Apoptosis In SW480 Cells Research Articles

Polyphyllin I Promotes Autophagic Cell Death and Apoptosis of Colon Cancer Cells via the ROS-Inhibited AKT/mTOR Pathway

Colon cancer is a common malignant tumor of the digestive tract, and it is considered among the biggest killers. Scientific and reasonable treatments can effectively improve the survival rate of patients if performed in the early stages. Polyphyllin I (PPI), a pennogenyl saponin isolated from Paris polyphylla var. yunnanensis, has exhibited strong anti-cancer activities in previous studies. Here, we report that PPI exhibits a cytotoxic effect on colon cancer cells. PPI suppressed cell viability and induced autophagic cell death in SW480 cells after 12 and 24 h, with the IC50 values 4.9 ± 0.1 μmol/L and 3.5 ± 0.2 μmol/L, respectively. Furthermore, we found PPI induced time-concentration-dependent autophagy and apoptosis in SW480 cells. In addition, down-regulated AKT/mTOR activity was found in PPI-treated SW480 cells. Increased levels of ROS might link to autophagy and apoptosis because reducing the level of ROS by antioxidant N-acetylcysteine (NAC) treatment mitigated PPI-induced autophagy and apoptosis. Although we did not know the molecular mechanism of how PPI induced ROS production, this is the first study to show that PPI induces ROS production and down-regulates the AKT/mTOR pathway, which subsequently promotes the autophagic cell death and apoptosis of colon cancer cells. This present study reports PPI as a potential therapeutic agent for colon cancer and reveals its underlying mechanisms of action.

Diprenylated flavonoids from licorice induce death of SW480 colorectal cancer cells by promoting autophagy: Activities of lupalbigenin and 6,8-diprenylgenistein

Ethnopharmacological relevanceLicorice is a well-known herbal medicine, and we previously found that several licorice prenylated flavonoids could cause death of SW480 colorectal cancer cells by promoting autophagy. Given many kinds of prenylated flavonoids in licorice, the activities of other compounds deserve further investigation. In addition, the contribution of isoprenyl groups on the autophagy promotion activities has not been clarified. Aim of the studyThis study aimed to investigate whether lupalbigenin (LPB) and 6,8-diprenylgenistein (DPG), two licorice diprenylated flavonoids, could induce autophagic cell death of SW480 cells, and clarify the contribution of isoprenyl groups. Materials and methodsCytotoxic activities of LPB and DPG were tested by using an MTT method, and apoptosis induction effects were evaluated by PI-Annexin V staining-based flow cytometry and protein levels of caspase-3 and PARP-1. Autophagy promotion effects of LPB and DPG were assessed by protein levels of LC3, p62, Akt and mTOR as well as number of autophagosomes in cells, and autophagy inhibitor chloroquine (CQ) was involved to identify the role of autophagy on LPB or DPG-caused death of SW480 cells. In addition, two groups of structurally similar diprenylated, mono-prenylated and free flavonoids were obtained from licorice, which were used to investigate the contribution of isoprenyl groups on their autophagy promotion activities. ResultsBoth LPB and DPG significantly induced apoptosis of SW480 cells with strong cytotoxic activities, and meanwhile, they also promoted autophagy probably through the Akt/mTOR signaling pathway. Further studies indicated that LPB and DPG could induce autophagic cell death of SW480 cells. Moreover, isoprenyl groups contributed mainly to the cytotoxic and autophagy promotion activities of licorice prenylated flavonoids. ConclusionThis study reported for the first time that licorice diprenylated flavonoids LPB and DPG induced death of SW480 cells by promoting autophagy, which was mainly attributed to the isoprenyl groups. The results provided theoretical basis for researches on anti-colorectal cancer drugs and their structural modification.

Structurally diverse diterpenoids from Isodon oresbius and their bioactivity

Twelve new diterpenoids, isoresbins A–L (1–12), together with twenty-eight known ones, were isolated from the aerial parts of Isodon oresbius. Their diverse structures included 6,7-seco-ent-kaurane, 7,20-epoxy-ent-kaurane, 6,7:8,15-diseco-ent-kaurane, and abietanes skeletons, which were elucidated by spectroscopic data interpretation, single-crystal X-ray diffraction, and quantum chemical calculation. Isoresbins A (1) and B (2) possessed a new rearranged 15(8 → 11)-abeo-6,7-seco-ent-kaurane skeleton. 1 and 5 promoted lysosomal function, which was evaluated by LysoTracker Red staining and DQ-ovalbumin dequenching assay. 1 showed cytotoxicity against six human tumor cell lines with IC50 values in 2.07–4.04 μM range. Moreover, 1 induced damage of mitochondrial membrane potential, G2/M cell cycle arrest and apoptosis in SW480 cells.

