bluuebunny/crossref_metadata_2025_split huggingface dataset:
Targeting the Anaphase Promoting Complex to Prevent Lung Cancer Cell Growth
Authors: Wainscott Nathan W , Hoffman J. Mason , Taylor B. Frazier , Trent John O. , States J. Christopher 
Published: November 2023

BackgroundCurrent chemotherapeutic agents widely used for the treatment of lung cancer include mitotic inhibitors such as taxanes. Taxanes stabilize the mitotic spindle, maintaining the activity of the Spindle Assembly Checkpoint (SAC), thus preventing the metaphase to anaphase transition. Prolonged mitotic arrest results in cell death. Non‐small cell lung cancer (NSCLC) accounts for 80–85% of lung cancer cases. NSCLC may intrinsically be SAC deficient or acquire defects, making them resistant to these drugs. New treatments are needed for SAC deficient tumors. One potential new target for mitotic inhibition is the Anaphase Promoting Complex/Cyclosome (APC/C), an E3 ubiquitin ligase that drives the transition from metaphase to anaphase. The catalytic core of the APC/C consists of the subunits ANAPC2 and ANAPC11.HypothesisTargeting ANAPC2 to prevent binding of ANAPC11 to ANAPC2 will inhibit the Anaphase Promoting Complex/Cyclosome in lung cancer cells and induce cell death.MethodsFour lead compounds (3, 8, 10, and 11) were predicted by in silico analyses to bind the ANAPC2 subunit, and thus inhibit the APC/C by preventing binding of the catalytic subunit, ANAPC11. Compounds 8, 10 and 11 are 2‐amino‐propanol derivatives while compound 3 is a triazaspiro derivative. The compounds were tested for their ability to induce cytotoxicity in two human NSCLC cell lines (H460, A549) and a non‐transformed human bronchial epithelial cell line (HBEC). Cell viability following treatment was evaluated by morphological change by microscopy, Alamar Blue fluorescence assays, and colony forming assays.ResultsMorphological assessment of cell viability indicated a concentration‐ and time‐dependent increase in the number of mitotic cells with increasing concentration for compounds 3, 8 and 10 in all three cell lines, and evidence of cell death at higher concentrations in A549 and H460 cells. The Alamar Blue assays and colony forming assays confirmed that these compounds lead to a concentration–dependent decrease in A549 and H460 cell viability and proliferation. Compounds 3, 10 and 11 were more effective at inducing cell death than compound 8. Compound 11 was toxic at all concentrations tested in all three cell lines.ConclusionsThese results indicate that the APC/C could be a viable target for development of novel chemotherapeutic drugs to treat NSCLC.Support or Funding InformationASPET‐Summer Undergraduate Research Fellowship (NCW); KY Lung Cancer Research Program (JCS,); KY Science and Engineering Foundation (JCS); Kosair Charities Pediatric Oncology Research Program (JOT); NIH P30GM106396 (JOT)
Aggressive Breast Cancer Cell Line and Role of Translocator Protein
Authors: Mukherjee Shyamali , Sharan Chakradhari , Mitra Piyali , Gopal Srirupa Hari , Das Salil K 
Published: November 2023

We have already established that translocator protein (TSPO) is a breast cancer biomarker. It is also established that translocator protein shows low expression in ER positive breast cancer cells than ER negative breast cancer cells. It is also reported that African American (AA) women have higher grade breast tumors, higher incidence of triple negative breast cancers (TNBCs), higher proliferation indices, and node positive disease than their Caucasian (C) counterparts. Mortality rate is also higher in AA than C women. Based on these facts our hypothesis is that expression of TSPO is higher in TNBC than non‐TNBC and that higher mortality rate among breast