Therapy For Colorectal Carcinoma Research Articles

Research development of the relationship between thymidine phosphorylase expression and colorectal carcinoma.

Thymidine phosphorylase (TP) is a key enzyme that contributes to the composition and decomposition of pyrimidine nucleotides. TP seems homologous to platelet-derived endothelial cell growth factor, and its effects on inducing vascularization and anti-apoptosis are closely related to growth and metastasis of colorectal carcinoma. In addition, TP is a key enzyme that catalyzes the transformation from 5-fluorouracil (FU) prodrugs of 5′-deoxy-5-fluorouridine (5′-DFUR) to 5-FU. The activity of TP is closely related to the sensitivity of colorectal carcinoma cells to fluorouracil drugs and targeted therapy. Given the important functions of TP in growth, metastasis, tumor treatment, and prognosis, determining its expression mechanism is significant. This article summarizes the research development of TP expression in colorectal carcinoma, tumor neovascularization, cytotoxicity activation of 5′-DFUR, and colorectal carcinoma therapy.

Acetate-induced apoptosis in colorectal carcinoma cells involves lysosomal membrane permeabilization and cathepsin D release

Colorectal carcinoma (CRC) is one of the most common causes of cancer-related mortality. Short-chain fatty acids secreted by dietary propionibacteria from the intestine, such as acetate, induce apoptosis in CRC cells and may therefore be relevant in CRC prevention and therapy. We previously reported that acetic acid-induced apoptosis in Saccharomyces cerevisiae cells involves partial vacuole permeabilization and release of Pep4p, the yeast cathepsin D (CatD), which has a protective role in this process. In cancer cells, lysosomes have emerged as key players in apoptosis through selective lysosomal membrane permeabilization (LMP) and release of cathepsins. However, the role of CatD in CRC survival is controversial and has not been assessed in response to acetate. We aimed to ascertain whether LMP and CatD are involved in acetate-induced apoptosis in CRC cells. We showed that acetate per se inhibits proliferation and induces apoptosis. More importantly, we uncovered that acetate triggers LMP and CatD release to the cytosol. Pepstatin A (a CatD inhibitor) but not E64d (a cathepsin B and L inhibitor) increased acetate-induced apoptosis of CRC cells, suggesting that CatD has a protective role in this process. Our data indicate that acetate induces LMP and subsequent release of CatD in CRC cells undergoing apoptosis, and suggest exploiting novel strategies using acetate as a prevention/therapeutic agent in CRC, through simultaneous treatment with CatD inhibitors.

Expression of the cannabinoid type I receptor and prognosis following surgery in colorectal cancer

The cannabinoid system has been considered to be a potential target of colorectal carcinoma therapy. The aim of this study was to address the correlation between cannabinoid type 1 (CB1) receptor expression and disease severity/outcomes in patients with colorectal cancer (CRC). CB1 receptor expression was analyzed by immunohistochemistry using tissue microarrays in consecutive patients who underwent surgical resection (n=534). CB1 receptor expression was categorized as a high (≥66%) vs. low (<66%) immunopercentage as a median split, and was analyzed in relation to disease severity and overall survival. CB1 receptor expression was observed in 409 patients (76.6%). Low CB1 receptor expression was more frequently identified in stage IV than in stage I/II or III cancer (P<0.01 for both). In stage IV CRC, high vs. low CB1 expression was correlated with a statistically significant poorer overall survival (P=0.033) that was independent of age, R0 resection, tumor differentiation and chemotherapy [hazard ratio (HR), 1.805; 95% confidence interval (CI), 1.042–3.094; P=0.035]. However, CB1 expression was not observed to be correlated with patient survival following surgery in stage I/II or III cancer. The high immunoreactivity of the cannabinoid type 1 receptor is a significant prognostic factor following surgery in stage IV CRC.

