KRAP regulates mitochondrial Ca2+ uptake by licensing IP3 receptor activity and stabilizing ER-mitochondrial junctions.

Inositol 1,4,5-trisphosphate (IP3) receptors (IP3Rs) are high-conductance channels that allow the regulated redistribution of Ca2+ from the endoplasmic reticulum (ER) to the cytosol and, at specialized membrane contact sites (MCSs), to other organelles. Only a subset of IP3Rs release Ca2+ to the cytosol in response to IP3. These 'licensed' IP3Rs are associated with Kras-induced actin-interacting protein (KRAP, also known as ITPRID2) beneath the plasma membrane. It is unclear whether KRAP regulates IP3Rs at MCSs. We show, using simultaneous measurements of Ca2+ concentration in the cytosol and mitochondrial matrix, that KRAP also licenses IP3Rs to release Ca2+ to mitochondria. Loss of KRAP abolishes cytosolic and mitochondrial Ca2+ signals evoked by stimulation of IP3Rs via endogenous receptors. KRAP is located at ER-mitochondrial membrane contact sites (ERMCSs) populated by IP3R clusters. Using a proximity ligation assay between IP3R and voltage-dependent anion channel 1 (VDAC1), we show that loss of KRAP reduces the number of ERMCSs. We conclude that KRAP regulates Ca2+ transfer from IP3Rs to mitochondria by both licensing IP3R activity and stabilizing ERMCSs.

Exploring In-Silico, In-Vitro Antioxidant, and Cytotoxic Potential of Valerian wallichii by On Cervical Epithelial Carcinoma (HeLa) Cell Lines.

Traditional medicinal practices often utilize herbal remedies for treating various diseases. This study focuses on exploring the phytochemical constituents, in-silico, in-vitro antioxidant, and anticancer activities of Valerian wallichii root extracts on human cervical epithelial carcinoma (HeLa) cell lines. The molecular docking approach was employed to predict the ligand molecule's orientation within the receptor like Epidermal growth factor receptor tyrosine kinase domain (PDB ID: 1M17) using Schrodinger's GLIDE model. Among the selected phytocompounds, hesperidin exhibited promising inhibitory activity against EGFR (Epidermal Growth Factor Receptor) domain with -8.701 kcal/mol docking score and interactions with MET 769, ASP 831, ASP776, LEU694 and ASN818 residues as compared to standard doxorubicin with -7.6 kcal/mol docking score and interactions with ASP 831, ASN818 and ASP776 residues and further, various antioxidant activity was assessed and In-vitro anticancer activity against HeLa cell lines was evaluated by hydroalcoholic root extracts demonstrated antioxidant capacities, and selective cytotoxicity was observed, with IC50 : 45.759±0.42 µg/mL for the overall extract and IC50 : 30.245±0.58 µg/mL for the EAF fraction as compared to standard doxorubicin with IC50 : 25.9891±0.25 µg/mL, respectively. The present study concluded that Valerian wallichii L possesses potential human cervical epithelial carcinoma cell line inhibition properties based on the computer aided drug design models and in-vitro activity.

Invasion of HeLa Cells by Shigella Species Clinical Isolates Recovered from Pediatric Diarrhea.

Shigella is considered a major public health concern, especially for children younger than 5 years of age in developing countries. The pathogenicity of Shigella is a complex process that involves the interplay of multiple genes located on a large, unstable virulence plasmid as well as chromosomal pathogenicity islands. Since various factors (including virulence and antibiotic resistance genes) are associated with the severity and duration of shigellosis, in this article, we aim to evaluate whether the invasion of HeLa cells is affected by Shigella spp. isolates with different characteristics (including serogroups, virulence gene profiles, and antibiotic resistance patterns) recovered from pediatric patients in Tehran, Iran. Cell invasion ability of 10 Shigella isolates with different serogroups (Shigella flexneri and Shigella sonnei), gene profiling (virA, sen, ipgD, ipaD, ipaC, ipaB, and ipaH), and antibiotic resistance phenotyping (ampicillin, azithromycin, ciprofloxacin, nalidixic acid, trimethoprim-sulfamethoxazole, cefixime, cefotaxime, minocycline, and levofloxacin) were measured by plaque-forming assay in HeLa cell lines. The results show that all the selected Shigella spp. isolates recovered from pediatric patients were able to invade HeLa cells, but the total number and average size of plaques were different between the isolates. The higher invasion ability of S. flexneri isolates in HeLa cells compared to S. sonnei isolates was attributed to the presence of particular virulence genes; however, the role of each of these virulence factors remains to be determined.

