Human Melanoma Cell Lines Research Articles

In‐Vitro Combo Treatments of Human Melanoma (1205Lu) Cell line with Curcumin, Vitamins and Steroids

Our previous work on combo treatments of human melanoma (BLM) cell line pointed to the two combos A+RU and Cur+P+RU as the best combination to decrease cell growth, a basal secretion profile of pro‐inflammatory cytokines and a decrease in the major pro‐inflammatory cytokine IL‐8 secretion by the cells. In order to confirm these findings, experiments were repeated on the 1205Lu cell line. Cells were incubated in a 96 well plate along with a combination of compounds for 48 hrs. After 48 hrs, cell growth was assayed using MTT and supernatants were collected for Elisarray and IL‐8 quantitation by Elisa. Single or solo compound treatment resulted in cell growth in the range of 37 to 59% compared to untreated control 1205Lu cell growth at 100%. A double combo of compounds such as Cur+D3, Cur+P, Cur+RU, D3+P, D3+RU, P+RU, A+D3, A+P, A+RU resulted in cell growth in the range of 17 to 37%. A triple combo of compounds such as CUR+P+RU, Cur+A+D3, Cur+D3+RU resulted in cell growth in the range of 19 to 30%. Elisarray of the supernatants from various treatments pointed to A+RU, A+D3, Cur+D3+RU, Cur+P+RU as the combos having a basal secretion profile of pro‐inflammatory cytokines. Quantitation of IL‐8 by Elisa suggested A+RU and Cur+P+RU as the two combos having a minimal amount of IL‐8 secretion 17.5 pg/ml and 8.0 pg/ml respectively.ConclusionThese experiments suggested that the combination of compounds really decreased cell growth and IL‐8 secretion. Two combos A+RU and Cur+P+RU were found to work well on both (BLM and 1205Lu) cell lines. These experiments will be repeated on other human melanoma cell lines for the confirmation of the above findings.

IFN-γ and TNF Induce Senescence and a Distinct Senescence-Associated Secretory Phenotype in Melanoma.

Immune checkpoint blockade (ICB) therapy is a central pillar of melanoma treatment leading to durable response rates. Important mechanisms of action of ICB therapy include disinhibition of CD4+ and CD8+ T cells. Stimulated CD4+ T helper 1 cells secrete the effector cytokines interferon-gamma (IFN-γ) and tumor necrosis factor alpha (TNF), which induce senescence in tumor cells. Besides being growth-arrested, senescent cells are metabolically active and secrete a large spectrum of factors, which are summarized as senescence-associated secretory phenotype (SASP). This secretome affects the tumor growth. Here, we compared the SASP of cytokine-induced senescent (CIS) cells with the SASP of therapy-induced senescent (TIS) cells. Therefore, we established in vitro models for CIS and TIS in melanoma. The human melanoma cell lines SK-MEL-28 and WM115 were treated with the cytokines IFN-γ and TNF as CIS, the chemotherapeutic agent doxorubicin, and the cell cycle inhibitor palbociclib as TIS. Then, we determined several senescence markers, i.e., growth arrest, p21 expression, and senescence-associated β-galactosidase (SA-β-gal) activity. For SASP analyses, we measured the regulation and secretion of several common SASP factors using qPCR arrays, protein arrays, and ELISA. Each treatment initiated a stable growth arrest, enhanced SA-β-gal activity, and—except palbociclib—increased the expression of p21. mRNA and protein analyses revealed that gene expression and secretion of SASP factors were severalfold stronger in CIS than in TIS. Finally, we showed that treatment with the conditioned media (CM) derived from cytokine- and palbociclib-treated cells induced senescence characteristics in melanoma cells. Thus, we conclude that senescence induction via cytokines may lead to self-sustaining senescence surveillance of melanoma.

Role of Amino Acid Transporter SNAT1/SLC38A1 in Human Melanoma.