cancer in AA women is mechanistically linked to higher incidence of TNBC cells. To test this hypothesis we have studied four TNBC (AA: HCC‐1806 and MDA‐MB‐468; C: BT‐549 and MDA‐MB‐231) and five non‐TNBC (C: MCF‐12A, MCF‐12F, MCF‐7, T47D and ZR‐75‐1) human cell line obtained from ATCC for TSPO expression. We analyzed the cultured cells for gene expression by RT‐PCR and protein by western blot. Both gene expression and protein levels were significantly higher in TNBC cells than non‐TNBC cells. We did not find any significant variation in either protein level or gene expression in TNBC cells of AA and C. TSPO Binding capacity (Bmax) was significantly higher in TNBC cell lines than normal cell lines. Proliferation of both TNBC and non‐TNBC cell lines is inhibited by PK 11195, antagonist to TSPO, in a dose‐dependent manner. However, inhibition efficiency was higher in TNBC cell line than non‐TNBC cell line.Support or Funding InformationMeTRC NIH grant 5U54MD007593
Structure‐Activity Relationship Studies of Oxypropanolamines: The Importance of Hydrophobicity About the Amine for Maintaining Anti‐Proliferative Effects in Lung Cancer Cells
Authors: Hoffman J. Mason , Wainscott Nathan W , Taylor B. Frazier , Burlison Joseph A. , Trent John O. , States J. Christopher 
Published: November 2023

BackgroundMitosis‐inhibiting chemotherapeutics are frequently used to treat cancer. Successful therapy is often hindered by drug‐resistant tumors, neurotoxicity, and limited supply. There is high demand for new drugs with diverse cellular targets. The anaphase promoting complex/cyclosome (APC/C) is a multi‐subunit E3 ubiquitin ligase that promotes cell cycle progression, especially mitosis, by marking key proteins for degradation. Thus, the APC/C offers great potential as an alternative target for anti‐mitotic drug development. The APC/C catalytic core consists of an ANAPC2/ANAPC11 dimer. Homology models of the ANAPC2/ANAPC11 subunits were analyzed in silico to identify lead compounds predicted to bind the ANPAC11‐binding domain of ANAPC2, and prevent ANAPC11 association and inhibit APC/C function. Among these leads were oxypropanolamine‐containing compounds. Preliminary studies indicate several of these compounds possess anti‐proliferative effects in various cancer cell lines. Based on this finding, a series of amine derivatives were synthesized to probe the effect of steric bulk and hydrophobicity in this region.ObjectiveWe aim to determine how modification of the amine substituent groups influences the ability to inhibit proliferation in several lung cancer cell lines in vitro.MethodsTo quantify the anti‐proliferative effect of compounds, AlamarBlue and colony forming assays were performed in A549 and H460 lung cancer cells and HBEC‐3KT immortalized non‐transformed bronchial epithelial cells.ResultsOutcomes of AlamarBlue and colony forming assays indicate that derivatives containing a secondary amine, but not tertiary, cause a decrease in cell proliferation. This anti‐proliferative effect is strengthened by increased steric bulk and hydrophobicity of the substituent amine.SummaryThese results provide structural insights to guide development of ANAPC2‐targeting compounds and are consistent with the hypothesis that the APC/C is a promising target for development of novel chemotherapeutics.Support or Funding InformationKY Lung Cancer Research Program (JCS,); KY Science and Engineering Foundation (JCS); ASPET Summer Undergraduate Research Fellowship (NCW); Kosair Charities Pediatric Oncology Research Program (JOT); NIH P30GM106396 (JOT)
Screening of a Small Flavonoid Library for Anti‐Cancer Activity Against Glioblastoma
Authors: Faler Ashton D. , Stang Christopher R.T. , Khupse Rahul S. , Schneider Ryan A. 
