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The facile generation of the α-acyloxy carboxamide radical is hereby reported for the first time, utilizing a photoredox catalyzed reaction of Passerini adducts synthesized using a 4-formyl-1,4-dihydropyridine as the carbonyl component. This radical effectively engages in a Giese reaction with a range of olefins, ultimately leading to the synthesis of novel Passerini-derived products not previously amenable to direct aldehyde-based transformations. Consequently, the resulting strategy, developed both in batch and in flow, offers a promising opportunity to expand the chemical space accessible through the Passerini reaction, virtually incorporating "impossible" aldehydes., (© 2024 Wiley-VCH GmbH.)

A substantial number of patients are at high-risk of intra- or post-operative complications or both. Most perioperative deaths are represented by patients who present insufficient physiological reserve to meet the demands of major surgery. Recognition and management of critical high-risk surgical patients require dedicated and effective teams, capable of preventing, recognize, start treatment with adequate support in time to refer patients to the satisfactory ICU level provision. The main task for health-care planners and managers is to identify and reduce this severe risk and to encourage patient's safety practices. Inadequate tissue perfusion and decreased cellular oxygenation due to hypovolemia, heart dysfunction, reduced cardiovascular reserve, and concomitant diseases are the most common causes of perioperative complications. Hemodynamic, respiratory and careful sequential monitoring have become essential aspects of the clinical practice both for surgeons and intensivists. New monitoring techniques have changed significantly over the past few years and are now able to rapidly identify shock states earlier, define the etiology, and monitor the response to different therapies. Many of these techniques are now minimally invasive or non-invasive. Advanced hemodynamic and respiratory monitoring combines invasive, non-invasive monitoring skills. Non-invasive ultrasound has emerged during the last years as an essential operative and perioperative evaluation tool, and its use is now rapidly growing. Perioperative management guided by appropriate sequential clinical evaluation combined with respiratory and hemodynamic monitoring is an established tool to help clinicians to identify those patients at higher risk in the attempt to reduce the complications rate and potentially improve patient outcomes. This review aims to provide an update of currently available standard concepts and evolving technologies of the various respiratory and hemodynamic monitoring systems for the high-risk surgical patients, highlighting their potential usefulness when integrated with careful clinical evaluation., Competing Interests: Competing interestsNone of the authors have any conflicts of interests to declare. The authors received no pharmaceutical or industrial support for this study. No further direct or indirect financial support or other assets were transferred to the authors or their family members for this study., (© The Author(s). 2019.)

Background: Refractory septic shock (RSS) requiring major vasopressor support is associated with high mortality, especially in Gram-negative infections. The study aim was to describe hemodynamics, organ failure, and clinical outcomes in high-dose vasopressor therapy (HDVT) RSS patients treated with Polymyxin B hemoperfusion (PMX-HP) as rescue therapy., Methods: We retrospectively analyzed 52 patients, unresponsive to conventional therapy, treated with two sessions of PMX-HP requiring HDVT (norepinephrine and/or epinephrine requirement (NEP+EP) ≥ 0.5 µg/kg/min), ≥ 2 organ failures, and suspected/confirmed Gram-negative infection from any source., Results: At baseline, mean arterial pressure (MAP) was 80 ± 13 mmHg and NEP + EP requirement was 1.11 ± 0.56 µg/kg/min. After two PMX-HP sessions, at 72 h, MAP significantly increased and NEP + EP requirement decreased respectively by 12% and 76%. Pulmonary and renal function also improved significantly. Thirty patients (58%) showed a ≥ 50% reduction in NEP + EP dose within only 24 h after the first PMX-HP session (early responders), and 22 did not or died from irreversible shock in the same time frame (early non-responders). The 30-day hospital mortality was 29%; it was 16% in early responders and 45% in early non-responders. On multivariate analysis, SAPS II score, vasopressin, and central venous pressure significantly affected 30-day hospital mortality., Conclusion: This is the first study describing the use of PMX-HP as a rescue therapy in RSS patients with HDVT and MOF. Our results suggest a possible role for PMX-HP in improving hemodynamics, organ function, and mortality in RSS, with a 30-day survival of up to 70%.

Determination of incorporation of the thymidine analog 5-bromo-2'-deoxyuridine (BrdU) into DNA is a widely used method to analyze the cell cycle. However, DNA denaturation is required for BrdU detection with the consequence that most protein epitopes are destroyed and their immunocytochemical detection for multiplex analysis is not possible. A novel assay is presented for identifying cells in active S-phase that does not require the DNA denaturation step but nevertheless detects BrdU. For this purpose, cells were pulsed for a short time by 5-ethynyl-2'-deoxyuridine (EdU) which is incorporated into DNA. The nucleotide-exposed ethynyl residue was then derivatized by a copper-catalyzed cycloaddition reaction ("click chemistry" coupling) using a BrdU azide probe. The resulting DNA-bound bromouracil moieties were then detected by commercial anti-BrdU monoclonal antibodies without the need for a denaturation step. This method has been tested using several cell lines and is more sensitive than traditional BrdU and allows multicolor and multiplex analysis in flow cytometry (FCM) and image-based cytometry., (Copyright © 2015 John Wiley & Sons, Inc.)

