The desymmetrization of p-peroxyquinols using a Brønsted acid-catalyzed acetalization/oxa-Michael cascade was achieved in high yields and selectivities for a variety of aliphatic and aryl aldehydes. ...Mechanistic studies suggest that the reaction proceeds through a dynamic kinetic resolution of the peroxy hemiacetal intermediate. The resulting 1,2,4-trioxane products were derivatized and show potent cancer cell-growth inhibition.
Abstract
Seven-membered nitrogen-containing heterocycles are considerably underrepresented in the literature compared to their five- and six-membered analogues. Herein, we report a relatively ...understudied photochemical rearrangement of
N
-vinylpyrrolidinones to azepin-4-ones in good yields. This transformation allows for the conversion of readily available pyrrolidinones and aldehydes to densely functionalized azepane derivatives in a facile two-step procedure.
Numerous American national committees have recommended the replacement of traditional labs with a more engaging curriculum that inspires inquiry and enhances scientific skills (examples include the ...President’s Council of Advisors on Science and Technology (PCAST)’s Engage to Excel program and American Association for the Advancement of Science (AAAS) Vision and Change, among others), due to a large body of evidence that shows significant enhancements in student learning and affective outcomes. The implementation of Course-Based Undergraduate Research Experiences (CUREs) is a creative way to scale up the deployment of authentic research experiences to students. Another highly regarded high-impact practice in postsecondary education is the addition of learning communities. The integration of a three-course learning community and authentic research experiences to laboratory courses adds both a community of scholarship and a development of scientific communication and process skills. This study describes a course that blends these two high-impact practices in higher education in order to promote greater post-course gains in essential elements of a CURE curriculum. This collaborative course shows large post-course gains in essential elements, such as scientific communication and working collaboratively.
Abstract only
Artemisinin is currently the frontline treatment for malaria and it has recently demonstrated potential as an anticancer drug. The key pharmacophore in artemisinin is an endoperoxide ...which generates ROS in the presence of reducing agents. It has been shown to work by decreasing cell proliferations, inducing apoptosis and cell death, and might involve ROS generation in intracellular compartments. In this study, in an effort to probe this reactivity and develop more potent endoperoxides, we developed a simple and economical synthesis of novel endoperoxide containing small molecules. Our unique methodology combines imines and peroxyquinols to form novel 1,2,4‐dioxazinanes in a one‐pot cascade reaction. 1,2,4‐dioxazinanes are a largely unexplored class of molecules with potential anticancer activity. One therapy to control cancer with minimal side effects would be to use non‐toxic natural products or their derivatives that could cause cell death by inducing apoptosis. Therefore, the efficacy of one of the synthetic derivatives, namely DMR‐1, to induce apoptosis in cancer cells was studied using a human colon carcinoma cell line, T84. Cells were grown to confluency in 12 well plates in a 1:1 mixture of Ham's F12 medium and Dulbecco's modified Eagle medium supplemented with 2.5 mM L‐‐glutamine and fetal bovine serum, 5%. Cells treated with increasing doses (5 – 100μM) of DMR‐1 in serum‐free media for 18 hrs were labeled with Annexin V and propidium iodide to assess apoptosis and cell death using an Accuri Flow cytometer. We report that DMR‐1 increased apoptosis at concentration as low as 5μM; however, there was no dose‐dependent increase in apoptosis AnnexinV positive cells, in %; Control: 9±1; 5 μM: 20±0.4; 10 μM: 15±2; 50 μM: 12±1; 100 μM: 11±1. Other artemisinin derivatives have been shown to be more effective in rapidly growing cell lines and at higher concentrations (>200 μM). T84 cells are slow growing cells and also have a tendency to secrete mucin, thereby requiring higher doses. Therefore, our future studies will include using higher doses of DMR‐1 and also studying the effect of DMR‐1 (and other derivatives synthesized in our laboratory) on rapidly growing cancer cells such as A549 lung cancer cells. If DMR‐1 is found to be effective in inducing apoptosis in these cells, the mechanism of action will be determined.