Published: November 2023

Flavonoids, including chalcones and flavanones, have been highly studied for their multitude of activities including antioxidant and anti‐cancer effects. We previously demonstrated that several of our chalcone derivatives had IC50 values in the low micromolar range in multiple distinct human cancer cell lines, but appeared to be most potent against A‐172 glioblastoma cells. The poor prognosis of glioblastoma necessitates further investigations to identify more effective treatment options. To determine if the activity of these chalcones was comparable in additional glioblastoma lines, our small library of flavonoids was screened against T98G and U87 MG glioblastoma cells. Our library consisted of eight flavonoid derivatives; six of which were chalcones and two flavanones. IC50 values, as measured by the CyQuant NF or XTT cell proliferation assay, were noted to be in the low micromolar range for three of these compounds; RK6, RK7, and RK15. RK6 had an IC50 of 9 μM and 20 μM against the T98G and U87 MG cell lines respectively. RK7 displayed IC50 values of 33 μM and 51 μM against the T98G and U87 MG cell lines respectively. RK15 was the most potent chalcone derivative with IC50 values of 3 μM in T98G cells and 5 μM in U87 MG cells. A Caspase‐Glo 3/7 assay was utilized to investigate the mechanism by which these compounds exhibited their cytotoxic effects. A significant increase in effector caspase activity was noted following treatments with RK6, RK7, and RK15 in both T98G and U87 MG cells. Furthermore, partial reversal of apoptosis by the pan‐caspase inhibitor, Z‐VAD‐FMK, suggests that caspases are involved in the observed cell death by RK6, RK7, and RK15 in these glioblastoma cell lines. Further investigations are ongoing with these chalcones to determine the molecular mechanisms of cell death in glioblastoma cells and to optimize the structure activity relationship.Support or Funding InformationThis work was supported in part by an Individual Summer Undergraduate Research Fellowship sponsored by ASPET for the first author.
Opposite angiogenic outcome of curcumin against ischemia and lung cancer: in silico, in vitro and in vivo studies
Authors: Li Xuejun 
Published: November 2023

Since many diseases are caused by complex factors, we try to apply the network pharmacology techniques to study the targets and networks of drugs for complex diseases. We investigated the angiogenic effects of curcumin on an ischemia and lung cancer model. To induce ischemia combined with lung cancer models, unilateral femoral arteries of C57BL/6 mice were disconnected on one side of the mouse and Lewis lung carcinoma (LLC) cells were xenografted on the opposite side. Angiogenic effects and underlying mechanisms associated with curcumin were investigated. Molecular target(s), signaling cascades and binding affinities were detected by Western blot, two‐dimensional gel electrophoresis (2‐DE), computer simulations and surface plasmon resonance (SPR) techniques. The results showed that Curcumin promoted post‐ischemic blood recirculation and suppressed lung cancer progression in inbred C57BL/6 mice via regulation of the HIF1α/mTOR/VEGF/VEGFR cascade oppositely. Inflammatory stimulation induced by neutrophil elastase (NE) promoted angiogenesis in lung cancer tissues, but these changes were reversed by curcumin through directly reducing NE secretion and stimulating α1‐antitrypsin (α1‐AT) and insulin receptor substrate‐1 (IRS‐1) production. Meanwhile, curcumin dose‐dependently influenced endothelial cells (EC) tube formation and chicken embryo chorioallantoic membrane (CAM) neovascularization. In conclusion, Curcumin had opposite effects on blood vessel regeneration under physiological and pathological angiogenesis, which was effected through negative or positive regulation of the HIF1α/mTOR/VEGF/VEGFR cascade. Curcumin had the promise as a new treatment modality for both ischemic conditions and lung cancer simultaneously in the clinic.Support or Funding InformationThis work was supported by the National Natural Science Foundation of China (No. 81673453, 81473235, 91129727, 81673486) and Research Fund from Ministry of Education of China (111 Projects No. B07001)