Electrochemiluminescence sensing of HeLa cells labeled with biotinylated ruthenium complex using bipolar electrode based on microwell modified optical fiber.

Biotinylated ruthenium complexes exhibit improved photoluminescent (PL) properties when they bind with streptavidin, making them useful labels or probes in bio-related analysis. However, their ECL properties are still unknown to date. Herein, we reported the use of [Ru(bpy)2(biot-bpy)]2+ complexes as a new ECL luminophore, which was functionalized with biotin moiety and exhibited higher ECL efficiency after binding to streptavidin. Moreover, [Ru(bpy)2(biot-bpy)]2+ complexes could be attached to HeLa cells through the biotin-streptavidin binding. A microwell bipolar electrode (MBE) prepared at one end of an optical fiber bundle was applied to produce ECL of the labeled HeLa cells, which was remotely detected at the other end. The [Ru(bpy)2(biot-bpy)]2+-streptavidin binding effect together with the high surface/volume ratio of MBE promoted the ECL generation on HeLa cells, which was applied to sensitively detect HeLa cells with a linear range from 1.56 × 102 to 6.74 × 106 cells/mL and a detection limit of 83 cells/mL. Moreover, ECL images were successfully acquired to resolve the emission on each HeLa cell. Such cytosensor based on [Ru(bpy)2(biot-bpy)]2+ and MBE may extend the applications of ECL for cell detections.

Mitochondrial calpain-5 truncates caspase-4 during endoplasmic reticulum stress.

Calpains are cysteine proteases activated in response to intracellular calcium signaling. Activated calpains regulate various cellular functions by degrading substrate molecules in a site-specific manner. Although most calpains are localized in the cytosol, we previously reported that calpain-5 exists in the mitochondria. The mitochondrial calpain-5 is activated during endoplasmic reticulum (ER) stress. However, the substrate of calpain-5, as well as the physiological significance of calpain-5 activation, has not yet been elucidated. In the present study, we treated HeLa cells with A23187, tunicamycin, or hydrogen peroxide to induce intracellular calcium increase, resulting in cell death. The cells treated with A23187 or tunicamycin exhibited the activation of calpain-5 and truncation of caspase-4. The truncation of caspase-4 was inhibited by the repression of calpain-5 expression with the appropriate siRNA. Additionally, both calpain-5 and caspase-4 were observed in the mitochondria. Our study is the first to demonstrate that the activation of mitochondrial calpain-5 triggers the truncation of caspase-4, suggesting that mitochondrial calpain-5 regulates the downstream pathway of caspase-4, including cell death and the inflammatory cascade. The results of the present study provide new insights into ER-stress-related diseases such as Alzheimer's disease and cancer. These perspectives allow us to propose new therapeutic strategies such as the development of inhibitors or activators of calpain-5, which may be useful in the development of treatment for ER-stress-related diseases.

Exogenous interleukin-1 beta promotes the proliferation and migration of HeLa cells via the MEK/ERK signaling pathway.

OBJECTIVE: Interleukin-1 beta (IL-1ß) is a crucial cytokine that has been implicated in cancer and metastasis development. However, its possible mechanistic role in cervical cancer remains unclear. This study aimed to investigate the functions of exogenous IL-1ß in cervical cancer cell proliferation and migration. METHODS: HeLa cell proliferation and migration were measured using MTT and Transwell assays. A lentivirus-mediated packaging system was used to construct an IL-1ß overexpressing cell line. MEK/ERK signal transduction was inhibited by pretreatment with the MEK inhibitor PD98059. qRT-PCR and Western blotting were used to test the expression of relevant genes. RESULTS: Exogenous IL-1ß promoted the proliferation and migration of HeLa cells. In addition, overexpression of IL-1ß in HeLa cells promoted cell proliferation. Mechanistically, exogenous IL-1ß increased the phosphorylated MEK and ERK levels in HeLa cells and the expression of JUN, RELB, and NF-κB2. Alternatively, blockade of MEK inhibited the promoting proliferation effects of IL-1ß and the expression of JUN, RELB, and NF-κB2. CONCLUSIONS: Our data suggest that exogenous IL-1ß regulates HeLa cell functions by regulating the MEK/ERK signaling pathway and by targeting JUN, RELB, and NF-κB2. Our study uncovered a potential association across IL-1ß, cervical tumor development, and cancer progression.

