Colorectal Carcinoma Cell Lines Research Articles

Activation of Epithelial-Mesenchymal Transition and Altered β-Catenin Signaling in a Novel Indian Colorectal Carcinoma Cell Line.

Colorectal cancer is the third major cause of cancer-related mortality worldwide. The upward trend in incidence and mortality rates, poor sensitivity to conventional therapies and a dearth of early diagnostic parameters pose a huge challenge in the management of colorectal cancer in India. Due to the high level of genetic diversity present in the Indian population, unraveling the genetic contributions toward pathogenesis is key for understanding the etiology of colorectal cancer and in reversing this trend. We have established a novel cell line, MBC02, from an Indian colorectal cancer patient and have carried out extensive molecular characterization to unravel the pathological alterations in this cell line. In-depth molecular analysis of MBC02 revealed suppression of E-cadherin expression, concomitant with overexpression of EMT related molecules, which manifested in the form of highly migratory and invasive cells. Loss of membrane-tethered E-cadherin released β-catenin from the adherens junction resulting in its cytoplasmic and nuclear accumulation and consequently, upregulation of c-Myc. MBC02 also showed dramatic transcriptional upregulation of β-catenin. Remarkably, we observed significantly elevated proteasome activity that perhaps co-evolved to compensate for the unnaturally high mRNA level of β-catenin to regulate the increased protein load. In addition, there was substantial misregulation of other clinically relevant signaling pathways that have clinical relevance in the pathogenesis of colorectal cancer. Our findings pave the way toward understanding the molecular differences that could define pathogenesis in cancers originating in the Indian population.

4-(2-(1H-Benzo[d]imidazol-2-ylthio)acetamido)-N-(substituted phenyl)benzamides: design, synthesis and biological evaluation

BackgroundDihydrofolate reductase (DHFR) is an important target for antimetabolite class of antimicrobials because it participates in purine synthesis. 2-mercaptobenzimidazole (2MBI) has similar structural features as purine nucleotides. Given that benzimidazole and similar heteroaromatics have been broadly examined for their anticancer potential, so, we hereby report the design, synthesis and biological studies (i.e. antimicrobial and anticancer studies) of 2MBI derivatives.MethodologyThe antimicrobial activity of synthesized 2MBI derivatives were evaluated against Gram positive and Gram negative bacterial species as well as fungal species by tube dilution technique whereas their anticancer activity was assessed against human colorectal carcinoma cell line (HCT116) by Sulforhodamine B (SRB) assay. They were also structurally characterized by IR, NMR, MS and elemental analyses.Results, discussion and conclusionThe antimicrobial activity findings revealed that compound N1 (MICbs,st,ca = 1.27, 2.54, 1.27 µM), N8 (MICec= 1.43 µM), N22 (MICkp,an= 2.60 µM), N23 and N25 (MICsa= 2.65 µM) exhibited significant antimicrobial effects against tested strains, i.e. Gram-positive, Gram-negative (bacterial) and fungal strains. The anticancer screening results demonstrated that compounds N9, N18 (IC50 = 5.85, 4.53 µM) were the most potent compounds against cancer cell line (HCT116) even more than 5-FU, the standard drug (IC50 = 9.99 µM).

Synthesis and Biological Evaluation of Some Novel Thiazole-Based Heterocycles as Potential Anticancer and Antimicrobial Agents.

A novel series of thiazole-based heterocycles was synthesized using 1,3-dipolar cycloaddition reactions in the presence of chitosan-grafted-poly(vinylpyridine) as an eco-friendly biopolymeric basic catalyst. The molecular structure of the synthesized compounds was illustrated by spectroscopic and elemental analysis. Various in vitro biological assays were performed to explore the potential antitumor, antimicrobial and hepatoprotective activities of the newly synthesized compounds. The cytotoxic activities were assessed against human hepatocellular carcinoma (HepG-2), colorectal carcinoma (HCT-116) and breast cancer (MCF-7) cell lines and results revealed that all compounds displayed antitumor activities with the chlorine-containing derivatives, 11c and 6g, being the most potent. The majority of the tested thiazole derivatives exhibited satisfactory antibacterial activity towards the used gram positive and gram-negative bacterial species. Moreover, many derivatives showed weak hepatoprotective activity against CCl4-induced hepatotoxicity.

