Progression Of Human Colorectal Cancer Research Articles

MiR-133b, a muscle-specific microRNA, is a novel prognostic marker that participates in the progression of human colorectal cancer via regulation of CXCR4 expression

BackgroundMicroRNA-133b (miR-133b), which is a muscle-specific microRNA, has been reported to be downregulated in human colorectal carcinoma (CRC) when compared to adjacent non-tumor tissue. However, its diagnostic value and role in CRC have yet to be described. CXC chemokine receptor-4 (CXCR4), which participates in multiple cell processes such as cell invasion-related signaling pathways, was predicted to be a potential target of miR-133b. The aim of this study was to investigate the associations and functions of miR-133b and CXCR4 in CRC initiation and invasion.MethodsMature miR-133b and CXCR4 expression levels were detected in 31 tumor samples and their adjacent, non-tumor tissues from patients with CRC, as well as in 6 CRC cell lines, using real-time quantitative RT-PCR (qRT-PCR). Luciferase reporter assays and Western blots were used to validate CXCR4 as a putative target gene of miR-133b. Regulation of CXCR4 expression by miR-133b was assessed using qRT-PCR and Western blot analysis, and the effects of exogenous miR-133b and CXCR4 on cell invasion and migration were evaluated in vitro using the SW-480 and SW-620 CRC cell lines.ResultsA significant downregulation of miR-133b was observed in 93.55% of CRC tissues, and the expression of miR-133b was much lower in metastatic tumors (stage C and D, stratified by the Modified Dukes Staging System) than in primary tumors (stage A and B). In contrast, CXCR4 protein expression significantly increased in 52.63% of CRC samples, and increased CXCR4 expression in CRC was associated with advanced tumor stage. CXCR4 was shown to be a direct target of miR-133b by luciferase reporter assays, and transfection of miR-133b mimics inhibited invasion and stimulated apoptosis of SW-480 and SW-620 CRC cells.ConclusionsOur study demonstrated that downregulated miR-133b contributed to increased cell invasion and migration in CRC by negatively regulating CXCR4. These findings may be significant for the development of therapy target for CRC.

RASSF2 hypermethylation is present and related to shorter survival in squamous cervical cancer

Ras association (RalGDS/AF-6) domain family member 2 (RASSF2) is a gene involved in the progression of several human cancers, including breast, colorectal and lung cancer. The aims of this study were to determine the hypermethylation of the gene in squamous cervical cancer and precursor lesions, along with that of RASSF1 and the recently described EPB41L3, and to analyze the potential prognostic role of these genes. Methylation-specific PCR and bisulfite sequencing were used to analyze the methylation status of RASSF2 and EPB41L3 gene in 60 squamous cervical cancer, 76 cervical intraepithelial neoplasias grade III, 16 grade II, 14 grade I and 13 cases of normal tissue adjacent to cervical intraepithelial neoplasia. RASSF2 expression was evaluated by immunohistochemistry and the re-expression of RASSF2 and EPB41L3 was analyzed by quantitative reverse-transcription PCR in HeLa, SiHa, C33A and A431 cell lines treated with 5-aza-2′-deoxycytidine and/or trichostatin. RASSF1 hypermethylation and human papillomavirus type were also analyzed in all the cases by methylation-specific PCR and reverse line blot, respectively. RASSF2 hypermethylation was predominant in squamous cervical cancer (60.9%) compared with cervical intraepithelial neoplasias (4.2%) and was associated with a lower level of RASSF2 expression and vascular invasion in squamous cervical cancer. EPB41L3 and RASSF1 hypermethylations were also more frequent in cancer than in precursor lesions. Patients with RASSF2 hypermethylation had shorter survival time, independent of tumor stage (hazard ratio: 6.0; 95% confidence interval: 1.5–24.5). Finally, the expressions of RASSF2 and EPB41L3 were restored in several cell lines treated with 5-aza-2′-deoxycytidine. Taken together, our results suggest that RASSF2 potentially functions as a new tumor-suppressor gene that is inactivated through hypermethylation in cervical cancer and is related to the bad prognosis of these patients.