Simple SummaryMalignant melanoma originates from melanocytes. Due to its high metastatic potential and its increasing incidence, it is one of the most aggressive types of cancer. Cancer cells generally exhibit an elevated metabolism, consequently adapting their expression of transport proteins to meet the increased demand of nutrients, such as amino acids. The aim of this study was to analyze the expression and function of the amino acid transporter SNAT1 in human melanoma. In addition, we wanted to determine its role in development and progression of malignant melanoma. We revealed that SNAT1 is overexpressed in melanoma tissue samples, as well as primary and metastatic cell lines. Moreover, we were able to show that SNAT1 plays an important role in forcing proliferation, colony formation, migration and invasion, and inhibiting senescence of melanoma cells. Amino acid transporters like SNAT1 are therefore promising targets for the development of novel therapeutic strategies against melanoma.The tumor metabolism is an important driver of cancer cell survival and growth, as rapidly dividing tumor cells exhibit a high demand for energetic sources and must adapt to microenvironmental changes. Therefore, metabolic reprogramming of cancer cells and the associated deregulation of nutrient transporters are a hallmark of cancer cells. Amino acids are essential for cancer cells to synthesize the necessary amount of protein, DNA, and RNA. Although cancer cells can synthesize glutamine de novo, most cancer cells show an increased uptake of glutamine from the tumor microenvironment. Especially SNAT1/SLC38A1, a member of the sodium neutral amino acid transporter (SNAT) family, plays an essential role during major net import of glutamine. In this study, we revealed a significant upregulation of SNAT1 expression in human melanoma tissue in comparison to healthy epidermis and an increased SNAT1 expression level in human melanoma cell lines when compared to normal human melanocytes (NHEMs). We demonstrated that functional inhibition of SNAT1 with α-(methylamino) isobutyric acid (MeAIB), as well as siRNA-mediated downregulation reduces cancer cell growth, cellular migration, invasion, and leads to induction of senescence in melanoma cells. Consequently, these results demonstrate that the amino acid transporter SNAT1 is essential for cancer growth, and indicates a potential target for cancer chemotherapy.

Cytotoxic effects of chitosan nanoparticles containing Zataria multiflora essential oil against human breast and melanoma cells

BackgroundBreast cancer is the most common cancer among women, and melanoma incidence increases worldwide. The emergence of drug resistance and side effects of chemotherapy drugs has led to a great deal of attention being paid to the development of natural medicines, especially using essential oil. The preparation of essential oil-based nanoformulation has thus recently received more attention.ResultsIn this study, chitosan nanoparticles (ChiNPs) containing Zataria multiflora essential oil with a particle size of 177 ± 10 nm, a narrow particle size distribution (SPAN 0.96), and a cubic-like shape were first prepared. IC50 values of the prepared nanoformulation against human melanoma (A-375) and breast cancer cell lines (MCF-7 and MDA-MB-468) were obtained as 32 (12–84), 46 (32–67), and 105 (85–131) µg/mL. Besides, an electrospun polycaprolactone–polyethylene oxide scaffold was prepared as a dressing after treatment with the nanoformulation. Fourier transform infrared analysis confirmed the scaffold's preparation as well as successful loading of the essential oil in chitosan nanoparticles. Furthermore, the scaffold did not show a cytotoxic effect on A-375, MCF-7, and MDA-MB-468, and its surface was hydrophobic as the water contact angle with the surface was 136.5°.ConclusionsThe prepared prototype with natural ingredients and high efficacy could be considered for further consideration in vivo study or complementary medicine.Graphical abstract

Optimization of some parametric values of MTT for the determination of human melanoma (SK-Mel-30) cell viability

