bluuebunny/crossref_metadata_2025_split huggingface dataset:
Rolapitant is not a mechanism‐based inactivator or tight‐binding inhibitor of CYP2D6 in vitro
Authors: Glass Sarah , Furge Laura Lowe 
Published: November 2023

Rolapitant is a NK1 receptor antagonist that was approved by the FDA in September 2015 as a treatment for chemotherapy‐induced nausea and vomiting. In vivo, rolapitant is primarily metabolized by CYP3A4, a member of the P450 superfamily that is responsible for the metabolism of 45% of pharmaceuticals. In clinical studies, it was shown that rolapitant inhibits the activity of another major drug metabolizing P450, CYP2D6, for at least a week after just one dose. Because the inhibition time is long, we hypothesized that rolapitant might be a mechanism‐based inactivator that irreversibly inhibits CYP2D6. Such inactivation could lead to adverse drug‐drug events. Interactions of rolapitant with both CYP3A4 and CYP2D6 were investigated utilizing AutoDock molecular modeling, spectral binding analysis, mass spectrometry metabolite studies, and in vitro inhibition assays. The results were compared with a known mechanism‐based inactivator, SCH 66712, and a tight‐binding inhibitor, quinidine. From this study, it appears that rolapitant is neither a mechanism‐based inactivator nor a tight‐binding inhibitor. Because of this, the in vivo inhibitory potency could be due to the drug's long mean elimination half‐life (7 days). Understanding the mechanism of inhibition by rolapitant has clinical relevance in preventing drug‐drug interactions caused by CYP2D6 inhibition during polypharmacy often occurring during cancer treatment.Support or Funding InformationNIH 1R15‐GM086767‐02
High‐Throughput Virtual Screening to Identify Novel Inhibitors of 6‐Phosphogluconate Dehydrogenase in Plasmodium vivax
Authors: Weber Nicole Brooke , Trieu Carolyn , Beckham Joshua 
Published: November 2023

The discovery of novel drugs is warranted in the face of rising resistance developed by Plasmodium vivax parasites to a number of malaria medicines. Therefore, the principal aim of this research study is to discover unknown inhibitors for 6‐phosphogluconate dehydrogenase (Pv6PG) in the protozoan parasite P. vivax. First, the full Pv6PG coding DNA sequence was synthesized with overlapping oligo‐primers, amplified via Polymerase Chain Reaction (PCR) methods, and cloned into a pET expression vector. Following transformation into E. coli BL21(DE3), the protein was expressed and then purified with Nickel Affinity Chromatography and size exclusion chromatography. Differential Scanning Fluorimetry (DSF) assays were performed to assess the ligand‐binding interactions through shifts in the denaturation temperature. One best ranking compound from the Maybridge HitFinder library was docked using GOLD and scored 74.19, but did not exhibit a significant shift of the melting curve. To identify potential inhibitors, virtual high‐throughput screening was used. First, a homology model of the target Pv6PG was created using ICM. This docking algorithm made binding mode predictions of a ChemBridge library of over 100,000 ligands. The best ranking compounds will next be tested in wet lab using spectrophotometric assays to determine if they any have significant ability to inhibit the enzymatic functionality of Pv6PG. Successful isolation of compounds that inhibit this protein will play a role in hindering the vital biochemical pathways in the malaria‐causing organism Plasmodium vivax and could lead to greater progress in the pursuit of a novel treatment for this infectious disease.Support or Funding InformationHHMI
Landscape of the Regulatory Elements for Lysine 2‐Hydroxyisobutyrylation Pathway
Authors: Huang He , Luo Zhouqing , Qi Shankang , Huang Jing , Dai Lunzhi , Dai Junbiao , Zhao Yingming 
Published: November 2023

