure risk under a prevailing easterly wind, especially when their doors or windows that connected to the corridor were open. Given that the floorplan and dimension of Luk Chuen House are similar to those of many hotels, our findings provide a probable explanation for COVID-19 outbreaks in quarantine hotels. Positive pressure and sufficient ventilation in the corridor would help to minimise such cross-corridor infections.

Insufficient ventilation led to a probable long-range airborne transmission of SARS-CoV-2 on two buses.

Uncertainty remains on the threshold of ventilation rate in airborne transmission of SARS-CoV-2. We analyzed a COVID-19 outbreak in January 2020 in Hunan Province, China, involving an infected 24-year-old man, Mr. X, taking two subsequent buses, B1 and B2, in the same afternoon. We investigated the possibility of airborne transmission and the ventilation conditions for its occurrence. The ventilation rates on the buses were measured using a tracer-concentration decay method with the original driver on the original route. We measured and calculated the spread of the exhaled virus-laden droplet tracer from the suspected index case. Ten additional passengers were found to be infected, with seven of them (including one asymptomatic) on B1 and two on B2 when Mr. X was present, and one passenger infected on the subsequent B1 trip. B1 and B2 had time-averaged ventilation rates of approximately 1.7 and 3.2 L/s per person, respectively. The difference in ventilation rates and exposure time could explain why B1 had a higher attack rate than B2. Airborne transmission due to poor ventilation below 3.2 L/s played a role in this two-bus outbreak of COVID-19.

Trifluoromethylselenolation and N-acylation of indoles with [Me4N][SeCF3].

An efficient method for oxidative trifluoromethylselenolation/N-acylation of indoles with excess [Me4N][SeCF3] in the presence of acyl peroxides and their derivatives is described. The reaction is easy to handle, proceeds smoothly at room temperature under metal-free conditions, and shows advantages such as good functional group tolerance, excellent regioselectivity, and compatibility of a number of substrates, producing 1-acyl and 3-trifluoromethylselanyl substituted indoles in good yields. Acyl peroxides and peroxycarboxylic acid behave as both oxidants and acyl sources in the transformation. This one-pot procedure provides a convenient access to a new class of indole derivatives, representing the first trifluoromethylselanyl bifunctionalization of indoles with the nucleophilic [Me4N][SeCF3] reagent.

Probable airborne transmission of SARS-CoV-2 in a poorly ventilated restaurant.

Although airborne transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been recognized, the condition of ventilation for its occurrence is still being debated. We analyzed a coronavirus disease 2019 (COVID-19) outbreak involving three families in a restaurant in Guangzhou, China, assessed the possibility of airborne transmission, and characterized the associated environmental conditions. We collected epidemiological data, obtained a full video recording and seating records from the restaurant, and measured the dispersion of a warm tracer gas as a surrogate for exhaled droplets from the index case. Computer simulations were performed to simulate the spread of fine exhaled droplets. We compared the in-room location of subsequently infected cases and spread of the simulated virus-laden aerosol tracer. The ventilation rate was measured using the tracer gas concentration decay method. This outbreak involved ten infected persons in three families (A, B, C). All ten persons ate lunch at three neighboring tables at the same restaurant on January 24, 2020. None of the restaurant staff or the 68 patrons at the other 15 tables became infected. During this occasion, the measured ventilation rate was 0.9 L/s per person. No close contact or fomite contact was identified, aside from back-to-back sitting in some cases. Analysis of the airflow dynamics indicates that the infection distribution is consistent with a spread pattern representative of long-range transmission of exhaled virus-laden aerosols. Airborne transmission of the SARS-CoV-2 virus is possible in crowded space with a ventilation rate of 1 L/s per person.

Tertiary-Amine-Initiated Synthesis of Acyl Fluorides from Carboxylic Acids and CF3 SO2 OCF3.

A convenient method for deoxyfluorination of aromatic and aliphatic carboxylic acids with CF3 SO2 OCF3 in the presence of a suitable base at room temperature has been developed. The reaction allows a straightforward access to a variety of acyl fluorides and proves that CF3 SO2 OCF3 is an effective deoxyfluorination reagent for carboxylic acids. The method features simplicity, expeditiousness, high efficiency, ease of handling, good functional group tolerance, a wide range of substrates, excellent yields of products, compatibility of many amine initiators, use of environmentally friendly reagents, and effortless removal of byproducts. This reaction represents the first utilization of trifluoromethyl trifluoromethanesulfonate as a fluorination reagent.

Oxidative trifluoromethylselenolation of 1,3-dicarbonyls with [Me4N][SeCF3].

Reactions of 1,3-ketoesters, -diesters, -diketones, and -ketoamides with [Me4N][SeCF3] in the presence of an appropriate oxidant provided a series of 2-trifluoromethylselenolated 1,3-dicarbonyls in moderate to good yields. The trifluoromethylselenolation featured simplicity, mildness, high efficiency, transition-metal-free conditions, and compatibility of various oxidants, and represented the first oxidative trifluoromethylselenolation of 1,3-dicarbonyl compounds with [Me4N][SeCF3]. This protocol was also applicable to the oxidative trifluoromethylthiolation of 1,3-dicarbonyls with [Me4N][SCF3]/NCS, and oxidative trifluoromethylchalcogenation with nucleophilic XCF3 (X = O, S, and Se) reagents were compared. The results demonstrated that these nucleophilic XCF3 salts showed different reaction profiles towards 1,3-dicarbonyls under oxidation conditions.