3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyltetrazoliumbromide (MTT) assay is a widely used assessment method for the determination of anticancerogenic effects of active compounds including plant secondary metabolites. Recently, some important plant active ingredients have been widely investigated for anticancerogenic properties on melanoma cancer lines which are the most lethal type of skin cancer. Although some methods including DNA assay, 3H-thymidine incorporation and flow cytometry have been recommended for counting cells in the culture, MTT is one of the most frequent method and therefore, MTT assay needs to be optimized for melanoma cell lines. In this study, the MTT analytical procedure for determination of cell viability of human melanoma cell line (SK-Mel-30) was divided into nine steps and various parameters in each step (reagent amount, incubation time, centrifugation, solvent type, waiting time before spectrophotometric analysis and spectrophotometric parameters) were optimized. Optimum amount of MTT reagent and incubation time after MTT addition were determined as 10 µL and 4 h for 96 well plate, respectively. Various solvents were evaluated for solubility effectiveness of the formed crystals and DMSO was found to be the best solvent to dissolve the crystals. Waiting time before spectrophotometric reading and Uv-vis spectrums were also evaluated. At the end of the study a flowchart, presented the best analytical conditions, was constructed. Obtained findings can be used for the determination of anticancerogenic properties of plant ingredients.

The Role of Transglutaminase 2 in the Radioresistance of Melanoma Cells.

Transglutaminase 2 (TG2) is a protein expressed in many tissues that exerts numerous, sometimes contradictory, intra- and extracellular functions, under both physiological and pathophysiological conditions. In the context of tumor progression, it has been found to be involved in cell adhesion, DNA repair mechanisms, induction of apoptosis, and mesenchymal transdifferentiation, among others. Here, we hypothesized that TG2 also contributes to the radioresistance of two human melanoma cell lines, A375 and MeWo, which can be seen to differ in their basal TG2 biosynthesis by examining their proliferation and clonal expansion after irradiation. For this purpose, cellular TG2 biosynthesis and TG2 activity were modulated by transfection-induced overexpression or TG2 knock-out and application of TG2-selective inhibitors. Proliferation and clonal expansion of TG2-overexpressing cells was not enhanced over wildtype cells, suggesting that increased TG2 biosynthesis does not further enhance the radioresistance of melanoma cells. Conversely, TG2 knock-out in A375 cells reduced their proliferation, as well as clonal and spheroidal expansion after irradiation, which indicates a contribution of TG2 to the radioresistance of melanoma cells. Since TG1, TG3, and partly also, TG6 biosynthesis was detectable in A375 and MeWo cells, it can be assumed that these other members of the TG family may exert a partially compensatory effect.

PPP6C, a serine-threonine phosphatase, regulates melanocyte differentiation and contributes to melanoma tumorigenesis through modulation of MITF activity

It is critical to understand the molecular mechanisms governing the regulation of MITF, a lineage specific transcription factor in melanocytes and an oncogene in melanoma. We identified PPP6C, a serine/threonine phosphatase, as a key regulator of MITF in melanoma. PPP6C is the only recurrently mutated serine/threonine phosphatase across all human cancers identified in sequencing studies and the recurrent R264C mutation occurs exclusively in melanoma. Using a zebrafish developmental model system, we demonstrate that PPP6C expression disrupts melanocyte differentiation. Melanocyte disruption was rescued by engineering phosphomimetic mutations at serine residues on MITF. We developed an in vivo MITF promoter assay in zebrafish and studied the effects of PPP6C(R264C) on regulating MITF promoter activity. Expression of PPP6C(R264C) cooperated with oncogenic NRAS(Q61K) to accelerate melanoma initiation in zebrafish, consistent with a gain of function alteration. Using a human melanoma cell line, we examined the requirement for PPP6C in proliferation and MITF expression. We show that genetic inactivation of PPP6C increases MITF and target gene expression, decreases sensitivity to BRAF inhibition, and increases phosphorylated MITF in a BRAF(V600E) mutant melanoma cell line. Our data suggests that PPP6C may be a relevant drug target in melanoma and proposes a mechanism for its action.

Activation of apoptosis and G0/G1 cell cycle arrest along with inhibition of melanogenesis by humic acid and fulvic acid: BAX/BCL-2 and Tyr genes expression and evaluation of nanomechanical properties in A375 human melanoma cell line.

