Fibroblast-derived exosomal microRNA regulates NKX3-1 expression in androgen-sensitive, androgen receptor-dependent prostate cancer cells.

Androgen deprivation therapy (ADT) targeting androgen production and androgen receptor (AR) signaling is the primary antihormonal therapy in the treatment of advanced prostate cancer (PCa). However, no clinically established molecular biomarkers have been identified to predict the effectiveness of ADT before starting ADT. The tumor microenvironment of PCa contains fibroblasts that regulate PCa progression by producing multiple soluble factors. We have previously reported that AR-activating factor-secreted fibroblasts increase the responsiveness of androgen-sensitive, AR-dependent PCa cells to ADT. Thus, we hypothesized that fibroblast-derived soluble factors may affect cancer cell differentiation by regulating cancer-related gene expression in PCa cells and that the biochemical characteristics of fibroblasts may be used to predict the effectiveness of ADT. Here, we investigated the effects of normal fibroblasts (PrSC cells) and three PCa patient-derived fibroblast lines (pcPrF-M5, -M28, and -M31 cells) on the expression of cancer-related genes in androgen-sensitive, AR-dependent human PCa cells (LNCaP cells) and three sublines showing different androgen sensitivities and AR dependencies. The mRNA expression of the tumor suppressor gene NKX3-1 in LNCaP cells and E9 cells (which show low androgen sensitivity and AR dependency) was significantly increased by treatment with conditioned media from PrSC and pcPrF-M5 cells but not from pcPrF-M28 and pcPrF-M31 cells. Notably, no upregulation of NKX3-1 was observed in F10 cells (AR-V7-expressing, AR-independent cells with low androgen sensitivity) and AIDL cells (androgen-insensitive, AR-independent cells). Among 81 common fibroblast-derived exosomal microRNAs that showed 0.5-fold lower expression in pcPrF-M28 and pcPrF-M31 cells than in PrSC and pcPrF-M5 cells, miR-449c-3p and miR-3121-3p were found to target NKX3-1. In only LNCaP cells, the NKX3-1 mRNA expression was significantly increased by transfection of an miR-3121-3p mimic but not that of the miR-449c-3p mimic. Thus, fibroblast-derived exosomal miR-3121-3p may be involved in preventing the oncogenic dedifferentiation of PCa cells by targeting NKX3-1 in androgen-sensitive, AR-dependent PCa cells.

HER2-positive adenocarcinoma arising from heterotopic pancreas tissue in the duodenum: A case report.

BACKGROUND: Adenocarcinoma originating from heterotopic pancreas tissue is a rare disease. Furthermore, to our knowledge, no HER2-positive cases in the duodenum have been reported in the scientific literature nor has the efficacy of trastuzumab treatment for the disease been reported. CASE SUMMARY: A 65-year-old woman whose clinical diagnosis was unresectable advanced duodenal cancer with HER2 overexpression responded well to trastuzumab chemotherapy. The main tumor in the duodenum reduced drastically. The patient underwent pancreaticoduodenectomy and lymph node dissection. A small number of cancer cells remained in the submucosal layer of the duodenum and pancreas head. After histological and immunohistochemical examination, the patient was diagnosed with duodenal adenocarcinoma originating from heterotopic pancreas tissue. CONCLUSION: Trastuzumab treatment is effective in HER2-positive adenocarcinoma originating from heterotopic pancreas tissue in the duodenum.

Cancer-related gene mutations and intratumoral genetic heterogeneity in human epidermal growth factor receptor 2 heterogeneous gastric cancer.

Human epidermal growth factor receptor 2 (HER2) protein overexpression is associated with HER2 gene amplification, a critical driver oncogenetic change in gastric cancer. HER2 heterogeneity in advanced gastric cancer is associated with a poor prognosis and affects the clinical efficacy of trastuzumab. However, the mechanisms of HER2 heterogeneity are not fully understood. Here, we examined whether HER2 heterogeneous gastric cancer exhibited intratumoral genetic heterogeneity in other cancer-related genes. Two cases of advanced gastric cancer with HER2 heterogeneity were selected, and samples of HER2-positive and HER2-negative areas in each case were analyzed using a cancer-associated multiple gene panel. In both cases, TP53 mutations were observed in both HER2-positive and HER2-negative areas, whereas many of the potential driver and passenger mutations differed between HER2-positive and HER2-negative areas. Overall, our findings demonstrated that HER2 heterogeneous gastric cancer exhibited intratumoral genetic heterogeneity in other cancer-related genes and that the molecular mechanisms could differ between HER2-positive and -negative areas.