Support or Funding Information
This research is supported by NSF ‐ MRI: DBI‐1427937 and Institutional Funds, Ben. U. to JS and to DMR
Bile acid (BA)‐induced diarrhea affects one‐third of patients with chronic diarrhea, but the underlying mechanism is not clear. We recently reported that exposure to CDCA (500μM) altered tight ...junction (TJ) pore function by decreasing transepithelial resistance (TER) and leak function by increasing paracellular permeability across T84 monolayers; this involved release of reactive oxygen species (ROS) and inflammatory cytokines. The secondary BA lithocholic acid (LCA) and ROS inhibitors attenuate leak, but not pore, TJ function (Physiol Reports, ‘17, 5: e13294). We also showed that CDCA alters TJs only when added to the apical side (AS) of the monolayer and increases Cl− secretion only when added to the basolateral side (BLS) (AJP Cell, 2013, 305: C447‐56). We hypothesize that luminal BAs alter TJ permeability and travel paracellularly to reach the BLS.
To test this, we synthesized a novel fluorescent BA, CDCA‐FA, by converting the carboxylic acid of CDCA to an acid chloride and then adding the FA tag. This was purified via column chromatography and the structure, molecular formula and mass (C45 H53N O7; 719.38) confirmed by NMR and mass spectroscopy. Functional studies were performed on confluent T84 cells grown on Transwells (TER >1000Ωcm2), treated AS with DMSO (CTRL), 500μM CDCA or CDCA‐FA ± 50μM LCA for 0.5–18 hours (H). Cell viability was measured by propidium iodide staining, fluorescence microscopy and Image J analysis. CDCA‐FA effects on TJs were assessed by examining: i) pore function measured as TER; ii) leak function measured as apparent permeability of CDCA‐FA (PappCDCA‐FA) compared to CDCA‐induced Fluorescein Isothiocyanate‐10 kDa dextran (F10D) flux across the monolayer. The effect of ROS scavenger, N‐acetyl cysteine (NAC; 1mM), and LCA (50μM) on CDCA‐FA's effects on TJ functions were also assessed.
Overnight exposure to 500μM CDCA‐FA, like CDCA, caused a small (~18%) decrease in cell viability that was not altered by LCA (n=6). Again, similar to CDCA, CDCA‐FA showed a time‐dependent decrease in TER (in Ωcm2; t=0 H: CTRL: 927 ± 130; CDCA‐FA: 926 ± 154; t =0.5 H: CTRL: 836 ± 54; CDCA‐FA: 222 ± 28*; t=18 H; CTRL: 731 ± 10; CDCA‐FA: 107 ± 18*; n=4; *p<0.05). Furthermore, CDCA‐FA's effect on TER was neither altered by NAC nor LCA (% decrease vs. CTRL; 18 H CDCA‐FA: 85±2; CDCA‐FA+NAC: 88±5; CDCA‐FA+LCA: 87±3; CDCA‐FA+LCA+NAC: 85±6, n=3). In contrast, CDCA‐FA permeability increased over time; this increase was attenuated by LCA and further decreased by NAC (18 H, Papp CDCA‐FA, cm/sec, CDCA‐FA: 57±2; CDCA‐FA+LCA: 24±3; CDCA‐FA+NAC: 20±4; CDCA‐FA+LCA+NAC: 8±2; p<0.05, n≥3). This action of CDCA‐FA on the leak function is similar to that of CDCA on F10D flux (18 H, Papp F10D, cm/sec: CTRL: 8±1; CDCA: 165±12; p<0.05, n=6). Thus, using a novel fluorescent BA probe, CDCA‐FA, we demonstrate that, like CDCA, it increases TJ pore and leak functions in T84 cells. We conclude that CDCA from the apical side traverses to the basolateral side by a ROS‐dependent increase in paracellular permeability and that LCA limits the movement of CDCA through the TJs. Identifying ways to prevent BA's access to the BLS may be an effective therapeutic tool for the treatment of BA‐induced diarrhea.
Support or Funding Information
NSF ‐ MRI: DBI‐1427937 to JS and Institutional Funds, Ben. U. to JS and DMR; Institutional Funds, UIC to MCR
This is from the Experimental Biology 2018 Meeting. There is no full text article associated with this published in The FASEB Journal.