Short‐chain fatty acids and their corresponding acyl‐CoAs sit at the crossroads of multiple metabolic pathways and play important roles in diverse cellular processes. They are also precursors for protein posttranslational lysine acylation pathways. A noteworthy example is the newly identified lysine 2‐hydroxyisobutyrylation (Khib) that is derived from 2‐hydroxyisobutyrate and 2‐hydroxyisobutyryl‐CoA. Histone Khib has shown association with active gene expression in spermatogenic cells. However, key elements for regulating this physiology‐relevant pathway remain unknown, hindering characterization of mechanisms by which this modification exerts its biological functions. Here we report the first global profiling of Khib proteome in mammalian cells, identifying 6548 Khib sites on 1725 substrate proteins. We showed that Esa1p in budding yeast and its homologue TIP60 in human could add Khib both in vitro and in vivo, respectively. We further confirmed TIP60's in vivo activity for Khib by identifying 497 potential Tip60‐regulated Khib substrate sites. In addition, we identified HDAC3 as the major enzyme to remove Khib. Our study thus discovered both the “writer” and “eraser” for histone Khib marks, and major Khib protein substrates. These results not only illustrate the landscape of this new lysine acylation pathway, but also open new avenues for studying diverse functions of cellular metabolites associated with this pathway.Support or Funding InformationOur work was supported by National Institutes of Health (NIH) award GM105933, DK107868, and GM115961 (Z.Y.). This work was also supported by Research Fund for the Doctoral Program of Higher Education of China 20120002110022 (J.D.)
Targeting Integrin‐linked Kinase with Small‐interfering RNA Suppresses Invasion and Metastasis in Cisplatin‐Resistant Ovarian Cancer
Authors: Reyes‐González Jeyshka M. , Báez Perla , Valiyeva Fatma , Vivas‐Mejía Pablo E. 
Published: November 2023

Ovarian cancer has the highest mortality rate among gynecological cancers. It continues to be characterized by metastatic bulky disease burden, which presents a major clinical challenge. Advanced disease is often treated with surgery, followed by platinum‐based chemotherapy such as cisplatin. Unfortunately, the majority of ovarian cancer patients become resistant to cisplatin, and relapse. Therefore, the identification of targets that link metastasis and chemoresistance is urgently needed to develop novel and effective strategies for ovarian cancer treatment. Integrin‐linked kinase (ILK), a serine/threonine kinase and adaptor protein, has been shown to play crucial roles in cancer progression, epithelial‐mesenchymal transition (EMT), angiogenesis, and metastasis. Our previous findings indicate that cisplatin‐resistant ovarian cancer cells (A2780CP20 and OV90CIS) express higher levels of phosphorylated ILK (p‐ILK) protein when compared to their parental counterparts (A2780 and OV90, respectively). Targeting ILK with small‐interfering RNAs (siRNAs) significantly reduced the invasive ability of A2780CP20 and OV90CIS cells in vitro (85.5% reduction; p<0.0001 and 61.6% reduction; p<0.0001, respectively). Therefore, to investigate the effects of ILK silencing in vivo, Control‐siRNA or ILK‐siRNA were encapsulated into DOPC‐DSPE‐PEG‐2000‐nanoliposomes. First, to assess the efficacy of the nanoliposomal siRNAs to silence their target in vivo, cisplatin‐resistant ovarian cancer cells (A2780CP20) were intraperitoneally injected into nude mice. Three weeks after tumor implantation, mice were treated twice with either Control‐siRNA or ILK‐siRNA nanoliposomes. ILK protein levels were significantly downregulated by ILK‐siRNA compared to Control‐siRNA, as evident in Western blot analysis (40% reduction; p<0.05). Next, to assess the involvement of ILK in metastasis in vivo, A2780CP20 cells were intraperitoneally injected into nude mice. Seven days after cell inoculation, mice received a weekly dose of either Control‐siRNA or ILK‐siRNA nanoliposomes. At the end of the treatment period, ILK‐siRNA significantly reduced the number of tumor nodules compared to Control‐siRNA (48% reduction; p<0.01). No obvious toxicity during therapy was observed. These results indicate that silencing ILK is a promising therapeutic approach for advanced and chemoresistant ovarian cancer. Further studies are required to investigate the biologic and molecular roles of ILK downstream targets in cisplatin‐resistant ovarian cancer.Support or Funding InformationThis project was supported in part by the RTRN (U54MD008149) grant to PEVM, institutional seed funds from the University of Puerto Rico, Comprehensive Cancer Center (PEVM), and National Institute of Health, Minority Biomedical Research Support (MBRS) RISE Grant Number R25‐GM061838 (JMRG).
Evaluation of the High Mobility Group A1 proteins (HMGA1) as a key mediate in the anticancer activity of ‐EF24.
Authors: Diaz Madelyn Nicole , Travis Lauren , Barber Sommer , Sumter Takita Felder 
Published: November 2023

