Patient-Derived Xenograft Models of Epithelial Ovarian Cancer for Preclinical Studies / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Patient-derived tumor xenografts (PDXs) can provide more reliable information about tumor biology than cell line models. We developed PDXs for epithelial ovarian cancer (EOC) that have histopathologic and genetic similarities to the primary patient tissues and evaluated their potential for use as a platform for translational EOC research. MATERIALS AND METHODS: We successfully established PDXs by subrenal capsule implantation of primary EOC tissues into female BALB/C-nude mice. The rate of successful PDX engraftment was 48.8% (22/45 cases). Hematoxylin and eosin staining and short tandem repeat analysis showed histopathological and genetic similarity between the PDX and primary patient tissues. RESULTS: Patients whose tumors were successfully engrafted in mice had significantly inferior overall survival when compared with those whose tumors failed to engraft (p=0.040). In preclinical tests of this model, we found that paclitaxel-carboplatin combination chemotherapy significantly deceased tumor weight in PDXs compared with the control treatment (p=0.013). Moreover, erlotinib treatment significantly decreased tumor weight in epidermal growth factor receptor–overexpressing PDX with clear cell histology (p=0.023). CONCLUSION: PDXs for EOC with histopathological and genetic stability can be efficiently developed by subrenal capsule implantation and have the potential to provide a promising platform for future translational research and precision medicine for EOC.

Proton Pump Inhibition Enhances the Cytotoxicity of Paclitaxel in Cervical Cancer / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: This study was conducted to investigate whether a proton pump inhibitor (PPI) could enhance chemosensitivity via the inhibition of vacuolar-type H⁺ ATPase (V-ATPase) in cervical cancer. MATERIALS AND METHODS: The expression of V-ATPase was evaluated in 351 formalin-fixed, paraffin-embedded human cervical cancer tissues using immunohistochemistry and compared with clinicopathologic risk factors for disease prognosis. The influence of cell proliferation and apoptosis following V-ATPase siRNA transfection or esomeprazole pretreatment was assessed in cervical cancer cell lines using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide and enzyme-linked immunosorbent assay, respectively. RESULTS: Immunohistochemical analysis revealed that V-ATPase was expressed in about 60% of cervical cancer tissue samples (211/351), and the expression was predominantly found in adenocarcinoma histology (p=0.016). Among patients with initially bulky cervical cancer (n=89), those with V-ATPase expression had shorter disease-free survival (p=0.005) and overall survival (p=0.023). Co-treatment with V-ATPase siRNA or esomeprazole with paclitaxel significantly decreased the cell proliferation of cervical cancer cell lines, including HeLa and INT407, compared to cell lines treated with paclitaxel alone (p < 0.01). Moreover, V-ATPase siRNA or esomeprazole followed by paclitaxel significantly increased the expression of active caspase-3 in these cells compared to cells treated with paclitaxel alone (both, p < 0.05). CONCLUSION: V-ATPase was predominantly expressed in cervical adenocarcinoma, and the expression of V-ATPases was associated with poor prognosis. The inhibition of V-ATPase via siRNA or PPI (esomeprazole) might enhance the chemosensitivity of paclitaxel in cervical cancer cells.

The effects of selenium on tumor growth in epithelial ovarian carcinoma / 부인종양

OBJECTIVE: Epidemiological studies suggest that selenium protects against the development of several cancers. Selenium (sodium selenite) has been reported to interfere with cell growth and proliferation, and to induce cell death. In this study, we tested whether selenium could have growth-inhibiting effect in ovarian cancer cells and an orthotopic animal model. METHODS: Cell growth in selenium-treated cells was determined in human ovarian cancer cells, A2780, HeyA8, and SKOV3ip1 using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) assay. Animal experiment of selenium with paclitaxel was performed using SKOV3ip1 cells in nude mice to evaluate their inhibiting effect for tumor growth. In addition, another animal experiment of paclitaxel with or without selenium was performed to assess the effect of survival and food intake in mice. RESULTS: The in vitro growth of selenium-treated cells was significantly decreased dose-dependently in A2780, HeyA8, and SKOV3ip1 cells. Therapy experiment in mice was started 1 week after injection of the SKOV3ip1 cells. Treatment with selenium (1.5 mg/kg, 3 times/week) and paclitaxel injection showed no addictive effect of the inhibition of tumor growth. However, combination of selenium and paclitaxel showed the slightly increased food intake compared with paclitaxel alone. CONCLUSION: Although selenium has growth-inhibiting effect in ovarian carcinoma cells in vitro, there is no additive effect on tumor growth in mice treated with combination of paclitaxel and selenium. However, food intake is slightly higher in selenium-treated mice during chemotherapy.