FOXO3 expression during colorectal cancer progression: biomarker potential reflects a tumour suppressor role

Background:In previous studies, the Forkhead/winged-helix-box-class-O3 (FOXO3) transcription factor has displayed both tumour suppressive and metastasis-promoting properties.To clarify its role in human colorectal cancer (CRC) progression, we examined in vivo FOXO3 expression at key points of the metastatic cascade.Methods:Formalin-fixed paraffin-embedded resection specimens from normal colon, adenomas, primary CRC specimens of different pathological stage and CRC specimens with matched liver metastases were used to generate three separate custom-designed tissue microarray (TMA) representations of metastatic progression. Triplicate cores, immunostained for FOXO3 were scored semiquantitatively by two investigators.Results:The FOXO3 expression is significantly reduced in CRC specimens compared with normal tissue, and progressive FOXO3 downregulation is associated with advancing pathological stage. In addition, recurrent stage I/II primary tumours show a significantly lower FOXO3 expression compared with stage-matched non-recurrent tumours. When stratified according to high and low FOXO3 expression, mean disease-free survival in the low-expressing group was 28 months (95% CI 15.8–50.6) compared with 64 months (95% CI 52.9–75.4) in the high-expressing group.Conclusion:We have demonstrated an association between low FOXO3 expression and CRC progression in vivo using purpose-designed TMAs. Forkhead/winged-helix-box-class-O3 may represent a novel biomarker of nodal and distant disease spread with clinical utility in CRC.

Cancer‐associated upregulation of histone H3 lysine 9 trimethylation promotes cell motility in vitro and drives tumor formation in vivo

Global histone modification patterns correlate with tumor phenotypes and prognostic factors in multiple tumor types. Recent studies suggest that aberrant histone modifications play an important role in cancer. However, the effects of global epigenetic rearrangements on cell functions remain poorly understood. In this study, we show that the histone H3 lysine 9 (H3K9) methyltransferase SUV39H1 is clearly involved in regulating cell migration in vitro. Overexpression of wild-type SUV39H1, but not enzymatically inactive SUV39H1, activated migration in breast and colorectal cancer cells. Inversely, migration was reduced by knockdown of SUV39H1 or chemical inhibition by chaetocin. In addition, H3K9 trimethylation (H3K9me3) was specifically increased in invasive regions of colorectal cancer tissues. Moreover, the presence of H3K9me3 positively correlated with lymph node metastasis in colorectal cancer patients. Furthermore, overexpression of SUV39H1 drove tumorigenesis in mouse, resulting in a considerable decrease in survival rate. These data indicate that H3K9 trimethylation plays an important role in human colorectal cancer progression, possibly by promoting collective cell invasion.

Abstract LB-326: Zeb1 regulates inflammation-induced invasion and self-renewal of colon cancer cells.

Abstract Up-regulation of inflammatory cytokines such as IL-1β in tumor microenvironment has been associated with progression of human colorectal cancer, although the exact molecular mechanisms are still unclear. To understand how IL-1β influences colon cancer cell behaviors, we used a human colon cancer cell line HCT-116 as well as primary human colon cancer cells derived from a patient, which more closely reflect the properties of the primary tumor than established cell lines. Here, we demonstrate that IL-1β promotes epithelial-mesenchymal transition (EMT) and cancer stem cell (CSC) development in colon cancer. IL-1β treatment resulted in loss of E-cadherin and gain of a mesenchymal phenotype in both HCT-116 cells and primary colon cancer cells. Among E-cadherin suppressors, Zeb1 was up-regulated in IL-1β-induced EMT cells. Consistent with EMT properties, IL-1β-treated HCT-116 cells exhibited higher cell migratory ability. Moreover, IL-1β induced expression of stem-cell marker Bmi-1 in both HCT-116 cells and primary colon cancer cells. IL-1β increased these cells’ self-renewal ability to form spheres in serum-free conditions and chemo-resistance; properties associated with colon CSCs. Taken together, these results indicate that IL-1β can induce EMT and promote generation of CSCs in colon cancer. More importantly, we found that ZEB1, but not other E-cadherin transcriptional repressors (such as Zeb2, Snail and Twist), was consistently up-regulated in IL-1β-induced EMT cells and sphere cells. This suggests that Zeb1 links IL-1β-induced CSC self-renewal and EMT in colon cancer cells. Moreover, knockdown of Zeb1 in HCT-116 cells using shRNAs reversed the IL-1β effects on these two processes, further indicating that Zeb1 is essential for IL-1β-induced stem cell and EMT phenotypes in colon cancer cells. Our finding indicates that IL-1β promotes colon tumor growth and invasion through activation of CSC self-renewal and EMT, while Zeb1 plays a critical role in activation of these two processes. Thus, targeting tumor microenvironment cytokines or Zeb1 might form the basis of a promising treatment for colon cancer. Citation Format: Lei Wang, Yijing Li, Amy Beckley, Lorretta Pappan, Jishu Shi. Zeb1 regulates inflammation-induced invasion and self-renewal of colon cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr LB-326. doi:10.1158/1538-7445.AM2013-LB-326 Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.

