bluuebunny/crossref_metadata_2025_split huggingface dataset:
Biochemical and cellular monitoring of the activity of the ecto‐5′‐nucleotidase (CD73), a key cancer modulator using HTS‐formatted Bioluminescent Technology
Authors: Goueli Said 
Published: November 2023

The ecto‐5′‐nucleotidase (CD73) is glycosyl phosphatidylinositol (GPI) anchored cell surface protein that is involved in switching on adenosinergic signaling. Its enzymatic and nonenzymatic activities (via its interaction with extracellular matrix components) are involved in cancer associated processes and not completely independent of each other. It catalyzes the hydrolysis of AMP into adenosine and phosphate, where adenosine plays important role in tumor immune escape. It is overexpressed in many types of cancer cell lines and patient's biopsies including breast, colon, ovarian, gastric, ovarian, etc. In addition to being important as clinical and prognostic marker in cancer patients, overexpression of CD73 is associated with resistance to antitumor agents, and inhibition of CD73 activity or knocking it down by SiRNA reversed chemoresistant phenotype of glioblastoma multiforme cells. Because of the significant role of CD73, it is of importance to develop an assay that monitor the activity of CD73 in order to develop modulators of its activity. Towards this goal, we have developed a bioluminescent assay to monitor the activity of the enzyme in biochemically pure as well cellular anchored enzyme forms. The assay is homogenous and formatted for HTS screening research, very sensitive, and robust as indicated by the high Z’. We also show that the use of inhibitors such as adenosine 5′‐(a‐b‐methylene) diphosphate (APCP) generates data with biochemically pure enzyme similar to the cellular bound form. We also demonstrate that cells that are enriched in CD73 can be easily identified from those that have low CD73. Thus, the current assay is robust, sensitive, and easy to use making it an ideal assay for HTS screening for new modulators of CD73 and generation of potential novel cancer therapeutics.Support or Funding InformationPromega Corp.
Enhancing Molecular Targets in Cancer through the use of Very Low Level Radiation
Authors: Branly Rolando , Oseni Saheed O , Kumi‐Diaka James , Tulp Orien 
Published: November 2023

Some recent studies show increased levels of apoptosis in cancer cell cultures utilized for phytonutrient effect studies when a low dose of X‐rays is initially given. We consider some of the effects of electromagnetic radiation in the production of intra‐cellular acoustical waves and the implications for altered probabilities of Biochemical Pathways. Monte Carlo simulation methods are employed with initial coarse grid models based on in vitro studies using LNCaP and PC3 cancer cell lines in combination therapy studies with X‐rays. Very Low Dose Radiation with X‐rays appears to enhance apoptosis mechanisms in the laboratory. The models presented serve as useful tools for temporal effect studies.Support or Funding InformationWe are thankful and acknowledge institutional support
Overcoming Chemoresistance in Pancreatic Cancer by Targeting the Anti‐Apoptotic Mcl‐1 Protein
Authors: Badejo Adeleke , McCreary Jordan , True Heather , Weisenburger Alexis , Davis Madison , Malhotra Ashim 
Published: November 2023

