Growth Of Human Colon Cancer Research Articles

MTOR and PDGF Pathway Blockade Inhibits Liver Metastasis of Colorectal Cancer by Modulating the Tumor Microenvironment

Tumor growth and metastasis are not determined by cancer cells alone but also by a variety of stromal cells, and platelet-derived growth factor receptors (PDGF-Rs) are overexpressed by various stromal cell populations. Activation of PI3K-AKT-mTOR signaling is frequently observed in many cancer types. We investigated whether the mTOR inhibitor everolimus, alone or in combination with the PDGF-R tyrosine kinase inhibitor nilotinib, can inhibit growth and metastasis of human colon cancer. The effects of nilotinib and everolimus on tumor growth and metastasis were examined in an orthotopic mouse model of human colon cancer and a model of liver metastasis. After treatment with nilotinib (versus distilled water), the stromal reaction of xenografts growing in the cecal wall and liver was significantly decreased. After treatment with everolimus, the stromal reaction did not decrease, but tumor cell proliferation and microvessel density decreased. With the two drugs in combination, both stromal reaction and tumor cell proliferation decreased and apoptosis of tumor cells increased, resulting in remarkable inhibition of tumor growth at both the orthotopic and the metastatic site. Concurrent inhibition of tumor cells and activated stromal cells by a PDGF-R tyrosine kinase inhibitor and an mTOR inhibitor used in combination may represent a novel therapeutic approach for colorectal cancer.

Downregulation of Msi1 suppresses the growth of human colon cancer by targeting p21cip1.

Musashi1 (Msi1), a member of the RNA-binding protein (RBP) family, is highly expressed in neural progenitor or stem cells for the maintenance of stemness as well as in various cancers. Emerging studies have demonstrated that it regulates cell processes by translational activation or suppresses specifically bound mRNA. In the present study, we initially reported remarkably increased expression of Msi1 in colon cancer tissues compared with adjacent non-tumor tissues. Knockdown of Msi1 significantly suppressed the proliferation, colony formation, tumorsphere formation and the progression of implanted colon cancers, and induced cell cycle attest at G0/G1 phase, along with the upregulated expression of p21(cip1). Reporter assays using a chimeric mRNA that combined luciferase and the 3'-UTR of p21(cip1) revealed that Msi1 decreased the reporter activity through the specific motif. Thus, the current results suggested that downregulation of Msi1 could inhibit the growth of colon cancers and Msi1 may be a promising therapeutic target molecule for human colon cancers.

Acetylamine derivative of diospyrin, a plant-derived binaphthylquinonoid, inhibits human colon cancer growth in Nod-Scid mice

Anticancer activity of diospyrin and its derivatives (1-5) was evaluated against thirteen human cell lines. Compared to diospyrin (1), the acetylamine derivative (4) exhibited increase in cytotoxicity, particularly in HT-29 colon cancer cells, showing GI50 values of 33.90 and 1.96 μM, respectively. Also, enhanced toxicity was observed when cells, pre-treated with compound 4, were exposed to radiation. In vivo assessment of 4 was undertaken on tumour-bearing Nod-Scid mice treated at 4 mg/kg/day. Significant reduction in relative tumour volume (~86-91 %) was observed during the 12th-37th days after drug treatment. Increased caspase-3 activity and DNA ladder formation was observed in HT-29 cells after treatment with 4, suggesting induction of apoptotic death after drug treatment. Moreover, flow cytometric determination of Annexin V- FITC positive and PI negative cells demonstrated 17.4, 26.4, and 27.9 % of early apoptosis, respectively, upon treatment with 5, 10 and 25 μM of 4. HT-29 cells after treatment with 4 (1-25 μM) revealed ~2.5- 3- folds generation of ROS. Furthermore, concentration dependent decrease of mitochondrial trans-membrane potential (∆ψm), and expression of Bcl-2/Bax and other marker proteins suggested involvement of mitochondrial pathway of cell death. Overall, our results demonstrated the underlying cell-death mechanism of the plant-derived naphthoquinonoid (4), and established it as a prospective chemotherapeutic 'lead' molecule against colon cancer.