Suppression of colitis-associated colorectal cancer by scutellarin through inhibiting Hedgehog signaling pathway activity

BackgroundColitis-associated colorectal cancer (CAC) is a specific type of colorectal cancer (CRC) and mainly develops from long-term intestinal inflammation. Mounting evidence reveals that activated Hedgehog signaling pathway plays a vital role in the pathogenesis of CRC. Scutellarin is a type of phytochemical flavonoid with a powerful efficacy on various malignancies, including CRC. AimHere, we studied the therapeutic effect of scutellarin on CRC and its direct regulating targets. MethodsThe CAC model in mice was established by azomethane oxide (AOM) and sodium dextran sulfate (DSS), followed by detection of the efficacies of scutellarin on the carcinogenesis, apoptosis, inflammation, Hedgehog signaling cascade and complicated inflammatory networks in CAC tissues of mice. In CRC SW480 cells, the effects of scutellarin on malignant phenotype, apoptosis and Hedgehog signaling were examined. In TNF-α-stimulated IEC-6 intestinal epithelial cells, the actions of scutellarin on inflammatory response and Hedgehog signals were assessed as well. ResultsScutellarin significantly ameliorated AOM/DSS-caused CAC in mice and induced apoptosis in CAC tissues of mice, by inhibiting NF-κB (nuclear factor kappa B) -mediated inflammation and Hedgehog signaling axis. RNA-seq and transcriptome analysis indicated that scutellarin regulated complicated inflammatory networks in mouse CAC. Also, scutellarin suppressed the proliferation, migration, colony formation, and induced apoptosis of SW480 cells by down-regulation of Hedgehog signaling pathway activity. Additionally, scutellarin lessened NF-κB-mediated inflammatory response in TNF-α-stimulated IEC-6 cells, by attenuating Hedgehog signaling cascade. ConclusionScutellarin potently ameliorates CAC by suppressing Hedgehog signaling pathway activity, underpinning the promising application of scutellarin to CRC in clinical settings.

Role of ubiquitin-specific peptidase 22 in multidrug resistance of colorectal cancer and its correlation with multidrug resistance gene P-gp

Objective: To investigate the role of ubiquitin-specific peptidase 22 (USP22) in colorectal cancer multidrug resistance and its correlation with multidrug resistance genes P-gp and MRP1, and to preliminarily explore the mechanism of USP22 affecting colorectal cancer resistance. Methods: USP22 over-expression plasmid was transfected into colorectal cancer cells (RKO, SW480)with low expression of USP22. Cell counting kit (CCK-8) assay was used to detect the effect of USP22 on oxaliplatin resistance in colorectal cancer cells. The cells were treated with oxaliplatin of the same concentration. Western blot method was used to detect the expression of apoptosis-related proteins cleaved-caspase3, Bcl-2, and drug resistance proteins MRP1, P-gp in the cells. The cell efflux test was used to detect the effect of up-regulated USP22 on Calcein-AM and rhodamine123. Immunohistochemical methods were used to detect the expressions of USP22 and P-gp in the oxaliplatin chemotherapy-sensitive group and the drug-resistant group and to analyze the correlation between USP22 and MRP1, P-gp. Results: CCK-8 assay showed that the IC50 values of SW480-USP22 (SW480 cells overexpressing USP22) treated with oxaliplatin for 24 h and 48 h was (4.62±0.05)μmol/L and (2.32±0.04)μmol/L respectively; which was 2.7 times and 3.0 times higher than that in control cells, respectively. After treating with 1.25 μmol/L oxaliplatin for 48 h, USP22 overexpression can inhibit SW480 cells apoptosis. The fluorescence intensity of calcein-AM and rhodamine123 in the SW480-USP22 group were significantly increased when compared with that in the control cells (both P<0.01). The protein expression levels of MRP1 and P-gp in SW480-USP22 cells were significantly increased when compared with that in the control cells(both P<0.01). Immunohistochemistry showed that the positive expression rates of USP22, MRP1, and P-gp in the oxaliplatin chemotherapy-sensitive group were significantly lower than those in the chemotherapy-resistant group, the difference was statistically significant (all P<0.05), and USP22 was positively correlated with the expressions of MRP1 and P-gp in colorectal cancer tissues (r1=0.377, r2=0.423, both P<0.05). Conclusions: The up-regulation of USP22 is related to the acquired resistance of colorectal cancer cells to oxaliplatin. USP22 may be involved in the process of platinum-based chemotherapy resistance of colorectal cancer by regulating the expressions of P-gp and MRP1.