Reduced aquaporin-1 transcript expression in colorectal carcinoma is associated with promoter hypermethylation

ABSTRACTAquaporin-1 (AQP1) is a homo-tetrameric transmembrane protein that facilitates rapid movement of water and ions across cell membranes. The clinical significance of AQP1 expression in colorectal carcinoma (CRC) is controversial. The aim of this study was to investigate the prognostic significance of AQP1 transcript expression and the association between expression and promoter methylation in normal colonic mucosa, CRC tissues and cell lines. Analysis of publicly available datasets from The Cancer Genome Atlas revealed that AQP1 expression was significantly decreased in CRC compared to normal mucosa (12.7 versus 33.3 respectively, P < 0.0001). However, expression increased with advanced disease, being significantly higher in stage IV (17.6) compared to either stage I (11.8, P = 0.0039) or II (10.9; P = 0.0023), and in patients with lymph node metastasis compared to those without (13.9 versus 11.3 respectively, P = 0.0023). Elevated expression was associated with decreased overall survival with univariate (Cox Proportional Hazard Ratio 1.60, 95% confidence interval 1.05–2.42, P = 0.028), but not multivariable analysis when considering the confounders stage and age. Analysis of HumanMethylation450 data demonstrated that AQP1 promoter methylation was significantly increased in CRC compared to normal mucosa. Analysis of CRC tissues and cell lines strongly suggested that methylation was associated with decreased expression. BRAFV600E mutation alone did not explain the increase in methylation. In conclusion, AQP1 transcript expression was decreased in CRC compared to normal mucosa, and this was associated with AQP1 promoter hypermethylation. AQP1 transcript expression increased with advanced disease but was not an independent prognostic indicator.

A Novel Small Molecule p53 Stabilizer for Brain Cell Differentiation.

Brain tumor, as any type of cancer, is assumed to be sustained by a small subpopulation of stem-like cells with distinctive properties that allow them to survive conventional therapies and drive tumor recurrence. Thus, the identification of new molecules capable of controlling stemness properties may be key in developing effective therapeutic strategies for cancer by inducing stem-like cells differentiation. Spiropyrazoline oxindoles have previously been shown to induce apoptosis and cell cycle arrest, as well as upregulate p53 steady-state levels, while decreasing its main inhibitor MDM2 in the HCT116 human colorectal carcinoma cell line. In this study, we made modifications in this scaffold by including combinations of different substituents in the pyrazoline ring in order to obtain novel small molecules that could modulate p53 activity and act as differentiation inducer agents. The antiproliferative activity of the synthesized compounds was assessed using the isogenic pair of HCT116 cell lines differing in the presence or absence of the p53 gene. Among the tested spirooxindoles, spiropyrazoline oxindole 1a was selective against the cancer cell line expressing wild-type p53 and presented low cytotoxicity. This small molecule induced neural stem cell (NSC) differentiation through reduced SOX2 (marker of multipotency) and increased βIII-tubulin (marker of neural differentiation) which suggests a great potential as a non-toxic inducer of cell differentiation. More importantly, in glioma cancer cells (GL-261), compound 1a reduced stemness, by decreasing SOX2 protein levels, while also promoting chemotherapy sensitization. These results highlight the potential of p53 modulators for brain cell differentiation, with spirooxindole 1a representing a promising lead molecule for the development of new brain antitumor drugs.

Development of novel apoferritin formulations for antitumour benzothiazoles.