TAGLN suppresses proliferation and invasion, and induces apoptosis of colorectal carcinoma cells

In order to find the correlation between transgelin gene (TAGLN) and colorectal carcinoma occurrence, we investigated the expression of TAGLN in colorectal carcinoma tissue samples and colorectal carcinoma LoVo cells. Meanwhile, the effects of TAGLN on the characteristics of LoVo cells were also examined. The expressions of TAGLN in colorectal carcinoma tissues, adjacent normal tissues, and LoVo cells were detected by the Western blot method. The recombinant plasmid pcDNA3.1-TAGLN was established and transfected into LoVo cells with the help of Lipofectamine™ 2000. At the same time, the TAGLN siRNA was transfected into LoVo cells in another group. Forty-eight hours later, the expressions of TAGLN in all groups were assayed by Western blot, and the cell viability was analyzed by MTT (3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) assay. The cell cycle and cell apoptosis were examined by flow cytometry, and the cell invasive ability was analyzed by Transwell invasion experiment. The effect of TALGN on the expression of matrix metalloproteinase 9 (MMP9) was detected by Western blot. Western blot analysis showed that the expressions of TALGN in colorectal carcinoma tissues and LoVo cells were significantly decreased compared with colorectal carcinoma adjacent normal tissues (p < 0.01). In the overexpression or RNAi experiments, the plasmid pcDNA3.1-TAGLN significantly enhanced TALGN expression (p < 0.01), and TAGLN siRNA significantly decreased TAGLN expression (p < 0.01) in LoVo cells 48h after transfection. In addition, MTT assay indicated that the cell viability of LoVo cells in the pcDNA3.1-TAGLN transfection group was significantly lower than that in the untransfected control group (p < 0.05). Furthermore, the overexpression of TAGLN significantly lowered the cell proliferation index (p < 0.05) and improved cell apoptosis (p < 0.01) in LoVo cells. In Transwell invasive experiments, the cell number, which had migrated through the chamber membrane, significantly decreased in the pcDNA3.1-TAGLN transfection group (p < 0.05) and significantly increased in the TAGLN knockdown group (p < 0.05) compared to the untransfected control group. At the same time, the expression of MMP9 was notably inhibited in the pcDNA3.1-TAGLN transfection group (p < 0.01). The expressions of TAGLN were inhibited in colorectal carcinoma tissues and colorectal carcinoma LoVo cells. The study also demonstrated that TAGLN could attenuate the proliferation and invasive ability of LoVo cells and enhance LoVo cell apoptosis. Furthermore, the expression of MMP9 was also inhibited by TAGLN. All these results could bring us a new perspective for biological therapy in colorectal carcinoma.

Fibroblast Growth Factor Receptor 2 IIIc as a Therapeutic Target for Colorectal Cancer Cells

A high percentage of colorectal carcinomas overexpress a lot of growth factors and their receptors, including fibroblast growth factor (FGF) and FGF receptor (FGFR). We previously reported that FGFR2 overexpression was associated with distant metastasis and that FGFR2 inhibition suppressed cell growth, migration, and invasion. The FGFR2 splicing isoform FGFR2IIIb is associated with well-differentiated histologic type, tumor angiogenesis, and adhesion to extracellular matrices. Another isoform, FGFR2IIIc, correlates with the aggressiveness of various types of cancer. In the present study, we examined the expression and roles of FGFR2IIIc in colorectal carcinoma to determine the effectiveness of FGFR2IIIc-targeting therapy. In normal colorectal tissues, FGFR2IIIc expression was weakly detected in superficial colorectal epithelial cells and was not detected in proliferative zone cells. FGFR2IIIc-positive cells were detected by immunohistochemistry in the following lesions, listed in the order of increasing percentage: hyperplastic polyps < low-grade adenomas < high-grade adenomas < carcinomas. FGFR2IIIc immunoreactivity was expressed in 27% of colorectal carcinoma cases, and this expression correlated with distant metastasis and poor prognosis. FGFR2IIIc-transfected colorectal carcinoma cells showed increased cell growth, soft agar colony formation, migration, and invasion, as well as decreased adhesion to extracellular matrices. Furthermore, FGFR2IIIc-transfected colorectal carcinoma cells formed larger tumors in subcutaneous tissues and the cecum of nude mice. Fully human anti-FGFR2IIIc monoclonal antibody inhibited the growth and migration of colorectal carcinoma cells through alterations in cell migration, cell death, and development-related genes. In conclusion, FGFR2IIIc plays an important role in colorectal carcinogenesis and tumor progression. Monoclonal antibody against FGFR2IIIc has promising potential in colorectal carcinoma therapy.