The architectural chromatin binding proteins High Mobility Group A1 (HMGA1) are some of the most overexpressed proteins in malignant cancers and induce neoplastic transformation. The protein is increased as the last step of a prominent colon cancer progression pathway and mediates drug resistance, therefore correlating with a poor patient prognosis. HMGA1‐‐mediated chemoresistance results from a self‐protective process called cellular senescence. Analogs of the antioxidant, curcumin, when used in combination with traditional chemotherapeutic agents, are useful treatment options for drug resistant tumors. This study had two specific aims. The first being to investigate how colon cancer cells HCT116 respond to treatment with EF24. The second aim was to evaluate how hmga1 expression changed as a result of treatment with EF24. Our preliminary findings showed that cell viability decreased after 24‐hour treatment with low‐dose EF24, as indicated by an MTS assay, with notable discrepancies between cells that underwent a pulsed treatment regimen versus those that underwent continuous treatment. Furthermore, we demonstrated that cells exhibited fragmented DNA when treated with low‐dose EF24, which is characteristic of apoptotic cells. At higher, pulsed doses, senescence activity increased indicating the induction of a senescence pathway. Lastly, gene expression studies indicated that hmga1 was significantly down regulated in cells treated with continuous, low‐‐dose EF24. Further investigation of this pathway could lead to decreased toxicity and increased viability of combination cancer therapies.Support or Funding InformationThe research support is provided by the National Center for Research Resources (5 P20 RR16461), the National Institute of General Medical Sciences (8 P20 GM103499) from the National Institutes of Health, and the Winthrop McNair Scholars program.
AraL Phosphatase From Bacillus subtilis, a Member of the HAD Superfamily
Authors: Martin Cassandra , Armeli Jordan , Madaio Michael , Hill Jacqueline , O'Handley Suzanne 
Published: November 2023

The HAD (Haloacid Dehalogenase) superfamily is a diverse superfamily with a majority of the enzymes being phosphatases. One family of the HAD superfamily is the nitrophenyl phosphatase family, so named because the first family members were yeast enzymes found only to hydrolyze p‐nitrophenyl phosphate. Since that time, a number of enzymes with a variety of activities have been identified, including an enzyme from B. subtilis, AraL, originally designated as a sugar phosphatase. The AraL gene has been subcloned to incorporate a HisTag for nickel affinity chromatography, and the enzyme has been expressed, purified and characterized. Interestingly, it was found that although AraL does cleave phosphate from some sugar phosphates (ribose 5‐phosphate, ribulose 5‐phosphate, and arabinose 5‐phosphate), intermediates of glycolysis (glyceraldehyde 3‐phosphate, phosphoenolpyruvate, and dihydroxyacetone phosphate) are much better substrates for the enzyme. Since glycolytic intermediates are better substrates, AraL's role may be to regulate the glycolysis and gluconeogenesis pathways when arabinose is the carbon source. Arabinose is metabolized into intermediates that enter in the middle of the glycolysis pathway. The fact that intermediates after the arabinose point of entry are substrates, and intermediates before the arabinose point of entry are not substrates, suggests that AraL may inhibit glycolysis and activate gluconeogenesis.Support or Funding InformationNIH AREA GRANT
The Use of Next Generation DNA Sequence to Determine Fungal Diversity on Potato Tubers Treated with 1,4‐dimethylnaphthalene
Authors: Patel Ravi Narendra , Diaz Rene Alfonso , Campbell Michael A 
Published: November 2023

