上调NDRG1表达降低奥沙利铂耐药结肠癌细胞的存活率

目的观察上调NDRG1表达对结肠癌细胞及奥沙利铂耐药结肠癌细胞的毒性作用并探讨其机制。方法以重组真核表达质粒pEGFP-NDRG1-N3转染HCT 116及奥沙利铂耐药的OHP-HCT 116结肠癌细胞,以实时定量(qRT)-PCR法和Western blot法检测转染效率。MTT法检测奥沙利铂对不同结肠癌细胞的毒性及验证OHP-HCT 116细胞的耐药性。给予奥沙利铂干预转染pEGFP-NDRG1-N3的HCT 116细胞及OHP-HCT 116细胞,MTT法检测细胞存活率,流式细胞术检测细胞凋亡率,Western blot法检测凋亡蛋白Bcl-2及p53蛋白水平变化。结果不同浓度奥沙利铂干预下的3株结肠癌细胞中HCT 116细胞对奥沙利铂最敏感,OHP-HCT 116细胞对奥沙利铂耐药得到验证。以不同浓度梯度奥沙利铂干扰转染pEGFP-NDRG1-N3的HCT 116细胞及OHP-HCT 116细胞,与MOCK及siNC组比较,在HCT 116细胞及OHP-HCT 116细胞,转染pEGFP-NDRG1-N3细胞的存活率下降(P<0.05),凋亡率升高(P<0.05),Bcl-2表达降低(P<0.05),p53表达升高(P<0.05)。结论上调NDRG1表达通过调控p53表达改善结肠癌细胞奥沙利铂耐药,提高化疗敏感性。     

Role of nonsteroidal anti-inflammatory drug-activated gene-1 in docetaxel-induced cell death of human colorectal cancer cells with different p53 status

Nonsteroidal anti-inflammatory drug-activated gene-1 (NAG-1) expression is upregulated not only by NSAIDs such as sulindac sulfide, but also by several antitumorigenic dietary compounds, suggesting that NAG-1 is a specific target for the development of effective anticancer agents. Despite being a downstream target of p53, NAG-1 induction is both p53-dependent and p53-independent. It is not clear whether NAG-1 induction is the responsible factor in cancer cell apoptosis with mutated p53. In this study, we report that NAG-1 induction alone cannot determine apoptotic cell fate in colon cancer cells. Although docetaxel induced an increase in NAG-1 and apoptosis in both HCT-116 (wild-type p53) and HT-29 (mutant p53) colon cancer cells, NAG-1 knockdown with siRNA prevented docetaxel-induced cell death in only HCT-116 cells. Docetaxel decreased Bcl-2 in HCT-116 cells, which have functionally active p53, according to luciferase reporter gene analyses, and docetaxel-induced cell death and changes in Bcl-2 and NAG-1 expression were blocked by PFT-α, a p53 inhibitor. In HT-29 cells with functionally inactive p53, the docetaxel-induced Bcl-xL decrease, NAG-1 increase, and cell death were not blocked by PFT-α. On the other hand, sulindac sulfide at concentrations that significantly induced NAG-1 did not decrease cell viability comparable to docetaxel, and it did not affect the level of p53, Bax, Bcl-2, and Bcl-xL in either cell line. The present study demonstrates that p53-dependent NAG-1 induction is linked to cell death and that NAG-1 induction without accompanying alteration of antiapoptosis protein Bcl-2 family members may not lead to cancer cell death.     

Octahedral Pt(IV) complex K101 induces apoptosis via ERK1/2 activation and the p53 pathway in human colon cancer cells