Inhibition of autophagy enhances apoptosis induced by Ce6-photodynamic therapy in human colon cancer cells

ObjectiveTo evaluate the therapeutic effect of Chlorin e6 photodynamic therapy (Ce6-PDT) in human colorectal cancer cells and investigate the role of autophagy in Ce6-PDT. MethodsSW480 cells underwent Ce6-PDT with and without pretreatment with the autophagy inhibitor 3-methyladenine (3MA). Cell viability was assessed using 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Apoptosis was evaluated using an Annexin V assay, using a rhodamine 123 (RH123) assay to evaluate mitochondrial membrane potential (MMP), and by measuring Caspase-3 and Bcl-2 protein expression using western blotting. Autophagy was evaluated by directly visualizing acridine orange-stained acidic vesicular organelles (AVOs) using fluorescent microscopy and by measuring LC3Ⅰ/Ⅱand Atg5 expression using western blotting. ResultsCe6-PDT decreased SW480 viability in a dose-dependent manner. Ce6-PDT induced apoptosis in SW480 cells via the mitochondrial apoptosis pathway as indicated by decreased mitochondrial membrane potential, increased Annexin V staining, and increased Caspase-3 expression. Ce6-PDT was also shown to induce autophagy as demonstrated by increased acridine-orange stained AVOs as well as increased expression of the autophagy-associated proteins Atg5. Inhibition of autophagy with 3MA potentiated SW480 cell response to Ce6-PDT and increased the rate of apoptosis in the treated cells. ConclusionsCe6-PDT induces autophagy and apoptosis of SW480 cells in a dose-dependent manner. Inhibition of autophagy increases the apoptosis induced by Ce6-PDT. Modulation of autophagy may be a potential therapeutic target for colon cancer cells treated with Ce6-PDT.

Assessing the photodynamic therapeutic effects of 5‑aminolevulinic acid on human colon cancer cells using light‑emitting diodes

The minimal invasiveness and high selectivity of 5‑aminolevulinic acid (5‑ALA)‑based photodynamic therapy (PDT; 5‑ALA‑PDT) renders it a viable therapeutic option for the treatment of various types of cancer. Compared with conventional lasers, light‑emitting diodes (LEDs) are an inexpensive and convenient low‑energy light source option. Nevertheless, the scope of LEDs in the 5‑ALA‑PDT of colorectal cancer (CRC) has yet to be fully determined. Thus, the aim of the present study was to assess the efficacy of LEDs in the 5‑ALA‑PDT of colon cancer <em>in vitro</em> by evaluating cytotoxic activity. 5‑ALA‑treated human CRC cells (SW480) were irradiated with LEDs of varying wavelengths: Red (630 nm), green (515 nm), blue (456 nm) and violet (399 nm). An MTS assay was conducted to determine cell viability. Additionally, a concentration‑response experiment was conducted with the most therapeutic wavelength (violet) to examine the 5‑ALA pharmacodynamics <em>in vitro</em>. The results revealed that only violet light in 5‑ALA‑PDT produced antitumour activity; this combination alone produced a drug concentration‑ and energy‑related decrease in cell viability. The decrease in viability was partially reversed by 3‑methyladenine, but not by Z‑VAD(OMe)‑FMK, suggesting that 5‑ALA induced the autophagy, but not the apoptosis of SW480 cells. The nature of the multi‑well plates used markedly affected the effectiveness of PDT. Black‑walled plates appeared to protect approximately 25% of cells from the effects of PDT. By contrast, clear plates permitted light access to the wells, even when protected from direct PDT treatment. On the whole, the findings of the present study indicate that the use of LEDs in 5‑ALA‑PDT <em>in vitro</em> induce the fluence‑dependent tumour cell death of SW480 cells. The choice of multi‑well plates greatly affects the results obtained <em>in vitro</em>. The antitumour effect was high with violet light. Hence, the use of LEDs in 5‑ALA‑PDT may prove to be an effective potential treatment for CRC.