Pancreatic cancer is highly aggressive with high mortality and poor 5‐year prognosis. Treatment is challenging due to substantial chemoresistance. The ability of cancers to limit the apoptotic response to drug treatment emphasizes the urgent need to develop drugs that can overcome chemoresistance. Overexpression of the anti‐apoptotic Mcl‐1 protein is associated with the development of chemoresistance in some cancers. Using the human pancreatic cancer cell line MIA PaCa‐2, we investigated the effect of the drug raloxifene on the regulation of Mcl‐1 in this model. The rationale for the use of raloxifene was the observation that it can cause apoptosis in cancer cell lines. We treated MIA PaCa‐2 cells with 0.05, 0.5, 5, and 20 μM raloxifene for 72 hours, followed by the preparation of whole cell lysates, which we used for Western Blot analysis. Our data show that: 1) raloxifene disrupted the protein‐protein interaction between the anti‐apoptotic Mcl‐1 and the pro‐apoptotic Bax, suggesting Bax homodimerization at the mitochondria, and possible activation of the apoptotic or autophagy response. To explain this, we investigated 1) the effect of raloxifene on the protein expression of the alternative splice variants of Mcl‐1, Mcl‐1L or Mcl‐1S; and, 2) whether raloxifene‐induced protein mistrafficking of Mcl‐1. Our data showed no changes in Mcl‐1L and Mcl‐1S, although levels decreased in the Bax/Mcl‐1 complex, as indicated by co‐immunoprecipitation experiments. 4) Additionally, we investigated the effect of raloxifene on global transcription by screening its effect on NFκB, Nrf‐2, eif‐2α, phospho‐cdc2, and cyclin proteins. While raloxifene treatment did not induce any of these transcription factors, at high doses it induced the expression of SRSF‐5, but had no effect on the expression of SRSF‐1, which are proteins that regulate the differential splicing of the Mcl‐1 gene. It also induced cyclin proteins. To further investigate the specific role of raloxifene in the turnover of Mcl‐1 protein, we pre‐incubated MIA PaCa‐2 cells with 10 μM cycloheximide, a global translation inhibitor. Our data showed a reduction in Mcl‐1 in raloxifene treated samples compared to control, suggesting a role for estrogen receptor β in the de novo expression of Mcl‐1. 6) Additionally, raloxifene treatment induced the autophagy markers Beclin‐1 and LCIII‐B, indicating a correlation between the differential regulation of Mcl‐1 and autophagy. Taken together, these data suggest that the main mechanism for the action of raloxifene is competitive displacement of Mcl‐1 from the Bax/Mcl‐1 protein complex.Support or Funding InformationThis research was supported in part by Pacific University Research Incentive Grant and Collins Medical Trust grant to AM.
Pathomimetic Avatars for Live‐Cell Imaging of Tumor Proteolysis and Drug Screening
Authors: Sloane Bonnie F. , Ji Kyungmin , Chalasani Anita , Moin Kamiar , Mattingly Raymond R , Xu Yong 
Published: November 2023

To define protease‐related druggable pathways involved in malignant progression of cancer, our laboratory has pioneered novel techniques for functional live‐cell imaging of protease activity, initially concentrating on pathomimetic avatars for breast cancer. We analyze proteolysis in the context of proliferation and formation of structures by breast cells (benign and cancerous) in 3‐D cultures over time (4D). In order to recapitulate the cellular composition and architecture of breast tissue, we include other tumor‐associated cells (e.g., fibroblasts, myoepithelial cells, lymphatic endothelial cells). We also model non‐cellular aspects of the tumor microenvironment, e.g., an acidic pericellular pH. Use of these pathomimetic avatars in concert with various types of imaging probes has allowed us to image, quantify and follow the dynamics of proteolysis in the tumor microenvironment and to test interventions that impact directly or indirectly on proteolytic pathways. To facilitate use of the pathomimetic avatars for drug screening, we have designed culture chambers with multiple wells that are either individual or connected by a bridge to allow cells to migrate between wells. Optical glass microscope slides underneath an acrylic plate allow the cultures to be imaged with an inverted microscope. Fluid ports in the acrylic plate are at a level above the 3D cultures to allow introduction of culture media and test agents such as drugs into the wells and the harvesting of media conditioned by the cultures for immunochemical and biochemical analyses. Covers contain integrated gas exchange ports and sensors to monitor oxygen levels, pH and temperature over the extended time periods in culture and to insure maintenance of such experimental conditions as hypoxia and/or a pericellular acidic pH. We predict that the pathomimetic avatars will accelerate identification of druggable pathways, screening of drug and natural product libraries and the entry of validated drugs or natural products into clinical trials.Support or Funding InformationSupported in part by National Institute of Health R21 CA 131990 (BFS) and the President's Research Enhancement Fund of Wayne State University (BFS and YX). Live‐cell imaging was performed in the Microscopy, Imaging and Cytometry Resources Core of Wayne State University, which was supported in part by National Institutes of Health P30 CA22453 to the Karmanos Cancer Institute and by the Perinatology Research Branch of the National Institute of Child Health and Human Development.
Transglutaminase 2 Plays a Critical, but Context‐Specific Role in Glioblastoma Proliferation
Authors: Monteagudo Alina , Gundemir Soner , Akbar Abdullah , Keillor Jeffrey , Johnson Gail 
Published: November 2023

