Growth Of Human Colon Cancer Cells Research Articles

Metabolomics reveals that Curcuma longa and demethoxycurcumin inhibit HCT116 human colon cancer cell growth

Studies on the use of natural products to treat cancer are ongoing, and turmeric (Curcuma longa L.), a medicinal crop, is known for various effects including anticancer activity. In this study, the inhibitory effect of C. longa and demethoxycurcumin on cancer cell growth in a colorectal cancer cell line (HCT116) was investigated by using nuclear magnetic resonance (NMR) spectroscopy-based metabolomics. For this analysis, HCT116 cells were treated with doxorubicin (positive control), C. longa extract, or demethoxycurcumin (20, 40, and 60 μM). In the NMR spectra of the HCT116 cell extract, 45 metabolites were identified and quantified. The quantified metabolites were analyzed by biomarker analysis, and significantly changed metabolites were filtered by the area under the curve (AUC) of the receiver operator characteristic (ROC) curve. Multivariate statistical analysis of NMR spectra was conducted to confirm the distribution among groups. Through an S-line plot, it was possible to identify metabolites that contributed to the differences seen in the OPLS-DA score plot. Taken together, the results reveal that C. longa extract induces oxidative stress and changes the energy metabolism in HCT116 cells, and that demethoxycurcumin inhibits the energy metabolism strategy for the survival of cancer cells, escape from immune cells, and cancer cell proliferation, thereby enabling the survival of HCT116 cells.

Abstract 5949: Aspirins metabolites inhibit human colon cancer cell growth: Potential role in colorectal cancer prevention

Abstract Aspirin’s role in colorectal cancer (CRC) prevention is evidential from several epidemiological data. Numerous theories and targets have been proposed, a consensus has not been reached regarding its mechanism of action. One of the unexplored area in aspirin’s CRC prevention is the role of Dihydroxy benzoic acids (DHBAs), which are generated from aspirin’s metabolism by gut microflora and cytochrome P450 (CYP450). Out of the screened DHBAs, 2,3-DHBA and 2,5-DHBA showed concentration dependent inhibition of CDK1, CDK2, CDK4 and CDK6 enzyme activity in-vitro. We demonstrated that 2,3-DHBA and 2,5-DHBA inhibits CDK-1 enzyme activity beginning at 500 µM, while CDK2 and CDK4 activity was inhibited only at higher concentrations (>750 µM). 2,3-DHBA inhibited CDK6 enzyme activity from 250 µM, while 2,5-DHBA inhibited its activity >750 µM. Using colony formation assays, we further demonstrated the role of 2,3-DHBA and 2,5-DHBA in cancer cell cycle inhibition in 3 different cancer cell lines. Out of the 2 DHBAs, 2,5-DHBA was highly effective in inhibiting clonal formation in HCT-116 and HT-29 CRC cell lines (250-500 µM), and in the MDA-MB-231 breast cancer cell line (~100 µM); whereas both aspirin and salicylic acid failed to inhibit all four CDKs and colony formation. Based on the present results, it is suggested that 2,3-DHBA and 2,5-DHBA may contribute to the chemopreventive properties of aspirin, possibly through the inhibition of CDKs. The present data and the proposed mechanisms should open new areas for future investigations. Citation Format: Poojitha Tummala, Ranjini Sankaranarayanan, Rakesh Dachineni, Jayarama Gunaje Bhat. Aspirins metabolites inhibit human colon cancer cell growth: Potential role in colorectal cancer prevention [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5949.

Zingiber officinale Roscoe and Piper betleExtracts Enhanced its Chemopreventive Effect against Colon Cancer Cells by Targeting Caspases-Mediated Apoptosis