Curcumin Mediates the Proliferation and Apoptosis of Colorectal Cancer Cells by Downregulating the Expression of Interleukin-1β through the Nuclear Factor-κB Signaling Pathway

Background: Colorectal cancer (CRC) is a frequently occurring malignant tumor, which is mainly observed in elderly men with no significant symptoms at the early stage. Among the malignant tumors of the digestive system, the incidence and mortality of CRC rank second only to hepatic and gastric cancer. Curcumin is an antioxidant and anti-inflammatory compound extracted from the roots of Curcuma longa plant. The antitumor effects of curcumin have been widely reported for various types of cancers, including CRC. Objective: In this study, we aimed to elucidate the protective effects and mechanism of interleukin (IL)-1 β on curcumin-induced apoptosis in SW480 cells. Materials and Methods: Expression levels of IL-1 β in CRC tissues and cells were detected by the quantitative reverse transcription polymerase chain reaction and Western blot assays. Followed by the incubation of cells with curcumin, the effect on IL-1 β was measured. Moreover, after transfection, the effects of IL-1 β on curcumin-induced SW480 cellular processes were analyzed by cell counting kit-8 and flow cytometric analysis. Results: According to the results of this study, IL-1 β was significantly increased in CRC tissues and cells. However, after incubation of the cells with curcumin, IL-1 β was downregulated and overexpression of IL-1 β counteracted the antitumor functions of curcumin in SW480 cells. Further studies have shown that curcumin could promote apoptosis of SW480 cells by inhibiting nuclear factor-κB (NF-κB) signaling pathway. Conclusion: Our study validated that curcumin inhibits SW480 cell proliferation but promotes apoptosis by downregulating the expression of IL-1 β probably through NF-κB signaling pathway. IL-1 β may an important target for the treatment of CRC.

Tetramethylpyrazine inhibits proliferation of colon cancer cells in vitro.

BACKGROUNDColon cancer is one of the most common malignancies worldwide, and chemotherapy is a widely used strategy in colon cancer clinical therapy. However, chemotherapy resistance is a major cause of disease recurrence and progression in colon cancer, and thus novel drugs for treatment are urgently needed. Tetramethylpyrazine (TMP), a component of the traditional Chinese medicine Chuanxiong Hort, has been proven to exhibit a beneficial effect in tumors. AIMTo investigate the potential anticancer activity of TMP in colon cancer and its underlying mechanisms.METHODSColon cancer cells were incubated with different concentrations of TMP. Cell viability was evaluated by crystal violet staining assay and cell counting kit-8 assay, and cell apoptosis and cell cycle were assessed by flow cytometry.RESULTSTMP significantly inhibited the proliferation of colon cancer cells in a dose- and time-dependent manner. In addition, flow cytometry revealed that TMP induced cell cycle arrest at the G0/G1 phase. TMP treatment caused early stage apoptosis in SW480 cells, whereas it caused late stage apoptosis in HCT116 cells.CONCLUSIONOur studies demonstrated that TMP inhibits the proliferation of colon cancer cells in a dose- and time-dependent manner by inducing apoptosis and arresting the cell cycle at the G0/G1 phase. Our findings suggest that TMP might serve as a potential novel therapeutic drug in the treatment of human colon cancer.

Antrodia salmonea induces apoptosis and enhances cytoprotective autophagy in colon cancer cells.

A traditional Chinese medicinal fungus, Antrodia salmonea (AS), with antioxidant properties is familiar in Taiwan but anti-cancer activity of AS in human colon cancer is ambiguous. Hence, we explored the anti-cancer activity of AS in colon cancer cells. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay revealed that AS showed a remarkable effect on cell viability in colon cancer cells; SW620, HCT116, and HT29. Annexin V/propidium iodide (PI) stained cells indicated that AS induced both early/late apoptosis in SW620 cells. Additionally, cells treated with AS induced caspase-3 activation, poly (ADP-ribose) polymerase (PARP) cleavage, mitochondrial dysfunction, and Bcl-2 associated X (Bax)/B-cell lymphoma (Bcl-2) dysregulation. Microtubule- associated protein 1A/1B-light chain 3B (LC3-II) accumulation, sequestosome 1 (p62/SQSTM1) activation, autophagy related 4B cysteine peptidase (ATG4B) inactivation, acidic vesicular organelles (AVOs) formation, and Beclin-1/Bcl-2 dysregulation revealed that AS-induced autophagy. Interestingly, cells pretreated with 3-methyladenine (3-MA) strengthened AS-induced caspase-3/apoptosis. Suppression of apoptosis by z-Val-Ala-Asp fluoromethyl ketone (Z-VAD-FMK) did not however block AS-induced autophagy, suggesting that autophagy was not attenuated by the AS-induced apoptosis. Application of N-acetylcysteine (NAC) prevented AS-induced cell death, caspase-3 activation, LC3-II accumulation, and AVOs formation, indicating that AS-induced apoptosis and autophagy was mediated by reactive oxygen species (ROS). Furthermore, AS-induced cytoprotective autophagy and apoptosis through extracellular signal-regulated kinase (ERK) signaling cascades. Moreover, in vivo data disclosed that AS inhibited colitis-associated tumorigenesis in azoxymethane (AOM)-dextran sodium sulphate (DSS)-treated mice. For the first time, we report the anti-cancer properties of this potentially advantageous mushroom for the treatment of human colon cancer.

