Human Adenocarcinoma Cell Research Articles

Establishment of an Embryo Implantation Model In Vitro.

Embryo implantation is the first step in the establishment of a successful pregnancy. An in vitro model for embryo implantation is critical for basic biological research, drug development, and screening. This paper presents a simple, rapid, and highly efficient in vitro model for embryo implantation. In this protocol, we first introduce mouse blastocyst acquisition and human endometrial adenocarcinoma cells (Ishikawa)preparation for implantation, followed by the co-culture method for mouse embryos and Ishikawa cells. Finally, we conducted a study to assess the impact of varying concentrations of 17-β-estradiol (E2) and progesterone (P4) on embryo adhesion rates based on this model. Our findings revealed that high concentrations of E2 significantly reduced embryo adhesion, whereas the addition of progesterone could restore the adhesion rate. This model offers a simple and fast platform for evaluating and screening molecules involved in the adhesion process, such as cytokines, drugs, and transcription factors controlling implantation and endometrial receptivity.

Quantitative Analysis of Isopimpinellin from Ammi majus L. Fruits and Evaluation of Its Biological Effect on Selected Human Tumor Cells.

Ammi majus L. (Apiaceae) is a medicinal plant with a well-documented history in phytotherapy. The aim of the present work was to isolate isopimpinellin (5,8-methoxypsoralen; IsoP) from the fruit of this plant and evaluate its biological activity against selected tumor cell lines. The methanol extract obtained with the use of an accelerated solvent extraction (ASE) method was the most suitable for the quantitative analysis of coumarins in the A. majus fruit matrix. The coumarin content was estimated by RP-HPLC/DAD, and the amount of IsoP was found to be 404.14 mg/100 g dry wt., constituting 24.56% of the total coumarin fraction (1.65 g/100 g). This, along with the presence of xanthotoxin (368.04 mg/100 g, 22.36%) and bergapten (253.05 mg/100 g, 15.38%), confirmed A. majus fruits as an excellent source of these compounds. IsoP was isolated (99.8% purity) by combined liquid chromatography/centrifugal partition chromatography (LC/CPC) and tested for the first time on its antiproliferative activity against human colorectal adenocarcinoma (HT29, SW620), osteosarcoma (Saos-2, HOS), and multiple myeloma (RPMI8226, U266) cell lines. MTT assay results (96 h incubation) demonstrated a dose- and cell line-dependent decrease in cell proliferation/viability, with the strongest effect of IsoP against the Saos-2 cell line (IC50; 42.59 µM), medium effect against U266, HT-29, and RPMI8226 (IC50 = 84.14, 95.53, and 105.0 µM, respectively), and very weak activity against invasive HOS (IC50; 321.6 µM) and SW620 (IC50; 711.30 µM) cells, as well as normal human skin fibroblasts (HSFs), with IC50; 410.7 µM. The mechanistic study on the Saos-2 cell line showed that IsoP was able to reduce DNA synthesis and trigger apoptosis via caspase-3 activation. In general, IsoP was found to have more potency towards cancerous cells (except for HOS and SW620) than against healthy cells. The Selective Index (SI) was determined, underlining the higher selectivity of IsoP towards cancer cells compared to healthy cells (SI = 9.62 against Saos-2). All these results suggest that IsoP might be a promising molecule in the chemo-prevention and treatment of primary osteosarcoma.

Green Synthesis of Silver Nanoparticles Using Plant Extract Blends and Its Impact on Antibacterial and Biological Activity