Synthesis and physicochemical characteristics of Ag-doped hydroxyapatite nanoparticles, and their potential biomedical applications.

In this present scenario, hydroxyapatite (HAp) nanostructures were synthesized through green routes for biomedical applications, particularly remediation towards human pathogens and cancer cells. The present study aims at forming non-toxic and eco-friendly silver (Ag+) doped HAp using Polyethylene glycol (PEG), Cetyl Trimethyl ammonium bromide (CTAB) and curcumin. Ag+ doped HAp nanoparticles (NPs) were prepared by the sol-gel method with a cube and rod-like morphology. Ag-HApNPs showed a sharp and well-defined diffraction peak, which possesses the hexagonal crystalline structure with space group P63/m. The Fourier-transform infrared spectroscopy and Raman spectra confirmed the formation of Ag-HApNPs, and the bandgap values were obtained using UV-DRS analysis. The Ag-HApNPs with PEG, CTAB and curcumin might be fabricated materials were examined against antibacterial, antifungal, antioxidant, and cytotoxic activities, which provided exemplary biomedical applications. Overall, Ag-HApNPs can be used as potential drug delivery and perspectives to control multidrug-resistant pathogens.

Fungi fabrication, characterization, and anticancer activity of silver nanoparticles using metals resistant Aspergillus niger.

The purpose of this study was to assess the bio-fabrication possibilities of pre-isolated (from bauxite mine tailings) metal-tolerant Aspergillus niger biomass filtrate and the anticancer potential of synthesized silver nanoparticles (AgNPs) tested with a Human Cervical cancer cell line (HeLa cells: Henrietta Lacks cells). The nitrate reduction test demonstrated that A. niger has the ability to reduce nitrate, and filtrate derived from A. niger biomass efficiently fabricated AgNPs from AgNO3, as demonstrated by a visible color change from pale greenish to brownish. The UV-visible spectroscopy analysis revealed an absorbance peak at 435 nm, which corresponded to the AgNPs. These AgNPs have been capped and stabilized with several functional groups related to various bioactive molecules such as aldehyde, benzene rings, aldehydic, amines, alcohols, and carbonyl stretch protein molecules. Fourier-Transform Infrared Spectroscopy (FTIR) analysis confirmed the capping and stabilizing chemical bonding pattern. Scanning Electron Microscopy (SEM) revealed that the synthesized AgNPs were spherical, with an average size of 21.38 nm. This bio-fabricated AgNPs has in-vitro anticancer potential when tested against the HeLa cell line due to its potential size and shape. At 100 g mL-1 concentrations of this bio-fabricated AgNPs, the anticancer activity percentage was found to be 70.2%, and the IC50 value was found to be 66.32 g m-1. These findings demonstrated that the metal-tolerant A. niger cell filtrate could produce AgNPs with anticancer potential.

Folate Conjugated Polyethylene Glycol Probe for Tumor-Targeted Drug Delivery of 5-Fluorouracil.

A targeted delivery system is primarily intended to carry a potent anticancer drug to specific tumor sites within the bodily tissues. In the present study, a carrier system has been designed using folic acid (FA), bis-amine polyethylene glycol (PEG), and an anticancer drug, 5-fluorouracil (5-FU). FA and PEG were joined via an amide bond, and the resulting FA-PEG-NH2 was coupled to 5-FU producing folate-polyethylene glycol conjugated 5-fluorouracil (FA-PEG-5-FU). Spectroscopic techniques (UV-Vis, 1HNMR, FTIR, and HPLC) were used for the characterization of products. Prodrug (FA-PEG-5-FU) was analyzed for drug release profile (in vitro) up to 10 days and compared to a standard anticancer drug (5-FU). Folate conjugate was also analyzed to study its folate receptors (FR) mediated transport and in vitro cytotoxicity assays using HeLa cancer cells/Vero cells, respectively, and antitumor activity in tumor-bearing mice models. Folate conjugate showed steady drug release patterns and improved uptake in the HeLa cancer cells than Vero cells. Folate conjugate treated mice group showed smaller tumor volumes; specifically after the 15th day post-treatment, tumor sizes were decreased significantly compared to the standard drug group (5-FU). Molecular docking findings demonstrated importance of Trp138, Trp140, and Lys136 in the stabilization of flexible loop flanking the active site. The folic acid conjugated probe has shown the potential of targeted drug delivery and sustained release of anticancer drug to tumor lesions with intact antitumor efficacy.