Recently, the synthesized octahedral Pt(IV) compound trans,cis-Pt(acetato)2Cl2(1,4-butanediamine), K101, showed potent anti-tumor activity in vitro and in vivo. For the further investigation of K101-induced anti-cancer activity, we tested cytotoxicity against various cancer cell lines and performed the histoculture drug response assay (HDRA) against human colorectal tumor tissues in vitro. We investigated the signaling pathway of K101-induced apoptosis via expression of p53 and ERK1/2 in the human colon cell line HCT116. The cytotoxicity and the three-dimensional HDRA of K101 were evaluated using the MTT assay. To study the K101-induced apoptosis pathway, we performed FACS analysis and immunoblotting of p53, p21, Bax, Fas and ERK1/2 in HCT116 cells treated with or without K101. The cytotoxic IC50 values of K101 ranged from 1.15 to 2.38 micromol/l, compared to cisplatin ranging from 2.13 to 13.1 micromol/l. Among several cancer cell lines, K101 showed greater potency than cisplatin in colon cancer cell lines. In the HDRA, K101 showed 80.0-91.4% efficacy rates compared with 48.6% for cisplatin against colorectal cancer patient tissues. In the signaling pathway, the expression of p53 and phospho-ERK1/2 was increased in a time-dependent manner by treatment with K101 in the HCT116 cells. When K101 was treated with MEK inhibitor U0126, the cell death rate was increased. The octahedral Pt(IV) complex K101 could be an attractive candidate as a chemotherapeutic agent against colon cancer. ERK1/2 activation and the p53 pathway may play significant functions in mediating K101-induced apoptosis in human colon cancer cells.     

The effect of full length and mature NAG-1 protein overexpression on cytotoxicity of celecoxib, tamoxifen and doxorubicin in HT1080

BACKGROUND AND THE PURPOSE OF THE STUDY: Non-steroidal anti-inflammatory drug-activated gene-1 (NAG-1) is involved in inflammation, apoptosis/survival and tumorigenesis as well as resistance to chemotherapy. NAG-1 protein is synthesized as pro-peptide, cleaved and secreted as mature protein. Regulation of NAG-1 is not completely discovered and increased level of NAG-1 has been reported in many cancers. The expression of NAG-1 in cancer cells could affect the progression of tumor growth. In addition the secretion of full length and mature forms of NAG-1 can influence cell proliferation in other cells. In this study the role of full length and mature forms of NAG-1 on viability of HT-1080 and MCF-7 cells were evaluated, and the cytotoxicity of celecoxib, indomethacin, tamoxifen and doxorubicin in HT1080 cells stably expressing NAG-1 were also tested. METHODS: Full length and mature NAG-1 was cloned from cDNA library of HCT116 cells and stably transfected in HT1080 cells. Cells were treated with different concentrations of indomethacin, celecoxib, tamoxifen and doxorubicin and viability was assessed by MTT assay. The effect of conditioned medium of NAG-1 expressing cells on proliferation of MCF-7 and HT1080 cells were also tested. RESULTS: The growth curves of HT1080 cells expressing full length and mature NAG-1 were not different. The viability of HT1080 cells expressing NAG-1 in the presence of indomethacin, doxorubicin and tamoxifen compared to untransfected cells was higher. The proliferation of HT1080 and MCF-7 cells were inhibited by conditioned medium of NAG-1 expressing cells in 24 and 48 hrs. MAJOR CONCLUSION: NAG-1 expression enhances drug resistance to tamoxifen, indomethacin and doxorubicin in HT1080. In addition, condition medium of NAG-1 expression cells inhibits proliferation in MCF-7 and HT1080 cells. Thus, NAG-1 expression can induce drug resistance in NAG-1 expressing cells and inhibition of viability in non expressing cells. Thus, NAG-1 is suggested as a marker for effective cancer chemotherapy and tumor progression.     

长链非编码RNA NR2F1-AS1靶向微小RNA-145-5p调控结肠癌细胞增殖、迁移和侵袭的分子机制

目的 探讨长链非编码RNA(lnc RNA)核受体亚族2F组成员1反义RNA(NR2F1-AS1)对结肠癌细胞增殖、迁移和侵袭的影响及作用机制.方法 收集2015年3月至2017年5月青海省第五人民医院外科手术切除并经病理证实为原发性结肠腺癌组织25例,另留取距肿瘤边缘3 cm的癌旁组织(正常结肠组织).实时荧光定量P...     