There is a strong interest in using green resources for synthesizing nanoparticles (NPs) of industrial and biomedical utility in a way to maintain desired material properties throughout use while not inducing any harmful effects. The use of various plant extracts as reducing, capping, or stabilizing agents is widely attempted in green nanotechnology. However, very little has been explored about incorporating plant extract blends into green NP synthesis routes. Here, we used the combination of tea and olive leaf extracts for the synthesis of silver NPs and evaluated the advantages it provided over both chemical and single-plant-mediated synthesis routes. Four different reducing agents (tannic acid, black tea leaves extract, olive leaves extract and their blend) were used to synthesize silver NPs (Ag NP) from silver nitrate (AgNO[Formula: see text]. The synthesized Ag NP was characterized by scanning electron microscopy (SEM), dynamic light scattering (DLS), and ultraviolet-visible (US-Vis) spectroscopy. The antimicrobial properties of Ag NP were assessed against Escherichia coli (E. Coli) and Staphylococcus aureus (S. Aureus) using the colony-forming unit (CFU) assay and the minimum inhibitory concentration (MIC) assay. The cytotoxic potential of Ag NP on human colorectal adenocarcinoma (Caco-2) cells was assessed by the WST-1 assay. Results showed that Ag NP synthesized using plant extract mixtures had a primary particle size of 40[Formula: see text]nm and were very effective antibacterial agents, with the MIC values ranging from [Formula: see text]g/mL to 10[Formula: see text][Formula: see text]g/mL. While the particle size obtained in chemical synthesis was slightly lower, the resultant Ag NP did not serve as an effective antibacterial agents at low doses. Further understanding of how best to integrate extracts of different plants into green NP synthesis routes will enable wider and safer biomedical applications.

Antiviral and cytotoxic activities and chemical profiles of two species of Abies nordmanniana from Türkiye.

Abies is an important genus of the family Pinaceae, with about 50 species found in the highlands of Asia, Europe, North Africa, and North and Central America. The principal aim of the present work was to investigate the chemical content and biological potential of the resin and cone from Abies nordmanniana subsp. bornmulleriana and Abies nordmanniana subsp. equi-trojani, respectively. The flavonoid and phenolic contents of the resin and cones were evaluated using liquid chromatography-high resolution mass spectrometry (LC-HRMS). Additionally, the essential oil and fatty acid compositions were analyzed using gas chromatography-mass spectrometry (GC-MS) and gas chromatography-flame ionization detector (GC-FID), respectively. Cytotoxicity of the extracts and essential oils were screened against certain cancer cell lines, namely, human prostate adenocarcinoma cell line (PC3), human lung adenocarcinoma cell line (A549), human pancreatic cancer cell line (PANC-1), human hepatocellular carcinoma cell line (HepG2), human breast cancer cell line (MDA-MB231), and normal human lung fibroblast cell line (CCD-34-LU), with MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide] assay. According to the MTT results, hexane extracts of both cone (CH) and resin (RH), ethanol-water (CEW), dichloromethane (CD), and acetone (CA) extracts of the cone mostly inflict cytotoxicity in HepG2 cell line. Antiviral activities of Abies nordmanniana subsp. extracts at doses of 5 μg/g and 10 μg/g were also evaluated in ovo for their virucidal activity against avian coronavirus. Abies nordmanniana subsp. extracts exhibited concentration-dependent antiviral activity on specific pathogen-free embryonated chicken eggs. Significantly, cone acetone extract (CA), cone ethanol extract (CE), and cone dichloromethane extract (CD) of Abies nordmanniana subsp. exhibited strong inhibition of the virus at a concentration of 10 μg/g. The most potent virucidal activity was observed with ethanol-water extract of conifer form (CEW). According to these results, it was proved that Abies nordmanniana species could be a potential, sustainable, and renewable drug source, especially considering the impressive antiviral and significant cytotoxic activity potentials.

Role of transforming growth factor-β1 pathway in angiogenesis induced by chronic stress in colorectal cancer

ABSTRACT Background Chronic stress can induce stress-related hormones; norepinephrine (NE) is considered to have the highest potential in cancer. NE can stimulate the expression of hypoxia-inducible factor-1α (HIF-1α), which is associated with vascular endothelial growth factor (VEGF) secretion and tumor angiogenesis. However, the underlying mechanisms are poorly understood. Methods Tumor-bearing mice were subjected to chronic restraint stress and treated with normal saline, human monoclonal VEGF-A neutralizing antibody bevacizumab, or β-adrenergic receptor (β-AR) antagonist (propranolol). Tumor growth and vessel density were also evaluated. Human colorectal adenocarcinoma cells were treated with NE, propranolol, or the inhibitor of transforming growth factor-β (TGF-β) receptor Type I kinase (Ly2157299) in vitro. TGF-β1 in mouse serum and cell culture supernatants was quantified using ELISA. The expression of HIF-1α was measured using Real time-PCR and western blotting. Cell migration and invasion were tested. Results Chronic restraint stress attenuated the efficacy of bevacizumab and promoted tumor growth and angiogenesis in a colorectal tumor model. Propranolol blocked this effect and inhibited TGF-β1 elevation caused by chronic restraint stress or NE. NE upregulated HIF-1α expression, which was reversed by propranolol or Ly2157299. Propranolol and Ly2157199 blocked NE-stimulated cancer cell migration and invasion. Conclusions Our results demonstrate the effect of NE on tumor angiogenesis and the critical role of TGF-β1 signaling during this process. In addition, β-AR/TGF-β1 signaling/HIF-1α/VEGF is a potential signaling pathway. This study also indicates that psychosocial stress might be a risk factor which weakens the efficacy of anti-angiogenic therapy.