Tumor suppressor DLC-1 induces apoptosis and inhibits the growth and invasion of colon cancer cells through the Wnt/β-catenin signaling pathway

The aim of the present study was to investigate the biological role and molecular mechanism of the deleted in liver cancer-1 (DLC-1) gene in human colon cancer growth and invasion. Recombinant lentiviral vectors encoding the DLC-1 gene were constructed for transfection into the human colon cancer cell line SW480. Real-time quantitative polymerase chain reaction (real-time qPCR) and western blot analysis were employed to evaluate the expression of DLC-1, β-catenin, GSK-3β and c-myc in DLC-1-transfected cells. Moreover, cell proliferation assay, cell colony formation assay, cell cycle analysis, apoptosis analysis and cell migration and invasion assays were performed in order to elucidate the role of DLC-1 in colorectal cancer development and progression. Both real-time qPCR and western blot analyses showed that the DLC-1 gene and protein were overexpressed in the DLC-1-transfected SW480 cells. In addition, the expression of β-catenin and GSK-3β was upregulated and the expression of the c-myc gene was downregulated in the DLC-1-transfected SW480 cells. Furthermore, DLC-1 overexpression inhibited cell proliferation, colony formation, migration and invasion, and induced cell cycle arrest at the G1 phase with subsequent apoptosis. DLC-1 inhibits cell growth and invasion in human colon cancer, functioning as a tumor-suppressor gene, possibly through the regulation of the Wnt/β-catenin signaling pathway.

Kaempferol Induces Cell Cycle Arrest in HT-29 Human Colon Cancer Cells.

Background:A greater intake of vegetables and fruits has been linked to a reduced incidence of colon cancer. Flavonoids are polyphenolic compounds which are broadly distributed in fruits and vegetables and display a remarkable spectrum of physiological activities, including anti-carcinogenic effects. The objective of this study was to determine the mechanisms by which kaempferol, a flavonol present in tea, apples, strawberries, and beans, inhibits the growth of HT-29 human colon cancer cells.Methods:To examine the effects of kaempferol on cell cycle progression in HT-29 cells, cells were treated with various concentrations (0–60 μmol/L) of kaempferol. Cell proliferation and DNA synthesis were evaluated by MTT assay and [3H]thymidine incorporation assay, respectively. Fluorescence-activated cell sorting analyses were conducted to calculate cell cycle phase distribution. Western blot analyses and in vitro kinase assays were used to estimate the expression of proteins involved in the regulation of cell cycle progression and the activity of cyclin-dependent kinase (CDK)s, respectively.Results:Kaempferol decreased viable cell numbers and [3H]thymidine incorporation into DNA of HT-29 cells in a dose-dependent manner. Kaempferol induced G1 cell cycle arrest within 6 h and G2/M arrest at 12 h. Kaempferol inhibited the activity of CDK2 and CDK4 as well as the protein expression of CDK2, CDK4, cyclins D1, cyclin E, and cyclin A, and suppressed the phosphorylation of retinoblastoma protein. Additionally, kaempferol decreased the levels of Cdc25C, Cdc2, and cyclin B1 proteins, as well as the activity of Cdc2.Conclusions:The present results indicate that kaempferol induces G1 and G2/M cell cycle arrest by inhibiting the activity of CDK2, CDK4, and Cdc2. The induction of cell cycle arrest may be one of the mechanisms by which kaempferol exerts anti-carcinogenic effects in colon cancer cells.