Macrophage‐derived extracellular vesicles transfer inflammasome components to endothelial cells and induces endothelial injury
Authors: Zuo Ping , Lin Xingguang , Li Xiang , Zhang Yang 
Published: November 2023

Background and aimsRecent studies have demonstrated an important role of Nlrp3 inflammasome in the development of endothelial dysfunction and injury under various pathological conditions. Extracellular vesicles (EVs) play an important role in cell‐cell communication and functional regulation. The role of macrophage‐derived EVs in endothelial homeostasis remains largely unknown. Here, we test whether or not the inflammasome components can be secreted into EVs derived from macrophages and then EVs transfer these proteins into endothelial cells and causes endothelial injury.MethodsExosomes and microvesicles, two major subtypes of EVs, were isolated from the cell culture medium from J774 macrophages by sequential ultracentrifugation. Exosomes were detected by Western blotting analysis of marker protein CD63. Inflammasome components Nlrp3, ASC, and caspase‐1 were detected in EVs by Western blotting. Endothelial injury was examined by analyzing the endothelial cell death using LDH assay and flow cytometry analysis.Result and conclusionIn macrophages, lipopolysaccharide (LPS) treatment dose‐dependently increased the expression of inflammasome component Nlrp3 and pro‐caspase‐1 in isolated exosomes or microvesicles. Co‐stimulation of macrophages with palmitate and LPS did not further enhance the secretion of Nlrp3 or pro‐caspase‐1 into these EVs. Incubation of isolated exosomes or microvesicles from LPS‐treated macrophages with naïve microvascular endothelial cells (MVECs) resulted in up‐regulation of the protein expression for Nlrp3 and pro‐caspase‐1 in these cells. LPS alone did not increase Nlrp3 expression in MVECs. These data indicate that the EV‐induced increases in inflammasome components in naïve MVECs may be primarily due to a protein transfer between EVs and the naïve MVECs but not up‐regulation of inflammasome gene expression. Lastly, incubation of naïve MVECs with microvesicles or exosomes derived from macrophages treated with LPS‐ or LPS plus palmitate induced significantly cell death in MVECs, whereas LPS alone had no effects on these naïve cells. Together, these results suggest that intercellular EV transfer of inflammasome components from macrophages to endothelial cell may exacerbate inflammasome‐mediated endothelial injury during vascular inflammation in obesity (Supported by NIH grants HL122769 and HL122937).Support or Funding InformationSupported by NIH grants HL122769 and HL122937
Psychedelics As A New Anti‐Inflammatory Therapeutic For Atherosclerosis
Authors: Nichols Charles D , Sebastian Melaine , Flanagan Thomas 
Published: November 2023

Background and ObjectiveWe previously discovered that serotonin 5‐HT2A receptor activation with psychedelics has potent anti‐inflammatory activity in both cell culture and whole animals, which indicated potent anti‐inflammatory effects in vascular tissues among others. More recently we found that the psychedelic (R)‐DOI potently prevents the development of allergic asthma in a mouse model. The effects of (R)‐DOI were found to not result from a generalized anti‐inflammatory process, but due to specific inflammatory pathways inhibition in both innate and Th2 cells. In this work, we have examined the therapeutic effects of the psychedelic (R)‐DOI in the ApoE −/− high‐fat model of atherosclerosis.MethodsOsmotic minipumps were used to deliver very low doses of (R)‐DOI to male ApoE −/− mice that were divided into four treatment groups [Saline, normal chow; (R)‐DOI, normal chow; Saline, hi‐fat diet; (R)‐DOI, hi‐fat diet]. After 16 weeks, mice were euthanized and tissues collected for analysisResultsCalculated steady state levels of ~0.0013 mg/kg (R)‐DOI resulted in a significant reduction of mRNA