BackgroundThe benzothiazole structure is important in medicinal chemistry, and 5‐fluoro‐2‐(3,4‐dimethoxyphenyl) benzothiazole (GW 610) is of particular interest as it shows outstanding anticancer activity in sensitive breast and colorectal carcinoma cell lines via generation of lethal DNA adducts in sensitive cancer cells. Despite promising activity, poor water solubility limits its applications. The apoferritin (AFt) protein cage has been proposed as a robust and biocompatible drug delivery vehicle.AimsHere, we aim to enhance solubility of GW 610 by developing amino acid prodrug conjugates and utilizing the AFt capsule as drug delivery vessel.Methods and resultsThe potent experimental antitumour agent, GW 610, has been successfully encapsulated within AFt with more than 190 molecules per AFt cage. The AFt‐GW 610 complex exhibits dose‐dependent growth inhibition and is more potent than GW 610 alone in 5/7 cancer cell lines. To enhance both aqueous solubility and encapsulation efficiency, a series of amino acid esters of GW 608 prodrug were synthesized via N,N′‐dicyclohexylcarbodiimide ester coupling to produce molecules with different polarity. A dramatic increase in encapsulation efficiency was achieved, with more than 380 molecules of GW 608‐Lys molecules per AFt cage. Release studies show sustained release of the cargo over 12 hours at physiologically relevant pH. The AFt‐encapsulated amino acid modified GW 608 complexes are sequestered more rapidly and exhibit more potent anticancer activity than unencapsulated agent.ConclusionThese results indicate that AFt‐encapsulation of GW 610 prodrug provides a biocompatible delivery option for this potent, selective experimental antitumour agent and for amino acid‐modified GW 608. Of particular interest is the encapsulation efficiency and in vitro antitumour activity of AFt‐GW 608‐Lys, which warrants further preclinical evaluation.

Polysaccharides from Nostoc commune Vaucher activate macrophages via NF-κB and AKT/JNK1/2 pathways to suppress colorectal cancer growth in vivo.

Colorectal carcinoma (CRC) is an aggressive malignancy with very limited therapeutic options. As a result, both morbidity and mortality of this malignancy are very high. It is well-established that Nostoc commune Vaucher is a type of healthy food and has been used as a traditional medicine against human cancers. However, the underlying mechanism(s) by which Nostoc commune Vaucher inhibits CRC was never described. Here we reported that polysaccharides from Nostoc commune Vaucher are a potent inhibitor of CRC growth by activating macrophages. Specifically, we purified polysaccharides from Nostoc commune and two fractions of these polysaccharides are able to inhibit the proliferation and growth of five human CRC cell lines with divergent genetic backgrounds. Through activating NF-κB and AKT/JNK1/2 signaling pathways, one fraction of polysaccharides significantly activated macrophages, which was reflected by the enlarged size of macrophages, enhanced phagocytosis and expression of TNF-α and IL-1β. In summary, this study demonstrated that the polysaccharides from Nostoc commune Vaucher are a potent activator of macrophages, which subsequently contributes to the inhibition of CRC.

Melatonin Sensitizes Human Colorectal Cancer Cells to γ-ray Ionizing Radiation In Vitro and In Vivo.

Colorectal cancer is the most commonly reported gastrointestinal malignancy, with a recent, rapid increase of the annual incidence all over the world. Enhancing the radiosensitivity of cancer cells while preserving the health of normal cells is one of the most important tasks in clinical radiobiology. However, resistance to radiotherapy for colorectal cancer greatly decreases the therapeutic outcome. Melatonin (N-acetyl-5-methoxytryptamine), a natural secretory product that the pineal gland in the brain normally produces, has been reported to have anticancer properties. In the study, we investigated the combination of melatonin with radiotherapy as a treatment for colorectal cancer. We firstly explored the anti-tumor activity of melatonin combined with ionizing radiation (IR) against colorectal carcinoma in vitro. It was found that melatonin effectively inhibited human colorectal carcinoma cell line HCT 116 cellular proliferation, colony formation rate and cell migration counts following IR. Increasing the radiosensitivity of colorectal cancer cells by melatonin treatment was found to be associated with cell cycle arrest in the G2/M phase, downregulation of proteins involved in DNA double-strand break repair and activation of the caspase-dependent apoptotic pathway. Moreover, we also investigated the combined effect of IR and melatonin on colorectal tumor in vivo. Results from a tumor xenograft showed that melatonin plus IR treatment significantly suppressed tumor cell growth compared with melatonin or IR alone, resulting in a much higher tumor inhibition rate for the combined treatment. The data suggested that melatonin combined with IR could improve the radiosensitivity of colorectal cancer and thus enhance the therapeutic effect of the patients, implying melatonin could function as a potential sensitizer in tumor radiotherapy.