Objective(s):Humic acid (HA) and Fulvic acid (FA) are major members of humic substances, which are extracted from organic sources including soil and peat. The pro-apoptotic and anti-melanogenic effects of HA and FA at the cellular and molecular levels in the A375 human melanoma cell line were examined in this study. Materials and Methods:The cytotoxicity effect of HA and FA were evaluated by cell viability assay. Apoptosis and cell cycle were investigated by flow cytometry. Real-time PCR was carried out to measure the expression of BAX, BCL-2, and Tyr genes. Moreover, the changes in nanomechanical properties were determined through atomic force microscopy (AFM). Results:It was found that HA and FA decrease cell viability with an IC50 value of 50 µg/ml (dose-dependent) for 14 hr, arrested cells in the G0/G1 phase, and increased the sub-G1 phase (induce apoptosis). Based on the AFM analysis, Young’s modulus and adhesion force values were increased, also ultrastructural characteristics of cells were changed. Results of Real-time PCR revealed that HA and FA lead to a decrease in the expressions of BCL-2 and Tyr genes, and increase the BAX gene expression.Conclusion:These results exhibited that HA and FA possess pro-apoptotic effects through increasing the BAX/ BCL-2 expression in A375 cells. These molecular reports were confirmed by cellular nanomechanical assessments using AFM and flow cytometry. In addition, HA and FA inhibited melanogenesis by decreasing the expression of the Tyr gene. It is worthwhile to note that, HA and FA can be regarded to design new anti-cancer and anti-melanogenesis products.

A stochastic mathematical model of 4D tumour spheroids with real-time fluorescent cell cycle labelling.

In vitro tumour spheroids have been used to study avascular tumour growth and drug design for over 50 years. Tumour spheroids exhibit heterogeneity within the growing population that is thought to be related to spatial and temporal differences in nutrient availability. The recent development of real-time fluorescent cell cycle imaging allows us to identify the position and cell cycle status of individual cells within the growing spheroid, giving rise to the notion of a four-dimensional (4D) tumour spheroid. We develop the first stochastic individual-based model (IBM) of a 4D tumour spheroid and show that IBM simulation data compares well with experimental data using a primary human melanoma cell line. The IBM provides quantitative information about nutrient availability within the spheroid, which is important because it is difficult to measure these data experimentally.

Effect of Citrullus colocynth fruit extract on pro and anti apoptotic molecules in human melanoma cell line (A375).

Melanoma remains a major cause of death worldwide. Chemotherapy might be used to treat advanced Melanoma which is associated with some side effects. Hence, providing proper treatment or proper employment of lower toxic substances is essential. Citrullus colocynthis, a medicinal plant, seems a potential anticancer herbal medicine against Melanoma via various efficient compounds. Therefore, it is of interest to analyze the effect of Citrullus colocynthis fruit extract on pro and anti apoptotic molecules in the human melanoma cell line (A375). Cell viability test was done using MTT assay. mRNA expression of Bax and Bcl2 was done by real-time PCR. The data was analyzed statistically by one way analysis of variance and Duncan multiple range test with graph prism version 5. p<0.05 was considered significant. Citrullus colocynthis extract caused a marked increase in cell death in a dose-dependent manner. At the end of 48 hours, maximum inhibition (50%) was at 400 and 500µg/ml. The study demonstrated that the effect of Citrullus colocynthis extract has elevated the Bax mRNA expression on human melanoma cell lines. Data shows that Citrullus colocynthis fruit extract has an anticancer effect on the human melanoma cell line (A375) by increasing the expression of Bax and Bcl2 and this could be because of the phytochemicals present in it.

In Vitro Anticancer and Cancer-Preventive Activity of New Triterpene Glycosides from the Far Eastern Starfish Solaster pacificus.