Aberrant activation of the mitogen-activated protein kinase (MAPK)-mediated pathway components, RAF-MEK-ERK, is frequently observed in human cancers and clearly contributes to oncogenesis. As part of a project aimed at finding inhibitors of B-Raf, a key player in the MAPK cascade, we originally identified a thiazole derivative endowed with high potency and selectivity, optimal in vitro ADME properties, and good pharmacokinetic profiles in rodents, but that suffers from elevated hERG inhibitory activity. An optimization program was thus undertaken, focused mainly on the elaboration of the R(1) and R(2) groups of the scaffold. This effort ultimately led to N-(4-{2-(1-cyclopropylpiperidin-4-yl)-4-[3-(2,5-difluorobenzenesulfonylamino)-2-fluorophenyl]thiazol-5-yl}-pyridin-2-yl)acetamide (20), which maintains favorable in vitro and in vivo properties, but lacks hERG liability. Besides exhibiting potent antiproliferative activity against only cell lines bearing B-Raf V600E or V600D mutations, compound 20 also intriguingly shows a weaker "paradoxical" activation of MEK in non-mutant B-Raf cells than other known B-Raf inhibitors. It also demonstrates very good efficacy in vivo against the A375 xenograft melanoma model (tumor volume inhibition >90% at 10 mg kg(-1) ); it is therefore a suitable candidate for preclinical development., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Mixtures of ascorbate and copper used in certain click chemistry experimental conditions act as oxidizing agents, catalyzing the formation of reactive oxygen species through Fenton and related reactions. Hydroxyl radicals act as chemical nucleases, introducing DNA strand breaks that can be exploited for BrdU immunostaining in place of acid denaturation. This procedure is readily applicable to high content analysis and flow cytometry assays, and provides results comparable to click chemistry EdU cycloaddition and classical BrdU immunodetection. Importantly, this approach allows preservation of labile epitopes such as phosphoproteins. This unit describes an optimized method that successfully employs Fenton chemistry for simultaneous detection of phosphoproteins and BrdU in intact cells., (Copyright © 2015 John Wiley & Sons, Inc.)

Ascorbic acid (AA) and copper have been increasingly employed in flow cytometry (FCM) and high content analysis (HCA) since the introduction of "click chemistry" as a non-destructive alternative to classical 5-bromo-2'-deoxyuridine (BrdU) immunodetection for DNA synthesis and proliferation assays. Mixtures of ascorbate and catalytic copper, under certain experimental conditions, act as oxidizing agent, catalyzing the formation of reactive hydroxyl radicals through hydrogen peroxides decomposition via Fenton reaction. We developed a procedure for BrdU incorporation detection based on the use of AA and cupric ions as DNA damaging agents. Optimal DNA damaging conditions were identified and found to provide results comparable with "click" 5-ethynyl-deoxyuridine (EdU) cycloaddition approach and classical BrdU immunodetection. Scavenger agents were found to prevent hydroxyl-induced DNA damages, providing the proof-of-concept for the use of this procedure for DNA denaturation prior to BrdU detection. We demonstrated hydroxyl radicals' reaction to be readily applicable to HCA and FCM assays, for both classical BrdU immunostaining and EdU cycloaddition procedure. This technique was successfully employed for BrdU pulse-chase experiments and in multiparametric immunofluorescence assays for the simultaneous detection of labile phosphoproteins in intact cells. The use of AA/Cu prior to immunodetection for BrdU incorporation assays is a viable alternative to chemical/physical DNA denaturing agents (acids or heat), since it allows preservation of labile epitopes such as phosphoproteins, and over enzymatic agents (digestion with DNases) for its lower cost., (© 2013 International Society for Advancement of Cytometry.)

From the second half of the 19th century up to modern times, the tremendous contribution of Italian chemists to the development of science resulted in the discovery of a number of innovative chemical transformations. These reactions were subsequently christened according to their inventors' name and so entered into the organic chemistry portfolio of "named organic reactions". As these discoveries were being conceived, massive social, political and geographical changes in these chemists' homeland were also occurring. In this review, a brief survey of known (and some lesser known) named organic reactions discovered by Italian chemists, along with their historical contextualization, is presented.

Since Gomberg's discovery of radicals as chemical entities, the interest around them has increased through the years. Nowadays, radical chemistry is used in the synthesis of 75% of all polymers, inevitably establishing a close relationship with Solid-Phase Organic Synthesis. More recently, the interest of organic chemists has shifted towards the application of usual "in-solution" radical chemistry to the solid-phase, ranging from the use of supported reagents for radical reactions, to the development of methodologies for the synthesis of small molecules or potential libraries. The aim of this review is to put in perspective radical chemistry, moving it away from its origin as a synthetic means for solid supports, to becoming a useful tool for the synthesis of small molecules.