Food regimens and herbs have been a target of scientific research in treating cancer. This study aimed to elucidate the effect of Zingiber officinale (ginger) and Piper betle (PB) alone or in combination on the colorectal cancer cell lines HCT116 and HT29. The chemopreventive effect of these extracts was determined by performing cell viability assay, cell cycle analysis, apoptosis analysis, and caspase 3 and 8 activity assays in HCT116 and p53-deficient HT29 cells. Ginger and PB extracts inhibited the proliferation of both cancer cell lines dose-dependently, but the combined extracts inhibited the growth of cancer cells synergistically with an IC50 PB plus ginger < IC50 PB < IC50 ginger. Similarly, the effect of the combined extracts on apoptosis of both cancer cell lines was higher than that of each individual extract. Consistent with the enhancement of caspase activity, both extracts increased the expression of BAX protein, while Bcl-2 protein was decreased. Flow cytometry analysis indicated that the combined extracts arrested a higher percentage of colon cancer cells at the G0/G1 phase with a concomitant decrease in cells in the S phase, indicating that the combined extracts inhibited human colon cancer cell growth by cell cycle arrest at the G0/G1 phase. The inhibitory effect of these combined extracts on cellular proliferation and the induction of apoptosis was synergistic, showing a significant combination index (CI) value < 1.0. The combined extracts of ginger and PB may serve as a potential chemopreventive agent against colon cancer through the induction of caspase-mediated apoptosis.

A teleost bactericidal permeability-increasing protein-derived peptide that possesses a broad antibacterial spectrum and inhibits bacterial infection as well as human colon cancer cells growth

The bactericidal permeability-increasing protein (BPI) is a multifunctional cationic protein produced by neutrophils with antibacterial, antitumor, and LPS-neutralizing properties. In teleost, a number of BPIs have been reported, but their functions are very limited. In this study, an N-terminal peptide, BO18 (with 18 amino acids), derived from rock bream (Oplegnathus fasciatus) BPI, was synthesized and investigated for its antibacterial spectrum, action mechanism, immunoregulatory property as well as the inhibition effects on bacterial invasion and human colon cancer cells growth. The results showed that BO18 was active against Gram-positive bacteria Bscillus subiilis, Micrococcus luteus, and Staphylococcus aureus, as well as Gram-negative bacteria Vibrio alginolyticus, Vibrio litoralis, Vibrio parahaemolyticus and Vibrio vulnificus. BO18 treatment facilitated the bactericidal process of erythromycin and rifampicin by enhancing the permeability of the outer membrane. During its interaction with V. alginolyticus, BO18 exerted its antibacterial activity by destroying cell membrane integrity, penetrating into the cytoplasm and binding to genomic DNA and total RNA. In vitro analysis indicated BO18 could enhance the respiratory burst ability and regulate the expression of immune related genes of macrophages. In vivo detection showed the administration of fish with BO18 before bacterial infection significantly reduced pathogen dissemination and replication in tissues. In addition, BO18 exerted a cytotoxic effect on the growth of human colon cancer cells HT-29. Together, these results add new insights into the function of teleost BPIs, and support that BO18 is a novel and broad-spectrum antibacterial peptide with potential to apply in fighting pathogenic infection in aquaculture.

Uncovering the Anticancer Potential of Murine Cytomegalovirus against Human Colon Cancer Cells

Human cytomegalovirus (HCMV) components are often found in tumors, but the precise relationship between HCMV and cancer remains a matter of debate. Pro-tumor functions of HCMV were described in several studies, but an association between HCMV seropositivity and reduced cancer risk was also evidenced, presumably relying on recognition and killing of cancer cells by HCMV-induced lymphocytes. This study aimed at deciphering whether CMV influences cancer development in an immune-independent manner. Using immunodeficient mice, we showed that systemic infection with murine CMV (MCMV) inhibited the growth of murine carcinomas. Surprisingly, MCMV, but not HCMV, also reduced human colon carcinoma development in vivo. In vitro, both viruses infected human cancer cells. Expression of human interferon-β (IFN-β) and nuclear domain (ND10) were induced in MCMV-infected, but not in HCMV-infected human colon cancer cells. These results suggest a decreased capacity of MCMV to counteract intrinsic defenses in the human cellular host. Finally, immunodeficient mice receiving peri-tumoral MCMV therapy showed a reduction of human colon cancer cell growth, albeit no clinical sign of systemic virus dissemination was evidenced. Our study, which describes a selective advantage of MCMV over HCMV to control human colon cancer, could pave the way for the development of CMV-based therapies against cancer.