P42.3: A promising biomarker for the progression and prognosis of human colorectal cancer

As a novel cell cycle-related gene, p42.3 has been shown to play a key role in the cell proliferation and tumorigenicity of gastric cancer. To date, the association between p42.3 and colorectal cancer (CRC) has not been reported. This study investigated the expression of p42.3 and its potential role in human colorectal cancers. Real-time polymerase chain reaction and western blotting were used to evaluate p42.3 mRNA and protein expression in 14 pairs of fresh frozen CRC samples, matched with adjacent normal mucosa. The p42.3 protein was evaluated by immunohistochemistry using CRC tissue microarrays, which included 212 CRC specimens and corresponding normal colorectal mucosa. The expression profiles of p42.3 in CRC tissues were analyzed against clinicopathological factors and post-surgical survival status. The expression profiles of p42.3 were also investigated in six human colon carcinoma cell lines. p42.3 was demonstrated to be over-expressed in colorectal cancer tissues compared with normal mucosa in the 14 tissue pairs (P = 0.011) and was significantly higher in patients with poor tumor differentiation (P = 0.045); patients positive for p42.3 expression had a poorer prognosis than those not expressing this protein (P = 0.033). In a multivariate survival analysis, p42.3 expression was identified as an independent prognostic factor for CRC patients (P = 0.030). The results indicated that p42.3 might play an important role in the progression of CRC, and it has a great value for assessing CRC patient prognosis after surgery.

Expression and potential role of apolipoprotein D on the death–survival balance of human colorectal cancer cells under oxidative stress conditions

Inverse correlations of apolipoprotein D (ApoD) expression with tumor growth have been shown, therefore proposing ApoD as a good prognostic marker for diverse cancer types, including colorectal cancer (CRC). Besides, ApoD expression is boosted upon oxidative stress (OS) in many pathological situations. This study aims at understanding the role of ApoD in the progression of human CRC. Samples of CRC and distant normal tissue (n = 51) were assayed for levels of lipid peroxidation, expression profile of OS-dependent genes, and protein expression. Three single-nucleotide polymorphisms in the ApoD gene were analyzed (n = 139), with no significant associations found. Finally, we assayed the effect of ApoD in proliferation and apoptosis in the CRC HT-29 cell line. In CRC, lipid peroxides increase while ApoD messenger RNA and protein decrease through tumor progression, with a prominent decrease in stage I. In normal mucosa, ApoD protein is present in lamina propia and enteroendocrine cells. In CRC, ApoD expression is heterogeneous, with low expression in stromal cells commonly associated with high expression in the dysplastic epithelium. ApoD promoter is basally methylated in HT-29 cells but retains the ability to respond to OS. Exogenous addition of ApoD to HT-29 cells does not modify proliferation or apoptosis levels in control conditions, but it promotes apoptosis upon paraquat-induced OS. Our results show ApoD as a gene responding to OS in the tumor microenvironment. Besides using ApoD as marker of initial stages of tumor progression, it can become a therapeutic tool promoting death of proliferating tumor cells suffering OS.