The Role of Ayahuasca in Colorectal Adenocarcinoma Cell Survival, Proliferation and Oxidative Stress.

The psychedelic beverage ayahuasca is originally obtained by Banisteriopsis caapi (B. caapi) (BC) and Psychotria viridis (P. viridis) (PV). However, sometimes these plant species are replaced by others that mimic the original effects, such as Mimosa hostilis (M. hostilis) (MH) and Peganum harmala (P. harmala) (PH). Its worldwide consumption and the number of studies on its potential therapeutic effects has increased. This study aimed to evaluate the anticancer properties of ayahuasca in human colorectal adenocarcinoma cells. Thus, the maximum inhibitory concentration (IC50) of decoctions of MH, PH, and a mixture of these (MHPH) was determined. The activities of caspases 3 and 9 were evaluated, and the cell proliferation index was determined through immunocytochemical analysis (Ki-67). Two fluorescent probes were used to evaluate the production of oxidative stress and the activity of the antioxidant enzymes superoxide dismutase (SOD) and glutathione peroxidase (GPx) was also evaluated. It was demonstrated that exposure to the extracts significantly induced apoptosis in Caco-2 cells, while decreasing cell proliferation. MH and MHPH samples significantly reduced oxidative stress and significantly increased glutathione peroxidase activity. No significant differences were found in SOD activity. Overall, it was demonstrated that the decoctions have a potential anticancer activity in Caco-2 cells.

Unfolding the role of cell state on stress granule heterogeneity and function in pancreatic cancer.

e16357 Background: A major impediment to cancer therapy is the reality that cancer is a heterogeneous disease. A single tumor can consist of many genotypic or phenotypic profiles, which are marked by degrees of sensitivity to different therapeutics. Identifying and targeting origins of cancer cell heterogeneity is critical for predicting patient outcome and enhancing treatment efficacy. We observe a novel source of cancer cell heterogeneity that involves one of the largest stress-sensing platforms described to date: the stress granule (SG). Stress granules are non-membranous, cytoplasmic organelles that assemble in response to stress, and promote cell survival. Deregulation of SGs has been linked to several pathological conditions, including cancer, neurodegeneration, ischemia, and viral infections. SGs have been shown to promote tumor growth and metastasis, as well as contribute to mechanisms of chemo-resistance. Thus, these signaling hubs are emerging as a potential target in cancer. Methods: To induce SG formation, human pancreatic ductal adenocarcinoma (PDAC) cell lines were treated with sodium arsenite, thapsigargin, or exposed to hypoxic conditions. Immunoflourescent staining of core SG proteins was then conducted. To investigate cell cycle-driven mechanisms of SG heterogeneity, cell cycle stage was identified by expression of a Fluorescent ubiquitination-based cell cycle indicator (Fucci) lentiviral system. To investigate cPLA2’s role in SG formation, the chemical inhibitor Pyrrophenone, an shRNA targeting cPLA2, and an exogenous full-length or cleaved form of cPLA2, were used. In vivo, ES149 cells expressing Luciferase and the Fucci system were orthotopically implanted into C57BL/6 mice. Established tumors were treated every 3 days with Vehicle, Oxaliplatin, Pyrrophenone, or a combination of the latter. Tumor size was tracked via IVIS Spectrum. Fucci flourescence was used to identify cell cycle phase. SGs and cell death were assessed by immunoflourescent staining of tumors. Results: Cancer cells show a high degree of heterogeneity in their production of SGs. Cell cycle phase dictates SG heterogeneity. G2 phase cells have significantly higher levels of SG formation as compared cells in G1/S. This process is regulated by cleavage of the cPLA2 protein. Oxaliplatin administration increases G2 phase content in tumors. Co-treatment with Oxaliplatin and Pyrrophenone significantly decreases SG formation, increases cell death, and leads to tumor regressions in vivo. Conclusions: G2 phase cancer cells develop higher levels of SGs when exposed to stressful stimuli, as compared to G1/S phase counterparts. The cPLA2 protein is cleaved into a catalytically dead form that is present in G1/S phases, but absent in G2 phase. Oxaliplatin treatment in vivo leads to an increase in G2 phase tumor cells. When paired with cPLA2 inhibition, this results in increased cancer cell death and decreased tumor growth.