Study of the antitumor mechanisms of apiole derivatives (AP-02) from Petroselinum crispum through induction of G0/G1 phase cell cycle arrest in human COLO 205 cancer cells

BackgroundApiole was isolated from the leaves of various plants and vegetables and has been demonstrated to inhibit human colon cancer cell (COLO 205 cells) growth through induction of G0/G1 cell cycle arrest and apoptotic cell death. This study further explored the antitumor effects of apiole derivatives AP-02, 04, and 05 in COLO 205 cancer cells.MethodsHuman breast (MDA-MB-231, ZR75), lung (A549, PE089), colon (COLO 205, HT 29), and hepatocellular (Hep G2, Hep 3B) cancer cells were treated with apiole and its derivatives in a dose-dependent manner. Flow cytometry analysis was subsequently performed to determine the mechanism of AP-02-induced G0/G1 cell cycle arrest. The in vivo antitumor effect of AP-02 (1 and 5 mg/kg, administered twice per week) was examined by treating athymic nude mice bearing COLO 205 tumor xenografts. The molecular mechanisms of AP-02-induced antitumor effects were determined using western blot analysis.ResultsAP-02 was the most effective compound, especially for inhibition of COLO 205 colon cancer cell growth. The cytotoxicity of AP-02 in normal colon epithelial (FHC) cells was significantly lower than that in other normal cells derived from the breast, lung or liver. Flow cytometry analysis indicated that AP-02-induced G0/G1 cell cycle arrest in COLO 205 cells but not in HT 29 cells (< 5 μM for 24 h, **p < 0.01). Tumor growth volume was also significantly inhibited in AP-02 (> 1 mg/kg)-treated athymic nude mice bearing COLO 205 tumor xenografts compared to control mice (*p < 0.05). Furthermore, G0/G1 phase regulatory proteins (p53 and p21/Cip1) and an invasion suppressor protein (E-cadherin) were significantly upregulated, while cyclin D1 was significantly downregulated, in AP-02-treated tumor tissues compared to the control group (> 1 mg/kg, *p < 0.05).ConclusionsOur results provide in vitro and in vivo molecular evidence of AP-02-induced anti-proliferative effects on colon cancer, indicating that this compound might have potential clinical applications.

166-LB: Canagliflozin but Not Empagliflozin Activates AMPK and Suppresses Colon Cancer Cell Growth

Background: Recent increase in the relative proportion of cancer death in diabetic patients has become a major issue. A hypoglycemic agent, sodium-glucose cotransporter (SGLT) 2 inhibitor has attracted attention because of its potency in reducing cardiovascular risks. Several lines of evidence has emerged suggesting a potential anti-oncotic effect. Here we address potential effects of canagliflozin (Cana) on human colon cancer cell growth and underlying mechanisms and compared with empagliflozin (Empa). Material and Methods: We examined the effects of Cana and Empa on 1) cell proliferation using colon cancer cell lines, SW480 and HCT116 by MTT assay, 2) DNA synthesis by flow cytometry-based analysis of bromodeoxyuridine (BrdU) uptake. We further examined 3) the expression of SGLT1 and 2 in these cells and 4) investigated molecules of signal transduction pathways with established roles in cancer cell growth such as AMPK, 4EBP1, mTOR by western blotting. Finally, we analyzed 5) time-dependent changes of metabolomic signatures after the treatment with Cana. Results: While SGLT1 and 2 mRNA and protein expression was detected in both cell lines, Cana but not Empa significantly inhibited cell proliferation and in a dose-dependent manner. Cana suppressed BrdU positive cells and enhanced AMPK phosphorylation with a consequent suppression of 4EBP1 and mTOR activity. Metabolomic profiling revealed that, as a function of time, intermediate metabolites of glycolysis especially phosphoenolpyruvate was decreased with an acute rise in ADP and gradual decrease in ATP and GSSG. Conclusion: We demonstrate that Cana deprives human colon cancer cells of energy and anti-oxidative capacity possibly through an inhibition of SGLT1-mediated glucose influx, and thereby activates AMPK and inhibits cancer cell growth. Our data herein suggest a potential beneficial effect of Cana in suppressing colon cancer progression especially in patients with diabetes. Disclosure T. Yagi: None. T. Tanaka: Research Support; Self; Japan Society for the Promotion of Science. Speaker's Bureau; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Daiichi Sankyo Company, Limited, Kissei Pharmaceutical Co., Ltd., Kowa Pharmaceutical Europe Co. Ltd., Mitsubishi Tanabe Pharma Corporation, Novartis Pharmaceuticals Corporation, Taisho Toyama Pharmaceutical Co. Ltd., Takeda Pharmaceutical Company Limited. E. Kubota: None. H. Koyama: None. T. Guo: None. K. Ogawa: None. D. Aotani: None. H. Kataoka: None.