Prevention of sprouting in stored potato tubers is a major concern for the agricultural and food industries. The compound 1,4‐dimethylnaphthalene (DMN) has been shown to prevent premature sprouting when fogged onto the surface of potato tubers. Treatment of Saccharomyces cerevisiae cultures with DMN resulted in a temporary suppression of growth. Based on that data we speculate that treatment of stored potato tubers with DMN would result in sprout suppression as well as alteration of fungal communities on the surface of potato tubers. The treatment of potatoes was done with DMN or sterile water (control) for two days. The surface of the potatoes consisted of 4.2 ppm of DMN residue after treatment. Periderm (2 cm) samples were quickly frozen in liquid N2 and stored at −80°C after the treatment. DNA was isolated from frozen tissues using a Power Soil DNA Isolation Kit (mobio.com). Total DNA was measured and prepared for PCR using primers specific to the ITS regions of fungi (F). PCR products were evaluated using gel electrophoresis and sequenced using Illumina technology (Penn State Nucleic Acid Facility). A total of 3.7 million DNA sequences were analyzed using the Mothur (v1.36) software package to determine fungal diversity by alignment to the silva.eukarya.fasta database. Among all samples there were 1124 operational taxonomical units (OTU). It was determined that there is a greater fungal diversity on the control tissue than the DMN treated tubers. This data suggests the DMN alters the fungal diversity on the surface of potato tubers.Support or Funding InformationPenn State Behrend, 1,4‐Group
Sirtuin 3‐ and Diet‐Mediated Regulation of Mitochondrial Function During Aging
Authors: Dhillon Rashpal S. , van Ginkel Paul R. , Fu Vivian , Prolla Tomas A. , Denu John M. 
Published: November 2023

Aging is a multifactorial process that results in the gradual damage of a number of cellular systems. Specifically, the decline of mitochondrial function due to hyperacetylation has been implicated in numerous metabolic defects, including senescence. The NAD+‐dependent mitochondrial deacetylase, sirtuin 3 (SIRT3) has a marked impact on mitochondrial acetylation status and may be responsible for attenuating oxidative stress. Furthermore, caloric restriction, which induces SIRT3 expression, has been linked to increased longevity. However, the impact of acetylation on mitochondrial function over the lifespan of an organism remains largely unknown. Here, we used a multi‐tissue approach to establish the role of SIRT3 and acetylation status in governing aging phenotypes by comparing mitochondrial function and structure in aging mice.C57BL/6 mice in the SIRT3+/+ and −/− condition were fed a control or calorically‐restricted diet, and sampled at 5, 15, and 25 months of age. Gastrocnemius, brain, liver and heart were measured for respiratory capacity, complex enzyme kinetics, mitochondrial morphology, mtDNA content, and the protein expression of key SIRT3 targets. Metabolomic profiling of the liver was used to assess the impact of electron transport impairments on substrate utilization and accumulation. To determine possible mechanisms for the aging phenotypes observed in the present study, we examined expression levels of ATP‐dependent mitochondrial proteases. Generally, all tissues had markedly different respiratory responses to the treatment condition. However, oxygen flux through complex II revealed a significant SIRT3‐CR “rescue” effect at old age in most tissues. These results suggest that the deficiencies in oxidative phosphorylation may be driven by an impaired function at respiratory complexes caused by reduced protein content or complex activity. Mitochondrial structure, as determined by electron microscopy, revealed differences amongst groups, dependent on both genotype and diet. Overall, our findings suggest that SIRT3 in conjunction with caloric restriction may play a pivotal role in the age‐related response of the mitochondrial respiratory chain to aging in a tissue‐specific manner.Support or Funding InformationNational Institute of Health, National Institute on Aging
Combination Therapy to Enhance the Efficacy of Recombinant Immunotoxins: Evaluating the Role of Mitochondrial Translation in Toxin‐Mediated Cell Death
Authors: Zhu Yuyi , Weldon John 
Published: November 2023