Artemisinin (ART), a standard treatment for malaria, is also known to have anti‐cancer properties. Our goal is to develop economical and novel ART analogs that can induce apoptosis in cancer cells at ...low doses with minimal toxicity. We previously showed that DMR1, a trioxane ART analog, induced apoptosis in T84 human colon cancer cells (Faseb J, 2017:31, 774.24). To explore better analogs, we synthesized HSM2, a novel dioxazinane ART analog with potentially improved pharmacokinetic properties over DMR1, and examined its efficacy in inducing apoptosis in cancer cells. The endoperoxide of ART analogs is the key pharmacophore, and we hypothesize that reactive oxygen species (ROS) generation in cancer cells is responsible for inducing cell death. Therefore, we investigated its role in DMR1/HSM2‐induced cell death.
T84 and A549 (lung cancer) cells were grown to confluency in 12‐well plates in fetal bovine serum‐supplemented media. Cells treated (18 hrs) with 0 – 100μM of DMR1 or HSM2 ± 1mM NAC (N‐acteyl cysteine; ROS scavenger) in serum‐free media were stained with FITC‐Annexin V and propidium iodide to measure apoptosis and cell death or CellRox Green and Sytox Red to measure mitochondrial and nuclear ROS and viability, respectively. Samples were assayed by flow cytometry (BD Accuri) and/or imaged using fluorescence microscopy (ZOE‐BioRad). Cytotoxicity was measured by a lactate dehydrogenase (LDH) colorimetric assay.
Flow cytometry analysis suggested that DMR1 was more effective in inducing apoptosis in A549 than in T84 cells (% Annexin V+ cells at 18 hr: A549 (n≥3): DMSO: 15 ±1; DMR1: 5μM: 45±4.5, 50μM: 46±4; vs.T84 (n=4) DMSO: 9±1; DMR1: 5 μM: 20±0.4; 10 μM: 15±2; 50 μM: 12±1). Fluorescence imaging revealed that the new ART derivative, HSM2, had a greater apoptotic effect than DMR1 on A549 cells (Image J analysis, in RGB mean pixel intensity 18 hr: DMSO: 1; HSM2: 5 μM: 38±5; 10 μM: 51±14; 50 μM: 29±8, n=3; vs. DMR1: 5 μM: 8±1; 10 μM: 35±2; 50 μM: 21±7, n=3). Similarly, HSM2 had a greater effect on T84 cell death at lower doses than DMR1. In marked contrast, DMR1 and HSM2 (5 – 50μM) did not induce apoptosis in the non‐cancerous BEAS2B cells, a normal bronchial cell line immortalized by viral transformation. At 10 μM, neither DMR1 nor HSM2 caused LDH release in T84 cells (% cytotoxicity, DMSO: 1.3±0.3; DMR1: 1.9±0.2; HSM2; 2.6±0.5; +ve control, 500 μM CDCA: 19.9±0.5; n=3). Flow cytometry measurements demonstrated that DMR1 increased ROS in T84 cells (% CellRox+ cells, DMSO: 7.5±0.2; 10 μM: 29±2; 100 μM: 28±4). Scavenging ROS with NAC (1mM) confirmed its role in ART analog‐induced cell death (Image J Analysis in RGB mean pixel intensity 18 hr: DMSO: 1.4±0.4; NAC: 3.7±0.8; HSM2, 10 μM: 24±6; DMR1, 10 μM: 32±3; HSM2+NAC: 3.2±1, DMR1+NAC: 2.5±1, n=3). Thus, ART analogs are minimally cytotoxic and induce cell death via apoptosis, but not necrosis, in cancer cells. In colon cancer cells, they cause cell death by increasing ROS. We are currently elucidating the mechanisms by which DMR1 and HSM2 cause apoptosis specifically in cancer cells. Synthesizing additional novel anti‐cancer ART analogs with minimal toxicity and understanding their mode of action will help develop potent therapeutic drugs to treat this disease that accounts for ~ 9 million deaths annually.
Support or Funding Information
NSF ‐ MRI: DBI‐1427937 to JS and Institutional Funds, Ben. U. to JS, DMR and MP
This is from the Experimental Biology 2018 Meeting. There is no full text article associated with this published in The FASEB Journal.
Photolysis of 1-(3-alkynoxy)-9,10-anthraquinones in deoxygenated methanol leads to moderate yields (35−45%) of 3-alkynals along with the unexpected formation of diacetals. Reaction of these ...3-alkynals with Grignard and Wittig reagents occurs nearly quantitatively without rearrangement to their 2,3-dienal isomers.
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