Innovative Therapeutic Strategies Targeting Colorectal Cancer Stem Cells

Colorectal cancer is the fourth most common cause of cancer-related death. Although many advances in the treatment of this disease have been made, a large number of patients develop metastasis and resistance to current therapies. The current evidence indicates that cancer stem cells (CSCs) and epithelial-to-mesenchymal transition (EMT) have crucial roles in colorectal carcinogenesis and metastasis. It is also very important to understand the mechanisms that allow the survival of CSCs, such as metabolic reprogramming, which permits them to obtain specific properties or the activation of alternative signaling pathways in response to first-line therapies. In this review, we discuss the failure of conventional therapies for colorectal cancer and provide a brief analysis of new therapeutic strategies for targeting non-responsive CSC. We highlight the use of combination therapies, such as horizontal or vertical targeting, as the most efficient strategy for eradicating these subpopulations of cancer cells.     

结直肠癌干细胞和肿瘤微环境的相互影响及靶向策略

肠癌干细胞是结直肠癌中少量的具有恶性表型特征的未分化致瘤细胞,具有自我更新和分化克隆等独特的特征,被认为是肿瘤复发、耐药、转移的主要原因。肠癌干细胞与肿瘤微环境中各成分之间互相依存,互相影响,了解肿瘤微环境中肿瘤干细胞和其他成分之间的联系,可能对结直肠癌的治疗具有重要作用。本文就肠癌干细胞和肿瘤微环境的关系及靶向治疗进行综述。     

Targeting Cancer Stem Cells by Phytochemicals: a Multimodal Approach to Colorectal Cancer

Cancer stem cells (CSCs) exist within a tumor as a rare subpopulation, with the capacity of self-renewal and the ability to differentiate into heterogeneous population of cancer cells. CSCs are increasingly being implicated in tumor recurrence thereby further augmenting the menace of the malignant disease. Characterization of CSCs has unearthed their pivotal role in all the hallmarks of cancer including tumorigenesis, angiogenesis, metastasis and drug resistance, thereby designating cancer as a “stem cell disease.” Here, we discuss the limitations of current therapeutic strategies that spare CSCs thereby failing to achieve complete cure of colorectal cancer, and elucidate the role of multimodal CSC-targeted treatment strategies, using natural phytochemicals and their derivatives. With emerging evidences identifying the molecular targets of phytochemicals in colorectal CSCs, development of better therapeutic strategies uprooting CSCs, the root of all evils, can be envisaged.     

Targeting Notch Signaling in Colorectal Cancer

The activation of Notch signaling is implicated in tumorigenesis in the colon due to the induction of pro-survival signaling in colonic epithelial cells. Chemoresistance is a major obstacle for treatment and for the complete eradication of colorectal cancer (CRC); hence, the inhibition of Notch is an attractive target for CRC and several groups are working to identify small molecules or monoclonal antibodies that inhibit Notch or its downstream events; however, toxicity profiles in normal cells and organs often impede the clinical translation of these molecules. Dietary agents have gained momentum for targeting several pro-survival signaling cascades, and recent studies demonstrated that agents that inhibit Notch signaling result in growth inhibition in preclinical models of CRC. In this review, we focus on the importance of Notch as a preventive and therapeutic target for colon cancer and on the effect of WA on this signaling pathway in the context of colon cancer.     

miR-16对大肠癌细胞增殖能力的影响

目的研究miR-16在大肠癌组织中的表达及其在大肠癌细胞株HCT116和LoVo细胞增殖中的作用。方法运用Realtime PCR法检测miR-16在33对配对肠癌及癌旁正常组织中的表达。HCT116和LoVo细胞转染miR-16 抑制剂、阴性对照剂和模拟剂;CCK8法检测细胞增殖;流式细胞仪进行细胞周期检测;Western blot法检测细胞中cyclinD1、CDK6和GAPDH蛋白表达。结果（1）Realtime PCR结果显示miR-16在大肠癌组织中低表达（P=0.047）。（2）miR-16过表达可抑制HCT116和LoVo细胞增殖,诱导细胞G₀/G₁期阻滞,降低cyclinD1和CDK6蛋白表达。结论miR-16过表达可以抑制大肠癌细胞的增殖能力。     