The changes of Th17 cells and the related cytokines in the progression of human colorectal cancers

BackgroundThe role of Th17 cells in colorectal tumorigenesis and development still remains unclear, despite the fact that it has been established in the pathogenesis of autoimmune diseases.MethodsWe first analyzed Th17 cells and Treg cells using flow cytometry in the circulation of colorectal adenoma (CRA) and colorectal carcinoma (CRC) patients and healthy controls, and the frequency of Th17 cells in peripheral blood mononuclear cells (PBMCs) stimulated by anti-CD3 plus anti-CD28 and treated by IL-1β, IL-6, and TGF-β in different concentrations. We then detected cytokines IL-1β, IL-6, IL-17A, IL-21, IL-23 or TGF-β by ELISA in sera and supernatants from both normal and tumor tissues cultured ex vivo.ResultsIt was found that the percentage of Th17 and Treg cells increased in the circulation of both CRA and CRC patients; the increase of Th17 cells in the circulation occurred in early stages, whereas the increase of Treg cells in the circulation and the increase of Th17 cells in tumor tissues occurred in advanced stages. The subsequent cytokine profiling showed that, along CRC progression, IL-1β, IL-17A and IL-23 underwent a similar change, while IL-6 in CRC exhibited an opposite change, with Th17 cells. In addition, high levels of TGF-β and IL-17A were detected in tumor tissues rather than in normal mucosa. The in vitro experiment further demonstrated that IL-1β, IL-6 or TGF-β modulated Th17 cell expansion in PBMC.ConclusionsOur study reveals a unique change of Th17 cells, which is regulated possibly by IL-1β, IL-6 and TGF-β in the progression of CRC.

Abstract LB-199: ZEB factors link IL-1β-induced EMT and stem cell phenotypes in colon cancer

Abstract Up-regulation of inflammatory cytokines such as IL-1β in tumor microenvironment has been associated with progression of human colorectal cancer. However, the exact molecular mechanisms by which inflammatory cytokines promote colon tumor growth are still unclear. Cancer stem cells (CSCs), a distinct subpopulation with self-renewal ability, drug resistance and high oncogenic potential, are thought to be responsible for tumor initiation and progression. Epithelia-mesenchymal transition (EMT), characterized by the repression of E-cardherin expression and increased cell mobility, is associated with enhanced metastasis. The objective of this study is to determine whether IL-1β may promote cancer progression by inducing EMT and colon CSC development. Here, we showed that IL-1β promoted E-cadherin repression in both human colon cancer cell line HCT-116 cells and primary colon cancer cells. Expression of E-cadherin transcription repressors ZEB1 and ZEB2 was up-regulated in IL-1β-treated HCT-116 and primary colon cancer cells, respectively. In addition, IL-1β-treated HCT-116 cells exhibited higher cell migratory ability. Moreover, IL-1β induced the expression of stem-cell marker Bmi-1 in both HCT-116 cells and primary colon cancer cells. IL-1β increased these cells’ ability of sphere formation in serum-free conditions and chemo-resistance; properties associated with colon CSCs. Taken together, these results indicate that IL-1β can induce EMT and promote generation of colon CSCs. More importantly, we found that ZEB1 and ZEB2, but not other E-cadherin transcriptional repressors (such as Snail and Twist), were consistently up-regulated in HCT-116 cells and primary colon cancer cells, respectively, during both processes of IL-1β-induced EMT and sphere formation. This suggests that ZEB factors may mediate IL-1β-induced EMT and generation of colon CSCs. Currently, this hypothesis is under evaluation using shRNAs to knock down ZEB1 in HCT-116 cells. Understanding the role of ZEB factors in IL-1β-mediated EMT and CSC development may lead to the identification of novel molecular targets for colon cancer therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-199. doi:1538-7445.AM2012-LB-199