Apis mellifera syriaca Venom: Evaluation of Its Anticoagulant Effect, Proteolytic Activity, and Cytotoxicity along with Its Two Main Compounds-MEL and PLA2-On HeLa Cancer Cells.

Bee venom (BV) is one of the most remarkable natural products that has been a subject of studies since ancient times. Recent studies have shown that Apis mellifera syriaca venom possesses antibacterial as well as cytotoxic effects on cancer cell lines. The venom contains a variety of bioactive molecules-mainly melittin (MEL) and phospholipase A2 (PLA2), as well as other compounds that are not well characterized. In this work, we continue the biological characterization of A. mellifera syriaca venom by testing its anticoagulant effect on human plasma using the prothrombin time (PT) test, as well as assessing its proteolytic activity. In addition, the cytotoxicity of the crude venom-and of its two main components, MEL and PLA2-was tested on HeLa cancer cell lines for the first time. The results obtained showed the capacity of A. mellifera syriaca venom to increase clotting time, thereby proving its anticoagulant effect. Moreover, the venom did not demonstrate a significant proteolytic activity unless administrated at concentrations ≥ 5 mg/mL. Finally, we showed that crude A. mellifera syriaca venom, along with MEL, exhibit a strong in vitro cytotoxic effect on HeLa cancer cell lines, even at low concentrations. In summary, our findings could serve as a basis for the development of new natural-based drug candidates in the therapeutic field.

Cesium Treatment Depresses Glycolysis Pathway in HeLa Cell.

BACKGROUND/AIMS: Cesium (Cs) is an alkali metal element that is of no essential use for humans; it has no known beneficial function that is verified by clinical research. When used as an alternative cancer therapy, it even causes toxicity in high doses. Thus, before using Cs as treatment in clinical settings, it is important to clearly determine its biological effects on cells. However, Cs was found to suppress the proliferation of human cervical cancer cells in a dose-dependent manner, and it was assumed that Cs inhibits the glycolysis pathway. In this study, we clearly determined the step of the glycolysis pathway that is affected by Cs. METHODS: The glycolytic enzyme expressions, activities, and metabolite concentrations in HeLa cells were measured by PCR, western blotting, and enzymatic methods, after treating the cells with Cs for 3 days. RESULTS: Cs treatment decreased transcriptional and expression levels of hexokinase, glyceraldehyde-3-phosphate dehydrogenase, pyruvate kinase (PK), and lactate dehydrogenase and the activity of PK. Analysis of glycolysis pathway metabolites revealed that Cs treatment reduces lactate level and increases the level of nicotinamide adenine dinucleotide (oxidized form, NAD+); however, it did not affect the levels of pyruvate and nicotinamide adenine dinucleotide (reduced form, NADH). Increase of the [NAD+]/[NADH] ratio and decrease of the [lactate]/[pyruvate] ratio indicate that Cs treatment inhibits the aerobic glycolysis pathway. CONCLUSION: Cs treatment inhibits PK activity and increases the [NAD+]/[NADH] ratio. Hence, Cs has been determined to inhibit glycolysis, especially the aerobic glycolysis pathway. These results suggest that suppression of HeLa cell proliferation following Cs treatment was caused by inhibition of aerobic glycolysis by Cs.

The propagation of HSV-1 in high autophagic activity.