hTERT启动子核心序列的克隆及其在肺癌细胞中靶向转录活性的研究

背景与目的构建人端粒酶催化亚单位(hTERT)启动子核心片段调控的绿色荧光蛋白(EGFP)基因表达质粒,并初步研究hTERT启动子在肺癌细胞和正常细胞中     

细胞内钙在人胃癌细胞缺氧诱导因子-1表达及转录激活中的作用

 目的 观察细胞内钙变化对氧调节性亚单位HIF-1α表达及其HIF-1转录激活的影响。方法 常氧时,采用通透性细胞内钙的螯合剂BAPTA-AM或钙的离子载体Ionomycin降低或升高细胞内钙,用Western-blot检测其对SGC7901细胞中HIF-1α蛋白表达的影响,然后采用间接免疫荧光法、双荧光素酶报告系统及ELISA法检测改变细胞内钙对HIF-1α转位、HIF-1转录活性及其靶基因血管内皮生长因子（Vascular endothelial growth factor,VEGF）分泌的影响。结果 用BAPTA-AM降低细胞内钙,能诱导HIF-1α的表达,促进HIF-1α的转位,增强HIF-1的转录活性,增加HIF-1调节基因VEGF的分泌。用Ionomycin升高细胞内钙,虽然也有微弱的促HIF-1α稳定及转位作用,但对HIF-1转录活性及VEGF的分泌并无明显影响。结论 螯合细胞内钙能诱导胃癌细胞中HIF-1α的表达及HIF-1的转录激活,提示细胞内钙变化可能在胃癌细胞的缺氧信号转导过程中发挥了重要作用。     

Circulating Tumor Cells and Colorectal Cancer

The significance of circulating tumor cells (CTCs) has been discussed for more than a century. The advent of modern technology has allowed for more reliable detection of CTCs, and recent studies have provided compelling evidence that CTCs predict clinical response in metastatic colorectal cancer (mCRC). Combination of CTC analysis with independent prognostic factors has demonstrated powerful synergy in some studies. The ability of CTCs to predict metastasis and therapy-specific response has high potential clinical utility, with early studies showing promising results in colorectal cancer (CRC). Reliable CTC detection has also allowed for examination of tumor cell dissemination during surgery, and there appears to be a heavy dependence on the approach chosen. This review discusses the evidence for CTC significance, with particular focus on detection methods, novel markers, and clinical outcomes in CRC. Numerous opportunities exist for preclinical, clinical, and translational studies to explore molecular determinants within CTCs, as well as the value of CTCs in directing targeted therapeutics.     

Ad-wtp53对p53状态不同结直肠癌细胞生长的影响

目的：探讨胃癌组织中MAGE－1基因启动子B'B区去甲基化状态及其与病理分级及临床分期的关系。方法：取胃癌组织标本40例，另外取同患者相应的癌旁组织40例作为对照。采用分子生物学技术－甲基化敏感性内切酶酶切及PCR扩增技术，研究了胃癌组织中MAGE－1启动子B'B区的去甲基化状态。结果：在所检测的胃癌组织标本中MAGE－1基因启动子B'B区去甲基化的发生率为25％（10／40）。而在癌旁组织中发生率为0，两者发生率的差别具有显著的统计学意义（P＜0．01）。在低分化腺癌中MAGE－1基因的B'B区去甲化发生率为50％（6／12），中分化腺癌中发生率为18．7％（3／16），高分化腺癌中发生率为8．3（1／11）。其发生率的差异有统计学意义（P＜0．05）。在早期胃癌组织中B'B区的去甲基化的发生率为16．7％，在晚期胃癌组织中发生率为28．6％（P＜0．05），差别有统计学意义。结论：胃癌组织中MAGE－1基因启动子的B'B区存在去甲基化。该区的去甲基化发生率与胃癌组织的病理分级以及与临床分期有关。     

血管内皮生长因子反义核酸对宫颈癌Hela细胞的放射增敏作用

目的：血管内皮生长因子（Vascular endothelial growth factor，VEGF）在人类多数实体肿瘤中均有表达，射线可诱导其表达增加，产生放射抗拒。本实验目的在于探讨血管内皮生长因子反义寡核苷酸干涉对高转移宫颈腺癌Hela细胞株的放射增敏作用。方法：实验组通过脂质体介导将VEGF基因反义寡核苷酸瞬时转染入Hela细胞，设VEGF正义寡核苷酸组和空白对照组作比较，各组均予6MV-X线照射，剂量为0Gy、2Gy、4Gy和6Gy，用RT-PCR检测照射前后Hela细胞中VEGFmRNA表达；用流式细胞术检测细胞凋亡；平板克隆形成试验检测克隆形成率的变化。结果：与对照组比较，VEGF反义寡核苷酸不仅可抑制Hela细胞中内源性VEGFmRNA的表达（P〈0．01），亦可显著抑制辐射诱导的VEGF表达的增加（P〈0．01）；VEGF表达下调使射线诱导的细胞凋亡率明显升高（P〈0．01）．细胞克隆数量明显降低（P〈0．01）;结论：针对人VEGF基因反义寡核苷酸干涉可抑制射线诱导的VEGF的高表达，增强射线对肿瘤细胞的杀伤作用。提高细胞的放射敏感性。     