Glioblastoma multiforme (GBM) is the most aggressive and common form of adult primary brain tumors. GBMs are characterized by resistance to chemotherapy and radiation, and exhibit a high propensity for reoccurrence. Therefore, it is imperative to define molecular mechanisms that contribute to GBM tumor survival and proliferation. One possible protein that may contribute to GBM proliferation is the multifunctional protein transglutaminase 2 (TG2). TG2 has been shown to play a critical role in proliferation in other types of cancers, and therefore we investigated its role in GBMs. Given the heterogeneity of GBM tumors, we used various GBM cell lines as well as non‐serum containing GBM neurospheres to determine if the role of TG2 in GBMs was broadly applicable, or just relevant to certain populations. Additionally, the neurospheres used represent three different GBM subtypes: mesenchymal, proneural and classical. By the use of novel inhibitors and genetic manipulations of TG2, results show that in most, but not all, cell lines TG2 plays a critical role in the proliferative process of GBM cells. Given the heterogeneity of GBM tumors, it is not surprising that TG2 facilitates the proliferative process in only certain GBM subtypes. Nonetheless, as the understanding of signaling mechanisms that contribute to GBM proliferation increases, TG2 inhibitors may in the future be effective therapeutic agents for certain GBMs, as determined by specific genetic determinants.Support or Funding InformationNIH RO1 NS065825
Pharmacological inhibition of soluble epoxide hydrolase ameliorates experimental acute pancreatitis in mice
Authors: Bettaieb Ahmed , Chahed Samah , Bachaalany Santana , Griffey Stephen , Hammock Bruce D. , Haj Fawaz G. 
Published: November 2023

BackgroundAcute pancreatitis (AP) is an inflammatory disease, and is one of the most common gastrointestinal disorders worldwide. Soluble epoxide hydrolase (sEH; encoded by Ephx2) deficiency and pharmacological inhibition have beneficial effects in inflammatory diseases. Ephx2 whole‐body deficiency mitigates experimental AP in mice but the suitability of sEH pharmacological inhibition for treating AP remains to be determined.ResultsWe investigated the effects of sEH pharmacological inhibition on cerulein‐ and arginine‐induced AP using the selective sEH inhibitor 1‐trifluoromethoxyphenyl‐3‐(1‐propionylpiperidin‐4‐yl) urea (TPPU) which was administered before and after induction of pancreatitis. Serum amylase and lipase levels were lower in TPPU‐treated mice compared with controls. In addition, pancreatic mRNA of the inflammatory cytokines TNFα, IL‐1β and IL‐6 was reduced in TPPU‐treated mice. Moreover, sEH pharmacological inhibition before and after induction of pancreatitis was associated with decreased cerulein‐ and arginine‐induced NF‐κB inflammatory response and endoplasmic reticulum stress.ConclusionssEH pharmacological inhibition before and after induction of pancreatitis mitigated cerulein‐ and arginine‐induced AP. This work suggests that sEH pharmacological inhibition may be of value in the therapy of acute pancreatitis.Support or Funding InformationThis work was supported by research grants from the JDRF (1‐2009‐337) and NIH (R56DK084317, R01DK090492 and RO1DK095359) to F.G.H.; K99DK100736 to A.B. and by NIH ES02710 and Superfund Basic Research Program Grant P42 ES04699 from NIEHS, and HL059699 Grant from NHLBI and U24DK097154 (to B.D.H.).
Pharmacokinetics of MAT9001, an Omega‐3 Fatty Acid Medication, Compared with Eicosapentaenoic Acid Ethyl Esters in Hypertriglyceridemic Subjects
Authors: Maki Kevin C , Keane William F , Bouhajib Mohammed , Pop Radu , Bobotas George 
Published: November 2023