expression for inflammatory markers like Il6 in vascular tissue, reduced levels of glucose, and a reduction in circulating cholesterol in the high fat fed animals. Additional ongoing studies are examining arterial plaque size and heart pathology.SummaryExtremely low levels of the psychedelic (R)‐DOI were sufficient to significantly block the development of vascular inflammation, normalize glucose homeostasis, and prevent the increase in cholesterol associated with a hi‐fat ‘western’ high diet. Activation of serotonin 5‐HT2A receptor therefore represents a powerful new strategy to treat inflammatory‐related vascular disease.Support or Funding InformationNIH/NIGMS P30GM106392
NACHT, LRR and PYD Domains‐containing Protein 3 (NLRP3) Crucial Role of the Cavernous Tissue Tone Modulation in Mice
Authors: Fais Rafael Sobrano , Rodrigues Fernanda Luciano , Tostes Rita Cassia , Carneiro Fernando Silva 
Published: November 2023

INTRODUCTIONErectile function depends on the tone of the cavernous tissue (CT), and increased contraction and/or decreased relaxation responses contribute to erectile dysfunction. Erectile dysfunction and cardiovascular diseases share several mechanisms, like inflammation. NLRP3 is an intracellular receptor of the innate immune system, responsible for the release of pro‐inflammatory cytokines that induce inflammation and has been implicated in vascular dysfunction. However, the role of NLRP3 in the modulation of vascular and CT tonus is unknown.HYPOTHESISNLRP3 modulates the basal tonus of the CT.AIMTo determine the role of NLRP3 in the modulation of contraction and relaxation pathways in CT.METHODSCT strips were obtained from male C57BL/6 (WT), deficient to NLRP3 (NLRP3−/−) and caspase1/11 (caspase1/11−/−) mice, with 10 to 12 weeks. Reactivity of CT and western blot were performed to evaluate the function of the contraction and relaxation pathways.RESULTSThere was no difference in contraction to phenylephrine (PE) (Emax WT: 836,5 ± 93,3, n=5; NLRP3−/−: 813.0 ± 75,4, n=5; caspase1/11−/−: 925,4 ± 73.7, n=4), to electric field stimulation (EFS, frequencies from 0,2 to 32 Hz) and relaxation to acetylcholine (Ach) (Emax WT: 71,6 ± 7,84, n=5; NLRP3−/−: 75,9 ± 6,8 n=5; caspase1/11−/−: 91,25 ± 8,8, n=4) or non‐adrenergic non‐cholinergic (NANC) stimulation. However, L‐NAME and atropine incubation promoted a significant increase in the contraction to PE (Emax WT: 865,2 ± 76,6, n=5; NLRP3−/−: 1202,0 ± 85,9, n=5; caspase1/11−/−: 1160,0 ± 70, n=4) and EFS (mean WT: 329,2 ± 24,2, n=5; NLRP3−/−: 705,7 ± 130,1, n=5; caspase1/11−/−: 594,9 ± 73,1, n=4, at 16 Hz) and (mean WT: 554,0 ± 50,4, n=5; NLRP3−/−: 847,5 ± 157,1, n=5; caspase1/11−/−: 985,7 ± 124,8, n=4, at 32 Hz) Hz in the CT strips of NLRP3−/− and caspase1/11−/− and reduced relaxation to sodium nitroprusside in NLRP3−/− strips (Emax WT: 82,0 ± 2,75, n=5; NLRP3−/−: 63,9 ± 3,78 n=5; caspase1/11−/−: 91,5 ± 7,8, n=4). The eNOS activity was increased in the NLRP3−/− and caspase1/11−/− CT strips. In addition, there was an increase in tyrosine hydroxylase expression. Furthermore, there was an increase in the activity of MYPT‐1 in the NLRP3−/−, and PKCα in the caspase1/11−/− strips. The expression of the soluble guanylate cyclase alpha was reduced in NLRP3−/− and caspase1/11−/− CT strips.CONCLUSIONThe NLRP3 constitutively modulates the CT tonus, and its absence leads to a series of compensatory alterations in the contraction and relaxation pathways.Support or Funding InformationFAPESP, CNPQ, CAPES, CRID, FAEPA
Doxorubicin Dose Dependently Attenuates Cyclooxygenase‐2 in Female Human Coronary Vascular Smooth Muscle Cells
Authors: Tat Trinh , So Me , Vijayavel Nirmal , Raman Puneet , Dickinson Jared M , Angadi Siddartha S , Gonzales Rayna J 
Published: November 2023