Sea stars or starfish (class Asteroidea) and holothurians or sea cucumbers (class Holothuroidea), belonging to the phylum Echinodermata (echinoderms), are characterized by different sets of glycosidic metabolites: the steroid type in starfish and the triterpene type in holothurians. However, herein we report the isolation of eight new triterpene glycosides, pacificusosides D–K (1–3, 5–9) along with the known cucumarioside D (4), from the alcoholic extract of the Far Eastern starfish Solaster pacificus. The isolated new compounds are closely related to the metabolites of sea cucumbers, and their structures of 1–3 and 5–9 were determined by extensive NMR and ESIMS techniques. Compounds 2, 5, and 8 have a new type of tetrasaccharide chain with a terminal non-methylated monosaccharide unit. Compounds 3, 6, and 9 contain another new type of tetrasaccharide chain, having 6-O-SO3-Glc as one of the sugar units. The cytotoxic activity of 1–9 against non-cancerous mouse epidermal cells JB6 Cl41 and human melanoma cell lines SK-MEL-2, SK-MEL-28, and RPMI-7951 was determined by MTS assay. Compounds 1, 3, 4, 6, and 9 showed potent cytotoxicity against these cell lines, but the cancer selectivity (SI > 9) was observed only against the SK-MEL-2 cell line. Compounds 1, 3, 4, 6, and 9 at the non-toxic concentration of 0.1 μM significantly inhibited neoplastic cell transformation of JB6 Cl41 cells induced by chemical carcinogens (EGF, TPA) or ionizing radiation (X-rays and UVB). Moreover, compounds 1 and 4 at the non-toxic concentration of 0.1 µM possessed the highest inhibiting activity on colony formation among the investigated compounds and decreased the colonies number of SK-MEL-2 cells by 64% and 70%, respectively. Thus, triterpene glycosides 1 and 4 can be considered as prospective cancer-preventive and anticancer-compound leaders.

Vasculogenic mimicry structures in melanoma support the recruitment of monocytes

ABSTRACT The progression of cancer is facilitated by infiltrating leukocytes which can either actively kill cancer cells or promote their survival. Our current understanding of leukocyte recruitment into tumors is largely limited to the adhesion molecules and chemokines expressed by conventional blood vessels that are lined by endothelial cells (ECs). However, cancer cells themselves can form their own vascular structures (a process known as vasculogenic mimicry (VM)); but whether they actively participate in the recruitment of leukocytes remains to be elucidated. Herein, we demonstrate that VM-competent human melanoma cell lines express multiple adhesion molecules (e.g. CD44, intercellular adhesion molecule (ICAM)-1 and junction adhesion molecules (JAMs)) and chemokines (e.g. CXCL8 and CXCL12) relevant for leukocyte recruitment. Microfluidic-based adhesion assays revealed that similar to ECs, VM-competent melanoma cells facilitate the rolling and adhesion of leukocytes, particularly monocytes, under conditions of shear flow. Moreover, we identified ICAM-1 to be a key participant in this process. Transwell assays showed that, similar to ECs, VM-competent melanoma cells facilitate monocyte transmigration toward a chemotactic gradient. Gene expression profiling of human melanoma patient samples confirmed the expression of numerous leukocyte capture adhesion molecules and chemokines. Finally, immunostaining of patient tissue microarrays revealed that tumors with high VM content also contained higher numbers of leukocytes (including macrophages). Taken together, this study suggests an underappreciated role of VM vessels in solid tumors via their active participation in leukocyte recruitment and begins to identify key adhesion molecules and chemokines that underpin this process.

The effect of ω-6 fatty acid on WT1 and WIF-1 genes expression and inducing apoptosis in A375 melanoma cell line

BackgroundAn exciting question as to whether nutritional linoleic acid (LA, Omega-6, or ω-6) fatty acid interferes with tumorigenesis is remained to be entirely determined in well experimentally qualified investigations. To elucidate the particular roles of LA in tumorigenesis of melanoma skin cancer, we evaluated the effect of this fatty acid on a melanoma cell line, A375, in terms of tumor growth characteristics by MTT and Flow Cytometry. In particular, we have analyzed the expression levels of two genes (WIF-1 and WT1) involved during Wnt signaling pathway as a molecular base for the formation and progression of many cancer types. MethodsIn order to analyze the cell viability, MTT assay was performed following the treatment of cultured human melanoma cell line, A375, by various concentrations of ω-6 in 24 h, 48 h, and 72 h time intervals, and apoptosis was determined using Flow Cytometry. The expression of WIF-1 and WT1 genes were measured using quantitative real-time PCR, which were further analyzed by Rest 2.0.13 Software. ResultsOur findings revealed that during treatment of cells with all the concentrations of ω-6 in various time intervals, the expression of the WIF-1 gene was significantly (p < 0.05) increased in contrast to decreasing WT1 expression levels in a time- and concentration-dependent manner. The cell viability was also shown to be decreased with an IC50 value of 1000 μM. The differences were more substantial following 72 h of treatment. DiscussionOur results supported the contributory role of ω-6 on the tumor growth of melanoma cells, highlighting the potential of this fatty acid as an influencing element which at least in part exerts its inhibitory roles by interfering with Wnt signaling molecules WIF-1 and WT1.