봉독의 HIF-1α 발현감소를 통한 혈관신생 억제효과

봉독은 동양의학에서 관절염, 류마티즘 및 각종 암을 포함하여 다양한 질병을 치료하기 위하여 이용되었다. 최근 봉독의 신생혈관 억제효과에 대한 연구가 보고되었으나 정확한 분자메커니즘에 대해서는 보고가 미흡하다. 따라서, 본 연구는 봉독이 인간결장암세포인 HCT116세포에서 신생혈관생성과 종양진행에 중요한 역할을 하는 HIF-<TEX>$1{\alpha}$</TEX>와 VEGF 발현 억제효과를 조사하였다. 그 결과 봉독은 <TEX>$CoCl_2$</TEX>로 유도한 저산소 상태에서 VEGF와, HIF-<TEX>$1{\alpha}$</TEX>의 발현을 감소시키며 HIF-<TEX>$1{\alpha}$</TEX>의 promoter 영역인 HRE 활성을 억제하였다. 이러한 봉독의 HIF-<TEX>$1{\alpha}$</TEX> 발현억제효과는 ERK1/2의 인산화 조절을 통한 것이며, 봉독은 p38, JNK, AKT의 인산화에는 영향을 끼치지 않았다. 또한 봉독의 효과를 나타내는 단일물질 탐색을 위해 봉독의 생리활성 물질로 알려진 아파민과 멜리틴을 조사한 결과, HIF-<TEX>$1{\alpha}$</TEX>와 VEGF 억제효과는 아파민에 기인하는 것이라고 예상 할 수 있었다. 이와 같은 결과를 통하여 본 연구에서는 봉독의 혈관신생 억제에 대한 새로운 신호전달기전 및 인간 결장암세포 전이 억제제로서의 잠재성을 확인하였다. Bee venom (BV) has been used in medicine to treat a variety of diseases including arthritis, rheumatism, and various cancers. Recent reports indicate that BV has anti-angiogenic effects, but the precise molecular mechanism underlying the effects of BV against colorectal cancer remains to be elucidated. We examined the effects of BV and its major components (melittin and apamin) on tumor angiogenesis and found that BV significantly decreased protein levels of hypoxia-inducible factor-<TEX>$1{\alpha}$</TEX> (HIF-<TEX>$1{\alpha}$</TEX>), an important factor involved in angiogenesis and tumor progression, in human colorectal carcinoma HCT116 cells. BV also suppressed the transcription of HIF-<TEX>$1{\alpha}$</TEX> under hypoxia, leading to a decrease in the expression of vascular endothelial growth factor (VEGF), a major target gene of HIF-<TEX>$1{\alpha}$</TEX>. We also found that these effects were mainly elicited by apamin, but not melittin. BV specifically inhibited the phosphorylation of ERK1/2 without changing the total levels of this protein, but had no effect on kinases of p38/JNK and AKT. Our results suggest that BV may inhibit human colorectal cancer progression and angiogenesis by inhibiting HIF-<TEX>$1{\alpha}$</TEX> and VEGF expression, thereby providing a novel potential mechanism for the anticancer action of BV.

Tumor budding as a potential histopathological biomarker in colorectal cancer: Hype or hope?

Colorectal cancer (CRC), the third most commonly diagnosed type of cancer in men and women worldwide is recognized as a complex multi-pathway disease, an observation sustained by the fact that histologically identical tumors may have different outcome, including various response to therapy. Therefore, particularly in early and intermediate stage (stages II and III, respectively) CRC, there is a compelling need for biomarkers helpful of selecting patients with aggressive disease that might benefit from adjuvant and targeted therapy. Histopathological examination shows that likely other solid tumors the development and progression of human CRC is not only determined by genetically abnormal cells, but also by intricate interactions between malignant cells and the surrounding microenvironment. This has led to reconsider the features of tumor microenvironment as potential predictive and prognostic biomarkers. Among the histopathological biomarkers, tumor budding (i.e., the presence of individual cells and small clusters of tumor cells at the tumor invasive front) has received much recent attention, particularly in the setting of CRC. Although its acceptance as a reportable factor has been held back by a lack of uniformity with respect to qualitative and quantitative aspects, tumor budding is now considered as an independent adverse prognostic factor in CRC that may allow for stratification of patients into risk categories more meaningful than those defined by tumor-node-metastasis staging alone, and also potentially guide treatment decisions, especially in T2-T3 N0 (stage II) CRCs.