Inhibition of G1/S transition potentiates oxaliplatin-induced cell death in colon cancer cell lines

In a series of colorectal cancer cell lines, both necrosis and apoptosis were induced upon exposure to oxaliplatin, and enhanced by co-administration of the Hsp90 inhibitor 17-AAG. We analyzed the effects of these interventions on the cell cycle, and found that oxaliplatin treatment caused G1 and G2 arrest in HCT116 cells, and S-phase accumulation in two p53-deficient cell lines (HT29 and DLD1). Addition of 17-AAG enhanced cell cycle effects of oxaliplatin in HCT116, and induced G1 arrest and decrease in S-phase population in the other cell lines. Analysis of cell cycle proteins revealed that the major difference between the cell lines was that in HCT116, 17-AAG resulted in profound inhibition of expression and phosphorylation of late G1 proteins cyclin E and cdk2, with no effect on p21/WAF1 induction. Consistent with these, an HCT116 p53(-/-) line, lacking p21, showed resistance to oxaliplatin, failure to enter apoptosis, and an accumulation of cells in S-phase. Introduction of p21 in these cells caused reversal of that phenotype, including restoration of the G1 block and re-sensitization to oxaliplatin. Inhibition of G1/S progression using cdk2 inhibitor also enhanced oxaliplatin cytotoxicity. We conclude that in colon cancer cells with impaired p53 function, interventions directed to cycle arrest in G1 may potentiate oxaliplatin activity.     

甲磺酸阿帕替尼通过p53抑制结肠癌细胞HCT116增殖的作用及机制

摘要：目的 探讨阿帕替尼对人结肠癌细胞HCT116及敲除野生型p53的HCT116细胞抑制作用的差异并探讨 其机制。方法 以0、15、30、60 μmol/L阿帕替尼处理HCT116 p53+/+、HCT116 p53-/-细胞24、48 h,以CCK-8法检测阿 帕替尼对两株细胞增殖的抑制作用;流式细胞术（Annexin V/PI法）检测0、15、30 μmol/L阿帕替尼处理细胞24 h后凋 亡率变化;实时荧光定量PCR法和Western blot法检测0、15、30 μmol/L阿帕替尼处理细胞24 h后,p53、NF-κB p65、 Caspase-3 mRNA和蛋白表达变化。结果 CCK-8结果显示,阿帕替尼能抑制HCT116 p53+/+、HCT116 p53-/-细胞的增 殖,抑制作用具有剂量依赖性（P＜0.01）。流式细胞术结果显示,阿帕替尼干预后,HCT116 p53+/+、HCT116 p53-/-细胞 凋亡率升高（P＜0.01）。与HCT116 p53+/+细胞相比,阿帕替尼处理后HCT116 p53-/-细胞的凋亡率较低（P＜0.05）。阿 帕替尼处理后,HCT116 p53+/+、HCT116 p53-/-细胞中 p53、NF-κB p65、capsase-3 mRNA 和 Caspase-3 蛋白表达升高 （P＜0.05）。阿帕替尼干预后,HCT116 p53+/+细胞中 p53、NF-κB p65 蛋白表达下调而 HCT116 p53-/-细胞中 NF-κB p65蛋白表达上调（P＜0.01）。结论 阿帕替尼可能通过p53/NF-κB通路抑制HCT116 p53+/+、HCT116 p53-/-细胞增殖 并促进细胞凋亡,结肠癌细胞可能通过野生型p53对阿帕替尼产生耐药。     

LncRNA-MALAT1通过miR-142-3p/TEAD1分子轴调控结直肠癌细胞增殖与凋亡#br#

目的　探讨长链非编码RNA-肺腺癌转移相关转录子1（LncRNA-MALAT1）对结直肠癌细胞增殖与凋亡的影响及相关作用机制。方法　通过Real-time PCR与Western blot实验检测结直肠癌细胞株与人正常结肠上皮细胞中LncRNA-MALAT1、miR-142-3p基因以及TEA结构域转录因子1（TEAD1）蛋白的表达;使用si-MALAT1转染HCT116细胞,在此基础上共转染miR-142-3p inhibitor,利用Real-time PCR与Western blot检测细胞中LncRNA-MALAT1与miR-142-3p基因以及TEAD1、Bax、Bcl-2与Cyclin D1蛋白的表达,通过CCK-8实验检测细胞的增殖水平,通过流式细胞术检测细胞的凋亡水平,通过双荧光素酶实验检测LncRNA-MALAT1与miR-142-3p以及miR-142-3p与TEAD1的结合。结果　与人正常结肠上皮细胞相比,结直肠癌细胞株中LncRNA-MALAT1基因与TEAD1蛋白呈高表达,miR-142-3p基因呈低表达（P＜0.05）;沉默LncRNA-MALAT1能够促进miR-142-3p基因与Bax蛋白表达,抑制LncRNA-MALAT1基因以及Bcl-2、Cyclin D1与TEAD1蛋白表达,抑制细胞增殖,并促进细胞凋亡;在此基础上沉默miR-142-3p能够对上述调控作用实现部分逆转;双荧光素酶实验结果显示,LncRNA-MALAT1与miR-142-3p以及miR-142-3p与TEAD1能够结合。结论　LncRNA-MALAT1能够结合miR-142-3p,促进miR-142-3p靶基因TEAD1表达,进而促进结直肠癌细胞增殖,抑制其细胞凋亡,促进结直肠癌的病理进程。     