Expression of TLR-4 in epithelial serous ovarian cancer / 부인종양

OBJECTIVE: This study was to determine the expression of Toll-like receptor 4 (TLR-4) in ovarian serous adenocarcinoma tissues. METHODS: TLR-4 expression was evaluated at the RNA level by real-time quantitative RT-PCR, in 24 fresh frozen ovarian serous adenocarcinoma tissues and 9 normal ovarian tissues. TLR-4 expression was also evaluated by immunohistochemistry (IHC) in each three ovarian carcinoma tissues and normal ovarian tissues. RESULTS: Positive immunoreactivity for TLR-4 was observed in the normal ovarian tissues but not in the ovarian carcinoma tissues. The staining was localized in the cytoplasm as well as on the cell surface. Real-time quantitative RT-PCR revealed that TLR-4 expression was significantly lower in tumors than in normal ovarian tissues (p=0.0003). There were no significant correlations between clinical parameters and the expression level of TLR-4 mRNA in ovarian serous adenocarcinomas. However, tumors without LN metastasis (p=0.068) and lower grade (p=0.075) showed trends of higher TLR-4 mRNA expression. CONCLUSION: TLR-4 expression was significantly lower in ovarian serous adenocarcinoma tissues than in normal ovarian tissues, and further studies on TLR-4 signaling pathway in ovarian carcinoma are needed.

Microarray versus magnetic resonance imaging prediction of lymph node metastasis in patients with cervical squamous cell carcinoma / 부인종양

OBJECTIVE: We investigated whether microarray-based gene expression profiling of primary tumor biopsy material could be used to predict lymph node (LN) metastasis in patients with uterine squamous cell carcinoma by comparing this approach with magnetic resonance imaging. METHODS: Forty three primary cervical cancer samples (16 with LN metastasis and 27 without LN metastasis) from radical hysterectomy with pelvic LN dissection were obtained, RNA was isolated, and oligonucleotide gene chips (Macrogen, Seoul, Korea) were hybridized. The samples were randomly divided into training (31 samples) and test (12 samples) sets. A prediction model for LN metastasis from the training set was developed by support vector machine methods using a 10-fold cross-validation and it was tested for its prediction accuracy by applying it to the test set. We evaluated pelvic LN status by MRI with newly designed criteria in these patients and compared the accuracy of MRI with microarray. In addition, we created a new approach by a combination of both. RESULTS: The "LN prediction model" derived from the signature of 156 distinctive genes had a prediction accuracy of 83% when applied to the independent test set. MRI showed an accuracy (69%) for the prediction of LN metastasis. The combination model with MRI findings and microarray improved prediction accuracy over MRI alone but the improvement was not statistically significant (74% and 69%, respectively; p=0.688). CONCLUSION: Current data show that the prediction of LN metastasis can be allowed by DNA microarray of the primary tumor biopsy, alone or in combination with MRI.

Promoter hypermethylation and loss of heterozygosity of FHIT genes in squamous cell carcinoma of uterine cervix / 부인종양

OBJECTIVE: This study was to investigate the status of hypermethylation and loss of heterozygosity (LOH) in chromosome 3p tumor-suppressor gene for cervical carcinoma. METHODS: We examined the promoter methylation status of the chromosome 3p gene, fragile histidine triad (FHIT), in 37 samples of cervical squamous cell carcinoma and corresponding noncancerous tissues using a methylation-specific polymerase chain reaction. We also analyzed the 37 paired samples for LOH at two loci on chromosome 3p. RESULTS: Promoter hypermethylation in FHIT was detected in 24% of tumors, whereas no hypermethylation was detected in the corresponding noncancerous tissues. LOH in the regions of FHIT was observed in 10% of informative cases. There were no correlations between LOH and promoter hypermethylation for the gene. FHIT hypermethylation was associated with small tumors and, when adjusted for tumor size, correlated significantly with more frequent lymph node metastasis. CONCLUSION: Promoter hypermethylation and LOH of FHIT gene may play a role in cervical carcinogenesis. In addition, hypermethylation of FHIT may be associated with the status (aggressiveness) of cervical carcinoma.

Increased expression of pAKT is associated with radiation resistance in cervical cancer / 부인종양

OBJECTIVE: The aim of this study was to investigate the association of phosphorylated AKT (pAKT) expression and radiation resistance in cervical cancer. METHODS: A retrospective review was made of the records of 25 women who received primary radiation therapy due to locally advanced cervical cancer (LACC) with FIGO stage IIB-IVA. Nine patients regarded as radiation resistant developed local recurrences with a median progression free interval of 10 months. Sixteen patients did not show local recurrences, and were regarded as a radiation sensitive group. Using pretreatment paraffin-embedded tissues, we evaluated pAKT expression by immunohistochemistry. RESULTS: A significant association was found between the level of pAKT expression and local recurrence. Immunohistochemical staining for pAKT was significantly more frequent in the radiation resistant than in the radiation sensitive group (p=0.007). The mean progression free survival (PFS) was 84 months for patients with pAKT negative staining (17 cases) and 44 months for patients with pAKT positive expression (8 cases)(p=0.015). CONCLUSION: These results suggest that signaling from PI3K/pAKT can lead to radiation resistance in LACC.