MicroRNA‑25/ATXN3 interaction regulates human colon cancer cell growth and migration.

The present study aimed to investigate the function of microRNA-25 (miR-25) in human colon cancer cell viability and migration in addition to the underlying possible mechanisms. miR-25 expression was upregulated in patients with colon cancer compared with the control group. Reverse transcription-quantitative polymerase chain reaction and gene chip technology were used to analyze the alterations of miR-25 in patients with colon cancer. Cell viability and cell migration were analyzed using MTT and wound healing assays, respectively, apoptosis was analyzed using flow cytometry, and western blot analysis was conducted to determine the protein expression of ataxin-3 (ATXN3), apoptosis regulator Bax (Bax) and cyclin D1. Overexpression of miR-25 increased cell viability and migration, decreased apoptosis, decreased caspase-3/9 activity level in addition to decreased Bax protein expression, and increased cyclin D1 protein expression in colon cancer cells. Furthermore, miR-25 was demonstrated to target ATXN3 and suppress ATXN3 protein expression. Downregulation of miR-25 induced apoptosis of colon cancer cells via increased expression ATXN3. Small interfering-ATXN3 inhibited the anti-cancer effects of miR-25 downregulation in colon cancer. Collectively, the present results demonstrated that miR-25 promoted human colon cancer cell viability and migration by regulating ATXN3 expression.

Expression of Concern: Aldose reductase inhibition prevents hypoxia-induced increase in hypoxia-inducible factor-1 α (HIF-1α) and vascular endothelial growth factor (VEGF) by regulating 26 S proteasome-mediated protein degradation in human colon cancer cells

Aldose Reductase Inhibition Prevents Hypoxia-induced Increase in Hypoxia-inducible Factor-1α (HIF-1α) and Vascular Endothelial Growth Factor (VEGF) by Regulating 26 S Proteasome-mediated Protein Degradation in Human Colon Cancer CellsJournal of Biological ChemistryVol. 286Issue 27PreviewThe development of intratumoral hypoxia, a hallmark of rapidly progressing solid tumors, renders tumor cells resistant to chemotherapy and radiation therapy. We have recently shown that inhibition of aldose reductase (AR), an enzyme that catalyzes the reduction of lipid aldehydes and their glutathione conjugates, prevents human colon cancer cell growth in culture as well as in nude mouse xenografts by inhibiting the NF-κB-dependent activation of oxidative stress-mediated inflammatory and carcinogenic markers. Full-Text PDF Open Access VOLUME 286 (2011) PAGES 24089–24100 The publisher of the Journal of Biological Chemistry is issuing an Expression of Concern to inform readers that credible concerns have been raised regarding some of the data and conclusions in the article listed above. The Journal of Biological Chemistry will provide additional information as it becomes available.

Magnetic cellulose nanocrystal stabilized Pickering emulsions for enhanced bioactive release and human colon cancer therapy.