Doxorubicin (Dox), an anthracycline‐based chemotherapeutic agent, is highly effective at reducing recurrence and mortality in breast cancer patients. However, this anti‐cancer drug elicits dose‐dependent cardiotoxicity. Multiple mechanisms have been implicated in its pathogenesis, one of which involves the development and progression of inflammation mediated by activation of the TLR4/NFkappaB pathway. However, we recently demonstrated that chronic in vivo Dox treatment reduced expression of the NFkappaB downstream proinflammatory mediator, cyclooxygenase‐2 (COX‐2), in cerebral and peripheral vasculatures isolated from ovariectomized female rats. Therefore, in the current study we tested whether Dox similarly alters vascular proinflammatory enzyme expression in the human vasculature and whether this response is dose dependent. We further investigated whether Dox alters upstream mediators of COX‐2 expression by assessing levels of expression and activation of NFkappaB and an additional downstream gene product, TLR4. Primary human coronary vascular smooth muscle cells of female origin were studied at passage 6 and treated with vehicle or Dox (0.3, 1.0, and 5 μM). Following drug treatment, cells were isolated, homogenized, and lysates (whole cell; nuclear and cytosolic) analyzed for NFkappaBp65, COX‐2, and TLR4 protein levels via immunoblotting. Our results demonstrate that while total NFkappaB levels were unaffected following Dox, enhanced translocation of NFkappaB to the nucleus was observed. However, even in the presence of Dox‐induced NFkappa translocation, basal expression of its downstream mediator, COX‐2, was dose dependently attenuated. Similar to COX‐2, a trend for a decrease in TLR4 levels was observed. Dox had no effect on cell morphology or density, as visualized via light microscopy. In conclusion, although others have suggested the involvement of NFkappaB activation during the development and progression of Dox‐induced inflammation in cardiac muscle, our studies demonstrate a possible novel action for the anticancer agent targeting important downstream mediators of inflammation in the female vasculature. Future studies will evaluate the impact of Dox on COX‐2 at the transcriptional or translational level to determine the mode of molecular regulation.Support or Funding InformationUniversity of Arizona Sarver Heart Center (RG and MS); Mayo Clinic Dept. of Medicine Extramural Chair Award in collaboration with Arizona State University (SA)
Expression and Characterization of Transcription factor EB in Coronary Arterial Smooth Muscle Cells
Authors: Bao Renyue , Zuo Ping , Li Xiang , Li Pin‐Lan , Zhang Yang 
Published: November 2023

BackgroundWe recently demonstrated that lysosome injury associated with abnormal sphingolipid metabolism causes impaired autophagic flux in coronary arterial smooth muscle cells (CASMCs). Under atherogenic stimulation, such impaired autophagic flux accelerates dedifferentiation of CASMCs to a synthetic phenotype. In the present study, we hypothesized that transcription factor EB (TFEB), a master regulator of lysosome biogenesis and autophagy signaling is involved in lysosome injury‐associated CASMC dedifferentiation.MethodsCASMCs were primarily cultured from mouse coronary arterial arteries and used in this study. The mRNA and protein expression of TFEB in CASMCs were detected by real time quantitative PCR and Western blot, respectively. The activation of TFEB was determined by nuclear translocation of TFEB using immunofluorescence microscopy.Results and conclusionLysosome functional inhibitors increased TFEB expression and activation in CASMCs, which was associated with up‐regulation of lysosome and autophagy markers. These results indicate that TFEB are expressed and functioning in CASMCs. Genetic deficiency of acid sphingomyelinase (ASM) results in enhanced cell proliferation and migration of CASMCs upon platelet‐derived growth factor (PDGF) stimulation. Such phenotypic changes in ASM‐deficient CAMSCs were associated with decreased nuclear translocation of TFEB compared to that in wild‐type cells. These results suggest that ASM/TFEB signaling axis is involved in the regulation of cell cycle in CASMCs implicating a novel role of TFEB in vascular smooth muscle plasticity.Support or Funding InformationSupported by NIH grants HL057244, HL122769 and HL122937