SPRY4 suppresses proliferation and induces apoptosis of colorectal cancer cells by repressing oncogene EZH2.

Colorectal cancer (CRC), a common malignant tumor in the digestive tract, is a leading cause of cancer-related death. SPRY4 has been reported to act as a tumor suppressor gene in various tumors. This study aims to assess the role of SPRY4 in colorectal cancer (CRC) and uncover its underlying mechanisms. Firstly, the expression levels of SPRY4 were measured in CRC cell lines. SPRY4-overexpressing or silencing plasmids were transfected into CRC cells to regulate its expression level. CCK-8, colony formation, EdU assay, wound-healing and Transwell assays were performed to determine cell proliferation, invasion and migration abilities. Then, apoptosis was measured by flow cytometry analysis, and the expression of apoptosis-related protein was analyzed by western-blotting. Next, the in vivo tumorigenesis assay was performed in nude mice. According to the results, there was a lower expression of SPRY4 in CRC cell lines compared with normal cell line, and the overexpression of SPRY4 significantly suppressed cell proliferation, migration and invasion, and promoted apoptosis in SW480 cells. Moreover, the enhanced proliferation, invasion and migration upon SPRY4 silencing was reversed by EZH2 inhibition. In addition, we found that the overexpression of SPRY4 inhibited tumorigenesis in vivo by diminishing the size and weight of the tumors. Our study indicates that SPRY4 might be a potential tumor suppressor gene and prognostic factor for patients with CRC.

Long non-coding RNA LINC01106 regulates colorectal cancer cell proliferation and apoptosis through the STAT3 pathway

To investigate the expression of LINC01106 in colorectal cancer and its role in regulating the proliferation and apoptosis of colorectal cancer cells. We analyzed the data of LINC01106 expression levels in tumor tissues and normal tissues of patients with colorectal cancer in TCGA database and explored the association of LINC01106 expression level with the prognosis of the patients. Colorectal cancer SW480 cell lines with LINC01106 knockdown or overexpression were established, and their proliferation and apoptosis relative to the parental cells were evaluated using CCK-8 assay and flow cytometry, respectively. The expressions of p-STAT3, STAT3, and Bcl-2 in the cells were detected by immunoblotting. Nude mouse models bearing xenografts of SW480 cells with LINC01106 knockdown or na?ve SW480 cells were established to observe the effect of LINC01106 knockdown on the growth of SW480 cells in vivo. Analysis of the data from TCGA database showed that the expression level of LINC01106 was significantly higher in colorectal cancer tissues than in normal tissues, and LINC01106 expression level was significantly related to the prognosis of the patients (P < 0.05). Knockdown of LINC01106 significantly inhibited the proliferation and promoted apoptosis of SW480 cells (P < 0.05), while LINC01106 overexpression significantly promoted proliferation of the cells. LINC01106 knockdown in SW-480 cells obviously lowered the expressions of p- STAT3 and Bcl-2 and suppressed the growth of the xenograft in nude mice. LINC01106 is significantly up-regulated in colorectal cancer tissue and is related to the prognosis of the patients. LINC01106 can regulate the proliferation and apoptosis of SW480 cells through STAT3/Bcl-2 signaling and may serve as a potential marker for the diagnosis and prognostic evaluation of colorectal cancer.