PPAR-β基因对人结肠癌细胞裸鼠成瘤的影响及机制研究

目的：探讨过氧化物酶体增殖物激活受体-β (peroxisome proliferator-activated receptor-β, PPAR-β)对人结肠癌细胞裸鼠移植瘤生长的影响及机制。方法：分别将PPAR-β特异性沉默和野生型的人结肠癌细胞KM12C种植于裸鼠背部皮下，动态监测移植瘤体积，20天后取瘤体称重，免疫组化比较二者分化程度；荧光定量PCR(RT- PCR)、免疫组化、Western blot检测体内外人结肠癌细胞血管内皮生长因子(VEGF)表达；此外，用PPAR-β特异性激动剂GW501516处理离体细胞，检测VEGF与剂量的相关性。结果：PPAR-β沉默组的移植瘤生长速度明显快于对照组，并且低分化肿瘤的比例、VEGF表达水平显著高于对照组。特异性激活沉默组的PPAR-β并没有改变其VEGF的表达，在对照组中则观察到VEGF水平与激活程度呈剂量相关性降低。结论：PPAR-β沉默在移植瘤中发挥了促瘤作用，提示PPAR-β可能是结肠癌细胞的抑癌基因，并可伴有阻止VEGF表达的作用。To explore the effect of peroxisome proliferator-activated receptor-β (PPAR-β) gene on human colon cancer growth in vivo and the molecular mechanism. Methods: The models of colon cancer xenografts in nude mice were established by respectively subcutaneous injected human colon cancer cell line KM12C with PPAR β knockdown or untreated. The xenografts were measured daily and weighted 20 days later. The results of two groups were compared by immunohistochemisty (IHC). Vascular endothelial growth factor (VEGF) was detected by real-time reverse-transcription (RT-PCR), IHC, Western blot. Meanwhile, KM12C cell was treated in vitro by using PPAR-β ago- nist GW501516 and then the relativity of the dose and VEGF was analyzed. Result: The xenografts in PPAR β-knock- down group grew significantly earlier and larger and they were more frequently less-differentiated compared to those in control group. Meanwhile, the expression of VEGF, either in xenografts or in KM12C cell line, was significantly higher in PPAR β-knockdown group than that in control group. Moreover, specific activation of PPAR β didn’t change the expression of VEGF in the KM12C cells with PPAR β knockdown, while yielded a dose-dependent decrease of VEGF in those without PPAR β knockdown. Conclusion: These findings indicate that PPAR β may induce the differentiation and inhibit the growth of colon cancer, which may be associated with its inhibition effect on VEGF in colon cancer cells.

IL‐32γ enhances TNF‐α‐induced cell death in colon cancer

Interleukin (IL)-32 is a recently discovered cytokine that appears to play an important role in human colon cancer growth. We investigated that IL-32γ in combination with TNF-α remarkably inhibited cell growth of human colon cancer cells (HCT116 and SW620) and tumor growth in xenograft-bearing nude mice. The transient enforced overexpression of IL-32γ potentiated the inhibitory effect of TNF-α on DNA synthesis, cell number and protein content, and enhanced apoptosis in colon cancer cells. We also found that knockdown of IL-32γ by siRNA showed the abolishment of cell growth inhibitory effect of TNF-α. The IL-32γ-overexpressing colon cancer cells further increased TNF-α-mediated expression of p38 MAPK as well as that of Bax, cleaved caspase-3 and -9, but decreased that of antiapoptotic proteins such as Bcl-2, cellular inhibitor of apoptosis protein (IAP) and X chromosome IAP. In xenograft model, the lipopolysaccharide (LPS)-injected (1.25 mg/kg) mice inoculated with IL-32γ-transfected HCT116 colon cancer cells were more decrease tumor volume and weight than inoculated with vector. Tumor tissues isolated from LPS-injected mice inoculated with IL-32γ-overexpressing colon cancer cells potentiated the expression levels of pro-apoptotic proteins such as cleaved caspase-3, 9 and Bax, but decreased that of Bcl-2. Furthermore, the mice increased IL-10 production, but decreased IL-6 levels in serum. In conclusion, our results suggest that IL-32γ may potentiate TNF-α-induced cell growth inhibition through activation of p38 MAPK pathways.