Proposed roles of the immune response regulator- ThPOK in human colorectal cancer progression

Solid tumours are commonly infiltrated by several immune cells [1-3]. In cancer, immune cells play conflicting roles with both the potentials to eliminate or to promote malignancy. In contrast to infiltration of cells responsible for chronic inflammation, the presence of high numbers of lymphocytes, especially T cells, has been reported to be important as indicator of good prognosis in many types of cancer [4-7]. The thorough knowledge of both manners and pathways with which tumors are able to evade immune-mediated attack, once established, is therefore of crucial importance. The strategies to escape anti-tumor immune responses include the limited priming or differentiation of antitumor T cells and the role of tumor microenvironment in order to prevent infiltration or activation of effector phase functions. We proposed to evaluate the role of Th inducing POZ-Kruppel Factor (ThPOK), a transcriptional regulator of T cell fate, in tumour-induced immune system plasticity during colorectal carcinogenesis. Data were collected on the amounts of CD4+, CD8+ and CD56+ as well as on ThPOK+ cells infiltrated in normal colorectal mucosa (NM), in dysplastic aberrant crypt foci (microadenomas, MA, the earliest detectable lesions in colorectal carcinogenesis) and in colorectal carcinomas (CRC); moreover, the colocalization of ThPOK with the above-mentioned markers of immune cells was evaluated using confocal microscopy. Interestingly, ThPOK showed a prominent increase since MA. A strong colocalization of ThPOK with CD4 both in NM and in MA was observed, weaker in carcinomas. Surprisingly, there was a peak in the colocalization levels of ThPOK with CD8 in MA, which was evident, although to a lesser extent, also in carcinomas. In conclusion, according to the data of the present study, ThPOK may be considered a central regulator of the earliest events in the immune system during colorectal cancer development. The novelty of the present study is the proposed role of ThPOK in influencing the immune response against cancer cells.

Suppression of tumour-specific CD4+ T cells by regulatory T cells is associated with progression of human colorectal cancer

BackgroundThere is indirect evidence that T cell responses can control the metastatic spread of colorectal cancer (CRC). However, an enrichment of CD4+Foxp3+ regulatory T cells (Tregs) has also been documented.ObjectiveTo...

Colorectal cancer mouse models: Integrating inflammation and the stroma

Sequences of molecular events that initiate and advance the progression of human colorectal cancer (CRC) are becoming clearer. Accepting that these events, once they are in place, accumulate over time, rapid disease progression might be expected. Yet CRC usually develops slowly over decades. Emerging insights suggest that the tumor cell microenvironment encompassing fibroblasts and endothelial and immune cells dictate when, whether, and how malignancies progress. Signaling pathways that affect the microenvironment and the inflammatory response seem to play a central role in CRC. Indeed, some of these pathways directly regulate the stem/progenitor cell niche at the base of the crypt; it now appears that the survival and growth of neoplastic cells often relies upon their subverted engagement of these pathways. Spurned on by the use of gene manipulation technologies in the mouse, dissecting and recapitulating these complex molecular interactions between the tumor and its microenvironment in the gastrointestinal (GI) tract is a reality. In parallel, our ability to isolate and grow GI stem cells in vitro enables us, for the first time, to complement reductionist in vitro findings with complex in vivo observations. Surprisingly, data suggest that the large number of signaling pathways underpinning the reciprocal interaction between the neoplastic epithelium and its microenvironment converge on a small number of common transcription factors. Here, we review the separate and interactive roles of NFκB, Stat3, and Myb, transcription factors commonly overexpressed or excessively activated in CRC. They confer molecular links between inflammation, stroma, the stem cell niche, and neoplastic cell growth.

Cancer Chemopreventive Effects of Lycopene: Suppression of MMP-7 Expression and Cell Invasion in Human Colon Cancer Cells