Antitumor 1-nitroacridine derivative C-1748, induces apoptosis, necrosis or senescence in human colon carcinoma HCT8 and HT29 cells

C-1748 is a DNA-binding agent with potent antitumor activity, especially towards prostate and colon carcinoma xenografts in mice. Here, we elucidated the nature of cellular response of human colon carcinoma HCT8 and HT29 cells to C-1748 treatment, at biologically relevant concentrations (EC₉₀ and their multiplicity). Cell cycle analysis showed gradual increase in HCT8 cells with sub-G1 DNA content (25% after 72 h) considered as apoptotic. Hypodiploid cell population increased up to 60% upon treatment with 4× EC₉₀ concentration of the drug. Compared with HCT8 cells, the fraction of sub-G1 HT29 cells did not exceed 14%, even following 4-fold dose escalation. Morphological changes and biochemical markers such as: phosphatydylserine externalization, apoptotic DNA breaks, mitochondrial dysfunction and caspase activation confirmed the presence of considerable amount of apoptotic HCT8 cells but only a low amount of apoptotic HT29 cells. Next, we demonstrated that HCT8 cells surviving after exposure to C-1748 were in the state of senescence, based on altered cell morphology and expression of a pH 6-dependent β-galactosidase. On the contrary, no β-galactosidase staining was observed in HT29 cells after C-1748 treatment. Moreover, prolonged drug incubation (up to 168 h) resulted in massive detachment of cells from culture plates, which together with Annexin V/PI results, indicated that necrosis was the main response of HT29 cells to C-1748 treatment. We also determined the ability of C-1748 to induce reactive oxygen species (ROS) in colon cancer cells and demonstrated, that generation of ROS was not essential for C-1748-induced apoptosis and cytotoxic activity of this drug.     

New benzimidazole acridine derivative induces human colon cancer cell apoptosis in vitro via the ROS-JNK signaling pathway

Aim:

To investigate the mechanisms underlying anticancer action of the benzimidazole acridine derivative N-{(1H-benzo[d]imidazol-2-yl)methyl}-2-butylacridin-9-amine(8m) against human colon cancer cells in vitro.

Methods:

Human colon cancer cell lines SW480 and HCT116 were incubated in the presence of 8m, and then the cell proliferation and apoptosis were measured. The expression of apoptotic/signaling genes and proteins was detected using RT-PCR and Western blotting. ROS generation and mitochondrial membrane depolarization were visualized with fluorescence microscopy.

Results:

8m dose-dependently suppressed the proliferation of SW480 and HCT116 cells with IC50 values of 6.77 and 3.33 μmol/L, respectively. 8m induced apoptosis of HCT116 cells, accompanied by down-regulation of Bcl-2, up-regulation of death receptor-5 (DR5), truncation of Bid, cleavage of PARP, and activation of caspases (including caspase-8 and caspase-9 as well as the downstream caspases-3 and caspase-7). Moreover, 8m selectively activated JNK and p38 without affecting ERK in HCT116 cells. Knockout of JNK1, but not p38, attenuated 8m-induced apoptosis. In addition, 8m induced ROS production and mitochondrial membrane depolarization in HCT116 cells. Pretreatment with the antioxidants N-acetyl cysteine or glutathione attenuated 8m-induced apoptosis and JNK activation in HCT116 cells.

Conclusion:

The new benzimidazole acridine derivative, 8m exerts anticancer activity against human colon cancer cells in vitro by inducing both intrinsic and extrinsic apoptosis pathways via the ROS-JNK1 pathway.     