Stroma‐directed imatinib therapy impairs the tumor‐promoting effect of bone marrow‐derived mesenchymal stem cells in an orthotopic transplantation model of colon cancer

Bone marrow-derived mesenchymal stem cells (MSCs) are reported to contribute to formation of tumor-promoting stromal cells. We reported recently that, in an orthotopic nude mice model of colon cancer, MSCs traveled to tumor stroma, where they differentiated into carcinoma-associated fibroblast (CAF)-like cells. We also found that CAFs express platelet-derived growth factor receptor (PDGFR) at a high level and that imatinib therapy targeting PDGFR in CAFs inhibits growth and metastasis of human colon cancer. These findings led us to examine whether the tumor-promoting effect of MSCs is impaired by blockade of PDGFR signaling achieved with imatinib. Orthotopic transplantation and splenic injection of human MSCs along with KM12SM human colon cancer cells, in comparison with transplantation of KM12SM cells alone, resulted in significantly greater promotion of tumor growth and liver metastasis. The KM12SM + MSC xenograft enhanced cell proliferation and angiogenesis and inhibited tumor cell apoptosis. When tumor-bearing animals were treated with imatinib, there was no significant increase in primary tumor volume or total volume of liver metastases, despite the KM12SM+MSC xenograft, and survival in the mixed-cell group was prolonged by imatinib treatment. Moreover, the ability of MSCs to migrate to tumor stroma was impaired, and the number of MSCs surviving in the tumor microenvironment was significantly decreased. In in vitro experiments, treatment with imatinib inhibited migration of MSCs. Our data suggest that blockade of PDGF signaling pathways influences the interaction between bone marrow-derived MSCs and tumor cells in the tumor microenvironment and, hence, inhibits the progressive growth of colon cancer.

Combination of gastrin-releasing peptide antagonist with cytotoxic agents produces synergistic inhibition of growth of human experimental colon cancers

We investigated the efficacy of a powerful antagonist of bombesin/gastrin-releasing peptide (BN/GRP) RC-3940-II administered as a single agent or in combination with cytotoxic agents on the growth of HT-29, HCT-116 and HCT-15 human colon cancer in vitro and in vivo. GRP-receptor mRNA and protein were found in all three cell lines tested. Exposure of HT-29 cells to 10 μM RC-3940-II led to an increase in the number of cells blocked in S phase and G2/M and cells with lower G0/G1 DNA content. Similar changes on the cell cycle traverse of HT-29 cells could also be seen at lower concentrations of RC-3940-II (1 μM) after pretreatment with 100 nM GRP (14–27), indicating a dose-dependent mechanism of action based on the blockage of BN/GRP induced proliferation of tumor cells at lower concentrations. Daily in vivo treatment with BN/GRP antagonist RC-3940-II decreased the volume of HT-29, HCT-116 and HCT-15 tumors xenografted into athymic nude mice by 25 to 67% (p < 0.005). Combined treatment with RC-3940-II and chemotherapeutic agents 5-FU and irinotecan resulted in a synergistic tumor growth suppression of HT-29, HCT-116 and HCT-15 xenografts by 43% to 78%. In HT-29 and HCT-116 xenografts the inhibition for the combinations of RC-3940-II and irinotecan vs. single substances (p < 0.05) was significantly greater. These findings support the use of RC-3940-II as an anticancer agent and may help to design clinical trials using RC-3940-II in combinations with cytotoxic agents.

Norcantharidin Anti-Angiogenesis Activity Possibly through an Endothelial Cell Pathway in Human Colorectal Cancer

The present study was based on the unexpected discovery that norcantharidin exerted anti-angiogenesis activity when effects on growth of human colon cancer were studied. The aim was to further verify this finding and explore possible mechanisms using a tumor xenograft model in nude mice. We confirmed that norcantharidin (5 or 15 mg/kg) could inhibit angiogenesis of human colon cancer in vivo. In vitro, crossing river assay, cell adhesion assay and tube formation assay indicated that NCTD could reduce the migration, adhesion and vascular network tube formation ability of HUVECs. At the same time, the expression levels of VEGF and VEGFR-2 proteins which play important roles in angiogenesis were reduced as examined by western blotting analysis. Taken together, the results firstly showed NCTD could inhibit angiogenesis of human colon cancer in vivo, probably associated with effects on migration, adhesion and vascular network tube formation of HUVECs and expression levels of VEGF and VEGFR-2 proteins.