Ciprofloxacin is a novel anti-ferroptotic antibiotic

Cancer patients undergoing chemotherapy are susceptible to various bacterial infections, necessitating prompt and precise antimicrobial treatment with antibiotics. Ciprofloxacin is a clinically utilized broad-spectrum antimicrobial agent known for its robust antiseptic activity. While ferroptosis, an oxidative form of cell death, has garnered attention as a promising avenue in cancer therapy, the potential impact of ciprofloxacin on the anticancer effects of ferroptosis remains unclear. This study seeks to investigate the potential influence of antibiotics on ferroptosis in human pancreatic ductal adenocarcinoma (PDAC) cells. Here, we report a previously unrecognized role of ciprofloxacin in inhibiting ferroptosis in human PDAC cells. Mechanistically, ciprofloxacin suppresses erastin-induced endoplasmic reticulum (ER) stress through the activating transcription factor 6 (ATF6) and ER to nucleus signaling 1 (ERN1) pathway. Excessive ER stress activation can trigger glutathione peroxidase 4 (GPX4) degradation through autophagic mechanisms. In contrast, ciprofloxacin enhances the protein stability of GPX4, a crucial regulator that suppresses ferroptosis by inhibiting lipid peroxidation. Thus, our study demonstrates the anti-ferroptotic role of ciprofloxacin, highlighting the importance of careful consideration when contemplating the combination of ciprofloxacin with specific ferroptosis inducers in PDAC patients.

Screening and verification of outer membrane proteins of Cronobacter sakazakii with potential adhesion capability to human ileocecal adenocarcinoma cells

Cronobacter sakazakii (C. sakazakii), a food-borne pathogen, can cause severe intestinal diseases, such as intestinal damage and necrotizing enterocolitis, and even death in neonates and infants through its adhesion to intestinal epithelium cells by protein-protein interaction and translocation across the intestinal barrier. Nevertheless, the mechanisms of adhesion, translocation, and pathogenesis remain unclear. This study aimed to screen and validate the outer membrane proteins of C. sakazakii capable of adhering to the surface of intestinal cells through biotinylated protein adhesion and LC-MS/MS identification. A total of 319 proteins were identified. Gene Ontology and KEGG analysis revealed significant enrichment of ligand proteins in metabolism-related processes and catalytic activity. The identified 37 outer membrane proteins and 41 proteins of unknown subcellular localization or function mainly enriched in five protein-protein interaction clusters. Ten of these outer membrane/unknown proteins were selected to test their roles on C. sakazakii adhesion and invasion of intestinal cells by silencing the encoding gene expression and nine of the ten proteins, Protein_26/30/34/38/42/98/104/194/282, were shown to be critical in this process. This study contributes to the understanding of proteins involved in the host adhesion process by C. sakazakii as well as the mechanisms for its translocation across the intestinal barrier.

Phytochemical profile and determination of cytotoxicity, acute oral toxicity, genotoxicity, and mutagenicity of aqueous and ethanolic extracts of Pseudobombax marginatum (A. St.-Hil.) A. Robyns