Chebulinic acid derived from triphala is a promising antitumour agent in human colorectal carcinoma cell lines

BackgroundTriphala is an Ayurvedic rasayana formulation reputed for its antitumour activities, and chebulinic acid and chebulagic acid, along with other phenolic acids, have been proposed to be responsible for its effects.MethodsIn this study, the anti-proliferative activities of these agents were evaluated in colorectal carcinoma cell lines with three phenotypes exposed to several batches of triphala samples with different quantities of chebulinic acid and chebulagic acid. The pro-apoptotic and anti-migratory activities and the probable antitumour mechanisms of the more potent anti-proliferative phytochemical were also investigated.ResultsThe results demonstrated that chebulinic acid, which exerts potent anti-proliferative, pro-apoptotic and anti-migratory effects, is a key molecule for maintaining the antitumour efficacy of triphala. The antitumour mechanism of chebulinic acid is probably related to the PI3K/AKT and MAPK/ERK pathways.ConclusionsChebulinic acid is not only a critical component of the anticancer activities of triphala but also a promising natural multi-target antitumour agent with therapeutic potential.

Design, synthesis and therapeutic potential of 3-(2-(1H-benzo[d]imidazol-2-ylthio)acetamido)-N-(substituted phenyl)benzamide analogues

BackgroundThe emergence of bacterial resistance is a major public health problem. It is essential to develop and synthesize new therapeutic agents with better activity. The mode of actions of certain newly developed antimicrobial agents, however, exhibited very limited effect in treating life threatening systemic infections. Therefore, the advancement of multi-potent and efficient antimicrobial agents is crucial to overcome the increased multi-drug resistance of bacteria and fungi. Cancer, which remains as one of the primary causes of deaths and is commonly treated by chemotherapeutic agents, is also in need of novel and efficacious agents to treat resistant cases. As such, a sequence of novel substituted benzamides was designed, synthesized and evaluated for their antimicrobial and anticancer activities.MethodologyAll synthesized compounds were characterized by IR, NMR, Mass and elemental analysis followed by in vitro antimicrobial studies against Gram-positive (Staphylococcus aureus), Gram-negative (Salmonella typhi and Klebsiella pneumoniae) bacterial and fungal (Candida albicans and Aspergillus niger) strains by the tube dilution method. The in vitro anticancer evaluation was carried out against the human colorectal carcinoma cell line (HCT116), using the Sulforhodamine B assay.Results, discussion and conclusionCompound W6 (MICsa, st, kp = 5.19 µM) emerged as a significant antibacterial agent against all tested bacterial strains i.e. Gram-positive (S. aureus), Gram-negative (S. typhi, K. pneumoniae) while compound W1 (MICca, an = 5.08 µM) was most potent against fungal strains (A. niger and C. albicans) and comparable to fluconazole (MIC = 8.16 µM). The anticancer screening demonstrated that compound W17 (IC50 = 4.12 µM) was most potent amongst the synthesized compounds and also more potent than the standard drug 5-FU (IC50 = 7.69 µM).

Tumor suppressive function of E2F-1 on PTEN-induced serrated colorectal carcinogenesis.