Stimuli-responsive drug release and controlled delivery play crucial roles in enhancing the therapeutic efficacy and lowering over-dosage induced side effects. In this paper, we report magnetically-triggered drug release and in-vitro anti-colon cancer efficacy of Fe3O4@cellulose nanocrystal (MCNC)-stabilized Pickering emulsions containing curcumin (CUR). The loading efficiency of CUR in the micron-sized (≈7 μm) MCNC-stabilized Pickering emulsions (MCNC-PE) template was found to be 99.35%. The drug release profiles showed that the exposure of MCNC-PE to external magnetic field (EMF) (0.7 T) stimulated the release of bioactive from MCNC-PE achieving 53.30 ± 5.08% of the initial loading over a 4-day period. The MTT assay demonstrated that the CUR-loaded MCNC-PE can effectively inhibits the human colon cancer cells growth down to 18% in the presence of EMF. The formulation also resulted in 2-fold reduction on the volume of the 3-D multicellular spheroids of HCT116 as compared to the control sample. The MCNC particle was found to be non-toxic to brine shrimp up to a concentration of 100 μg/mL. Our findings suggested that the palm-based MCNC-PE could be a promising yet effective colloidal drug delivery system for magnetic-triggered release of bioactive and therapeutics.

Glucuronorhamnoxylan from Capsosiphon fulvescens inhibits the growth of HT-29 human colon cancer cells in vitro and in vivo via induction of apoptotic cell death

Colorectal cancer is the third most diagnosed cancer and a leading cause of cancer death. Dissatisfaction with currently available anti-colorectal cancer drugs caused by unwanted side effects and low efficacy necessitates new therapeutic agents. In the present study, a sulfated glucuronorhamnoxylan polysaccharide (named SPS-CF) purified from a green alga Capsosiphon fulvescens was evaluated for its anti-cancer activity in vitro and in vivo against colorectal cancer. The SPS-CF treatment resulted in a dose-dependent inhibition of the HT-29 human colon cancer cell growth up to 40% at 500 μg/mL. This inhibitory activity was shown to be mediated by upregulation of the cleavage of caspase-8, -9, -3 and poly (ADP-ribose) polymerase (PARP), induction of DNA fragmentation, and disruption of mitochondrial membrane potential (MMP), demonstrating that SPS-CF causes apoptotic death of HT-29 cancer cells though activation of caspase-dependant pathway. Administration of SPS-CF to BALB/c-nude mice bearing HT-29 cell-xenograft tumor also reduced the tumor growth. The results of this study demonstrated that the SPS-CF effectively inhibits the colorectal tumor growth both in vitro and in vivo by induction of apoptotic death of tumor cells, suggesting that it can be a potent ingredient for health-beneficial foods or anti-cancer agents to prevent or ameliorate human colon cancer.

Annonaceous acetogenin mimic AA005 suppresses human colon cancer cell growth in vivo through downregulation of Mcl-1.

Annonaceous acetogenins are a well-established family of natural products with significant bioactivities, especially high cytotoxic and antitumor activities. AA005 is an annonaceous acetogenin mimic that has shown significant cytotoxicity against a variety of cancer cell lines, but its in vivo antitumor effects have not been demonstrated so far, and its anticancer mechanisms remain ambiguous. In this study, we investigated the effects of AA005 on human colon cancer cell lines in vivo. Human colon carcinoma cell line SW620 xenograft nude mice were treated with AA005 (5 mg/kg/day, i.p.) for 21 days. AA005 administration markedly inhibited the tumor growth via promoting nuclear translocation of apoptosis-inducing factor (AIF) and inducing AIF-dependent cell death. Subsequent studies in human colon carcinoma cell lines SW620 and RKO in vitro revealed that after the colon cancer cells exposed to AA005, downregulation of a B-cell lymphoma 2 family protein, myeloid cell leukemia-1 (Mcl-1), was an early event due to the inhibition of Mcl-1 mRNA level and protein synthesis in a time-dependent manner. Intriguingly, knockdown of Mcl-1 using small interfering RNA markedly accelerated the nuclear translocation of AIF and upregulation of receptor interacting protein-1, and enhanced AA005-mediated lethality, whereas ectopic expression of Mcl-1 substantially attenuated AA005-mediated lethality in the colon cancer cells. Finally, silencing Mcl-1 expression markedly enhanced AA005-induced lethality in SW620 xenograft nude mice, demonstrating a pivotal role of Mcl-1 downregulation in mediating the in vivo antitumor effects of AA005. Taken together, this study demonstrates for the first time the anticancer effects of AA005 against human colon cancer cell lines in vivo, which is mediated through the downregulation of Mcl-1.