Abstract 1319: Establishment and characterization of fresh primary human colorectal tumor implants

Abstract A personalized approach towards the therapy of colorectal carcinoma is critical for a successful outcome. In recent years, direct transfer xenograft models (or the “tumor graft”) have proven to be highly predictive and are being adapted in clinical practice. However, direct drug evaluation in tumor graft models for each individual patient is not feasible due to time and cost constraints. Predictive biomarker discovery is an extremely promising application of the tumor graft model, since the discovered biomarker signatures will direct therapy choice in a much broader patient population. In collaboration with the Catholic Health Initiatives Center for Translational Research we at Caliper Life Sciences have established a collection of tumor graft samples of primary human colorectal carcinomas. All samples were collected fresh from consenting patients following an IRB-approved protocol and implanted in vivo on the day of surgical tumor resection. Tumors were grafted either orthotopically or subcutaneously in female NIH-III mice. We present here the results of a comparative study of gene and protein expression profiles for the primary tumor (clinical samples) and the direct transfer xenografts. These data will be correlated with drug sensitivity profiles generated by using the same samples. The resulting data sets will be used to characterize predictive biomarker signatures that will be a valuable tool for selection of the most effective therapy tailored to the individual patient. 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 1319. doi:1538-7445.AM2012-1319

Gelatinases A and B Expression in Human Colorectal Cancer in Upper Egypt: A Clinicopathological Study

Aim: Prognosis of colorectal carcinoma depends on many factors, such as age and sex of patient; location; multiplicity; local extent and size of tumor, bowel obstruction, or perforation; as well as tumor microscopic type and grade; vascular and perineural invasion; and nodal and distant metastasis. The matrix metalloproteinases (MMPs) are a family of proteolytic enzymes strongly implicated in tumor invasion and metastasis, hence in tumor prognosis. The purpose of this study was to assess the role of MMP-2 and MMP-9 expression in colorectal tumorigenesis, invasion, and metastasis, hence their prognostic values.Method: Immunohistochemical analysis of MMP-2 and MMP-9 in colorectal cancer cells, an immunohistochemical score based on the intensity of immunoreactivity and proportion of immunoreactive cells that established for each MMP, and correlation of this expression with the established prognostic factors.Results: MMP-2 was expressed in 81.8% (strong expression in 40%) of cases, and MMP-9 was expressed in 72% (strong expression in 35%) of cases.Conclusions: MMP-2 and MMP-9 are widely expressed in colorectal carcinoma, suggesting significant diagnostic and prognostic values in these tumors. Increased levels of MMP-2 and MMP-9 protein expression in colorectal carcinoma tissues as compared to normal tissues suggest their association with colorectal tumor invasion and metastasis and that they could be targets for intervention and therapy in colorectal carcinoma.

Combining bacterial-immunotherapy with therapeutic antibodies: A novel therapeutic concept

Immunotherapeutic strategies become more and more important for cancer treatment. Therapeutic monoclonal antibodies (mAbs) like Panitumumab binding and blocking the EGF-receptor are in routine clinical use for the treatment of colorectal carcinoma (CRC). Also, bacterial therapy proved beneficial for experimental treatment of different tumor entities. The latter has been attributed to an activation of the immune system. Here, we describe a combination of both immunotherapeutic approaches in order to develop a novel targeted therapy for CRC. The therapeutic mAbs Trastuzumab and Panitumumab were conjugated to heat-inactivated bacteria expressing protein A or protein G.The potential of the conjugates was tested in comparison to the single components both in vitro and in vivo using a panel of patient-derived CRC cell lines. Antitumoral effects observed in vitro were strictly dependent on the presence of bacteria. Generally, effects could be enhanced by the addition of human lymphocytes. Detailed analysis of effector cells in autologous and allogeneic long-term stimulated lymphocyte cultures revealed the predominance of NK-cell-like cytolytic effectors. Reactivity was observed both against CRC target cells but also against the NK cell target K562. Similarly, in a subsequent in vivo study we observed substantial tumor growth delay accompanied by an increase in circulating NK cells. Contrary to this, the monotherapy with mAb alone caused only marginal effects and the treatment with bacteria was comparable to the mock-treated control.These data demonstrate successful targeting of CRC by bacteria/mAb conjugates. This novel concept may be interesting for future clinical approaches. Additionally, it illustrates the effectiveness of NK cells for cancer immunotherapy.