Autophagy is an intracellular process involving double-membrane vacuoles that ultimately merge with the lysosomes and play a key role in the inhibition of herpessimplex virus 1 (HSV-1) proliferation. This virus is an agent of some lethal neuronal diseases like encephalitis. HSV-1 requires the expression of its latent proteins, such as ICP34.5, to promote cell infection, which disrupts the autophagy process. In this study, we aimed to evaluate the effect of autophagy induction on HSV-1 replication in host cells at the early and late stages of its replication. Furthermore, we explored the consequences of autophagy induction before and after cell infection.Cells were transfected through Beclin-1-expressing plasmids. Autophagy induction was performed with microtubule-associated protein 1 light chain 3 (LC3-II) as an autophagosome formation marker by using flow cytometry. In the first stage, HSV-1 was inoculated into transfected cells 18 hours post-transfection. Next, viral DNA was extracted 18 and 48 hours post-infection, and eventually viral copies per milliliter were calculated through real-time polymerase chain reaction (PCR). For the second stage, the plasmid containing Beclin-1 was transfected to the cells following virus inoculation to examine the influence of autophagy induction after cell infection.Study results have shown that in neuroblastoma cells autophagy activation reduces virus yield from 4×10 5 copies/ml (control sample) to 9×10 4 copies/ml at 24 h postinfection and viral load after 48 h declines up to 1×10 6 copies/ml, which is less than that of the control sample about 5 logs. However, in HeLa cells, we observed a significant reduction in autophagy induction with reducing HSV-1 propagation. Despite these results, HSV-1 proliferation in both cell types increased and these viruses were able to maintain their ability to propagate even in high autophagic activity. Hyperactivation of autophagy can only slow the rate of virus replication. This study may provide new insight into the effect of autophagy on HSV-1 replication.

Selective optosensing of iron(III) ions in HeLa cells using NaYF4:Yb3+/Tm3+ upconversion nanoparticles coated with polyepinephrine.

Novel polyepinephrine-modified NaYF4:Yb,Tm upconversion luminescent nanoparticles (UCNP@PEP) were prepared via the self-polymerization of epinephrine on the surfaces of the UCNPs for selective sensing of Fe3+ inside a cell and for intracellular imaging. The proposed UCNP@PEP probe is a strong blue light emitter (λmax = 474 nm) upon exposure to an excitation wavelength of 980 nm. The probe was used for detecting Fe3+ owing to the complexation reaction between UCNP@PEP and Fe3+, resulting in reduced upconversion luminescence (UCL) intensity. The proposed probe has a detection limit of 0.2 µM and a good linear range of 1-10 µM for sensing Fe3+ ions. Moreover, the UCNP@PEP probe displays high cell viability (90%) and is feasible for intracellular imaging. The ability of the probe to sense Fe3+ in a human serum sample was tested and shows promising output for diagnostic purposes. The prepared UCNP@PEP probe was characterized by using UV-visible (UV-Vis) absorption spectrometry, fluorescence (FL) spectrometry, field emission scanning electron microscopy (FE-SEM), X-ray photoelectron spectroscopy (XPS), and Fourier transform infrared spectroscopy (FT-IR).

A study on MAPK/ERK and CDK2-Cyclin-E signal switch "on and off" in cell proliferation by bis urea derivatives of 1, 4-Diisocyanatobenzene.

A series of novel substituted bisurea 1,4-Diisocyanatobenzene compounds were designed, synthesized and introduced as potent anticancer compounds and screened for their in vitro anti-proliferative activities in human cancer cell lines. The structures of all titled compounds were characterized using Fourier-transform infrared mass spectra, nuclear magnetic resonance spectroscopy, elemental analysis and evaluated their sustainability using biological experiments. A selected group of ten derivatives were apprised for their anti-proliferative activity. The compounds 3d and 3e displayed potent anticancer activity with low IC50 value of 5.40, and 5.89 µM against HeLa cancer cell lines. The observed apoptosis data has demonstrated that compounds 3d and 3e induce the activaties of caspase-9 and caspase-3, the compounds 3d and 3e regulated fungal zone inhibition. Due to promising growth inhibitions, the all synthesized compounds were allowed to campaign includes quantum-polarized-ligand, quantum mechanical and molecular mechanical, docking experiments. The compounds 3d and 3e have exhibited a higher affinity for ERK/MAP kinase and CDK2 proteins. The molecular docking interactions have demonstrated two stage inhibition of cancer cells by binding with ERK/MAP kinase and CDK2 leads to inactivation of cell proliferation,cell cycle progression,cell divisionanddifferentiation, and hypo-phosphorylation of ribosome leading cells to restricts at point boundary of the G1/S phase. The long-range molecular dynamics, 150 ns, simulations were also revealed more consistency by 3d. Our study conclude good binding propensity for active-tunnel of ERK/MAP kinase and CDK2 proteins, by 3d (1,1'-(1,4-phenylene) bis(3-(2-chlorobenzyl)urea)), to suggest that the designed and synthesized 3d is to use as selective novel nuclei in anti-cancer chemotherapeutics.