Clinical studies indicate that high blood levels of leptin or matrix metalloproteinase-7 (MMP-7; matrilysin) proteins are associated with tumor progression of human colorectal cancer (CRC). Leptin could play an important role in cell migration and invasion of cancer cells. Our previous study indicated that lycopene could inhibit the proliferation of human colon cancer cells in vitro. However, the inhibitory effects of lycopene on the progression of human colon cancer cells have not been demonstrated yet. In this study, we investigated the inhibitory effects of lycopene on tumor progression including cell invasion and MMP-7 expression in leptin-stimulated human colon cancer cells in vitro. Our results demonstrated that lycopene significantly inhibited leptin-mediated cell invasion and MMP-7 expression in human colon cancer HT-29 cells. Lycopene could augment the expression and stability of E-cadherin proteins. Our results showed that MAPK/ERK and PI3K/Akt signaling pathways played important roles in leptin-mediated MMP-7 expression and cell invasion. Lycopene could effectively inhibit the phosphorylation of Akt, glycogen synthase kinase-3β (GSK-3β) and ERK 1/2 proteins. The molecular mechanisms of lycopene were in part through decreases in nuclear levels of AP-1 and β-catenin proteins. These novel findings suggested that lycopene could act as a chemopreventive agent to suppress MMP-7 expression and leptin-mediated cell invasion in human colon cancer HT-29 cells.

Prognostic Significance of TWEAK Expression in Colorectal Cancer and Effect of Its Inhibition on Invasion

Tumor necrosis factor-like weak inducer of apoptosis (TWEAK) has been implicated in tumor development and progression. The aim of this study was to investigate the role of TWEAK in colorectal cancer (CRC) progression. To investigate the involvement of TWEAK in the progression of human CRC, normal, and tumor specimens from 174 patients were analyzed immunohistochemically for the expression of TWEAK. TWEAK recombinant protein treatment, transfection of expression plasmids, and small interfering RNA to knockdown TWEAK expression were performed to test invasive ability with a Boyden chamber. The mRNA expression profile in recombinant TWEAK treatment was compared to a control group by microarray analysis. To identify downstream effectors, Raf kinase inhibitor (RKIP) and its correlation with TWEAK in vitro and in vivo were examined by quantitative real-time polymerase chain reaction and invasion assays. CRC patients whose tumors displayed high TWEAK expression had a statistically significantly higher overall survival and a disease-free advantage over those with a low TWEAK expression. In in vitro invasion assays, alterations in TWEAK expression in CRC cell lines inversely modulated their invasive ability. By means of integrated genomics, we identified RKIP as a downstream effector in TWEAK-mediated invasion inhibition. Knockout of RKIP expression in HCT116 cells by short hairpin RNA (shRKIP) resulted in increased invasiveness. Clinically, RKIP and TWEAK mRNA expression showed strong positive correlations in CRC patient samples. Our results implicate TWEAK as a key regulator of CRC invasion, and it appears to be a useful prognostic factor for patients with CRC.

HOXB7 as a Prognostic Factor and Mediator of Colorectal Cancer Progression

This study was to investigate the clinicopathologic significance and potential role of HOXB7 in the development and progression of colorectal cancer (CRC). The relationship between HOXB7 expression and clinical characteristics of CRC was analyzed in 224 paraffin-embedded archived CRC specimens by immunohistochemistry (IHC). The effects of HOXB7 on cell growth and proliferation, as well as on tumorigenesis, were examined both in vitro and in vivo, using MTT assay, colony formation assay, cell cycle analysis, soft agar assay, and tumorigenesis in nude mice. Western blotting and real-time reverse transcriptase-PCR were performed to examine the impact of HOXB7 on the PI3K/Akt and MAPK signaling pathways. HOXB7 protein level was significantly correlated with advanced Dukes stage (P < 0.001), T stage (P = 0.012), distant metastasis (P = 0.042), higher proliferation index (P = 0.007) and poor survival of patients (P = 0.005). Enforced expression of HOXB7 in CRC cell lines significantly enhanced cell growth, proliferation and tumorigenesis. Conversely, knockdown of HOXB7 caused an inhibition of cell growth, proliferation, and tumorigenesis. We also showed that HOXB7 accelerated G(0)-G(1) to S-phase transition concomitantly with upregulation of cyclin D1 and downregulation of p27Kip1. On the contrary, knockdown of HOXB7 caused G(1)-S-phase arrest, downregulation of cyclin D1 and upregulation of p27Kip1. Enforced expression of HOXB7 could enhance PI3K/AKT and MAPK pathway activity. Our findings suggest that HOXB7 protein, as a valuable marker of CRC prognosis, plays an important role in the development and progression of human CRC.