Recombinant immunotoxins (RITs) based on Pseudomonas exotoxin A (PE) are targeted cytotoxic proteins currently in clinical trials for the treatment of cancer. PE covalently modifies and inhibits elongation factor 2 (EF2), which halts protein synthesis within the cytosol and ultimately induces apoptotic cell death. Although PE inhibits translation in the cytoplasm, eukaryotes also have a translation system in mitochondria that is similar to the translation system in bacteria. Recent findings have highlighted the importance of mitochondrial stability and function in tumorigenesis, sustaining cancer cell proliferation, and maintaining cancer cell survival. By inhibiting protein synthesis within the mitochondria through the usage of antibiotics, mitochondria may become unable to properly function, resulting in the inability to meet the energy requirements of the cell and the activation of apoptotic cell death. I proposed that combination treatment with both PE‐based RITs and antibiotics that inhibit mitochondrial protein synthesis will reduce cancer cell survival compared to independent treatments of either therapeutic alone. To test this hypothesis, combination treatment of RITs and seven different antibiotics (tetracycline, chloramphenicol, streptomycin, linezolid, fusidic acid, doxycycline, and kanamycin) have been applied to CA46 and HEK293 cell lines followed by the assessment of cell viability through WST‐8 assay. No statistically relevant effects were observed for combination treatments in HEK293 cells, but the addition of a nontoxic level of chloramphenicol to RIT‐treated CA46 cells showed a ~50% enhancement in cell killing compared to RIT treatment alone (p=.05). In response, we are continuing to explore the usage of chloramphenicol in combination therapies with RIT. Though we conclude that targeting mitochondrial translation with antibiotics is not a general strategy for RIT combination therapy, combination therapy may be applicable in specific instances.Support or Funding InformationThis project was supported by funds from Towson University's Office of Undergraduate Research and the Fisher College of Science and Mathematics, as well as an undergraduate research award from the American Society for Biochemistry and Molecular Biology.
Buffer Therapy for Cancer
Authors: Pilot Christian Joseph , Gillies Robert 
Published: November 2023

BackgroundSolid tumors exhibit an increase in glucose metabolism when compared to normal tissue. The increased metabolism generates an acidic tumor environment, allowing for metastasis and local invasion of tumor cells to occur. Previous research has demonstrated that oral buffers can neutralize this acidity and prevent metastasis and local invasion. Buffer therapy utilizes the intrinsic buffering capacity of various GRAS (generally regarded as safe) compounds to raise the pH tumors. While the acidity and alkalinity of foods has been documented, this is not the same as their buffering capacities.MethodsA Mettler‐Toledo pH titration system was used to measure the buffering capacity of nutritional supplements and foods. The presence of titratable groups within the supplements and food sources led to their selection. Data were expressed as mmoles H+ consumed per 100 g (or per serving) to reach a pH of 4, which is the post‐prandial pH in the duodenum.Results/DiscussionThe buffering capacity of NaHCO3 has yielded promising results in mouse models and has undergone evaluation in phase I/II human trials. Clinically, compliance in buffer therapy remains problematic in patients, as the therapy requires ingestion of 40g‐50g NaHCO3 capsules daily. A plausible solution is supplement NaHCO3 with foods and supplements, expressed on a point‐based system wherein 1 point corresponds to 1g of NaHCO3. Research suggests that the therapeutic properties of buffer therapy occur when ~50 points are ingested daily. A compilation of common items and their corresponding point values has been created as a patient guide.Support or Funding InformationMoffitt Cancer Center
E‐cadherin regulates mitochondrial metabolism and induces cell growth through NF‐κB in E‐cadherin deficient AGS cells
Authors: Park Song Yi , Shin Jee‐Hye , Kee Sun‐Ho 
Published: November 2023

In this study the interrogation of Wnt regulator β‐catenin and an emerging cancer control molecule NF‐κB was explored in E‐cadherin expressed AGS gastric cancer cells (EC96 cells). It was found that E‐cadherin can increase cell proliferation and energy metabolism through activation of NF‐κB. EC96 cells treated with NF‐κB inhibitors showed a decrease in cell survival. Knockdown of E‐cadherin expression by shRNA could clearly reverse the E‐cadherin‐induced energy metabolism potential in EC96 cells. The application of NF‐kB inhibitors supported the evidence that NF‐kB act as an upstream signal of c‐myc and Glut1. Normally NF‐kB is present in the cytosol of resting cells. Our results showed that E‐cadherin expression led NF‐κB to translocate from cytosol to nucleus in AGS cells. The treatment of NF‐kB inhibitors or knockdown with E‐cadherin shRNA abolished the ability of E‐cadherin to translocate NF‐kB into nucleus in EC96 cells, which was accompanied with the reduction of cell growth. In conclusion, E‐cadherin expression generates cell proliferation signals through NF‐κB signal pathway as Wnt signal pathway was suppressed.Support or Funding Information[This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (NRF‐2015R1D1A1A09056775)]