Many human cancers present Phosphatase and tensin homolog (PTEN) deficiency and between 20 and 30% of colorectal tumors show PTEN loss. The transcription factor, E2 promoter binding factor 1 (E2F-1), exhibits tumor promoter or suppressive functions depending on cellular type and tissue context, but its role in the progression and development of colorectal carcinogenesis was largely unknown. Here, using a tamoxifen-inducible PTEN knockout mouse model, we have demonstrated that loss of PTEN leads to the development of colorectal tumorigenesis through the serrated pathway. Next, we studied PTEN loss-driven colorectal lesions in the context of E2F-1 deficiency in vivo. Our results revealed that monoallelic and biallelic absence of E2F-1 led to an increased incidence and progression of serrated tumorigenesis induced by PTEN loss. Finally, we investigated the mechanisms by which double PTEN/E2F-1 deficiency leads to enhanced tumorigenesis. We found that colorectal tumors from PTEN/E2F-1 double knockout mice and the human colorectal carcinoma cell line HT29 with shRNA-mediated downregulation of PTEN and E2F-1 exhibit hyperactivation of the RAS-MAPK pathway, accumulation of DNA damage and resistance to apoptosis. To date, this is the first preclinical study evaluating the effect of genetic deletion of E2F-1 in colorectal malignancies driven by PTEN deficiency. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Lactic acid bacteria isolated from fermented flour of finger millet, its probiotic attributes and bioactive properties

This study aims to isolate and identify lactic acid bacteria from fermented flour of selected finger millet varieties grown in Sri Lanka and to evaluate their probiotic attributes and bioactive properties in vitro. Fifteen lactic acid bacteria were isolated from three varieties of fermented finger millet flour namely ravi, raavana and oshadha. These isolates were screened for phenotypical and biochemical characteristics. The selected isolates were identified by 16 S rRNA sequencing as Bacillus cereus (five strains), Streptococcus lutetiensis, Lactobacillus plantarum, Lactobacillus fermentum (two strains), Brevibacillus borstelensis, Paenibacillus species, Lactococcus lactis subspecies lactis, Enterococcus faecium, Pediococcus acidilactici, and Enterococcus lactis, and their partial sequences were deposited in GenBank. Among them, five isolates including two isolates, L. plantarum MF405176.1 and L. fermentum MF033346.1 isolated from ravi; two isolates, L. lactis MF480428.1 and E. faecium MF480431.1 isolated from raavana; and P. acidilactici MF480434.1 isolated from oshadha varieties respectively, exhibited in vitro safety attributes and could tolerate acid, gastric juice, bile, salt, phenol, and temperature under simulated gastric conditions, and also were susceptible to antibiotics tested. Further, they demonstrated bactericidal activity against both drug-sensitive and multidrug-resistant pathogens. Among the selected isolates, L. plantarum MF405176.1 demonstrated highest hydrophobicity and adhesion to both colon colorectal adenocarcinoma and colon colorectal carcinoma cell lines. L. lactis subspecies lactis MF480428.1 exhibited the highest auto-aggregation and 2, 2, diphenyl-1-pricrylhydrazyl free radical scavenging activity. P. acidilactici MF480434.1 demonstrated the lowest IC50 values against HCT-116 and HT-29 cells. None of the LAB isolates could assimilate > 10% cholesterol in vitro.

The role of autophagy in the treatment of BRAF mutant colorectal carcinomas differs based on microsatellite instability status.