Role of anti-EGFR target therapy in colorectal carcinoma

Role of anti-EGFR target therapy in colorectal carcinoma

Role of anti-EGFR target therapy in colorectal carcinoma

The epidermal growth factor receptor (EGFR) has become an important target in cancer treatment. In consequence, drugs directed at this and other molecular targets are an increasingly important part of the treatment of numerous tumours. Cetuximab and panitumumab, two monoclonal antibodies that target EGFR, have proved to be effective in metastatic colorectal cancer treatment. However, some patients do not respond to treatment with EGFR inhibitors and, for this reason, interest in the identification of patients most likely to benefit from treatment with these agents has grown considerably. K-Ras, a member of the RAS family of signalling proteins plays an important role in EGFR- mediated regulation of cellular proliferation and survival. Patients with wild-type K-Ras were found to have significantly greater overall survival, progression-free survival and/or response rate compared with patients harbouring K-Ras mutations.

The significance of portal vein embolization in the treatment of colorectal liver metastases

The first aim of the present paper was to evaluate hypertrophy of liver parenchyma after portal vein embolization in patients after systemic chemotherapy for colorectal carcinoma metastases and planned extensive liver resections. The second aim was to study whether hypertrophy of the liver parenchyma remnant after could influence the postoperative course large liver resections in long-term chemotherapy within complex therapy of colorectal carcinoma.The prospective study comprised of 43 patients with colorectal hepatic metastases in whom liver resections of 4-5 segments were planned (Table 1). All patients underwent complex therapy of colorectal carcinoma, including chemotherapy consisting of 6-12 therapeutic cycles. Time interval between chemotherapy and liver resection was 2-24 months (mean interval of 8 months). Twenty patients whose presumed liver parenchyma remnant was less than 40% of total liver volume were indicated for portal vein embolization (mean liver parenchyma remnant of 29%). This was always embolization of the right portal branch. Twenty-three patients were primarily indicated to liver resection. Hypertrophy of the left liver lobe occurred in all 20 patients. After portal vein embolization, the volume of left liver increased on average from 476 ml (282-754) to 584 ml (380-892) (P < 0.05). Mean hypertrophy of left liver lobe after portal vein embolization was 28.5%. The measured parenchyma remnant after tumor resection increased from 29% up to 38% by hypertrophy. Mean values of ALT and AST in the postoperative period were significantly different in the groups in this study. The values of alkaline phosphatase (ALP) and gamma glutamyl transpeptidase (GMT) were lower in patients after portal vein embolization (P < 0.05). Significant differences were in postoperative level of serum bilirubin, bilirubin levels in patients after portal vein embolization were 2-3 times lower than in the group of patients after immediate surgery (P < 0.05). he values of prothrombin time were also significantly lower in patients who underwent surgery without previous portal vein embolization (P < 0.05).

Alteration of CXCR7 Expression Mediated by TLR4 Promotes Tumor Cell Proliferation and Migration in Human Colorectal Carcinoma

The link between inflammation and colorectal carcinoma has been acknowledged. However, the impact of bacterial lipopolysaccharide (LPS) binding to Toll-like receptor 4 (TLR4) on chemokine receptors in human colorectal carcinoma cells still remains to be elucidated. The present study shows that exposure to LPS elevated CXC chemokine receptor 7 (CXCR7) expression in colorectal carcinoma SW480 and Colo 205 cell lines expressing TLR4/myeloid differential protein (MD-2). CXCR7 is associated with SW480 cell proliferation and migration. However, exposure of SW480 and Colo 205 cells to LPS had no effect on CXCR4 expression. To further support the above results, the expression of TLR4, MD-2, and CXCR7 was analyzed in human colorectal carcinoma tissues. Higher rates of TLR4 (53%), MD-2 (70%), and CXCR7 (29%) expression were found in colorectal carcinoma tissues than in normal tissues. We demonstrated that the recombination of TLR4, MD-2 and CXCR7 strongly correlated with tumor size, lymph node metastasis and distant metastasis in colorectal carcinoma tissue samples (p = 0.037, p = 0.002, p = 0.042, resp.). Accordingly, simultaneous examination of the expression of TLR4, MD-2 and CXCR7 in cancer tissues of colorectal carcinoma may provide valuable prognostic diagnosis of carcinoma growth and metastasis. Interplay of TLR4, MD-2 and CXCR7 may be of interest in the context of novel immunomodulatory therapies for colorectal carcinoma.