Synthesis and characterization of TiO2 NPs by aqueous leaf extract of Coleus aromaticus and assess their antibacterial, larvicidal, and anticancer potential.

The frequent applications of synthetic chemical insecticides and drugs create resistance among insects and microbes, creating a new threat to human and environmental welfare. This investigation focused on evaluating the possibilities of fabricating and characterizing the titanium dioxide nanoparticles (TiO2 NPs) from titanium dioxide (TiO2) through the aqueous leaf extract of Coleus aromaticus. Their biological applications were studied against the larvae of Aedes aegypti human pathogenic bacteria, and cancer cell line. The results revealed that the aqueous leaf extract had the metal reducing proficiency to produce nanoparticles from TiO2. The synthesized TiO2 NPs were initially confirmed by visible color changes and Ultraviolet-Visible Spectrophotometer analysis that showed a predominant peak at 332 nm. Furthermore, the nanocrystals, structural alignment, functional groups and elemental compositions were studied by following standard operating protocol in XRD (X-ray Powder Diffraction), FTIR (Fourier Transform Infrared Spectroscopy), TEM (Transmission Electron Microscopy), and EDX (Energy-Dispersive X-ray Spectroscopy) techniques, respectively. The results attained from these techniques confirmed that the plant mediated and fabricated particles were in the nanoscale range (12-33 nm) with a hexagonal shape. The synthesized TiO2 NPs had an outstanding (1000 µg mL-1) larvicidal activity against the four stages of instars larvae of Ae. aegypti at 1000 µg mL-1. It also had an excellent antibacterial potential against E. faecalis (33 mm), followed by S. boydii (30 mm) at 30 mg L-1 concentration. The green fabricated TiO2 NPs had a fabulous (92.37%) cytotoxic activity on the HeLa cell line at 100 µg mL-1 dosage within one day of exposure. The entire results concluded that the C. aromaticus mediated TiO2 NPs have excellent biological applications and thus, could be considered for the welfare of human beings.

Ti4+-immobilized hierarchically porous zirconium-organic frameworks for highly efficient enrichment of phosphopeptides.

Ti4+-immobilized hierarchically porous zirconium-organic frameworks (denoted as THZr-MOFs) was prepared for phosphopeptide enrichment. The THZr-MOFs showed high specific surface area of 185.28 m2 g-1, wide pore-size distribution of 3 ~ 20 nm, good chemical stability and excellent hydrophilicity. Introduction of hierarchical pores in MOFs not only facilitated the accessibility of phosphopeptides to the internal metal affinity sites and reduce their mass transfer resistance, but also increased the exposure sites of metal affinity interaction and binding energies of Zr and Ti elements. Benefited from these advantages, the THZr-MOFs showed high adsorption capacity (79.8 µg mg-1) towards standard phosphopeptide. A low detection limit (0.05 fmol µL-1) and high enrichment selectivity (ß-casein/BSA with a molar ratio of 1:5000) were also obtained by MALDI-TOF MS. The THZr-MOFs were applied to analyze complex samples including nonfat milk, human serum, and HeLa cell lysate. In total, 1432 phosphopeptides derived from 762 phosphoproteins were identified from human HeLa cell lysate. Schematic representation of the application of Ti4+-immobilized hierarchically porous zirconium-organic frameworks (denoted as THZr-MOFs) in high-efficiency and selective enrichment of low-abundance phosphopeptides from the tryptic digest of human HeLa cell lysate.

No Intercellular Regulation of the Cell Cycle among Human Cervical Carcinoma HeLa Cells Expressing Fluorescent Ubiquitination-Based Cell-Cycle Indicators in Modulated Radiation Fields.