DNA damage response to the Mdm2 inhibitor Nutlin-3

Mdm2 inhibitors represent a promising class of p53 activating compounds that may be useful in cancer treatment and prevention. However, the consequences of pharmacological p53 activation are not entirely clear. We observed that Nutlin-3 triggered a DNA damage response in azoxymethane-induced mouse AJ02-NM₀ colon cancer cells, characterized by the phosphorylation of H2AX (at Ser-139) and p53 (at Ser-15). The DNA damage response was highest in cells showing robust p53 stabilization, it could be triggered by the active but not the inactive Nutlin-3 enantiomer, and it was also activated by another pharmacological Mdm2 inhibitor (Caylin-1). Quantification of γH2AX-positive cells following Nutlin-3 exposure showed that approximately 17% of cells in late S and G2/M were mounting a DNA damage response (compared to a ∼50% response to 5-fluorouracil). Nutlin-3 treatment caused the formation of double-strand DNA strand breaks, promoted the formation of micronuclei, accentuated strand breakage induced by doxorubicin and sensitized the mouse colon cancer cells to DNA break-inducing topoisomerase II inhibitors. Although the HCT116 colon cancer cells did not mount a significant DNA damage response following Nutlin-3 treatment, Nutlin-3 enhanced the DNA damage response to the nucleotide synthesis inhibitor hydroxyurea in a p53-dependent manner. Finally, p21 deletion also sensitized HCT116 cells to the Nutlin-3-induced DNA damage response, suggesting that cell cycle checkpoint abnormalities may promote this response. We propose that p53 activation by Mdm2 inhibitors can result in the slowing of double-stranded DNA repair. Although this effect may suppress illegitimate homologous recombination repair, it may also increase the risk of clastogenic events.     

Anticancer bioactive peptides suppress human colorectal tumor cell growth and induce apoptosis via modulating the PARP-p53-Mcl-1 signaling pathway

Aim:

We have reported novel anticancer bioactive peptides (ACBPs) that show tumor-suppressive activities in human gastric cancer, leukemia, nasopharyngeal cancer, and gallbladder cancer. In this study, we investigated the effects of ACBPs on human colorectal cancer and the underlying mechanisms.

Methods:

Cell growth and apoptosis of human colorectal tumor cell line HCT116 were measured using cell proliferation assay and flow cytometry, respectively. The expression levels of PARP, p53 and Mcl1A were assessed with Western blotting and immunohistochemistry. For evaluation of the in vivo antitumor activity of ACBPs, HCT116 xenograft nude mice were treated with ACBPs (35 μg/mL, ip) for 10 days.

Results:

Treatment of HCT116 cells with ACBPs (35 μg/mL) for 4–6 days significantly inhibited the cell growth. Furthermore, treatment of HCT116 cells with ACBPs (35 μg/mL) for 6–12 h significantly enhanced UV-induced apoptosis, increased the expression of PARP and p53, and decreased the expression of Mcl-1. Administration of ACBPs did not change the body weight of HCT116 xenograft nude mice, but decreased the tumor growth by approximately 43%, and increased the expression of PARP and p53, and decreased the expression of Mcl-1 in xenograft mouse tumor tissues.

Conclusion:

Administration of ACBPs inhibits human colorectal tumor cell growth and induces apoptosis in vitro and in vivo through modulating the PARP-p53-Mcl-1 signaling pathway.     

c-Jun N-terminal kinase 1 is required for cordycepin-mediated induction of G2/M cell-cycle arrest via p21WAF1 expression in human colon cancer cells

Cordycepin (3′-deoxyadenosine) has many anti-cancer properties. However, neither its molecular mechanism nor its molecular targets are well understood. In the present study, we investigated novel molecular mechanisms for the anti-tumor effects of cordycepin in human colon cancer HCT116 cells. After treatment of cells with cordycepin, dose-dependent cell growth inhibition was observed at an IC₅₀ value of 200 μM. Cordycepin treatment resulted in G2/M-phase cell-cycle arrest, which was associated with increased p21WAF1 levels and reduced amounts of cyclin B1, Cdc2, and Cdc25c in a p53-independent pathway. Moreover, cordycepin treatment induced activation of JNK (c-Jun N-terminal kinases). Pretreatment with SP600125, a JNK-specific inhibitor, abrogated cordycepin-mediated p21WAF1 expression, cell growth inhibition, and reduced cell-cycle proteins. Furthermore, JNK1 inhibition by small interfering RNA (siRNA) produced similar results: suppression of cordycepin-induced p21WAF1 expression, decreased cell growth, and reduced cell-cycle proteins. Together, these results suggest a critical role for JNK1 activation in cordycepin-induced inhibition of cell growth and G2/M-phase arrest in human colon cancer cells.     