Autophagy has been identified as a catabolic mechanism in cells but its’ role in cancer remains controversial. Autophagy has been characterized either as tumor suppressor or inducer mechanism in many tumor types. Monoclonal antibodies against EGFR (cetuximab and panitumumab) represent a major step in the treatment of mCRC. Several studies propose that cetuximab and panitumumab trigger autophagy which reveals a potential resistance mechanism to these agents. The last years immunotherapy appears to be a novel promising strategy for the treatment of patients with solid tumors, including colorectal cancer. Checkpoint inhibitors, such as anti-PD1 (nivolumab and pembrolizumab) and anti-CTLA-4 (ipilimumab) antibodies have already been developed and applied in mCRC patients with MSI-H phenotype. The association between mtBRAF and autophagy or MSI status has already been characterized. In our study, we identify the autophagy initiation through anti-EGFR monoclonal antibodies and checkpoint inhibitors in colorectal carcinoma cell lines according to microsatellite status. The combination of autophagy inhibition, anti-EGFR antibodies and checkpoint inhibitors as well as autophagy targeting, MEK inhibition and anti-EGFR antibodies or checkpoint inhibitors appears to be the best treatment approach for microsatellite instability high and stable colorectal cancer cell lines, respectively. Both combinatorial approaches reduce cell viability through the induction of apoptotic cell death. The findings of this study point out the importance of different approach for the treatment of BRAF mutant metastatic colorectal cancers based on their microsatelite instability phenotype.

Biological evaluations of newly-designed Pt(II) and Pd(II) complexes using spectroscopic and molecular docking approaches

To perform biological evaluations of newly-designed Pt(II) and Pd(II) complexes, the present study was conducted with targeted protein human serum albumin (HSA) and HCT116 cell line as model of human colorectal carcinoma. The binding of Pt(II) and Pd(II) complexes to HSA was analyzed using fluorescence spectroscopy and molecular docking. The thermal stability and alterations in the secondary structure of HSA in the presence of Pt(II) and Pd(II) complexes were investigated using the thermal denaturation method and circular dichroism (CD) spectroscopy. The cytotoxicity of the Pt(II) and Pd(II) complexes was studied against the HCT116 cell line using MTT assay. The binding analysis revealed that the fluorescence findings were well in agreement with docking results such that there is only one binding site for each complex on HSA. Binding constants of 8.7 × 103 M−1, 2.65 × 103 M−1, 0.3 × 103 M−1, and 4.4 × 103 M−1 were determined for Pd(II) and Pt(II) complexes (I–IV) at temperature of 25 °C, respectively. Also, binding constants of 1.9 × 103 M−1, 15.17 × 103 M−1, 1.9 × 103 M−1, and 13.1 × 103 M−1 were determined for Pd(II) and Pt(II) complexes (I–IV) at temperature of 37 °C, respectively. The results of CD and thermal denaturation showed that the molecular structure of HSA affected by interaction with Pt(II) and Pd(II) complexes is stable. Cytotoxicity studies represented the growth suppression effect of the Pt(II) and Pd(II) complexes toward the human colorectal carcinoma cell line. Therefore, the results suggest that the new designed Pt(II) and Pd(II) complexes are well promising candidates for use in cancer treatment, particularly for human colorectal cancer.Communicated by Ramaswamy H. Sarma

Radiosensitizing effect of the fucoidan from brown alga Fucus evanescens and its derivative in human cancer cells

Fucoidan from brown alga Fucus evanescens and its product of enzymatic hydrolysis have precisely established structure and possess significant biological activities.The aim of present study was to determine radiosensitizing activity of fucoidan from brown alga F. evanescens and its derivative in human melanoma, breast adenocarcinoma, and colorectal carcinoma cell lines and elucidate mechanism of their action.The fucoidan from F. evanescens and its derivative had a comparable radiosensitizing activity and increased the inhibiting effect of X-ray radiation on proliferation and colony formation of human cancer cells, with significant inhibition of melanoma cells. The molecular mechanism of this action was associated with the induction of apoptosis by activating the initiator and effector caspases, suppressing the expression of the anti-apoptotic protein, and enhancing the fragmentation of DNA. The obtained data confirm the prospects of using fucoidan’s derivative in combination with radiation therapy for the improvement of the schemes of cancer therapy.

CRP and LOX-1: a Mechanism for Increasing the Tumorigenic Potential of Colorectal Cancer Carcinoma Cell Line.