Anti-tumor Effects of Cationic Hybrid Liposomes against Colon Carcinoma along with Apoptosis &lt;i&gt;in Vitro&lt;/i&gt;

New-type three-component cationic hybrid liposomes (HLs) composed of dimyristoylphosphatidylcholine (DMPC), polyoxyethylene(21)dodecyl ether (C(12)(EO)(21)) and O,O'-ditetradecanoyl-N-(α-trimethylammonioacetyl) diethanolamine chloride (2C(14)ECl) were produced. Cationic HLs were smaller and more stable than pure DMPC liposomes. It is noteworthy that cationic HLs could remarkably inhibit the growth of human colon cancer (HCT116) cells along with apoptosis in vitro for the first time in this study.

Cladosporol a stimulates G1‐phase arrest of the cell cycle by up‐regulation of p21waf1/cip1 expression in human colon carcinoma HT‐29 cells

Cladosporols, purified and characterized as secondary metabolites from Cladosporium tenuissimum, display an antifungal activity. In this study, we tested the antiproliferative properties of cladosporol A, the main isoform of this metabolite family, against human cancer cell lines. By assessing cell viability, we found that cladosporol A inhibits the growth of various human colon cancers derived cell lines (HT-29, SW480, and CaCo-2) in a time- and concentration-dependent manner, specifically of HT-29 cells. The reduced cell proliferation was due to a G1-phase arrest, as assessed by fluorescence activated cell sorting analysis on synchronized HT-29 cells, and was associated with an early and robust over-expression of p21(waf1/cip1) , the well-known cyclin-dependent kinases inhibitor. This suggests that the drug may play a role in the control of cancer cell proliferation. Consistently, cyclin D1, cyclin E, CDK2, and CDK4 proteins were reduced and histone H1-associated CDK2 kinase activity inhibited. In addition to p21(waf1/cip1) , exposure to 20 µM cladosporol A caused a simultaneous increase of pERK and pJNK, suggesting that this drug activates a circuit that integrates cell cycle regulation and the signaling pathways both involved in the inhibition of cell proliferation. Finally, we showed that the increase of p21(waf1/cip1) expression was generated by a Sp1-dependent p53-independent stimulation of its gene transcription as mutagenesis of the Sp1 binding sites located in the p21 proximal promoter abolished induction. To our knowledge, this is the first report showing that cladosporol A inhibits colon cancer cell proliferation by modulating p21(waf1/cip1) expression.

Kallikrein-Related Peptidase 14 Acts on Proteinase-Activated Receptor 2 to Induce Signaling Pathway in Colon Cancer Cells

Serine proteinases participate in tumor growth and invasion by cleaving and activating proteinase-activated receptors (PARs). Recent studies have implicated PAR-1 and PAR-4 (activated by thrombin) and PAR-2 (activated by trypsin but not by thrombin) in human colon cancer growth. The endogenous activators of PARs in colon tumors, however, are still unknown. We hypothesize that the kallikrein-related peptidase (KLK) family member KLK14, a known tumor biomarker, is produced by colonic tumors and signals to human colon cancer cells by activating PARs. We found that i) KLK14 mRNA was present in 16 human colon cancer cell lines, ii) KLK14 protein was expressed and secreted in colon cancer cell lines, and iii) KLK14 (0.1 μmol/L) induced increases in intracellular calcium in HT29, a human colon cancer-derived cell line. KLK14-induced calcium flux was associated with internalization of KLK14-mediated activation of PAR-2. Furthermore, KLK14 induced significant extracellular signal-regulated kinases 1 and 2 (ERK1/2) phosphorylation and HT29 cell proliferation, presumably by activating PAR-2. A PAR-2 cleavage and activation-blocking antibody dramatically reduced KLK14-induced ERK1/2 signaling. Finally, ectopic expression of KLK14 in human colon adenocarcinomas and its absence in normal epithelia was demonstrated by IHC analysis. These results demonstrate, for the first time, the aberrant expression of KLK14 in colon cancer and its involvement in PAR-2 receptor signaling. Thus, KLK14 and its receptor, PAR-2, may represent therapeutic targets for colon tumorigenesis.