Targeting Hyaluronic Acid and Peritoneal Dissemination in Colorectal Cancer

Peritoneal metastasis (PM) from colorectal cancer (CRC) carries a significant mortality rate for patients and treatment is challenging. The development of PM is a multistep process involving detachment, adhesion, invasion and colonization of the peritoneal cavity. Cytoreductive surgery and HIPEC (hyperthermic intraperitoneal chemotherapy) for PM from CRC has some benefit but overall survival is poor and recurrence rates are high. Treatments to prevent the development of peritoneal metastasis could have the potential to improve CRC survival and disease-free outcomes.The ability of cancer cells to invade the peritoneum and become established as metastatic tumors is influenced by a multifactorial process. Hyaluronic acid (HA) has been shown to coat the mesothelial cells of the peritoneum and has been demonstrated to be utilized in various malignancies as part of the metastatic process in peritoneal dissemination. CD44, RHAMM (CD168) and ICAM-1 have all been shown to be binding partners for HA. Targeting HA-mediated binding may prevent adhesion to distant sites within the peritoneum through suppression of interaction of these molecules. Here we review the current literature and discuss key molecules involved with PM dissemination, with the potential to target these mechanisms in the delivery of future treatments.     

电离辐射后阿霉素对大肠癌HCT-8细胞毒活性的影响

[目的]研究不同剂量电离辐射后阿霉素对大肠癌细胞株HCT-8细胞毒活性的影响,旨在探讨逆转大肠癌多药耐药性的方法。[方法]体外培养大肠癌细胞株HCT-8,以400ng/ml阿霉素做为实验模型刺激浓度。实验模型分为以下5组:对照组;大剂量组(2Gy);0.05Gy 2Gy组;0.1Gy 2Gy组;0.2Gy 2Gy组。采用MTT法测定给予阿霉素后大肠癌细胞株HCT-8毒活性。[结果]与假照射组相比,2Gy照射组及0.2Gy 2Gy组照射后HCT-8细胞存活率明显降低(P<0.05),先给予低剂量照射(0.05Gy,0.1Gy)后,再给予大剂量照射,HCT-8细胞存活率降低更明显(P<0.01)。与单纯2Gy照射组比较,0.05Gy 2Gy组及0.1Gy 2Gy组HCT-8细胞生存率明显降低(P<0.05)。[结论]先给予低剂量照射后,再给予大剂量照射,阿霉素对HCT-8细胞存活率明显降低,提示低剂量照射可增强大肠癌细胞株对阿霉素的敏感性。     

MiR-1297 Regulates the Growth,Migration and Invasion of Colorectal Cancer Cells by Targeting Cyclo-oxygenase-2

Cyclo-oxygenase-2(Cox-2), a key regulator of inflammation-producing prostaglandins, promotes cellproliferation and growth. Therefore, a better understanding of the regulatory mechanisms of Cox-2 could leadto novel targeted cancer therapies. MicroRNAs are strongly implicated in colorectal cancer but their specificroles and functions have yet to be fully elucidated. MiR-1297 plays an important role in lung adenocarcinomaand laryngeal squamous cell carcinoma, but its significance in colorectal cancer (CRC) has yet to be reported.In our present study, we found miR-1297 to be down regulated in both CRC-derived cell lines and clinical CRCsamples, when compared with normal tissues. Furthermore, miR-1297 could inhibit human colorectal cancerLOVO and HCT116 cell proliferation, migration, and invasion in vitro and tumorigenesis in vivo by targetingCox-2. Moreover, miR-1297 directly binds to the 3`-UTR of Cox-2, and the expression level was drasticallydecreased in LOVO and HCT116 cells following overexpression of miR-1297. Additionally, Cox-2 expressionlevels are inversely correlated with miR-1297 expression in human colorectal cancer xenograft tissues. Theseresults imply that miR-1297 has the potential to provide a new approach to colorectal cancer therapy by directlyinhibiting Cox-2 expression.     

Inhibitory Effects of Cyrtopodion scabrum Extract on Growth of Human Breast and Colorectal Cancer Cells

Breast and colorectal cancers rank high in Iran as causes of mortality. Most of the current treatments areexpensive and non-specific. The potential anticancer properties of common home gecko, Cyrtopodion scabrum,were investigated in this study. The effects of C. scabrum extract on proliferation, viability and migration of thecolorectal cancer (SW-742), breast cancer (MCF-7) and normal (MSC) cell lines were investigated using MTTand in vitro wound healing assay. IC50 values calculated for the extract were 559±28.9μg/ml for MCF-7 and339±11.3μg/ml for SW-742. No toxic effects on the normal control cells were observed. MCF-7 and SW-742 cellgrowth was inhibited by 32.6% and 62%, under optimum conditions, compared to the untreated control cells.The extract also decreased the motility and migration ability of both cancer cell lines, with no significant effectson the normal control cells. Data suggest C. scabrum extract as a useful natural resource for targeting cancercells specifically.     