Expression and Clinical Significance of l-Plastin in Colorectal Carcinoma

Introductionl-plastin, an actin-binding protein, is upregulated in many tumours, including colorectal carcinoma. This study evaluated the expression of l-plastin in plasma and colorectal tumour tissue and analysed the correlation between clinicopathological staging and prognosis. Materials and MethodsEnzyme-linked immunosorbent assay was used to detect l-plastin in the plasma of 120 colorectal carcinoma patients and 40 control subjects. Immunohistochemistry analyses were also used. ResultsThe rate of positive l-plastin expression was significantly higher in colorectal carcinoma patients than in control subjects, and was significantly higher in tumour tissues than in the tissues surrounding the tumour. l-Plastin expression also is correlated with tumour grade and size, and lymph node metastasis. However, there was no correlation with the extent of tumour invasion or distant metastasis. Conclusionl-Plastin may be a useful marker for screening colorectal carcinoma and determining the prognosis of patients with colorectal carcinoma, and for genetic therapy and targeted therapy of colorectal carcinoma.

Suppression of lung cancer metastasis-related protein 1 (LCMR1) inhibits the growth of colorectal cancer cells

Lung cancer metastasis-related protein 1 (LCMR1) is a critical subunit of the mediator complex, and plays an important role in the elaborate regulation of gene transcription. However, the functional role of LCMR1 in colorectal carcinoma (CRC) has not been clarified. In this study, LCMR1 expression in CRC specimens was quantified by using the quantitative reverse transcription polymerase chain reaction method. The effect of downregulation of LCMR1 by lentivirus-mediated small hairpin RNA (shRNA) on CRC cell proliferation and tumorigenesis was explored. There was a higher expression of LCMR1 in CRC tissue in comparison with adjacent normal colon tissue (P<0.05). LCMR1 gene was effectively knocked down in human CRC RKO and DLD-1 cells that infected with lentivirus delivering shRNA against LCMR1, which resulted in inhibition of cell proliferation and augmentation of G0/G1 phase proportion. Moreover, the tumorigenicity of RKO cells was also dramatically inhibited after LCMR1 was knocked down. In conclusion, our results suggest that LCMR1 promotes CRC cell growth, and lentivirus-mediated silencing of LCMR1 may contribute to the gene therapy for CRC.

Structure and antitumor (LOVO) activity of Cortex moutan heteroglycan and Curcumin

The Box–Behnken design combined with response surface methodology was used to optimize the extraction of curcumin. The optimized results showed that the highest extraction yield of curcumin could arrive 1.12%. The suitability of the model equation for predicting the optimum response values was tested using the selected optimal conditions. The predicted extraction yield of heteroglycan was 1.13%, which was consistent with the practical extraction yield of heteroglycan of 1.17%. Cortex moutan heteroglycan (500mg) was applied to a column (3.2cm×32cm) of DEAE Sepharose CL-6B (Cl−) that was equilibrated with H2O. The two adsorbed fractions (Fraction I and Fraction II) were obtained as lyophilisate after dialysis The antitumor activities of the Cortex moutan heteroglycan and curcumin were evaluated in vitro. The results indicated that the Cortex moutan heteroglycan and curcumin could inhibit colorectal carcinoma (LOVO) cells growth and stimulate apoptosis. Moreover, antitumor activities of curcumin was stronger than that of Cortex moutan heteroglycan. This indicated that curcumin was useful for therapy of colorectal carcinoma.