Baicalein and baicalin inhibit colon cancer using two distinct fashions of apoptosis and senescence.

Baicalein and baicalin are active components of the Scutellaria baicalensis Georgi and both have broad anti-tumor activity. However, how and whether baicalein and baicalin inhibit colon cancer is unclear. Here we demonstrate that baicalein and baicalin can significantly inhibit human colon cancer cell growth and proliferation. Furthermore, both can induce cell cycle arrest, and suppress cancer cell colony formation and migration. The suppressive effects are mechanistically due to the induction of colon cancer cell apoptosis and senescence mediated by baicalein and baicalin, respectively. Furthermore, we revealed that baicalin-induced senescence in tumor cells is due to its inhibition of telomerase reverse transcriptase expression in tumor cells, and that MAPK ERK and p38 signaling pathways are causatively involved in the regulation of colon cancer cell apoptosis and senescence mediated by baicalein and baicalin. In addition, our in vivo studies using human colon cancer cells in humanized mouse xenograft models, further demonstrated that baicalein and baicalin can induce tumor cell apoptosis and senescence, resulting in inhibition of tumorigenesis and growth of colon cancer in vivo. These data clearly suggest that baicalein and baicalin have potent anti-cancer effects against human colon cancer and could be potential novel and effective target drugs for cancer therapy.

A metabolite of nobiletin, 4'-demethylnobiletin and atorvastatin synergistically inhibits human colon cancer cell growth by inducing G0/G1 cell cycle arrest and apoptosis.

Combining different chemopreventive agents is a promising strategy to reduce cancer incidence and mortality due to potential synergistic interactions between these agents. Previously, we demonstrated that oral administration of nobiletin (NBT, a citrus flavonoid) at 0.05% (w/w, in diet) together with atorvastatin (ATST, a lipid-lowering drug) at 0.02% (w/w, in diet) produced much stronger inhibition against colon carcinogenesis in rats in comparison with that produced by NBT (at 0.1% w/w in diet) or ATST (at 0.04% w/w in diet) alone at higher doses. To further elucidate the mechanism of this promising synergy between NBT and ATST, herein, we measured the levels of NBT, its major metabolites and ATST in the colonic tissue of rats fed NBT (0.05% w/w, in diet) + ATST (0.02% w/w, in diet), and determined the mode of interaction between the major NBT metabolite and ATST in inhibiting colon cancer cell growth. HPLC-MS analysis showed that 4'-demethylnobiletin (4DN) is the most abundant metabolite of NBT with a level about 5-fold as high as that of NBT in the colonic tissue, which indicated the potential significance of 4DN in mediating the biological effects of NBT in the colon. We found that co-treatments of 4DN/ATST at 2 : 1 concentration ratio produced much stronger growth inhibitory effects on human colon cancer HT-29 cells than 4DN or ATST alone, and isobologram analysis confirmed that this enhanced inhibitory effect by the 4DN/ATST combination was highly synergistic. The co-treatment of 4DN/ATST led to G0/G1 cell cycle arrest and induced extensive apoptosis in HT-29 cells. Furthermore, the 4DN/ATST co-treatment profoundly modulated key signaling proteins related to the regulation of the cell cycle and apoptosis. Our results demonstrated a strong synergy produced by the 4DN/ATST co-treatment in inhibiting colon cancer cell growth, which provided a novel mechanism by which NBT/ATST in combination synergistically inhibit colon carcinogenesis.

IL33 Promotes Colon Cancer Cell Stemness via JNK Activation and Macrophage Recruitment.