Mitochondrial Effects of Teucrium Polium and Prosopis Farcta Extracts in Colorectal Cancer Cells

Background: Teucrium Polium and Prosopis Farcta have been traditionally employed in cancer treatment. In this study we evaluated the effects of methanolic extracts of these two plants in HT-29 cells. Methods: IC50s of extracts were obtained via MTT assay and the levels of ROS production, cell death, collapse of mitochondrial membrane potential and Sirt3 enzyme activity were determined. Results: After 48 hours exposure, IC50s for Teucrium and Prosopis extracts were 3 and 2μg/ml, respectively. Extracts induced higher ROS production after 6 hours than after 12 hours. Mitochondrial membrane potential collapse and cell death rate were also increased; Teucrium caused greater cell death than Prosopis. Extracts from both plants increased Sirt3 activity in its normal form, but only Teucrium extract caused a significant increase in activity of Sirt3 enzyme isolated from cancer cells. Conclusion: Teucrium and Prosopis extracts exert anticancer activity via mitochondrial alterations, as exemplified by increased ROS levels, Sirt3 activity and cell death in HT-29 colorectal cancer cells.     

热休克诱导结直肠癌细胞外泌体的免疫效应

目的探讨热休克影响结直肠癌细胞外泌体诱导树突状细胞肿瘤相关蛋白因子的释放和细胞毒效应。方法临床经病理学确诊的结直肠管状腺癌细胞分离培养,43℃热休克1 h,四步离心获得细胞上清外泌体。诱导树突状细胞刺激细胞毒反应。酶联免疫测定树突状细胞释放TNF-α、MIP-1α、RANTES,MTT法测定对结直肠癌细胞的杀伤作用。结果结直肠癌细胞热休克获得的外泌体刺激DC分泌TNF-α、MIP-1α、RANTES,与对照组比较差异有统计学意义（P<0．05）。热休克促进外泌体诱导的结直肠癌细胞杀伤活性,与未经热休克培养获得的外泌体诱导组比较差异有统计学意义(P<0.05),与结直肠癌细胞裂解物诱导组比较差异具有统计学意义(P<0.01),未经热休克培养获得的外泌体诱导组与癌细胞裂解物诱导组比较差异有统计学意义（P<0.05）。100 μg/ml剂量诱导效应强于50 μg/ml组。结论热休克培养可以通过促进效应细胞肿瘤相关蛋白因子释放,增强对结直肠癌细胞的杀伤活性,有剂量依赖性,为开拓高效无细胞疫苗来源提供了实验基础。     

血管表皮生长因子对大肠癌细胞生长的影响机制研究

目的 探讨VEGF对大肠癌细胞生长的影响机制研究.方法 采用基因芯片方法,筛选大肠癌的相关因子.采用免疫组化方法检测大肠癌组织、癌旁组织和正常组织中VEGF的表达.采用免疫荧光方法检测三种组织内淋巴管和血管中VEGF的表达.结果 基因芯片筛选结果显示VEGF在大肠癌组织中呈高表达.免疫组化结果显示22例大肠癌组织中VEGF呈高表达,与正常组织比较,具有统计学意义,P＜0.05.免疫荧光检测结果显示在癌组织中呈高表达VEGF时其淋巴管和血管内均增高(相比于正常组时),且有统计学差异,(分别为P＜0.001和P＜0.05),在癌旁组织中VEGF呈低表达(相比于癌组织)时相对癌组织,淋巴管和血管内均降低,且有统计学差异(分别为P＜0.001和P＜0.05).结论 VEGF可以通过调节淋巴内皮细胞进行促进大肠癌细胞的生长.     

Clusterin and Chemotherapy Sensitivity Under Normoxic and Graded Hypoxic Conditions in Colorectal Cancer

Background and Aims  

In vitro studies have shown that clusterin modulates treatment sensitivity in a number of human cancers; however, the interaction between clusterin expression and hypoxia in controlling treatment response in CRC has not previously been examined. The aim of this study was to assess the effect of clusterin overexpression in CRC cells on sensitivity to 5-fluorouracil (5-FU), oxaliplatin and FOLFOX treatment under normoxic and graded hypoxic conditions.