lncRNA FOXCUT调控Notch通路抑制结肠癌细胞增殖、侵袭的实验研究#br#

目的　研究长链非编码RNA（lncRNA）FOXC1启动子临近转录体（FOXCUT）对结肠癌细胞（HCT116）增殖、侵袭的影响及作用机制。方法　向HCT116细胞转染FOXCUT siRNA（FOXCUT siRNA组）,以转染阴性siRNA为阴性对照组（FOXCUT siRNA-NC组）,未转染组为正常对照组（NC组）,通过qPCR法确定FOXCUT干扰效果,并检测结肠癌及癌旁组织、人结肠癌细胞株（SW480、SW620、HCT116、HT29）和人正常结肠上皮细胞（NCM460）中FOXCUT mRNA表达水平;通过CCK-8、集落形成实验和Transwell实验检测沉默FOXCUT对HCT116细胞增殖、集落形成能力及侵袭能力的影响;qPCR检测沉默FOXCUT后HCT116细胞中Notch1、Hes1 mRNA表达变化;蛋白免疫印迹（Western blot）实验检测HCT116细胞中Notch1、Hes1蛋白表达情况。结果　与癌旁组织比较,结肠癌组织中FOXCUT mRNA表达水平显著增高（P＜0.01）;与NCM460细胞比较,SW480、SW620、HCT116、HT29结肠癌细胞株中FOXCUT mRNA表达水平均显著增高（P＜0.05）,其中以HCT116细胞中表达水平最高;与FOXCUT siRNA-NC组比较,FOXCUT siRNA组HCT116细胞中FOXCUT mRNA表达水平显著降低（P＜0.01）;NC组、FOXCUT siRNA-NC组HCT116细胞中FOXCUT mRNA表达水平差异无统计学意义（P＞0.05）;沉默FOXCUT可显著抑制细胞增殖、集落形成能力及侵袭（P＜0.05）,并抑制Notch1、Hes1 mRNA及蛋白表达（P＜0.01）。结论　沉默FOXCUT可抑制结肠癌HCT116细胞增殖及侵袭,可能与抑制Notch通路有关。     

Low-dose BBR3610 toxicity in colon cancer cells is p53-independent and enhanced by inhibition of epidermal growth factor receptor (ERBB1)-phosphatidyl inositol 3 kinase signaling

We have examined the mechanisms by which the multinuclear platinum chemotherapeutic BBR3610 kills human colon cancer cells. BBR3610 more efficiently killed HCT116, DLD1, SW480, and HT29 cells than BBR3464, cisplatin, or oxaliplatin. The amount of platinum uptake per cell and its incorporation into DNA were identical for BBR3464 and BBR3610. BBR3610 lethality (IC(75)) was unaltered comparing HCT116 wild-type and p53-/- cells, was reduced in p21-/- cells, and was enhanced in K-RAS D13 null cells. Small molecule or molecular inhibition of epidermal growth factor receptor (ERBB1) or phosphatidyl inositol 3 kinase (PI3K) enhanced BBR3610 toxicity in HCT116, DLD1, and SW480 cells. Small molecule or molecular inhibition of caspase 8 function abolished the toxicity of BBR3610 and of BBR3610 + ERBB1 inhibitor treatments, whereas inhibition of caspase 9 suppressed the ability of ERBB1 inhibitors to enhance BBR3610 lethality. Treatment with BBR3610 reduced AKT activity; the expression of dominant-negative AKT enhanced and expression of constitutively active AKT suppressed, respectively, the toxicity of BBR3610 and of BBR3610 + ERBB1 inhibitor treatments. Treatment with BBR3610 reduced expression of c-FLIP-s and MCL-1, levels that were maintained in cells expressing constitutively active AKT. Overexpression of c-FLIP-s or loss of BID function suppressed BBR3610 toxicity, whereas overexpression of XIAP or Bcl-xL suppressed the potentiation of cell killing by ERBB1 inhibitors. Collectively, our data argue that BBR3610 promotes cell killing via a caspase 8-dependent mechanism, which can be enhanced by ERBB1/PI3K inhibitors that promote additional BBR3610-dependent cell killing via activation of BAX and caspase 9.     