Design, synthesis and evaluation of novel indirubin-based N-hydroxybenzamides, N-hydroxypropenamides and N-hydroxyheptanamides as histone deacetylase inhibitors and antitumor agents

Several novel indirubin-based N-hydroxybenzamides, N-hydropropenamides and N-hydroxyheptanamides (4a-h, 7a-h, 10a-h) were designed using a fragment-based approach with structural features extracted from several previously reported HDAC inhibitors, such as SAHA (vorinostat), MGCD0103 (mocetinostat), nexturastat A and PXD-101 (belinostat). The biological results reveal that our compounds showed excellent cytotoxicity toward three common human cancer cell lines (SW620, PC-3 and NCI-H23) with IC50 values ranging from 0.09 to 0.007 µM. The cytotoxicity of the compounds was equipotent or even up to 10-times more potent than adriamycin and up to 205-times more potent than SAHA. Among the series of N-hydroxypropenamides, compounds 10a-d were the most potent HDAC inhibitors as well as cytotoxicity toward the cell lines tested. In addition, the strong inhibitory activites toward HDAC of our compounds were observed with IC50 values of below-micromolar range. Especially, compound 4a inhibited HDAC6 with an IC50 value of 29-fold lower than that against HDAC2 isoform. Representative compounds 4a and 7a were found to significantly arrest SW620 cells at G0/G1 phase. Compounds 7a and 10a were found to strongly induce apoptosis in SW620 cells. Docking studies revealed some important features affecting the selectivity against HDAC6 isoform. The results clearly demonstrate the potential of the indirubin-hydroxamic acid hybrids and these compounds should be very promising for further development.

MiR-581/SMAD7 Axis Contributes to Colorectal Cancer Metastasis: A Bioinformatic and Experimental Validation-Based Study.

Metastasis is a well-known poor prognostic factor and primary cause of mortality in patients with colorectal cancer (CRC). Recently, with the progress of high through-put sequencing, aberrantly expressed non-coding RNAs (ncRNAs) were found to participate in the initiation and development of cancer. However, the mechanisms of ncRNA-mediated regulation of metastasis in CRC remain largely unknown. In this study, we systematically analyzed the expression network of microRNAs (miRNAs) and genes in CRC metastasis using bioinformatics, and discovered that the miR-581/SMAD7 axis could be a potential factor that drives CRC metastasis. A dual luciferase report assay and protein analysis confirmed the binding relationship between miR-581 and SMAD7. Further functional assays revealed that miR-581 inhibition could suppress cell proliferation and induce apoptosis in SW480 cells. Up-regulation or down-regulation of miR-581 could both affect cell invasion capacity and modulate epithelial to mesenchymal transition (EMT) via a SMAD7/TGFβ signaling pathway. In conclusion, our findings elucidated that miR-581/SMAD7 could be essential for CRC metastasis, and may serve as a potential therapeutic target for CRC patients.

Flurbiprofen suppresses the inflammation, proliferation, invasion and migration of colorectal cancer cells via COX2.

Colorectal cancer is an aggressive disease with a poor prognosis and low survival rate at the advanced stage, therefore new innovative targets are urgently required. Flurbiprofen has been reported to exhibit therapeutic effects in other types of cancer, such as esophageal cancer, breast cancer and colorectal cancer. Therefore, the present study aimed to investigate the function of flurbiprofen in colorectal cancer. SW620 colorectal cancer cells were treated with different concentrations of flurbiprofen to determine the optimum concentration. Subsequently, COX2 expression affected by flurbiprofen was tested using western blotting, reverse transcription-quantitative PCR and immunofluorescence. Enzyme-linked immunosorbent assay was used to determine the levels of tumor necrosis factor-α, interleukin (IL)-6 and IL-1β. Cell Counting Kit-8, colony formation and flow cytometry assays were used to assess the proliferation and apoptosis of SW620 cells in various groups. Western blotting was performed to investigate the expression of proliferation-, apoptosis- and migration-related proteins after different treatments. Wound healing and Transwell assays were performed to measure the invasion and migration of colorectal cancer cells, respectively. The results demonstrated that flurbiprofen inhibited colorectal cancer cell proliferation. Furthermore, it was identified that flurbiprofen inhibited the expression of COX2. Notably, flurbiprofen suppressed the expression of inflammatory factors by inhibiting COX2. Moreover, flurbiprofen inhibited the proliferation, invasion and migration of colorectal cancer cells by inhibiting COX2. In conclusion, the present study revealed that flurbiprofen inhibited COX2 expression in colorectal cancer, and affected the proliferation, invasion, migration and apoptosis of colorectal cancer cells. These results expand the understanding of the function of COX2 in colorectal cancer and the effect of flurbiprofen on COX2 expression.

Anticancer Effects of Fufang Yiliu Yin Formula on Colorectal Cancer Through Modulation of the PI3K/Akt Pathway and BCL-2 Family Proteins.