A novel series of 3-amino-1H-thieno[3,2-c]pyrazole derivatives demonstrating high potency in inhibiting Aurora kinases was developed. Here we describe the synthesis and a preliminary structure-activity relationship, which led to the discovery of a representative compound (38), which showed low nanomolar inhibitory activity in the anti-proliferation assay and was able to block the cell cycle in HCT-116 cell line. This compound demonstrated favorable pharmacokinetic properties and good efficacy in the HL-60 xenograft tumor model., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Quantification of BrdU incorporation into DNA is a widely used technique to assess the cell cycle status of cells. DNA denaturation is required for BrdU detection with the drawback that most protein epitopes are destroyed and classical antibody staining techniques for multiplex analysis are not possible. To address this issue we have developed a novel method that overcomes the DNA denaturation step but still allows detection of BrdU. Cells were pulsed for a short time by 5-ethynyl-2'-deoxyuridine, which is incorporated into DNA. The exposed nucleotide alkyne group of DNA was then derivatized in physiologic conditions by the copper (I)-catalyzed azide-alkyne cycloaddition (CuAAC) using BrdU azides. The resulting DNA-bound bromouracil moiety was subsequently detected by commercial anti-BrdU mAb without the need for a denaturation step. Continuous labeling with EdU showed a slightly increased anti-proliferative activity compared to BrdU. However, using a lower concentration of EdU for labeling can compensate for this. Alkynyl tags could be detected quickly by a highly specific reaction using BrdU azides. Fluorescence quenching by the DNA dye PI using both BrdU azides was negligible. Our labeling method is suitable for FCM and HCA and shows a higher signal to noise ratio than other methods. This method also allowed multiplex analysis by simultaneous detection of EdU-BrdU, caspase-3, and phospho-histone 3 mAbs, proving sensitivity and feasibility of this new technique. In addition, it has the potential for use in vivo, as exemplified for bone marrow studies. We have established a new method to determine the position of cells in the cell cycle. This is superior when compared to traditional BrdU detection since it allows multiplex analysis, is more sensitive and shows less quenching with PI. The method provides new opportunities to investigate changes in protein expression at different cell cycle stages using pulse labeling experiments., ((c) 2008 International Society for Advancement of Cytometry)

Determination of incorporation of the thymidine analog 5-bromo-2'-deoxyuridine (BrdU) into DNA is a widely used method to analyze the cell cycle (see UNIT 7.7). However, DNA denaturation is required for BrdU detection with the consequence that most protein epitopes are destroyed and their immunocytochemical detection for multiplex analysis is not possible. A novel assay is presented for identifying cells in active S-phase that does not require the DNA denaturation step but nevertheless detects BrdU. For this purpose, cells were pulsed for a short time by an alkenyl deoxyuridine (5-ethynyl-2'-deoxyuridine, EdU), which is incorporated into DNA. The nucleotide exposed ethynyl residue was then derivatized by a copper-catalyzed cycloaddition reaction ("click chemistry" coupling) using a BrdU azide probe. The resulting DNA-bound bromouracil moieties were then detected by commercial anti-BrdU monoclonal antibodies without the need for a denaturation step. This method has been tested using several cell lines and is preferred over traditional BrdU detection since it is more sensitive and allows multicolor and multiplex analysis in FCM and imaging.

A traceless solid-phase synthesis of oxazoles 4 via Robinson-Gabriel reaction of solid-supported alpha-acylamino ketones 2 has been achieved. The reaction requires that the cyclization precursor be linked to a benzhydrylic-type linker (compounds 2) and that trifluoroacetic anhydride be used as the cyclodehydrating agent. The solvent has a dramatic effect on the latter reaction, which goes to completion and follows a cyclative-type mechanism only when an ethereal solvent is used. Different synthetic routes have been investigated toward assembling compounds 2. The most straightforward one, which we have validated more extensively, comprises the reaction of Merrifield alpha-methoxyphenyl (MAMP) resin with an alpha-amino ketone to form compounds 1, which are, in turn, acylated. Other methodologies and strategies allowing for the synthesis of compounds 1 that have been investigated include direct alkylation of Rink amide resin; reductive amination of the latter with alpha-keto aldehydes; reaction of MAMP resin with alpha-amino alcohols, followed by oxidation; and protection of Rink amide resin with either 2,4-dinitrosulfonyl or allyl group, followed by alkylation and removal of protecting group. In addition, we disclose a novel variant of the Ugi four-component reaction that allows for the preparation of compounds 2 in a single synthetic step.

The study evaluates the cardiocirculatory and oxymetric effects of dobutamine infusion in doses between 5 and 15 g/kg/min for a period of 48 hours in 18 critical patients of whom 9 with a low cardiac index (mean 2.1) and 9 with a normal or increased cardiac index (mean 3.3). The beta mimetic effect of the drug is particularly evident in the 9 hypodynamic patients who benefited from a 28% increase in the cardiac index. Likewise, oxygen transport and consumption improved by 22% and 15% respectively. In overall terms, the variables which revealed significant modifications were: cardiac index (CI), systolic index (SI), mean pulmonary pressure (MPAP), pulmonary impaction pressure (PAWP), systemic and pulmonary vascular resistance (SVRO), PVRI), oxygen transport and consumption (DO2, VO2) (No major changes were observed in mean arterial pressure (MAP) and heart rate (HR). No tachyphylaxia was noted after 48 hours of continuous infusion.