ShRNA-mediated gene silencing of heat shock protein 70 inhibits human colon cancer growth

Heat shock protein 70 (Hsp70), a chaperone involved in tumor progression, is overexpressed in various human tumors. However, its role in colon cancer progression is not completely understood. In the present study, two shRNA plasmid vectors against Hsp70 were constructed and stably transfected into the colon cancer cell line HT29 to determine the effect of Hsp70 on cell proliferation, cell cycle distribution and cell apoptosis in HT29 cells in vitro, and its effect on xenograft tumor growth and apoptosis in vivo. Cell proliferation was determined using MTT assay. The results revealed that Hsp70 silencing efficiently inhibited the growth of HT29 cells in culture, induced cell cycle arrest at the G1 phase, and significantly increased apoptosis. Moreover, stable clones from the Hsp70 shRNA-2 vector suppressed xenograft tumor growth and enhanced apoptosis in vivo compared with a mock and vector control group. In conclusion, specific Hsp70 shRNA silencing may inhibit colon cancer growth, indicating that Hsp70 silencing is a potential therapeutic strategy for the treatment of colon cancer.

Abstract 4220: Inhibition of cancer cell growth by muscadine grape seed and grape skin extracts

Abstract The muscadine grape (Vitis rotundifolia) is native to the Southeastern United States and muscadine grape seed products are marketed as dietary supplements, based upon their antioxidant properties. Extracts from muscadine grapes contain a number of antioxidants, including resveratrol and ellagic acid; however, the natural antioxidants found in grape seed extracts are predominantly procyanidins, while the grape skin extracts contain more anthocyanins. We investigated the effect of muscadine grape seed extracts (MSE) and muscadine grape skin extracts (MSKE) on the growth of human lung, colon, prostrate, skin, brain and breast cancers as well as human leukemias. Cells were incubated with increasing concentrations (from 0.5 to 50 μg/mL) of aqueous extracts of either MSE or MSKE and the total number of cells was quantified after 7 days. Both the MSE and MSKE inhibited the growth of A549 and SK-LU-1 human lung adenocarcinoma cancer cells (81.8% inhibition at the highest concentration) and HT29 and HCT116 human colon cancer cells (80.5% inhibition at the highest dose). Similarly, extracts from both muscadine grape seeds and skins inhibited the growth of LNCaP and PC3 human prostate cancer cells. The growth of U87 and U373 human glioblastoma cells and RPMI 7951 and SKMEL28 human skin cancer cells was dose-dependently reduced by treatment with the MSE or MSKE. Human leukemia cells were also incubated with increasing concentrations of the extracts; the growth of THP-1 human acute monocytic leukemia cells, HL-60 human acute promyelocytic cells, and K562 human chronic myelogenous leukemia cells was reduced by both muscadine grape extracts (average of 74.2% inhibition at the highest dose). Both the MSE and MSKE inhibited the growth of estrogen receptor-dependent ZR-75-1, HER2 over-expressing SKBR3, and triple negative MDA-MB-231 human breast cancer cells. The inhibition of growth by either extract was highest in the triple negative breast cancer cells (92.6% inhibition at the highest concentration). The reduction in human breast cancer cell growth was accompanied by a significant decrease in the phosphorylation and activation of the mitogen-activated protein kinases ERK1 and ERK2; MAP kinase activities were reduced 91% by MSE and 80% by MSKE in ZR-75-1 estrogen receptor-dependent breast cancer cells and 73% by MSE and 66% by MSKE in MDA-MB-231 triple negative breast cancer cells. There were no significant differences between the effects of the grape seed extract compared to the grape skin extract with any of the cell lines. These results demonstrate that extracts from muscadine grape seeds and muscadine grape skins inhibit the growth of human lung, colon, prostate, breast, skin, brain and leukemia cells in vitro, suggesting that further studies are warranted to investigate their potential use in the prevention or treatment of cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4220. doi:10.1158/1538-7445.AM2011-4220