Chronic inflammation and dyslipidemia are associated with an increase in the incidence of colorectal cancer (CRC). Serum C- reactive protein (CRP) and oxidized low-density lipoprotein (oxLDL), as Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) ligands, increase during inflammation and dyslipidemia, respectively. To evaluate the effects of CRP on the expression of important genes involved in the development of CRC, the CRC cell line, LS174T, was treated with the commercial CRP. Based on the Real-time PCR data, in the presence of CRP, LOX-1, CEA, MMP1, and MMP2 mRNA expression significantly increased, compared to the control group. Moreover, in the presence of CRP, secretion, and expression of CEA in the cell lysate and conditioned media increased in a concentration-dependent manner. The results of flow cytometry showed that expression of LOX-1 receptors at the cell surface increased significantly in the presence of 10mg/L of CRP. However, inhibition of LOX-1 receptors with a specific monoclonal antibody reduced the effects of CRP on protein/mRNA expression. In conclusion, Increased CRP level, can potentially elevate the expression of important genes in CRC by stimulating LOX-1 receptors.

The Effect of Different Treatments of (–)-Epigallocatechin-3-Gallate on Colorectal Carcinoma Cell Lines

Backgroud: (-)-Epigallocatechin-3-gallate (EGCG), the major component of green tea, is well documented to induce apoptosis and cell cycle arrest in cancer by targeting multiple signal transduction pathways. However, EGCG is extremely unstable in general culture conditions and rapidly degraded. So, to what extent EGCG or which degradation products of EGCG play a role in anti-tumor is still unknown. In this study, we evaluated the effect of different treatments of EGCG on HCT116 cells.Design: MTT assay was applied to evaluated the inhibitory effect of different treatments of EGCG on HCT116 cells. Cell cycle and apoptosis were performed by flow cytometry. Finally, western blot analysis was used to elucidate the molecular mechanism associated with cell cycle arrest and apoptosis.Results: Compared with control, both EGCG and O-EGCG (i.e., EGCG being pre-incubated at 37°C for 3 h) significantly inhibited HCT116 cells growth. Surprisingly, we found that the inhibitory effect of O-EGCG was stronger than that of EGCG. The IC50 values of EGCG and O-EGCG were 8.75 and 5.40 μM, respectively. Cell cycle analysis showed that 20 μM of EGCG simultaneously caused cell cycle arrest at G1 and G2 phase in HCT116 cells, differing from O-EGCG which exclusively caused cell cycle arrest at G2. This result suggested that parent EGCG at the early treatment might cause cell cycle arrest at G1. As time went on, EGCG disappeared and degraded products of EGCG were formed which might cause cell cycle arrest at G2. Further studies revealed that EGCG induced cell cycle arrest at G1 by downregulation of cyclin E and cyclin D1 and upregulation of p21. On the other hand, O-EGCG induced HCT116 cells apoptosis mainly by increasing the expression of p53 and cleaved caspase-3, which might be the underlying reason why O-EGCG had stronger inhibitory effect on HCT116 cells line than EGCG.Conclusions: The pretreatment of EGCG may be an effective way to enhance its antitumor effect.

Different Cytotoxic Effects of Caper from Different Geographical Regions May Be Related to Changes in Mitochondrial Sirt3

Background and objectives: Beside its nutritional role, caper (Capparis spinosa) has long been used as an analgesic, anti-inflammatory, anti-diabetic and anti-cancer remedy. In the present study, we tested whether this plant can make effective changes in Sirt3 and mitochondrial function in colorectal carcinoma cell line since mitochondrial dysfunction has long been implicated in both cancer and diabetes and benefit confers by caper in these diseases might be due to mitochondrial alterations. Methods: Total flavonoids and phenolics were assayed using colorimetric tests. Cytotoxicity of a phenolic-flavonoid rich extract of caper collected from two different geographical regions (south and west) were mechanistically studied in HT-29 cell line. Activity of an essential mitochondrial enzyme, Sirt3 has also been evaluated along with other parameters. IC50 of extracts were determined by MTT cytotoxicity assay, cell death and mitochondrial membrane potential were evaluated via flow cytometric analysis. Also, at IC50 concentrations, Sirt3 activity was determined fluorimetrically. Results: The results showed that caper induced significant cytotoxicity in HT-29 cells followed by mitochondrial membrane potential collapse, ROS overproduction, Sirt3 activity alteration and cell death. Conclusions: The above-mentioned cytotoxic parameters were inversely proportional to the phenolic and flavonoid contents of the extract showing that other mechanisms beyond their antioxidant capacities may contribute to their anti-cancer effects. In other term, these results suggest that antioxidant capacity may not directly contribute to the anticancer property.