ABSTRACT Pseudobombax marginatum, popularly known as “embiratanha,” is widely used by traditional communities as anti-inflammatory and analgesic agent. This study aimed to determine the phytochemical profile as well as cytotoxicity, acute oral toxicity, genotoxicity, and mutagenicity attributed to exposure to aqueous (AqEx) and ethanolic (EtEx) extracts of embiratanha bark. Phytochemical screening was conducted using thin-layer chromatography (TLC). Cell viability was analyzed using MTT assay with human mammary gland adenocarcinoma (MDA-MB-231) and macrophage (J774A.1) cell lines, exposed to concentrations of 12.5, 25, 50, or 100 µg/ml of either extract. For acute oral toxicity, comet assay and micronucleus (MN) tests, a single dose of 2,000 mg/kg of either extract was administered orally to Wistar rats. TLC analysis identified classes of metabolites in the extracts, including cinnamic acid derivatives, flavonoids, hydrolyzable tannins, condensed tannins, coumarins, and terpenes/steroids. In the cytotoxicity assay, the varying concentrations of extracts derived from embiratanha induced no significant alterations in the viability of MDA-MB-231 cells. The lowest concentration of EtEx significantly increased macrophage J774A.1 viability. However, the higher concentrations of AqEx markedly lowered macrophage J774A.1 viability. Animals exhibited no toxicity in the parameters analyzed in acute oral toxicity, comet assay, and MN tests. Further, EtEx promoted a significant reduction in DNA damage index and DNA damage frequency utilizing the comet assay, while the group treated with AqEx exhibited no marked differences. Thus, data demonstrated that AqEx or EtEx of embiratanha may be considered safe at a dose of 2,000 mg/kg orgally under our experimental conditions tested.

Dual Antimicrobial-Anticancer Potential, Hydrolysis, and DNA/BSA Binding Affinity of a Novel Water-Soluble Ruthenium-Arene Ethylenediamine Schiff base (RAES) Organometallic

The need for a systematic approach in developing new metal-based drugs with dual anticancer-antimicrobial properties is emphasized by the vulnerability of cancer patients to bacterial infections. In this context, a novel organometallic assembly was designed, featuring ruthenium(II) coordination with p-cymene, one chlorido ligand, and a bidentate neutral Schiff base derived from 4-methoxybenzaldehyde and N,N-dimethylethylenediamine. The compound was extensively characterized in both solid-state and solution, employing single crystal X-ray diffraction, nuclear magnetic resonance, infrared, ultraviolet–visible spectroscopy, and density functional theory, alongside Hirshfeld surface analysis. The hydrolysis kinetic was thoroughly investigated, revealing the important role of the chloro-aqua equilibrium in the dynamics of binding with deoxyribonucleic acid and bovine serum albumin. Notably, the aqua species exhibited a pronounced affinity for deoxyribonucleic acid, engaging through electrostatic and hydrogen bonding interactions, while the chloro species demonstrated groove-binding properties. Interaction with albumin revealed distinct binding mechanisms. The aqua species displayed covalent binding, contrasting with the ligand-like van der Waals interactions and hydrogen bonding observed with the chloro specie. Molecular docking studies highlighted site-specific interactions with biomolecular targets. Remarkably, the compound exhibited wide spectrum moderate antimicrobial activity against Staphylococcus aureus, Pseudomonas aeruginosa, and Candida albicans, coupled with low micromolar cytotoxic activity against human colorectal adenocarcinoma cells and significant activity against human leukemic monocyte lymphoma cells. The presented findings encourage further development of this compound, promising avenues for its evolution into a versatile therapeutic agent targeting both infectious diseases and cancer.

Exploring the Antiproliferative Potential of Morpholine‐Functionalized Aurones: Design, Synthesis, SAR, DFT, Hirshfeld Surface, 3D Energy Frameworks and Molecular Docking Analysis

AbstractIn quest of generating a diverse array of potential cytotoxic candidates through molecular hybridization approach, herein we have designed a library of forty‐one morpholine‐functionalized aurones 7(a–ao), each with a distinct substitution pattern. These aurones were obtained by reacting 6‐hydroxyaurones, with 4‐(2‐chloroethyl)morpholine hydrochloride, characterized by various spectroscopic techniques including single crystal XRD (7b) and in‐vitro screened for their antiproliferative potential against human gastric adenocarcinoma (AGS) cells. Most of the synthesized compounds have exhibited excellent cytotoxic results. Twenty analogs were found to have cytotoxic potential better than the reference drug Leucovorin and analog 7j emerged as the most potent with IC50=5.98 μM nearly one fifth to the standard drug Leucovorin (IC50=30.8 μM). This library, consisting of twenty three novel and eighteen reported morpholine‐functionalized aurones, bearing diverse substitution pattern, further helped in making a concrete structure‐activity relationship for antiproliferative potential. Additionally, in‐silico molecular docking, Hirshfeld surface analysis and two dimensional fingerprint plots were also studied to understand the molecular interactions and various structural aspects. Further computational investigations of these compounds were conducted using the DFT/B3LYP method, for the determination of HOMO and LUMO energy values as well as the calculation of chemical softness, hardness, electronegativity and electrophilic index etc.