Sec23a inhibits the self-renewal of melanoma cancer stem cells via inactivation of ER-phagy

BackgroundThe genesis and developments of solid tumors, analogous to the renewal of healthy tissues, are driven by a subpopulation of dedicated stem cells, known as cancer stem cells (CSCs), that exhibit long-term clonal repopulation and self-renewal capacity. CSCs may regulate tumor initiation, growth, dormancy, metastasis, recurrence and chemoresistance. While autophagy has been proposed as a regulator of the stemness of CSCs, the underlying mechanisms requires further elucidation.MethodsThe CSC component in human melanoma cell lines M14 and A375 was isolated and purified by repetitive enrichments for cells that consistently display anchorage-independent spheroid growth. The stemness properties of the CSCs were confirmed in vitro by the expressions of stemness marker genes, the single-cell cloning assay and the serial spheroid formation assay. Subcutaneous tumor transplantation assay in BALB/c nude mice was performed to test the stemness properties of the CSCs in vivo. The autophagic activity was confirmed by the protein level of LC3 and P62, mRFP-LC3B punta and cytoplasmic accumulation of autolysosomes. The morphology of ER was detected with transmission electron microscopy.ResultsIn the present study, by employing stable CSC cell lines derived from human melanoma cell lines M14 and A375, we show for the first time that Sec23a inhibits the self-renewal of melanoma CSCs via inactivation of ER-phagy. Mechanistically, inhibition of Sec23a reduces ER stress and consequently FAM134B-induced ER-phagy. Furthermore, TCGA data mining and analysis show that Sec23a is a favorable diagnostic and prognostic marker for human skin cutaneous melanoma.ConclusionThis study has elucidated a new mechanism underlying the regulation of autophagy on stemness, i.e. CSCs can exploit the SEC23A/ER-stress/FAM134B/ER-phagy axis for the self-renewal. These observations provide new ideas for exploration of the regulatory network of CSC self-renewal to develop CSCs-based therapy strategies for malignant tumors.1GUvTquPwXFE-w4yWuDKxrVideo

Antagonism of the transient receptor potential melastatin‑2 channel leads to targeted antitumor effects in primary human malignant melanoma cells.

Melanoma continues to be the most aggressive and devastating form of skin cancer for which the development of novel therapies is required. The present study aimed to determine the effects of antagonism of the transient receptor potential melastatin‑2 (TRPM2) ion channel in primary human malignant melanoma cells. TRPM2 antagonism via use of the antifungal agent, clotrimazole, led to decreases in cell proliferation, as well as dose‑dependent increases in cell death in all melanoma cell lines investigated. The targeting of TRPM2 channels was verified using TRPM2 knockdown, where treatment with TRPM2 small‑interfering RNA led to similar levels of cell death in all melanoma cell lines when compared with clotrimazole treatment. Minimal effects on proliferation and cell death were observed following antagonism or knockdown of TRPM2 in non‑cancerous human keratinocytes. Moreover, characteristics of TRPM2 were explored in these melanoma cells and the results demonstrated that TRPM2, localized to the plasma membrane as a non‑specific ion channel in non‑cancerous cells, displayed a nuclear localization in all human melanoma cell lines analyzed. Additional characterization of these melanoma cell lines confirmed that each expressed one or more established multidrug resistance genes. Results of the present study therefore indicated that antagonism of the TRPM2 channel led to antitumor effects in human melanoma cells, including those that are potentially unresponsive to current treatments due to the expression of drug resistance genes. The unique cellular localization of TRPM2 and the specificity of the antitumor effects elicited by TRPM2 antagonism suggested that TRPM2 possesses a unique role in melanoma cells. Collectively, the targeting of TRPM2 represents a potentially novel, efficacious and readily accessible treatment option for patients with melanoma.