Abstract 780: Immune escape in colorectal carcinoma: role of the IFN-γ pathway

Abstract The presence of a Th1 adaptive immune response in colorectal carcinoma (CRC) has been associated with improved clinical outcome and survival of patients. The Th1 adaptive immune response is mediated by IFN-γ, a known anti-proliferative, pro-apoptotic and pro-immunogenic cytokine. We have previously identified the guanylate-binding protein 1 (GBP-1), one of the proteins the most highly induced by IFN-γ, as a sensitive marker for the detection of this immune response in CRC in vivo. In a tissue microarray study with 388 CRC tumors, 30% showed GBP-1 expression. Interestingly, in these tumors the expression of GBP-1 was only induced in the cells of the desmoplastic stroma and not observed in adjacent tumor cells. This suggested that CRC tumor cells might undergo an immune escape from the Th1 adaptive immune response, characterized by the loss of IFN-γ responsiveness. Therefore, we investigated the responsiveness to IFN-γ in sixth different CRC cell lines and in normal colon epithelial cells. After treatment with IFN-γ, three of the sixth CRC cell lines (DLD-1, SW480 and HCT116) failed to express GBP-1 as well as other IFN-induced proteins such as caspase-1 and MxA. On the contrary, a robust GBP-1 induction was present in normal colon epithelial cells, in T84, WiDr and HT29 cells. We then investigated whether the loss of expression of IFN-γ target genes was associated with phenotypic changes in CRC cells. In fact, we found that DLD-1, HCT116 and SW480 were resistant to IFN-γ-induced apoptosis. Furthermore, the loss of IFN-γ responsiveness correlated in SW480 and HCT116 with the absence of expression of the IFN-γ receptor alpha chain (IFNγRα). Tumor xenotransplant experiments using DLD-1 cells with reconstituted GBP-1 expression resulted in significantly reduced tumor growth as compared to xentotransplants with control vector transfected DLD-1 cells. This indicated that GBP-1 expression may mediate the anti-tumorigenic effects of the Th1 response. Overall, these data suggested that loss of IFN-γ responsiveness may be a common event in CRC and may protect tumor cells against the anti-tumorigenic (e.g. pro-apoptotic) effects of IFN-γ. The presence of an immune escape from the Th1 adaptive immune response may have important repercussions for prognosis and patients recruitment for immune-based therapy in 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 780. doi:10.1158/1538-7445.AM2011-780

Abstract 4919: Multiplexed EGFR signaling pathway analysis in FFPE tissue using quantitative mass spectrometry

Abstract The epidermal growth factor receptor (EGFR) is a drug target for both small molecule and antibody therapeutics and has been approved in non small-cell lung carcinoma (NSCLC) and colorectal carcinoma (CRC) among other indications. These drugs block receptor signaling though blockade of the tyrosine kinase domain, or through inhibition of ligand binding. Current genomic tests measure receptor amplification, RNA levels, the mutation status of receptor or pathway molecules (EGFR or kRAS mutations) but no current assay can directly assess the activation state of the EGFR or its downstream signaling pathway components. Indeed, the EGFR mutation positive NSCLC tumors (thought to be constitutively active) show a high response rate to TKI therapy, but the many non responders (50% or more) demonstrate the limitation of genomic analysis. Since activation of EGFR is necessary for the response to these targeted agents, it is critical to measure what levels of receptor activation and downstream signaling determines tumor responsiveness to EGFR targeted therapies in these patients. For this reason, we have developed a panel of new diagnostic assays which measure the activation of the EGFR and key downstream signaling proteins through quantitation of the phosphorylation state of these proteins. These assays are based on the Liquid Tissue®-SRM technology platform. This approach enables relative and absolute quantification of proteins and their phosphorylation status directly in formalin fixed paraffin embedded (FFPE) tissue. We preclinically validated the multiplexed Liquid Tissue® phospho-SRM assay on formalin fixed EGF stimulated A431 cells. We followed up these in vitro studies with phospho-SRM analysis of FFPE NSCLC xenograft explants where extensive independent histopathologic and molecular characterization had been performed, allowing us to benchmark our phospho-SRM analysis with standard diagnostic analyses. We have now extended these quantitation studies by measuring the expression of EGFR and phospho-EGFR in FFPE tissues obtained from relevant human clinical trial cohorts – Gefitinib treated NSCLC and Cetuximab treated CRC. It is hoped that we will be able to correlate EGFR expression, activation and signaling in these tumors with responsiveness to EGFR targeted therapy, and to validate this assay for use as a companion diagnostic to guide therapy in both NSCLC and 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 4919. doi:10.1158/1538-7445.AM2011-4919