Colorectal cancer (CRC) is one of the most common malignant tumors in China. Fufang Yiliu Yin (FYY) is a traditional Chinese medicine formula used in clinical practice for cancer treatment, but its effectiveness and mechanism of action in human CRC are unclear. In this study, we investigated the antitumor effect of FYY on HCT116 and SW480 human CRC cell lines in vitro and evaluated the underlying molecular mechanism. A subcutaneous xenograft mouse model was used to confirm the antitumor effect in vivo. The components and targets of FYY were collected from the Traditional Chinese Medicine Systems Pharmacology Database (TCMSP) database. CRC targets were collected via the GeneCards and OMIM databases. Protein–protein interactions were explored using the STRING platform. Cytoscape was used to construct drug–disease–target networks. KEGG and GO analyses were performed to investigate common FYY and CRC targets. FYY significantly inhibited cell proliferation and induced HCT116 and SW480 cell apoptosis. Cell proliferation was blocked at the G0/G1 phase, while cell apoptosis was promoted at the early stage. According to the network pharmacological analysis, quercetin and kaempferol were the most bioactive compounds of FYY. The key targets of FYY were cyclin-D1, MAPK8, and EGFR. GO analysis showed that core targets included the apoptotic signaling pathway, response to steroid hormone, and cellular response to organic cyclic compound. The KEGG pathway analysis showed that FYY may affect CRC through the PI3K/Akt pathway. In vitro, FYY significantly inhibited tumor growth. Pathway analysis confirmed that FYY induced cell apoptosis by modulating PI3K/Akt signaling and BCL-2 family proteins. Hence, our findings indicate that FYY may be a promising adjuvant therapy for CRC.

Ligustrazine inhibits the viability and motility of colon cancer cells.

BackgroundDespite the advances in the diagnosis and treatment of colon cancer, the disease remains a major threat worldwide. Ligustrazine (LSZ) is an alkaloid with anti-tumor activity. This study aimed to elucidate the anti-tumor qualities of LSZ on colon cancer cells in vitro and vivo, as well as its involved mechanism.MethodsIn this study, CCK-8, colony formation, transwell invasion assay and flow cytometry were employed to examined the cells behaviors. Moreover, the proteins related to the epithelial mesenchymal transformation (EMT) process were determined by RT-qPCR and western blotting. Then, the expression of PI3K, AKT, and mTOR were tested by western blotting. Furthermore, a xenograft mouse model was used to investigate the role of LSZ in vivo.ResultsLSZ treatment (0.1, 0.5 and 1.5 mM) significantly inhibited SW480 cells’ efforts to proliferate, migrate and invade and also induced SW480 cell apoptosis. In addition, LSZ inhibited EMT process. Meanwhile, Western blotting indicated that LSZ treatment administered on a dose-dependent basis inhibited the phosphorylation of PI3K, AKT, and mTOR. Furthermore, in our animal experiments, the weight of colon cancer was significantly lower in LSZ-treated mice than in untreated ones, and the expression of Ki67 and N-cadherin positive cells was significantly lower in the treated mice than in their control group counterparts.ConclusionsTogether, the results suggested that LSZ inhibited proliferation, motility, and EMT of colon cancer cells through the PI3K/AKT/mTOR pathway in vitro and in vivo.

Enhanced antitumor efficacy in colon cancer using EGF functionalized PLGA nanoparticles loaded with 5-Fluorouracil and perfluorocarbon