Cytokine and Chemokine Receptor Patterns of Human Malignant Melanoma Cell Lines.

Cytokine and chemokine receptors can promote tumor progression, invasion, and metastasis development by inducing different intracellular signaling pathways. The aim of this study was to determine the cytokine and chemokine receptor gene expression patterns in human melanoma cell lines. We found a large set of cytokine and chemokine receptor genes that were significantly differentially expressed between melanoma cell lines that originated from different subtypes of primary melanomas as well as cell lines that originated from melanoma metastases. The relative expressions of two receptor genes (CCR2 and TNFRSF11B) were positively correlated with the invasive potential of the cell lines, whereas a negative correlation was observed for the TNFRSF14 gene expression. We also found a small set of receptor genes that exhibited a significantly decreased expression in association with a BRAFV600E mutation. Based on our results, we assume that the analyzed cytokine and chemokine receptor collection may provide potential to distinguish the different subtypes of melanomas, helping us to understand the biological behavior of BRAFV600E-mutated melanoma cells.

Elucidation of the Metabolite Profile of Yucca gigantea and Assessment of Its Cytotoxic, Antimicrobial, and Anti-Inflammatory Activities.

The acute inflammation process is explained by numerous hypotheses, including oxidative stress, enzyme stimulation, and the generation of pro-inflammatory cytokines. The anti-inflammatory activity of Yucca gigantea methanol extract (YGME) against carrageenan-induced acute inflammation and possible underlying mechanisms was investigated. The phytochemical profile, cytotoxic, and antimicrobial activities were also explored. LC-MS/MS was utilized to investigate the chemical composition of YGME, and 29 compounds were tentatively identified. In addition, the isolation of luteolin-7-O-β-d-glucoside, apigenin-7-O-β-d-glucoside, and kaempferol-3-O-α-l-rhamnoside was performed for the first time from the studied plant. Inflammation was induced by subcutaneous injection of 100 μL of 1% carrageenan sodium. Rats were treated orally with YGME 100, 200 mg/kg, celecoxib (50 mg/kg), and saline, respectively, one hour before carrageenan injection. The average volume of paws edema and weight were measured at several time intervals. Levels of NO, GSH, TNF-α, PGE-2, serum IL-1β, IL-6 were measured. In additionally, COX-2 immunostaining and histopathological examination of paw tissue were performed. YGME displayed a potent anti-inflammatory influence by reducing paws edema, PGE-2, TNF-α, NO production, serum IL-6, IL-1β, and COX-2 immunostaining. Furthermore, it replenished the diminished paw GSH contents and improved the histopathological findings. The best cytotoxic effect of YGME was against human melanoma cell line (A365) and osteosarcoma cell line (MG-63). Moreover, the antimicrobial potential of the extract was evaluated against bacterial and fungal isolates. It showed potent activity against Gram-negative, Gram-positive, and fungal Candida albicans isolates. The promoting multiple effects of YGME could be beneficial in the treatment of different ailments based on its anti-inflammatory, antimicrobial, and cytotoxic effects.