Antitumor effects of the silencing of programmed cell death ligand1 in colorectal cancer via immunoregulation.

Activation of programmed cell death 1 (PD-1)/PD-ligand 1 (PD-L1) can promote immune suppression of the tumor microenvironment. However, the effects and mechanisms of PD-L1 silencing on colorectal cancer growth are largely unknown. In the present study, PD-L1 expression was compared in colorectal cancer and paracancerous tissues by immunofluorescence. A stable colorectal carcinoma cell line encoding PD-L1 short hairpin RNA (shRNA) was established. Thereafter, inoculated tumors were modeled in C57B/L6 mice. Experiments were divided into 3 groups: Control group, vector group, and PD-L1 silencing group (inoculated with the stable CT26 cell line encoding PD-L1 shRNA). Following decapitation of the mice, tumors were weighed and apoptosis of tumor cells was detected. The number and viability of cluster of differentiation (CD)4+ and CD8+ T cells were analyzed by flow cytometry and a cell counting kit assay, respectively. Compared with paracancerous tissue, colorectal cancer tissue extensively expressed PD-L1, RAC-α serine/threonine-protein kinase (AKT), and phosphatidylinositol 3-kinase (PI3K). Lymphocyte-activating gene 3 (LAG-3) expression was observed at the edge of tumor tissue, but rarely observed in paracancerous tissue. A stable CT26 cell line encoding PD-L1 shRNA was established, and lack of PD-L1 expression was confirmed by reverse transcription-polymerase chain reaction and western blotting. Compared with the control, the shPD-L1 group demonstrated reduced tumor growth, a high level of apoptosis in tumor cells, a low level of PI3K and AKT expression, and an increased number of cells and greater activity of CD4+ T and CD8+ T cells. Taken together, PD-L1 silencing promoted tumor cell apoptosis, at least in part, through the activation of CD4+ and CD8+ T cells.

Preparation, formation mechanism, photocatalytic, cytotoxicity and antioxidant activity of sodium niobate nanocubes.

A hydrothermal method was employed to prepare the sodium niobate (NaNbO3) nanocubes. We executed time dependent experiments to illustrate the formation mechanism of sodium niobate nanocubes. It was observed that the morphology of NaNbO3 nanocubes was dependent on the reaction time and 12hr reaction time was found to be suitable. Morphology, composition, structure and optical properties of sodium niobate nanocubes were evaluated by scanning electron microscope, X-ray energy-dispersive spectrometer, X-ray diffraction and UV-visible diffuse reflectance spectrometer. The photocatalytic activity of sodium niobate was studied for photocatalytic hydrogen production. It was anticipated that the sodium niobate (NaNbO3) cubes exhibited good photocatalytic activity under UV light irradiation using lactic acid as sacrificial agent. The cytotoxicity activity of sodium niobate nanocubes was studied as well at different concentrations (5 mg/mL, 3 mg/mL, 1 mg/mL, and 0.25 mg/mL) against human colon colorectal carcinoma cell line (HCT116) by MTT assay and EC50 was found to be 1.9 mg/mL. Sodium niobate proved to be a good DPPH free radical scavenging material, tested at different concentrations. It was noticed that peak intensity at 517 nm was decreased after 30 minute incubation, further supporting the antioxidant activity. This study will be useful for design and engineering of materials that can be used in biomedical applications and in photocatalysis.