1,1-Bis(3'-indolyl)-1-(p-substitutedphenyl)methanes are peroxisome proliferator-activated receptor gamma agonists but decrease HCT-116 colon cancer cell survival through receptor-independent activation of early growth response-1 and nonsteroidal anti-inflammatory drug-activated gene-1

1,1-Bis-(3'-indolyl)-1-(p-substitutedphenyl)methanes containing p-trifluoromethyl (DIM-C-pPhCF3), p-t-butyl (DIM-C-pPhtBu), and phenyl (DIM-C-pPhC6H5) substituents decrease survival of HCT-116 colon cancer cells and activate peroxisome proliferator-activated receptor (PPAR) gamma in this and other cancer cell lines. These PPARgamma-active compounds had minimal effects on expression of cell cycle proteins and did not induce caveolin-1 in HCT-116 cells. However, these compounds induced nonsteroidal anti-inflammatory drug-activated gene-1 (NAG-1) and apoptosis in HCT-116 cells, and in time-course studies, the PPARgamma agonists maximally induced early growth response-1 (Egr-1) protein within 2 h, whereas a longer time course was observed for induction of NAG-1 protein. These data, coupled with deletion and mutation analysis of both the Egr-1 and NAG-1 gene promoters, indicate that activation of NAG-1 by these compounds was dependent on prior induction of Egr-1, and induction of these responses was PPARgamma-independent. Results of kinase inhibitor studies also demonstrated that activation of Egr-1/NAG-1 by methylene-substituted diindolylmethanes (C-DIMs) was phosphatidylinositol 3-kinase-dependent, and this represents a novel receptor-independent pathway for C-DIM-induced growth inhibition and apoptosis in colon cancer cells.     

NES1基因在结肠癌细胞中表达及意义

目的：探讨NES1基因在结肠癌细胞中表达及意义。方法用RT-PCR方法检测NES1mRNA在正常结肠细胞、结肠癌细胞株HT29、HCT116表达情况。结果 NES1在结肠癌细胞株中表达较正常结肠黏膜细胞明显增高。结论 NES1基因在结肠癌的发生发展中起重要作用,这为选用NES1作为结肠癌治疗靶点提供了实验依据及理论基础。     

Antitumor effect of the angiogenesis inhibitor bevacizumab is dependent on susceptibility of tumors to hypoxia-induced apoptosis

Angiogenesis inhibition has been shown to enhance the therapeutic efficacy of cytotoxic chemotherapy in colorectal cancer. The basis of the contribution of this modality has not been defined fully. To determine the potential role of hypoxia-induced apoptosis, we studied a series of colon cancer cell lines with varying susceptibility to hypoxia. We exposed HT29 and HCT116 colon adenocarcinoma cell lines to sublethal periods of hypoxia three times weekly for 40 exposures, and derived cell lines both more resistant (from HT29) and more sensitive (from HCT116) to hypoxia-induced apoptosis. Both hypoxia-derived cell lines demonstrated more rapid growth than the parental lines when implanted subcutaneously in immunodeficient mice. Treatment of tumor-bearing mice with bevacizumab resulted in depletion of tumor microvasculature, upregulation of Hypoxia-inducible factor-1 alpha (HIF-1alpha), and increased pimonidazole staining, consistent with an anti-angiogenic effect and induction of hypoxia in tumors derived from all cell lines. The proportion of apoptotic cells was increased in all the treated tumors, and was most pronounced in the bevacizumab-treated HCT116-derived cells. The bevacizumab-treated tumors showed growth delay in HT29 and its derivative, and the parental HCT116. In the hypoxia-sensitive HCT116-derived tumors, marked tumor shrinkage and prolonged growth control occurred. Therefore, bevacizumab treatment is an effective inducer of a hypoxic environment, but the resulting cell kill and tumor shrinkage is determined by the susceptibility of the tumor to apoptosis. The induction of apoptosis by hypoxia may contribute to the benefits of such treatment in the clinical setting.