The non-targeted effects of radiation have been known to induce significant alternations in cell survival. Although the effects might govern the progression of tumor sites following advanced radiotherapy, the impacts on the intercellular control of the cell cycle following radiation exposure with a modified field, remain to be determined. Recently, a fluorescent ubiquitination-based cell-cycle indicator (FUCCI), which can visualize the cell-cycle phases with fluorescence microscopy in real time, was developed for biological cell research. In this study, we investigated the non-targeted effects on the regulation of the cell cycle of human cervical carcinoma (HeLa) cells with imperfect p53 function that express the FUCCI (HeLa-FUCCI cells). The possible effects on the cell-cycle phases via soluble factors were analyzed following exposure to different field configurations, which were delivered using a 150 kVp X-ray irradiator. In addition, using synchrotron-generated, 5.35 keV monochromatic X-ray microbeams, high-precision 200 µm-slit microbeam irradiation was performed to investigate the possible impacts on the cell-cycle phases via cell-cell contacts. Collectively, we could not detect the intercellular regulation of the cell cycle in HeLa-FUCCI cells, which suggested that the unregulated cell growth was a malignant tumor. Our findings indicated that there was no significant intercellular control system of the cell cycle in malignant tumors during or after radiotherapy, highlighting the differences between normal tissue and tumor characteristics.

Genetic Load and Potential Mutational Meltdown in Cancer Cell Populations.

Large genomes with elevated mutation rates are prone to accumulating deleterious mutations more rapidly than natural selection can purge (Muller's ratchet). As a consequence, it may lead to the extinction of small populations. Relative to most unicellular organisms, cancer cells, with large and nonrecombining genome and high mutation rate, could be particularly susceptible to such "mutational meltdown." However, the most common type of mutation in organismal evolution, namely, deleterious mutation, has received relatively little attention in the cancer biology literature. Here, by monitoring single-cell clones from HeLa cell lines, we characterize deleterious mutations that retard the rate of cell proliferation. The main mutation events are copy number variations (CNVs), which, estimated from fitness data, happen at a rate of 0.29 event per cell division on average. The mean fitness reduction, estimated reaching 18% per mutation, is very high. HeLa cell populations therefore have very substantial genetic load and, at this level, natural population would likely face mutational meltdown. We suspect that HeLa cell populations may avoid extinction only after the population size becomes large enough. Because CNVs are common in most cell lines and tumor tissues, the observations hint at cancer cells' vulnerability, which could be exploited by therapeutic strategies.

The synthesis and biological evaluation of nucleobases/tetrazole hybrid compounds: A new class of phosphodiesterase type 3 (PDE3) inhibitors.

Spired by the chemical structure of Cilostazol, a selective phosphodiesterase 3A (PDE3A) inhibitor, several novel hybrid compounds of nucleobases (uracil, 6-azauracil, 2-thiuracil, adenine, guanine, theophylline and theobromine) and tetrazole were designed and successfully synthesized and their inhibitory effects on PDE3A as well as their cytotoxicity on HeLa and MCF-7 cancerous cell lines were studied. Obtained results show the linear correlation between the inhibitory effect of synthesized compounds and their cytotoxicity. In some cases, the PDE3A inhibitory effects of synthesized compounds are higher than the Cilostazol. Besides, compared to a standard anticancer drug methotrexate, some of the synthesized compounds showed the higher cytotoxicity against the HeLa and MCF-7 cancerous cell lines.

Design, synthesis and biological evaluation of bicyclic carboxylic acid derivatives as IDO1 inhibitors.

Indoleamine 2,3-dioxygenase 1 (IDO1) plays a vital role in tumor immune escape and has emerged as a promising target for cancer immunotherapy. In this study, a novel series of bicyclic carboxylic acid derivatives were designed, synthesized and evaluated for inhibitory activities against IDO1. Among these, compound 9c exhibited strong IDO1 inhibitory activity (HeLa cellular IC50 = 2.6 nM, THP-1 cellular IC50 = 11.2 nM). Further anti-tumor studies in vivo have shown that compound 9c has a great inhibitory effect on tumor growth in mice CT26 model as a single agent or in combination with 5-fluorouracil (inhibition rate was 53.9% and 92.7%, respectively). These results indicate that compound 9c is a effective IDO1 inhibitor for further investigation.