SynopsisMAT9001 (Matinas BioPharma Inc., Bedminster, NJ) is a novel investigational omega‐3 fatty acid (OM3) medication containing predominantly eicosapentaenoic (EPA) and docosapentaenoic (DPA) acids under development as an adjunct to diet for the treatment of severe hypertriglyceridemia (≥500 mg/dL).PurposeThis randomized, crossover trial compared the bioavailability of MAT9001 and an OM3 EPA ethyl ester (EPA‐EE) agent (Vascepa; Amarin Pharmaceuticals, Bedminster, NJ) after single‐ and multiple‐dosing under fed (low‐fat) conditions in men and women with fasting triglycerides (TG) 200–400 mg/dL at screening without lipid‐altering therapy, or TG 200–350 mg/dL if on stable‐dose statin monotherapy.MethodsForty‐two subjects received 4 g/day MAT9001 and EPA‐EE in random order, while housed in a clinical research unit for 14‐day treatment periods, separated by ≥35 days of wash‐out. Baseline values were averaged from pre‐dose samples drawn on days −1 and 0. On‐treatment values were determined from plasma samples drawn on days 0 (single‐dose) and 13 (multiple‐dose). Total EPA, DPA, docosahexaenoic (DHA) and heneicosapentaenoic (HPA) acids were measured in plasma by a liquid chromatography–tandem mass spectrometry method using isotopically substituted OM3 compounds as internal standards. Total OM3 was calculated as the sum of molar concentrations of total EPA, DPA, DHA, and HPA for each time point. Systemic exposure (area under the curve [AUC]) and peak exposure (Cmax) were calculated based on the measured and baseline‐adjusted concentrations. The log‐transformed AUC and Cmax were analyzed using a statistical model corresponding to a 2‐way crossover design.ResultsAfter single dose administration, MAT9001 was superior to EPA‐EE for systemic bioavailability of EPA, DPA and total OM3, with baseline‐adjusted AUC and maximal concentrations approximately 8‐fold higher. Following multiple dose administration, the bioavailability over the dosing interval (AUCtau) and the steady state Cmax were approximately 6‐fold higher for MAT9001 compared to EPA‐EE for both total OM3 and EPA. At steady state conditions, AUCtau and maximal plasma concentration for DPA were 4‐fold and 5‐fold higher, respectively, for MAT9001 as compared to EPA‐EE. Each of these comparisons achieved a statistical significance with p<0.0001.ConclusionThese results suggest that MAT‐9001, comprised predominantly of EPA and DPA, dosed with low‐fat meals exhibits significantly increased bioavailability after both single and multiple dose administrations, as compared to EPA‐EE. This significantly higher systemic absorption of fatty acids from MAT9001 relative to EPA‐EE has also been shown to produce greater TG reduction. Further research is underway to investigate the pharmacodynamic effects of MAT9001 and to evaluate possible implications for cardiovascular risk.Support or Funding InformationFunded by Matinas BioPharma Inc., Bedminster, NJ
The Cholesterol‐Lowering Effect of Statins is Potentiated by Whole Grains Intake. The Polish Norwegian Study (PONS)
Authors: Vaidean Georgeta , Manczuk Marta , Vansal Sandeep , Griffith Jacqueline 
Published: November 2023

IntroductionAdequate whole grains (WG) dietary intake is advocated for cardiovascular health. Statins represent the cornerstone of medical control of hypercholesterolemia. Whether there is an interaction between WG intake and statins, has not been fully explored in ‘real life’ settings. Our objective was to assess the interaction between statin use and whole grains fiber intake on plasma lipids.MethodsWe used cross‐sectional data of 12754 participants, ages 45 to 64 years, in an ongoing cohort study in Poland. Statin use during the last 30 days was recorded by trained nurses. WG intake was estimated from a food frequency questionnaire assessing participants’ habitual food intake in the past year. Plasma total, LDL, HDL cholesterol, triglycerides and glucose were measured in a fasting state. Multivariable linear and logistic regression models were used for analyses.ResultsThere were 1728 (13.55%) statin users. High WG grain intake (at or above the recommended levels of ≥3 oz/d) was achieved by 2835 (22.23%) participants. Those taking statins, had lower mean (95%CI) plasma lipid levels if they had a high WG intake, even after adjusting for age, sex, education, total caloric intake, various dietary nutrients intake, physical activity, smoking, glucose levels, BMI and prevalent CVD: Cholesterol: 185.13 mg/dL (181.65–188.60) vs. 190.14 mg/dL (188.02–192.27); non‐HDL cholesterol: 127.26 mg/dL (123.83–130.71) vs. 133.02 mg/dL (130.92–135.12); LDL: 104.42 mg/dL (101.37–107.45) vs. 108.54 mg/dL(106.69–110.40). High vs. low WG intake did not have an effect among those not taking statins, nor on triglyceride or HDL levels. In fully adjusted models, statin users with a high WG intake had 33% lower odds of having a cholesterol level ≥ 240 mg/dL and 38% lower odds of having a non‐HDL cholesterol ≥130 mg/dL, compared to statin users with low WG intake.ConclusionWe found a potentiation of effect of statins by whole grain dietary intake on plasma lipid levels. Given that the current paradigm on controlling plasma lipids is that “lower is better” and that the uptitration of statins can be potentially toxic, the interaction with the dietary intake of WG has potential clinical significance.Support or Funding InformationFINANCIAL DISCLOSURES: The data collection was supported by a grant from the Polish‐Norwegian Research Fund (PNRF‐228‐AI‐1/07).
In Vivo Analysis of Cilia Function and Connections to Renal Disease and Physiology
Authors: Yoder Bradley K. 
Published: November 2023