Markovnikov-Type Hydrotrifluoromethylchalcogenation of Unactivated Terminal Alkenes with [Me4N][XCF3] (X = S, Se) and TfOH.

The first Markovnikov-type hydrotrifluoromethylselenolation of unactivated terminal alkenes with the readily accessible [Me4N][SeCF3] reagent and the superacid TfOH is reported. The reaction proceeded at room temperature under catalyst- and additive-free conditions to give the branched trifluoromethylselenolated products in good yields. This protocol is also applicable to the Markovnikov-type hydrotrifluoromethylthiolation of unactivated terminal alkenes using [Me4N][SCF3]/TfOH, but not to the hydrotrifluoromethoxylation with CsOCF3/TfOH under the same conditions. The successful hydrotrifluoromethylselenolation and hydrotrifluoromethylthiolation showed simplicity and high regioselectivity, taming the sensitive -XCF3 (X = Se, S) anions with TfOH, and offered a convenient method for the straightforward synthesis of branched trifluoromethyl selenoethers and thioethers from unactivated alkenes.

Impact of intervention methods on COVID-19 transmission in Shenzhen.

By March 31, 2020, COVID-19 had spread to more than 200 countries. Over 750,000 confirmed cases were reported, leading to more than 36,000 deaths. In this study, we analysed the efficiency of various intervention strategies to prevent infection by the virus, SARS-CoV-2, using an agent-based SEIIR model, in the fully urbanised city of Shenzhen, Guangdong Province, China. Shortening the duration from symptom onset to hospital admission, quarantining recent arrivals from Hubei Province, and letting symptomatic individuals stay at home were found to be the three most important interventions to reduce the risk of infection in Shenzhen. The ideal time window for a mandatory quarantine of arrivals from Hubei Province was between 10 January and January 17, 2020, while the ideal time window for local intervention strategies was between 15 and 22 January. The risk of infection could have been reduced by 50% if all symptomatic individuals had immediately gone to hospital for isolation, and by 35% if a 14-day quarantine for arrivals from Hubei Province had been introduced one week earlier. Intervention strategies implemented in Shenzhen were effective, and the spread of infection would be controlled even if the initial basic reproduction number had doubled. Our results may be useful for other cities when choosing their intervention strategies to prevent outbreaks of COVID-19.

Monitoring circulating tumor cells in vivo by a confocal microscopy system.

Circulating tumor cells (CTCs) play a key role in cancer metastasis but are very difficult to detect. in vivo monitoring CTCs has been recognized as an important technique for cancer research and clinical diagnosis. Recently, a noninvasive method, in vivo flow cytometry (IVFC) has been developed to enable continuous, real-time, and long-duration detection of CTCs in animal models by detecting CTC fluorescence in blood vessels excited by lasers. In this study, we present a simple optical scheme for direct noninvasive CTC detection using confocal microscopes. We demonstrate that line scanning of confocal microscopy can provide effective and quantitative CTC detection in live mice during cancer development. Rare CTC signals can be acquired at the early stage of the tumor development after implantation of subcutaneous tumor and monitored continuously to the end. Signals from CTC clusters can also be acquired and distinguished from single CTCs. Our results suggest confocal microscopy is a simple and reliable method for biologists and doctors to use for cancer research. © 2018 International Society for Advancement of Cytometry.

Concise and Additive-Free Click Reactions between Amines and CF3 SO3 CF3.

Trifluoromethyl trifluoromethanesulfonate has proved to be an excellent reservoir of difluorophosgene and a promising click ligation for amines in the preparation of urea derivatives, heterocycles, and carbamoyl fluorides under metal- and additive-free conditions. The reactions are rapid, efficient, selective, and versatile, and can be performed in benign solvents, giving products in excellent yields with minimal efforts for purification. The characteristics of the reactions meet the requirements of a click reaction. The use of trifluoromethyl trifluoromethanesulfonate as a click reagent is advantageous over other "CO" sources (e.g., TsOCF3 , PhCO2 CF3 , CsOCF3 , AgOCF3 , and triphosgene) because this reagent is readily accessible; easy to scale up; and highly reactive, even under metal- and additive-free conditions. It is anticipated that CF3 SO3 CF3 will be increasingly as important as SO2 F2 as a click agent in future drug design and development.

Cu-Mediated arylselenylation of aryl halides with trifluoromethyl aryl selenonium ylides.

An unprecedented arylselenylation of aryl halides with trifluoromethyl aryl selenonium ylides in the presence of copper is described. The reaction proceeded at 100-140 °C under ligand- and additive-free conditions for 3-20 h to form a variety of unsymmetrical diaryl selenides in good to high yields. Arylselenylation is easy to operate, has good functional group tolerance, and demonstrates the different reaction profiles of trifluoromethyl aryl selenonium ylides from the homologous trifluoromethyl aryl sulfonium ylides.