The Influence of Mesotrione on Human Colorectal Adenocarcinoma Cells and Possibility of Its Toxicity Mitigation by Cichoric Acid.

Mesotrione, as a widely used herbicide, is present in the environment in detectable amounts, causing serious damage. Here, we aimed to investigate the effect of mesotrione on Caco-2 cells and the possibility of its toxicity mitigation by cichoric acid. Therefore, we analyzed the cytotoxicity of both these compounds and the selected oxidative stress parameters, apoptosis and interaction of both the tested compounds with the cell membrane and their accumulation within the cells. In cytotoxicity studies, the stimulating activity of mesotrione was observed, and simultaneously, the inhibitory effect of cichoric acid was noticed. This effect was related to the results of oxidative stress analysis and apoptosis measurements. The activity level of key enzymes (glutathione peroxidase, catalase and superoxide dismutase) in Caco-2 cells exposed to cichoric acid was higher as compared to that of the control. The treatment with mesotrione did not induce apoptosis in the Caco-2 cells. The penetration of the studied compounds into the Caco-2 cells was measured by using an HPLC methodology, and the results indicate mesotrione's high penetration capacity. The distribution of charge on the surface of the cell membranes changed under the influence of both compounds. Considering the mutual interactions of beneficial and potentially toxic food ingredients, it should be noted that, despite the observed favorable trend, cichoric acid is not able to overcome the toxic and cancer-stimulating effects of this pesticide.

Nanotechnology-Based Strategy for Enhancing Therapeutic Efficacy in Pancreatic Cancer: Receptor-Targeted Drug Delivery by Somatostatin Analog

A novel nanotechnology-based drug delivery system (DDS) targeted at pancreatic cancer cells was developed, characterized, and tested. The system consisted of liposomes as carriers, an anticancer drug (paclitaxel) as a chemotherapeutic agent, and a modified synthetic somatostatin analog, 5-pentacarbonyl-octreotide, a ligand for somatostatin receptor 2 (SSTR2), as a targeting moiety for pancreatic cancer. The cellular internalization, cytotoxicity, and antitumor activity of the DDS were tested in vitro using human pancreatic ductal adenocarcinoma (PDAC) cells with different expressions of the targeted SSTR2 receptors, and in vivo on immunodeficient mice bearing human PDAC xenografts. The targeted drug delivery system containing paclitaxel exhibited significantly enhanced cytotoxicity compared to non-targeted DDS, and this efficacy was directly related to the levels of SSTR2 expression. It was found that octreotide-targeted DDS proved exceptionally effective in suppressing the growth of PDAC tumors. This study underscores the potential of octreotide-targeted liposomal delivery systems to enhance the therapeutic outcomes for PDAC compared with non-targeted liposomal DDS and Paclitaxel-Cremophor® EL, suggesting a promising avenue for future cancer therapy innovations.

Synthesis of selenophene-containing flavonols and 2-styrylchromones: Evaluation of their activities compared with selenophene-containing chalcones as potential anticancer agents.

Previously, we documented the synthesis and assessed the biological effects of chalcones containing selenium against HT-29 human colorectal adenocarcinoma cells, demonstrating their significant potential. As research on selenium-containing flavonoids remains limited, this article outlines our design and synthesis of three selenium-based flavonols and three 2-styrylchromones. We conducted evaluations of these compounds to determine their impact on human lung cancer cells (A549, H1975, CL1-0, and CL1-5) and their influence on normal lung fibroblast MRC5 cells. Additionally, we included selenium-based chalcones in our testing for comparative purposes. Our findings highlight that the simplest compound, designated as compound 1, exhibited the most promising performance among the tested molecules.