The role of galectins in colorectal cancer progression

Galectins constitute a family of 15 mammalian galactoside-binding proteins that share a consensus amino acid sequence in their carbohydrate binding sites. They are multi-functional molecules and are expressed widely in human tissues. Four galectins: galectin -1, -3, -4 and -8 are expressed in the human colon and rectum and their expressions show significant changes during colorectal cancer development and metastasis. These changes in galectin expression correlate with alterations in cancer cell growth, apoptosis, cell-cell and cell-matrix interactions and angiogenesis. This review summaries current knowledge of the expression and roles of these galectins in the progression of human colorectal cancer and discusses the relevance of galectins and their ligands as potential therapeutic targets for cancer treatment.

Abstract 1679: Increased expression of hCAS/CSE1L in colorectal cancer: Correlation with tumor progression

Abstract Introduction. The human cellular apoptosis susceptibility (hCAS) gene product has been shown to be involved in cellular proliferation, apoptosis, invasion and metastasis. CSE1 is a yeast gene with high homology to hCAS (CSE1-like or CSE1L) which has been shown to be involved in chromosome segregation during mitosis. hCAS has also been shown to be a key player in cytoplasm-to-nuclear transport by shuttling importin-alpha from the nucleus back out into the cytoplasm. We hypothesized that the expression of hCAS may be altered during the progression of human colorectal cancer (CRC). Methods. In order to examine hCAS at the gene expression level, we surveyed three different datasets from studies in which Affymetrix U133+gene chips had been performed on patients with CRC, as well as patients with adenomas and normal individuals. We examined relative gene expression levels for 3 probe sets on the U133+ gene chip that closely align to the refseq for the hCAS gene. hCAS protein expression level was semiquantitatively measured using immunohistochemistry (IHC) and the tissue microarray (TMA) technique. A CRC TMA had been previously created at the Moffitt Cancer Center using resection specimens of primary CRC of different stages, and inclusive of adenomas and normal tissues samples from the same patients. Results. hCAS gene expression was increased in all adenoma and CRC samples tested by microarray, compared to histologically normal colonic mucosa. However, the difference in hCAS mRNA levels between adenoma and CRC samples was not statistically different. No correlation was found between hCAS and cancer stage, grade, vascular invasion, sex and/or age. Interestingly, in the invasive CRCs, intense hCAS proteins expression was localized to both nucleus and cytoplasm. Conversely, early stage lesions showed predominantly cytoplasmic hCAS staining. Conclusions. Our results demonstrate that hCAS protein is expressed early and across all stages of CRC development. Because of its early expression hCAS may represent a marker for early detection of CRC. In addition, because of its involvement in a variety of important cellular processes, this gene product might represent a new potential target for prevention and treatment of CRC. 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 1679. doi:10.1158/1538-7445.AM2011-1679

Expression of KITENIN in human colorectal cancer and its relation to tumor behavior and progression

KAI1 COOH-terminal interacting tetraspanin (KITENIN) contributes to tumor invasion and metastasis in various cancers. The aim of current study was to evaluate whether KITENIN affects tumor cell invasion and prognosis in human colorectal cancers. We investigated the biologic role of KITENIN on tumor cell invasion by using small interfering RNA in Caco2, DLD1, and SW480. We evaluated the expression of KITENIN and activator protein-1 (AP-1) target genes in human colorectal cancer tissues. The tumor cell invasion was decreased by knockdown of KITENIN in three tested cell lines. The mRNA expression of cyclin D1 and COX-2 was decreased in KITENIN knockdown Caco2 and the mRNA expression of MMP-3 and COX-2 was decreased in KITENIN knockdown DLD1 and SW480. The extracellular-signal protein kinase 1/2 (ERK1/2) phosphorylation was decreased in KITENIN knockdown in three tested cell lines. Expression of KITENIN and AP-1 target genes was significantly increased in human colorectal cancer tissues. The ERK1/2, c-Jun N-terminal kinase (JNK) and p38 phosphorylations were increased in human colorectal cancer tissues. Expression of KITENIN was significantly associated with lymphovascular invasion, depth of invasion, lymph node metastasis, tumor stage and poor survival. These results indicate that KITENIN is associated with human colorectal cancer progression including invasion and metastasis.