Abstract 519: Blockade of PDGFR signaling impairs the tumor promoting effect of bone marrow-derived mesenchymal stem cells in colon cancer

Abstract Bone marrow-derived mesenchymal stem cells (MSCs) are reported to migrate to tumor stroma as well as injured tissue. We reported recently that, in an orthotopic nude mice model of colon cancer, MSCs traveled to tumor stroma, where they differentiated into carcinoma-associated fibroblasts (CAF)-like cells. We have also found that CAFs express platelet-derived growth factor receptor (PDGFR) at a high level and that imatinib therapy targeting PDGFR in CAFs combined with administration of a cytotoxic drug significantly inhibits growth and metastasis of human colon cancer. These findings led us to examine whether the tumor promoting effect of MSCs is impaired by blockade of PDGFR signaling achieved with imatinib. KM12SM colon cancer cells alone or KM12SM cells mixed with MSCs in a 1:2 ratio were transplanted into the cecal wall of nude mice. Orthotopic transplantation of KM12SM cells mixed with MSCs, in comparison to transplantation of KM12SM cells alone, resulted in tumors of greater weight. The survival rate was significantly lower in the mixed-cell group. Co-injection of MSCs with tumor cells promotes tumor growth and metastasis of colon cancer by enhancing angiogenesis and tumor migration and invasion and by inhibiting tumor cell apoptosis. When tumor bearing animals were treated with imatinib (by gavage once daily at 50 mg/kg for 35 days), there was no significant increase in primary tumor volume or number of metastases with the KM12SM+MSC xenograft and the lower survival rate in the mixed-cell group was prolonged by the treatment. Moreover, migratory ability of MSCs to tumor stroma was impaired, and MSCs surviving in the tumor microenvironment were significantly decreased. In in vitro experiments, treatment with imatinib inhibited migration and proliferation of MSCs. Our data suggest that blockade of PDGFR signaling pathways influences the interaction between bone marrow-derived MSCs and tumor cells in the tumor microenvironment and, hence, inhibits the progressive growth of colon cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 519. doi:10.1158/1538-7445.AM2011-519

Betulinic acid delivered in liposomes reduces growth of human lung and colon cancers in mice without causing systemic toxicity

Betulinic acid (BetA) is a plant-derived pentacyclic triterpenoid with potent anticancer capacity that targets the mitochondrial pathway of apoptosis. BetA has a broad efficacy in vitro against prevalent cancer types, including lung, colorectal, prostate, cervix and breast cancer, melanomas, neuroblastomas, and leukemias. The cytotoxic effects of the compound against healthy cells are minimal, rendering BetA a promising potential anticancer drug. However, because of the weak hydrosolubility of BetA, it has been difficult to study its efficacy in vivo and a pharmaceutical formulation is not yet available. We report the development of a liposome formulation of BetA and show its successful application in mice. Large liposomes, assembled without cholesterol to reduce their rigidity, efficiently incorporated BetA. Nude mice xenografted with human colon and lung cancer tumors were treated intravenously with the BetA-containing liposomes. Tumor growth was reduced to more than 50% compared with the control treatment, leading to an enhanced survival of the mice. Oral administration of the liposomal formulation of BetA also slowed tumor growth. Any signs of systemic toxicity caused by BetA treatment were absent. Thus, liposomes are an efficient formulation vehicle for BetA, enabling its preclinical development as a nontoxic compound for the treatment of cancers.