BackgroundTumor recurrence and metastasis occur at a high rate in patients with colon cancer. Identification of effective strategies for the treatment of colon cancer is critical. Recently, poly (lactic-co-glycolic acid) (PLGA) has been shown to have potential as a broad therapeutic drug delivery system. We designed a dual-loaded nanoparticle drug delivery system to overcome the limitations of chemotherapeutic drugs used to treat colon cancer.MethodsWe developed epidermal growth factor (EGF) functionalized PLGA nanoparticles (NPs) co-loaded with 5-fluorouracil (5Fu) and perfluorocarbon (PFC) (EGF-PLGA@5Fu/PFC) for targeted treatment of colon cancer. CCK-8 assay, Hoechst33342 staining and flow cytometry were performed to investigate the functions of EGF-PLGA@5Fu/PFC NPs in SW620 cells. Beside, animal experiment, histological analysis and immunofluorescence staining were adopted to further confirm the role of EGF-PLGA@5Fu/PFC NPs in vivo.ResultsThe findings showed that EGF-PLGA@5Fu /PFC NPs had an average size 200 nm and a 5Fu-loading efficiency of 7.29%. Furthermore, in vitro release was pH-sensitive. Targeted EGF-PLGA@5Fu/PFC NPs exhibited higher cellular uptake than non-targeted NPs into colon cancer cells. In addition, EGF-PLGA@5Fu/PFC NPs suppressed cell viability and induced apoptosis in SW620 cells to a greater extent than non-targeted NPs. In tumor xenografted mice, EGF-PLGA@5Fu/PFC NPs suppressed tumor growth more effectively than 5Fu, PLGA@5Fu or PLGA@5Fu/PFC NPs. Histopathological analysis further demonstrated that EGF-targeted NPs inhibited tumor growth to a greater extent than non-targeted or non-NP treatments. The improved therapeutic outcomes observed in this study were due to relief of tumor hypoxia by transport of oxygen by PFC to the tumors.ConclusionWe constructed a biocompatible nanodrug delivery system based on functionalized nanoparticles that provided a novel strategy for selective delivery of chemotherapy drugs to tumors.

Exosome-mediated transfer of miR-93-5p from cancer-associated fibroblasts confer radioresistance in colorectal cancer cells by downregulating FOXA1 and upregulating TGFB3

BackgroundCancer-associated fibroblasts (CAFs) have been intensively studied in recent studies with aims of finding more concrete evidence on their mechanism of involvement in tumor progression, which is currently unknown. CAFs can secrete exosomes which are loaded with proteins, lipids and RNAs, all of which affect tumor microenvironment. The present study identified microRNA-93-5p (miR-93-5p) as a novel exosomal cargo responsible for the pro-tumorigenic effects of CAFs on colorectal cancer (CRC).MethodsCAFs and normal fibroblasts (NFs) were isolated from cancerous tissues and matched with paracancerous tissues that had been surgically resected from CRC patients. The interaction among miR-93-5p, forkhead box A1 (FOXA1) and TGFB3 was identified through ChIP and dual luciferase reporter assays. The proliferation and apoptosis of SW480 cells co-cultured with CAFs-derived exosomes under irradiation were evaluated by CCK-8, colony formation, and flow cytometric assays. Tumorigenesis of SW480 cells in nude mice was assessed under the irradiation.ResultsFOXA1 was found to be associated with reduced radioresistance in CRC cells and was verified as a target of miR-93-5p. CAFs-derived exosomes contained higher miR-93-5p than those from NFs, which augmented SW480 cell proliferation and rescued them from radiation-induced apoptosis. miR-93-5p was identified as a mediator of the exosomal effects of CAFs on SW480 cells, possibly through downregulating FOXA1 and upregulating TGFB3. FOXA1 could bind to the promoter of TGFB3, thereby inhibiting nuclear accumulation of TGFB3. Also, CAFs-derived exosomes containing miR-93-5p increased the tumor growth of SW480 cells in irradiated nude mice.ConclusionThe present study identifies miR-93-5p as a specific exosomal cargo that rescues CRC cells against radiation-induced apoptosis.

Daphnane Diterpenoids from Daphne genkwa Inhibit PI3K/Akt/mTOR Signaling and Induce Cell Cycle Arrest and Apoptosis in Human Colon Cancer Cells.

Seven new daphnane-type diterpenoids, daphgenkins A-G (1-7), and 15 known analogues (8-22) were isolated from the flower buds of Daphne genkwa. Their structures and absolute configurations were elucidated by spectroscopic data and calculated ECD analyses. The cytotoxicities of all daphnane-type diterpenoids (1-22) obtained were evaluated against three human colon cancer cell lines (SW620, RKO, and LoVo). Compounds 1, 12, and 13 exhibited cytotoxic effects against the SW620 and RKO cell lines, with IC50 values in the range of 3.0-9.7 μM. The most active new compound, 1, with an IC50 value of 3.0 μM against SW620 cells, was evaluated further for its underlying molecular mechanism. Compound 1 induced G0/G1 cell cycle arrest, leading to the induction of apoptosis in SW620 cells. Also, it induced cancer cell apoptosis by an increased ratio of Bax/Bcl-2, activated cleaved caspase-3 and caspase-9, and upregulated PARP. Finally, compound 1 significantly inhibited PI3K/Akt/mTOR signaling in SW620 cells. Together, the results suggest that compound 1 may be a suitable lead compound for further biological evaluation.