Many cystic kidney diseases are caused by defects in proteins required for cilia assembly or activity. It has been proposed that cilia regulate cell proliferation, oriented cell division, responses to tissue injury, and as mechanosensors. The cilium is thought to mediate its effects by regulating the activity of several signaling proteins or channels including Polycystins 1 and 2, mTor, Stat3/6, Hedgehog, and purinergic receptors. Importantly, these signaling pathways are frequently altered when cilia are absent from cells and in cystic renal epithelium. However, the exact mechanisms connecting cilia dysfunction to cyst formation remains largely unknown. Recent data have raised concerns regarding the mechanosensory model for cystogenesis. Despite data from cell culture studies showing fluid flow mediated deflection of cilia results in changes in intracellular calcium levels, this has not been observed in vivo. Further, loss of cilia function in juvenile mice results in rapid cyst formation, but in adult mice cyst do not form for 6 to 8 months after cilia function is disrupted. This is not consistent with simple defects in mechanosensation. Our studies are trying to assess the functions of the cilium in vivo. We are addressing whether it has mechanosensory activities under different physiological conditions and how ciliary defects contribute to cyst formation. We are using conditional mutations in genes required for cilia formation and function along with a series of biosensors to analyze cilia function in live tissues. To begin addressing the in vivo roles of the cilium, we generated a CiliaGFP mouse allowing us to observe cilia in whole tissues of live animals. We found under normal conditions that cilia are consistently bent in the direction of fluid flow along the renal tubule. However, by altering physiological conditions, we show cilia oscillate or even reverse orientation, indicating that the behavior of cilia is dynamic and influenced by other systems in the body. We are determining how cilia react to changes in heart rate, blood pressure, glomerular filtration rates, restricted renal blood flow, during cyst formation, and after kidney injury. We will evaluate how changes in cilia behavior under different conditions influence Ca2+, Stat6, and mTor signaling activity and whether cilia morphology, length, or behavior is altered during cyst formation. Importantly, all of these studies will be performed in the kidney of a live mouse. Results will provide novel insights into how the cilium functions in the kidney and how defects in cilia are possibly involved in diseases.Support or Funding InformationP30DK074038R01DK065655
Preclinical Evaluation of the Phosphodiesterase‐4 Inhibitor, Rolipram, in an Infant Model of Sepsis‐Induced Cardiorenal Syndrome
Authors: Sims Clark R , Singh Sharda P , Mayeux Philip R 
Published: November 2023

Multi‐organ failure is a frequent complication of infant sepsis. Sepsis‐induced cardiorenal syndrome (type 5), defined as development of acute cardiac dysfunction and acute kidney injury, is a frequent complication of infant sepsis that dramatically increases mortality. Septic shock in adults typically presents as a hyperdynamic state with high cardiac output and low systemic vascular resistance. However, most infants with septic shock show low cardiac output and high systemic vascular resistance (cold shock). To define the apparently unique pathogenic mechanisms contributing to renal and cardiac failure during septic shock in infants, and identify targeted therapies to reverse them, we developed a rat pup (17–19 days old) model of infant cardiorenal syndrome induced by cecal ligation and puncture (CLP), a model of infant sepsis that replicates the key cardio‐renal abnormalities. Rolipram is a phosphodiesterase‐4 inhibitor shown to reduce renal vascular resistance and improve microvascular perfusion in an adult mouse model of sepsis. The therapeutic potential of rolipram was evaluated using intravital video microscopy (IVVM) and echocardiography at specific time points following CLP in our rat pup model. A dose‐finding study was completed using IVVM at 6h following CLP that showed that a dose of 0.1 mg/kg, administered at the time of CLP, protected the pups from sepsis‐induced renal hypoperfusion. This dose was then used in further studies. At 18h following CLP, the single dose of rolipram administered at the time of CLP still prevented renal hypoperfusion and also prevented sepsis‐induced decline in stroke volume, left ventricular velocity, fractional shortening, ejection fraction, and most importantly, cardiac output as measured by echocardiography. Rolipram was also able to prevent the increase in blood urea nitrogen, which is used clinically to assess renal function. The protection by rolipram on perfusion and cardiac output indicate that rolipram may be therapeutically beneficial in infant cardiorenal syndrome.Support or Funding InformationCRS supported by T32 GM106999, F31DK104533, AHA PRE20450050; PRM supported by AHA 15GRNT2508025 and UAMS Medical Research Foundation.