EGFR and COX-2 Dual Inhibitor: The Design, Synthesis, and Biological Evaluation of Novel Chalcones

For most researchers, discovering new anticancer drugs to avoid the adverse effects of current ones, to improve therapeutic benefits and to reduce resistance is essential. Because the COX-2 enzyme plays an important role in various types of cancer leading to malignancy enhancement, inhibition of apoptosis, and tumor-cell metastasis, an indispensable objective is to design new scaffolds or drugs that possess combined action or dual effect, such as kinase and COX-2 inhibition. The start compounds A1 to A6 were prepared through the diazo coupling of 3-aminoacetophenone with a corresponding phenol and then condensed with two new chalcone series, C7–18. The newly synthesized compounds were assessed against both COX-2 and epidermal growth factor receptor (EGFR) for their inhibitory effect. All novel compounds were screened for cytotoxicity against five cancer cell lines. Compounds C9 and G10 exhibited potent EGFR inhibition with IC50 values of 0.8 and 1.1 µM, respectively. Additionally, they also displayed great COX-2 inhibition with IC50 values of 1.27 and 1.88 µM, respectively. Furthermore, the target compounds were assessed for their cytotoxicity against pancreatic ductal cancer (Panc-1), lung cancer (H-460), human colon cancer (HT-29), human malignant melanoma (A375) and pancreatic cancer (PaCa-2) cell lines. Interestingly, compounds C10 and G12 exhibited the strongest cytotoxic effect against PaCa-2 with average IC50 values of 0.9 and 0.8 µM, respectively. To understand the possible binding modes of the compounds under investigation with the receptor cites of EGFR and COX-2, a virtual docking study was conducted.

Comparison of characteristics and tumor targeting properties of extracellular vesicles derived from primary NK cells or NK-cell lines stimulated with IL-15 or IL-12/15/18

NK cell-based therapies have shown promise for hematological cancer forms, but their use against solid tumors is hampered by their poor ability to infiltrate the tumor. NK cells release extracellular vesicles (EVs) containing cytolytic proteins, indicating that NK-cell derived EVs may have therapeutic potential. In this study, we compared the tumor-targeting potential of EVs derived from either primary NK cells or the NK cell lines NK-92 and KHYG-1 cultured in IL-15 alone or in combination with IL-12 and IL-18. Primary NK cells were also stimulated through the activating receptor CD16. Tumor cell apoptosis was measured using a panel of human colon, melanoma, glioblastoma, prostate, breast, and ovarian tumor cell line spheroids. NK cells or NK-92 cells stimulated with IL-12, IL-15, and IL-18 generated EVs with higher efficiency than EVs from resting cells, although similar amounts of EVs were produced under both conditions. Proteomic analysis indicated similar distribution of cytolytic proteins in EVs from primary NK cells and NK-92, but lower levels in KHYG-1 EVs that translated into poor capacity for KHYG-1 EVs at targeting tumor cell lines. Further, we show that CD16-stimulated NK cells release low amounts of EVs devoid of cytolytic proteins. Importantly, EVs from cytokine-stimulated NK cells penetrate into the spheroid core, and tumor spheroid susceptibility to NK-cell derived EVs was linked to differential expression of the NKG2D ligands MICA/B, which was blocked with an anti-NKG2D antibody. We conclude that EVs from activated primary NK cells or NK-92 cells has the best potential to infiltrate and target solid tumors.

Probe into DNA interaction, cell toxicity and antifungal activities of phyto-synthesized yttrium oxide (Y2O3) nanoparticles

Green synthesis is considered to be eco-friendly approach in comparison to chemical mediated synthesis and hence the study demonstrates facile Nyctanthes arbortristis mediated phyto-synthesis of biocompatible yttrium oxide (Y2O3) nanoparticles. X-ray diffraction (XRD) and Fourier Transform Infrared Spectroscopy (FTIR) were employed to confirm the formation of cubic Y2O3 nanoparticles. Further, the interaction of Y2O3 with calf thymus-DNA were scrutinized by UV-visible, fluorescence spectroscopy and agarose gel electrophoresis techniques and the results suggested the stabilization of CT-DNA by the biosynthesized Y2O3. Also, the synthesized Y2O3 does not exhibit any toxicity in SK-MEL, human melanoma cell lines. Further, the studies on antifungal activities of Y2O3 nanoparticles revealed significant inhibitory effects on the growth of plant pathogen Exserohilum rostratum causing Exserohilum leaf blight in pearl millet. Hence, the findings deliver the foundation for furtherbiological research investigations of Y2O3 and its application in phytopathogen control.