Folic acid-mediated enhancement of the diagnostic potential of luminescent europium-doped hydroxyapatite nanocrystals for cancer biomaging

Early and accurate cancer diagnosis is crucial for improving patient survival rates. Luminescent nanoparticles have emerged as a promising tool in fluorescence bioimaging for cancer diagnosis. To enhance diagnostic accuracy, ligands promoting endocytosis into cancer cells are commonly incorporated onto nanoparticle surfaces. Folic acid (FA) is one such ligand, known to specifically bind to folate receptors (FR) overexpressed in various cancer cells such as cervical and ovarian carcinoma. Therefore, surface modification of luminescent nanoparticles with FA can enhance both luminescence efficiency and diagnostic accuracy. In this study, luminescent europium-doped hydroxyapatite (EuHAp) nanocrystals were prepared via hydrothermal method and subsequently modified with (3-Aminopropyl)triethoxysilane (APTES) followed by FA to target FR-positive human cervical adenocarcinoma cell line (HeLa) cells. The sequential grafting of APTES and then FA formed a robust covalent linkage between the nanocrystals and FA. Rod-shaped FA-modified EuHAp nanocrystals, approximately 100 nm in size, exhibited emission peaks at 589, 615, and 650 nm upon excitation at 397 nm. Despite a reduction in photoluminescence intensity following FA modification, fluorescence microscopy revealed a remarkable 120-fold increase in intensity compared to unmodified EuHAp, attributed to the enhanced uptake of FA-modified EuHAp. Additionally, confocal microscope observations confirmed the specificity and the internalization of FA-modified EuHAp nanocrystals in HeLa cells. In conclusion, the modification of EuHAp nanocrystals with FA presents a promising strategy to enhance the diagnostic potential of cancer bioimaging probes.

Preclinical pharmacokinetic studies of villocarine A, an active Uncaria alkaloid.

Villocarine A is a bioactive indole alkaloid isolated from the Uncaria genus. It has demonstrated vasorelaxation activity and potential to protect the central nervous system. To identify the pharmacokinetic properties of villocarine A, a series of in vitro and in vivo studies have been performed. Villocarine A was found to be highly permeable (15.6 ± 1.6*10-6cm/s) across human colorectal adenocarcinoma cell monolayer with high protein binding (>91%) in both rat and human plasma. Hepatic extraction ratio of villocarine A was 0.1 in pooled rat liver and 0.2 in human liver microsomes and was found stable in rat plasma at 37°C. Due to the high permeability and low rate of metabolism properties, villocarine A was initially considered suitable for preclinical development and an ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for quantification (linearity: 1-150 ng/ml) in rat plasma was developed and validated for in vivo studies. Essential pharmacokinetic parameters included the volume of distribution and clearance of villocarine A, which were found to be 100.3 ± 15.6L/kg and 8.2 ± 1.1L/h/kg, respectively, after intravenous administration in rats. Following oral dosing, villocarine A exhibited rapid absorption as the maximum plasma concentration (53.2 ± 10.4ng/ml) occurred at 0.3 ± 0.1h, post-dose. The absolute oral bioavailability of villocarine A was 16.8 ± 0.1%. To our knowledge, this was the first pharmacokinetic study of villocarine A, which demonstrated the essential pharmacokinetic properties of villocarine A: large volume distribution, high clearance, and low oral bioavailability in rats.

PARP inhibitor synthetic lethality in ATM biallelic mutant cancer cell lines is associated with BRCA1/2 and RAD51 downregulation.

Ataxia telangiectasia-mutated (ATM) kinase is a central regulator of the DNA damage response (DDR) signaling pathway, and its function is critical for the maintenance of genomic stability in cells that coordinate a network of cellular processes, including DNA replication, DNA repair, and cell cycle progression. ATM is frequently mutated in human cancers, and approximately 3% of lung cancers have biallelic mutations in ATM, i.e., including 3.5% of lung adenocarcinomas (LUAD) and 1.4% of lung squamous cell carcinomas (LUSC). We investigated the potential of targeting the DDR pathway in lung cancer as a potential therapeutic approach. In this context, we examined whether ATM loss is synthetically lethal with niraparib monotherapy. This exploration involved the use of hATM knockout (KO) isogenic cell lines containing hATM homozygous (-/-) and heterozygous (+/-) generated via CRISPR/Cas9 gene knockout technology in DLD-1, a human colorectal adenocarcinoma cell line. Subsequently, we extended our investigation to non-small cell lung cancer (NSCLC) patient derived xenograft (PDX) models for further validation of poly ADP-ribose polymerase inhibitor (PARPi) synthetic lethality in ATM mutant NSCLC models. Here, we demonstared that biallelic hATM deletion (-/-) in DLD-1 impairs homologous recombination (HR) repair function and sensitizes cells to the PARPi, niraparib. Niraparib also caused significant tumor regression in one-third of the NSCLC PDX models harboring deleterious biallelic ATM mutations. Loss of hATM (-/-) was concomitantly associated with low BRCA1 and BRCA2 protein expression in both the hATM (-/-) DLD-1 cell line and PARPi-sensitive ATM mutant NSCLC PDX models, suggesting a downstream effect on the impairment of HR-mediated DNA checkpoint signaling. Further analysis revealed that loss of ATM led to inhibition of phosphorylation of MRN (Mre11-Rad50-NBS1) complex proteins, which are required for ATM-mediated downstream phosphorylation of p53, BRCA1, and CHK2. Taken together, our findings highlight that the synthetic lethality of niraparib in ATM-deficient tumors can be regulated through a subsequent effect on the modulation of BRCA1/2 expression and its effect on HR function.