MicroRNA-221 controls CDKN1C/P57 expression in human colorectal carcinoma

To investigate the expression of microRNA-221 (miR-221) and CDKN1C/P57 in colorectal carcinoma (CRC) and adjacent non-cancerous tissues. The effect of miR-221-specific inhibitor on cell proliferation and apoptosis in CRC cells was also assessed. The expression of miR-221 was detected by real-time RT-PCR. CDKN1C/P57 mRNA and corresponding protein expression pattern were detected by semi-quantitative RT-PCR and Western-blot. The specific 2'-methoxy-modified RNA oligonucleotide of miR-221(miRNA inhibitor,anti-miR-221) was designed, synthesized and transfected into Caco2 cell by liposome. Finally, the status of CRC cell proliferation and apoptosis were detected by MTT assay and flow cytometry. The expression of miR-221 was significantly up-regulated in CRC tissues as compared to the adjacent non-cancerous tissues(2.041±1.401 vs. 0.806±0.341, P<0.01). There was no significant difference in CDKN1C/P57 mRNA expression between CRC and non-cancerous tissues, whereas CDKN1C/P57 protein markedly decreased in CRC (3.019±1.708 vs. 0.972±0.316, P<0.01). miR-221-specific inhibitor significantly enhanced CDKN1C/P57 protein expression, inhibited proliferation of CRC cells and induced apoptosis of CRC cells(P<0.01). miR-221 inhibits CDKN1C/P57 expression by post-transcriptional gene silencing to promote CRC development and progression. miR-221-specific inhibitor potentially inhibits the growth of CRC cells. Therefore, it may be a new target for the biologic therapy for CRC.

The feasibility of Cep55/c10orf3 derived peptide vaccine therapy for colorectal carcinoma

In our previous study, we demonstrated that a peptide derived from the novel centrosome residing protein Cep55/c10orf3 can be targeted by the cytotoxic T lymphocytes (CTLs) in peripheral blood mononuclear cells (PBMCs) of breast carcinoma patients. In this report, we evaluated the feasibility of cancer immunotherapy using Cep55/c10orf3 peptide for colorectal carcinoma (CRC). To evaluate the expression of Cep55/c10orf3 in CRC tissues, we performed immunohistochemical staining of using anti-Cep55/c10orf3 monoclonal antibody. Sixty-three percent cases showed weak positive for Cep55/c10orf3 in total 70 CRC cases. The Cep55/c10orf3 expression intention was collated with high histological grade of CRC. Thus, we hypothesized that Cep55/c10orf3 can also be the target of CTLs in CRC cases. We generated CTLs from PBMCs of human leukocyte antigen (HLA)-A24-positive colorectal carcinoma patients using HLA-A24-restricted Cep55/c10orf3 peptides. Two of 6 colorectal cancer patients were reactive for the Cep55/c10orf3_193(10) peptide, which was the only immunogenic peptide in breast carcinoma patients. CTL clone specific for Cep55/c10orf3_193(10) recognized and lysed HLA-A24 (+) and Cep55/c10orf3 (+) colorectal carcinoma cell lines. In addition, 1 of 6 colorectal carcinoma patients was reactive for the Cep55/c10orf3_402(11) and Cep55/c10orf3_283(12) peptides, but not for Cep55/c10orf3_193(10) with the ELISPOT assay. These observations suggest that the antigenic peptide repertoire presented by HLA-A24 in colorectal carcinoma might be different from that in breast carcinoma. Thus, these peptide vaccination peptide mixture of Cep55/c10orf3_193(10), Cep55/c10orf3_402(11) and Cep55/c10orf3_283(12) might be more effective than a single peptide in the treatment of colorectal carcinoma patients.

Carboxyl-Terminated PAMAM-SN38 Conjugates: Synthesis, Characterization, and in Vitro Evaluation

In this work, carboxyl-terminated PAMAM G-3.5 was covalently attached to SN38 via glycine and β-alanine spacers. The conjugates were stable at pH 7.4 and moderately hydrolyzed in cell culture media and rat plasma. Similarly to SN38 but to a lesser extent, both conjugates inhibited proliferation of human colorectal cancer HCT-116 cells, arrested the cell cycle in the G(2)/M phase, and led to nuclear fragmentation. However, activity of the conjugate with glycine spacer (IC(50) = 129 nM) was higher compared to that of the β-alanine linked conjugate (IC(50) = 387 nM). These PAMAM-SN38 conjugates have the potential for targeted therapy of colorectal carcinoma.