The expression and biological role of IL33 in colon cancer is poorly understood. In this study, we show that IL33 is expressed by vascular endothelial cells and tumor cells in the human colon cancer microenvironment. Administration of human IL33 and overexpression of murine IL33 enhanced human and murine colon cancer cell growth in vivo, respectively. IL33 stimulated cell sphere formation and prevented chemotherapy-induced tumor apoptosis. Mechanistically, IL33 activated core stem cell genes NANOG, NOTCH3, and OCT3/4 via the ST2 signaling pathway, and induced phosphorylation of c-Jun N terminal kinase (JNK) activation and enhanced binding of c-Jun to the promoters of the core stem cell genes. Moreover, IL33 recruited macrophages into the cancer microenvironment and stimulated them to produce prostaglandin E2, which supported colon cancer stemness and tumor growth. Clinically, tumor IL33 expression associated with poor survival in patients with metastatic colon cancer. Thus, IL33 dually targets tumor cells and macrophages and endows stem-like qualities to colon cancer cells to promote carcinogenesis. Collectively, our work reveals an immune-associated mechanism that extrinsically confers cancer cell stemness properties. Targeting the IL33 signaling pathway may offer an opportunity to treat patients with metastatic cancer. Cancer Res; 77(10); 2735-45. ©2017 AACR.

Stereoisomers of Astaxanthin Inhibit Human Colon Cancer Cell Growth by Inducing G2/M Cell Cycle Arrest and Apoptosis.

Astaxanthin (AST) is a xanthophyll carotenoid with potential protective effects against carcinogenesis. Different stereoisomers of AST (ASTs) exist in a variety of food sources. Due to limited information on the bioactivities of ASTs, the present study investigated the inhibitory effects of ASTs on HCT116 and HT29 human colon cancer cells. ASTs investigated herein included 3S,3'S (S) from Haematococcus pluvialis, 3R,3'R (R) from Phaffia rhodozyma, and a statistical mixture (S: meso: R = 1:2:1) (M) from synthetic AST. Cell viability assay showed that ASTs all inhibited colon cancer cell growth in a time-dependent (24-72 h) and dose-dependent (4-16 μM) manner, and there was no significant difference among the IC50 values of ASTs (p > 0.05). Flow cytometry analysis indicated that ASTs induced G2/M cell cycle arrest and cellular apoptosis in cancer cells. The cell cycle arrest caused by ASTs was associated with increases in the expression levels of p21Cip1/Waf1, p27, and p53, as well as decreases in the levels of CDK4 and CDK6. Meanwhile, the apoptosis induced by ASTs was confirmed by activation of caspase-3 and PARP in the cancer cells. The results indicated that hydroxyl (OH) at C3 and C3' of terminal ring structure might not be the major factor that affects the anticancer activity of AST. This study revealed important information on the inhibitory effects of ASTs on human colon cancer cells, which provided a basis for using ASTs as chemopreventive agents for colon cancer.

The Heat Shock Protein 70 is a Novel Target for Nobiletin in Human Colon Cancer Cells

Nobiletin is a unique flavonoid mainly found in citrus fruits, and has been reported to inhibit colon carcinogenesis in multiple rodent models. However, the direct molecular targets of nobiletin are unknown. Heat shock protein 70 (HSP70) contributes to cancer cell survival and resistance to chemotherapies, thereby inhibition of HSP70 is a promising strategy in cancer chemoprevention. Using affinity chromatography, we demonstrated that nobiletin bound to HSP70 at its ATP‐binding domains in human colon cancer cells and specifically inhibited its ATPase activity. Down regulation of HSP70 by siRNAs significantly increased cancer cells’ sensitivity to nobiletin treatments, which were evidenced by increased apoptosis and decreased colony formation capacity of cancer cells. The association between HSP70 and HSP90 is critical for the stability of their client proteins, such as EGFR, AKT, and STAT3, which contributes to carcinogenesis. We found that nobiletin disrupted the association between HSP70 and HSP90, and combined treatment with nobiletin and HSP90 inhibitors promoted the degradation of those client proteins. Moreover, combined treatment with nobiletin and HSP90 inhibitors resulted in enhanced inhibition on cancer cell growth by inducing apoptosis and cell cycle arrest. Most interestingly, our results also demonstrated that the major colonic metabolites of nobiletin (generated in the colon of mice fed with nobiletin) showed similar inhibitory effects against HSP70‐mediated signaling pathways. Overall, our results demonstrated, for the first time, that HSP70 is a promising target of nobiletin and its colonic metabolites in inhibiting human colon cancer cell growth, which warrants further investigation on their role in colon cancer chemoprevention.Support or Funding InformationThis study was supported by funding from NIH, AICR and USDA.