Expression of the membrane tetraspanin claudin 18 on cancer cells promotes T lymphocyte infiltration and antitumor immunity in pancreatic cancer

Tumors weakly infiltrated by T lymphocytes poorly respond to immunotherapy. We aimed to unveil malignancy-associated programs regulating Tcell entrance, arrest, and activation in the tumor environment. Differential expression of cell adhesion and tissue architecture programs, particularly the presence of the membrane tetraspanin claudin (CLDN)18 as a signature gene, demarcated immune-infiltrated from immune-depleted mouse pancreatic tumors. In human pancreatic ductal adenocarcinoma (PDAC) and non-small cell lung cancer, CLDN18 expression positively correlated with more differentiated histology and favorable prognosis. CLDN18 on the cell surface promoted accrual of cytotoxic T lymphocytes (CTLs), facilitating direct CTL contacts with tumor cells by driving the mobilization of the adhesion protein ALCAM to the lipid rafts of the tumor cell membrane through actin. This process favored the formation of robust immunological synapses (ISs) between CTLs and CLDN18-positive cancer cells, resulting in increased Tcell activation. Our data reveal an immune role for CLDN18 in orchestrating Tcell infiltration and shaping the tumor immune contexture.

Simultaneous Delivery of Curcumin and Resveratrol via In Situ Gelling, Raft-Forming, Gastroretentive Formulations.

Curcumin and resveratrol are polyphenolic compounds that have been shown to exhibit synergistic therapeutic properties including anti-inflammatory, anticancer, and antiulcer activities, which may be exploited for the treatment of gastric diseases. However, both compounds have poor aqueous solubility and rapid metabolism, resulting in a low oral bioavailability. In situ gelling, liquid formulations were developed to produce a gastroretentive, raft-forming delivery vehicle to improve bioavailability. Solid dispersions containing a mixture of curcumin and resveratrol with Eudragit® EPO (Cur/Res-SD) were first prepared using solvent evaporation, to improve the solubility and dissolution of the compounds. Solid dispersions of a weight ratio of 1:10 curcumin/resveratrol to Eudragit® EPO were subsequently incorporated into in situ gelling, liquid formulations based on the gelling polymers, sodium alginate (low viscosity and medium viscosity), pectin, and gellan gum, respectively. Calcium carbonate and sodium bicarbonate were included to produce carbon dioxide bubbles in the gel matrix, on exposure to gastric fluid, and to achieve flotation. Moreover, the calcium ions acted as a crosslinking agent for the hydrogels. Optimized formulations floated rapidly (<60 s) in simulated gastric fluid (pH = 1.2) and remained buoyant, resulting in the gradual release of more than 80% of the curcumin and resveratrol content within 8 h. The optimized formulation based on medium-viscosity sodium alginate exhibited enhanced cytotoxic activity toward human gastric adenocarcinoma cell lines (AGS), compared with unformulated curcumin and resveratrol compounds, and increased anti-inflammatory activity against RAW 264.7 macrophage cells compared with the NSAID, indomethacin. These findings demonstrate that in situ gelling, liquid formulations, loaded with a combination of curcumin and resveratrol in the form of solid dispersions, show potential as gastroretentive delivery systems for local and systemic effects.