Effects of α-mangostin on apoptosis induction of human colon cancer

To investigate the effect of α-mangostin on the growth and apoptosis induction of human colon cancer cells. The three colorectal adenocarcinoma cell lines tested (COLO 205, MIP-101 and SW 620) were treated with α-mangostin to determine the effect on cell proliferation by MTT assay, cell morphology, chromatin condensation, cell cycle analysis, DNA fragmentation, phosphatidylserine exposure and changing of mitochondrial membrane potential. The molecular mechanisms of α-mangostin mediated apoptosis were further investigated by Western blotting analysis including activation of caspase cascade, cytochrome c release, Bax, Bid, p53 and Bcl-2 modifying factor. The highest inhibitory effect of α-mangostin on cell proliferation of COLO 205, MIP-101 and SW 620 were 9.74 ± 0.85 μg/mL, 11.35 ± 1.12 μg/mL and 19.6 ± 1.53 μg/mL, respectively. Further study showed that α-mangostin induced apoptotic cell death in COLO 205 cells as indicated by membrane blebbing, chromatin condensation, DNA fragmentation, cell cycle analysis, sub-G1 peak (P < 0.05) and phosphatidylserine exposure. The executioner caspase, caspase-3, the initiator caspase, caspase-8, and caspase-9 were expressed upon treatment with α-mangostin. Further studies of apoptotic proteins were determined by Western blotting analysis showing increased mitochondrial cytochrome c release, Bax, p53 and Bmf as well as reduced mitochondrial membrane potential (P < 0.05). In addition, up-regulation of tBid and Fas were evident upon treatment with α-mangostin (P < 0.01). α-Mangostin may be effective as an anti-cancer agent that induced apoptotic cell death in COLO 205 via a link between extrinsic and intrinsic pathways.

Antioxidant and antiproliferative properties of water extract from Mahonia bealei (Fort.) Carr. leaves

Mahonia bealei (Fort.) Carr. (Berberidaceae) leaves have been widely used as a tea leaf beverage south of the Qinling Mountains of China. In this study, the antioxidant and antiproliferative properties of M. bealei leaves were investigated. Our data showed that the water extract of M. bealei leaves (WML) exhibited extremely high antioxidant properties, which were demonstrated by its ability to scavenge 50% of 1,1-diphenyll-2-2-pricylhydrazyl (DPPH) free radicals at 60.46 μg/ml, and it eliminated approximately 71.19% of superoxide radicals at 500 μg/ml. In addition, the WML showed strong reducing abilities and provided protection against oxidative protein damage induced by hydroxyl radicals. Cellular proliferation and the induction of apoptosis were also examined by cellular proliferation assay, flow cytometry, and mRNA expression analysis. These results demonstrate that WML significantly inhibited the growth of human colon cancer (HT-29) cells in a concentration-dependent manner, and it gradually increased the proportion of apoptotic cells and reduced the expression of the survivin gene. The bioactivity-guided study of WML resulted in the isolation and identification of berberine, a known isoquinoline alkaloid. Berberine exhibited strong antiproliferative activity on HT-29 cells, with IC 50 values of 36.54 μM, suggesting it is, in part, responsible of the anticancer activity of WML.

Notch1 regulates the growth of human colon cancers

The aberrant activation of the Notch signaling has been associated with the development of colon cancers. However, the role of Notch1 in the pathogenesis of colon cancers is poorly understood. The expression of Notch1 in colon cancer tissues and nontumor tissues and in colon cancer cell lines was examined by Western blot analysis and immunohistochemistry. The impact of small interfering RNA (siRNA)-mediated Notch1 knockdown or Notch1 intracellular domain (NICD)-based transgene-induced Notch1 overexpression on the proliferation, cell cycling, apoptosis, colony formation, and tumorsphere formation in vitro and the development and growth of implanted tumors in vivo was characterized. Notch1 was overexpressed in colon cancer tissues, and the levels of Notch1 expression in different types of colon cancers were associated with the pathologic grade, progression, and metastasis of colon cancers. Furthermore, knockdown of Notch1 significantly inhibited the proliferation, colony formation, and tumorsphere formation of SW480 and HT-29 cells, induced apoptosis and cell cycle arrest at G0/G1 phase, and mitigated the development and growth of implanted colon cancers in vivo. In contrast, Notch1 overexpression promoted the proliferation, colony formation, cell cycling, and tumorsphere formation of colon cancer cells in vitro and the development and growth of implanted colon cancers in vivo, but it inhibited spontaneous apoptosis. The current results indicated that Notch1 signaling positively regulates the growth of colon cancers. Conceivably, the modulation of Notch1-related signaling may be a promising therapy for human colon cancers.