Synergistic inhibitory effect of cetuximab and tectochrysin on human colon cancer cell growth via inhibition of EGFR signal.

The purpose of this study was to evaluate the enhancing potency of tectochrysin, a flavonoid isolated from Alpinia oxyphylla Miquel by combining cetuximab, an anti-EGFR monoclonal antibody, on human colon cancer cell growth through further inhibition of EGFR pathway. HCT116 and SW480 colon cancer cells were treated with cetuximab (30μg/mL, 1/10 of IC50), tectochrysin (5μg/mL, 1/3 of IC50), or the combination of both agents. The growth inhibitory effect was examined using the MTT assay while apoptotic cell death was performed using TUNEL staining assays. The DNA binding activity of NF-kappa B and AP-1 was investigated by electrophoretic mobility shift assay. Protein expression was determined by Western blot. Cell proliferation was significantly inhibited by the combination of cetuximab and tectochrysin than treatment with cetuximab or tectochrysin alone (combination index: 0.572 and 0.533, respectively). Combination treatment of cells with cetuximab and tectochrysin significantly reduced the expressions of p-EGFR and COX-2 in both cell lines. Combination treatment also significantly inhibited activities of NF-kB and AP-1 compared to the single agent treatment. Our results indicate that combined therapy with lower concentration of cetuximab and tectochrysin could significantly enhance the cancer cell growth inhibitory effect through the inhibition of EGFR signaling.

The Histone Acetyltransferase GCN5 Expression Is Elevated and Regulated by c-Myc and E2F1 Transcription Factors in Human Colon Cancer.

The histone acetyltransferase GCN5 has been suggested to be involved in promoting cancer cell growth. But its role in human colon cancer development remains unknown. Herein we discovered that GCN5 expression is significantly upregulated in human colon adenocarcinoma tissues. We further demonstrate that GCN5 is upregulated in human colon cancer at the mRNA level. Surprisingly, two transcription factors, the oncogenic c-Myc and the proapoptotic E2F1, are responsible for GCN5 mRNA transcription. Knockdown of c-Myc inhibited colon cancer cell proliferation largely through downregulating GCN5 transcription, which can be fully rescued by the ectopic GCN5 expression. In contrast, E2F1 expression induced human colon cancer cell death, and suppression of GCN5 expression in cells with E2F1 overexpression further facilitated cell apoptosis, suggesting that GCN5 expression is induced by E2F1 as a possible negative feedback in suppressing E2F1-mediated cell apoptosis. In addition, suppression of GCN5 with its specific inhibitor CPTH2 inhibited human colon cancer cell growth. Our studies reveal that GCN5 plays a positive role in human colon cancer development, and its suppression holds a great therapeutic potential in antitumor therapy.

Diarylheptanoids from lesser galangal suppress human colon cancer cell growth through modulating Wnt/β-catenin pathway

The Wnt/β-catenin pathway plays a central role in cell growth and differentiation but abnormal activation causes colorectal carcinogenesis. Inhibiting the Wnt/β-catenin pathway can be a good strategy for chemoprevention and treatment of colorectal cancer. While screening for Wnt/β-catenin pathway inhibitors from food materials, we found that (E)-7-(4-hydroxy-3-methoxyphenyl)-1-phenylhept-4-en-3-one (compound 1), among six diarylheptanoids from lesser galangal (Alpinia officinarum), most potently suppressed Wnt3a-induced β-catenin/T-cell factor activity. Moreover, compound 1 suppressed proliferation of colon cancer cells by inhibiting β-catenin translocation to the nucleus by disrupting the β-catenin/galectin-3 complex. Furthermore, a structure–activity realtionship study implicated that the enone group in the linker is critical and the hydroxy substituent on the aromatic ring is generally preferred for activity. Our findings suggest that diarylheptanoid from lesser galangal exerts anti-colon cancer activity by down regulating the Wnt/β-catenin pathway. Bioactive diarylheptanoids and the basic understanding of their structure–activity relationship could be utilized to develop potential candidates for β-catenin-targeted cancer treatment.