Decreased expression of galectin-3 is associated with the progression of cervical neoplasia / 부인종양

OBJECTIVE: Galectin-3, a member of the beta-galactoside-binding proteins, is an intracellular and extracellular lectin that interacts with intracellular glycoproteins, cell surface molecules and extracellular matrix proteins. Galectin-3 is expressed widely in epithelial and immune cells and the level of expression varies in many cancer cells relative to the normal tissues from which they arise. We investigated whether the expression of galectin-3 is associated with the progression of cervical neoplasia. METHODS: The galectin-3 expression was evaluated by immunohistochemistry in 90 formalin-fixed paraffin-embedded cervical tissues: 10 normal cervical specimens, 20 low-grade squamous intraepithelial lesions (LSILs), 20 high-grade squamous intraepithelial lesions (HSILs), and 40 invasive squamous cell carcinomas (ISCCs). RESULTS: The immunohistochemical staining showed that the expression of galectin-3 was strong in all normal cervical squamous epithelia. Staining gradually decreased in accordance with the histopathologic grades from an LSIL to an HSIL and an ISCC (P<0.001). In particular, the expression of galectin-3 was significantly decreased in HSILs (P=0.001) and this down-regulation was more pronounced in ISCCs than normal tissues (P<0.001). CONCLUSION: These data constitute the first observation that the expression of galectin-3 is down-regulated in cervical cancer tissues and suggest the decreased expression of this galactoside-binding lectin is associated with the progression of cervical neoplasia.

Development of Tetracycline-regulated Adenovirus Expression Vector System

Recombinant adenovirus vector systems with strong promoters have been used to achieve high level production of recombinant protein. However, this overexpression system cause some problems such as disturbance of cell physiology and increment of cellular toxicity. Here, we showed a tetracycline-regulated adenovirus expression vector system. Our results showed that the expression level of transgene(p-53) was high and easily regulated by tetracycline. In addition, the maximal gene expresion level of the tetracycline-controlled gene expression system was higher than that of the wild type CMV promoter system. Therefore, tetracycline-regulated adenoviral vector system could be applicable for regulatory high-level expression of toxic gene. Also, this system will be useful for functional studies and gene therapy.

Expression of p73 in Null-p53 SKOV3 Ovarian Cancer Cell Line / 대한부인종양콜포스코피학회잡지

p73, a first p53 relative, has been identified at chromosome 1p36, a region that is deleted in variety of human cancers. This protein shares strong homology with p53 protein, suggesting functional similarities with p53. Indeed, p73 can activate p53 downstream genes inducing apoptosis or growth arrest in tumor cells lacking p53. This phenomenon leads us to investigate the function of p73 in ovarian cancer because aberrant p53 was very frequently found in this cancer. We hypothesize that DNA damaging agents trigger p53 dependent apoptotic pathway through p73 instead of p53 in ovarian cancer having aberrant p53. We selected SKOV3 ovarian cancer cell line having no p53 gene and treated this cell line with cisplatin. After the treatment, we examined the transcriptional level of p73 and p21. Moreover, to identify whether the status of p53 influence to the function of p73, we performed same experiment after inserting adenovirus mediated p53(Avp53) into cell line. We detected significantly increased transcripts of p73 whcn treated with cisplatin. But treated with Avp53 or combined treatment with cisplatin, the transcriptional levels were not changed. These data suggest that overexpression of p73 may be important to trigger apoptotic pathway when the p53 gene is lost, but not so important in cells having normal p53.

Effects of verapamil, tamoxifen and cyclosporin A for the modulation of multidrug resistance in human lung cancer lines / 대한암학회지

No abstract available.

Comparison of p53 gene mutations in paired primary and metastatic gastric tumor tissues

Our previous study revealed that mutations of the p53 gene were detected by cDNA sequencing in one of four (25%) primary gastric tumors and in five of six (83%) gastric cancer cell lines. It was of interest that all five cell lines established from metastatic lesions had p53 gene mutations, while the single cell line established from a primary tumor lacked an abnormality. Thus, the current study was initiated to determine the frequency of p53 mutations in 10 pairs of samples from primary gastric carcinomas and their lymph node metastases, in addition to morphologically normal gastric mucosa. In addition, we correlated the findings with other relevant molecular markers including the metastasis associated nm23-H1 gene and loss of heterozygosity (LOH) using multiple polymorphic markers for chromosome 17p and sequencing the entire open reading frame (ORF) of the p53 gene. Five of ten (50%) patients were constitutionally heterozygous for one or more 17p and/or p53 probes (pYNZ 22, BamHI RFLP; pMct35.1, Mspl RFLP; php53cl, Bg/II RFLP), while none had LOH at the 17p and/or p53. A Bg/II RFLP for analysis of possible nm23-H1 somatic allelic deletion revealed no LOH out of four informative cases. One paired sample demonstrated the substitution of valine for isoleucine at codon 41 (GTT to ATT) in both primary gastric tumor and metastasis. Another metastatic sample demonstrated the substitution of proline for threonine at codon 278 (CCT to C/ACT) in addition to a non-mutated codon, while only the wild-type p53 sequence was present in the paired primary gastric tumor tissue.(ABSTRACT TRUNCATED AT 250 WORDS)

p53 gene mutation in hepatocellular carcinoma from Korean patients and in established hepatocellular carcinoma cell lines / 대한암학회지

No abstract available.