SOX11-induced decrease in vimentin and an increase in prostate cancer cell migration attributed to cofilin activity.

SOX11 is a transcription factor in the SOX family of genes that regulate multiple cellular events by influencing the expression of key genes in developmental, physiological, and tumorigenic cells. To elucidate the role of SOX11 in prostate cancer cells, PC-3 prostate cancer cells were cloned (S6 and S9 cells) to highly express SOX11. We demonstrated that both S6 and S9 lose vimentin expression, acquiring epithelial marker proteins, which indicates the Epithelial state phenotype. S6 and S9 cells have cancer-promoting characteristics that include higher migratory properties compared with control cells. The mechanisms that are responsible for the enhanced migration are cofilin activity and keratin 18 expression. TCGA (The Cancer Genome Atlas) dataset analysis revealed that metastatic prostate cancer tumors tend to have more SOX11 gene amplification compared with primary tumors. These results suggest the tumor promotive role and epithelial protein induction of SOX11 in prostate cancer cell.

Association of HER2 gene amplification and tumor progression in early gastric cancer.

Overexpression of human epidermal growth factor receptor 2 (HER2) protein in association with HER2 gene amplification is found in 7-34% of gastric cancers. In breast cancer, HER2 overexpression is a prognostic factor in advanced cases and is associated with tumor progression in ductal carcinoma in situ. However, the biological and clinical significance of HER2 status in early gastric cancer is unknown. Here, we aimed to examine the correlation between HER2 gene amplification and tumor progression in early gastric cancer. The HER2 status was evaluated in 149 lesions from 141 consecutive patients with early gastric cancer who underwent endoscopic resection by immunohistochemistry and dual color in situ hybridization. HER2 gene amplification was detected in 35 (23.5%) of 149 lesions, and of those, 26 cases (74.3%) showed intratumoral heterogeneity. HER2 gene amplification was found in noninvasive carcinoma, and there was a significant correlation between HER2 status and T factor (P = 0.0290). Our study demonstrated that HER2 gene amplification occurred during the early stages of gastric cancer and showed heterogeneity in several cases. HER2 gene amplification may be involved in tumor progression in early gastric cancer.

Cytological features of adenocarcinoma admixed with small cell neuroendocrine carcinoma of the uterine cervix.

Adenocarcinoma admixed with neuroendocrine carcinoma of the uterine cervix is a rare malignancy with a poor prognosis, and few reports have described the cytological features of this carcinoma. To characterize the cytological features of this malignancy in cervical smears, we report a case of a 52-year-old Japanese woman with cervical adenocarcinoma admixed with small cell neuroendocrine carcinoma (SCNEC). Cytologically, there were two types of cells with different sizes. The smaller cells formed clusters, which showed a partially Indian file pattern, a high nuclear/cytoplasmic ratio, and hyperchromatic nuclei. In contrast, the larger cells showed cytological features of adenocarcinoma, indicating a glandular-like pattern. Histological examination of biopsy specimens revealed that the tumors were composed of almost equal areas of SCNEC and adenocarcinoma. Neuroendocrine differentiation was confirmed by immunohistochemistry for synaptophysin and CD56. Thus, when adenocarcinoma cells are detected in smears, attempts to search for SCNEC cells should be made by combined cytological and histological analyses in order to reach an accurate diagnosis of the carcinoma in the uterine cervix.

ASF-4-1 fibroblast-rich culture increases chemoresistance and mTOR expression of pancreatic cancer BxPC-3 cells at the invasive front in vitro, and promotes tumor growth and invasion in vivo.

Pancreatic cancer develops dense stromal tissue through the desmoplastic reaction. The aim of the present study was to assess the effects of a fibroblast-rich environment on the malignant potential of pancreatic cancer. Cells from the human pancreatic cancer cell line BxPC-3 were mixed at a ratio of 1:3 (fibroblast-rich) or 1:1 (fibroblast-poor) with cells from the human skin fibroblast line ASF-4-1. In the fibroblast-rich co-culture, tumor budding was observed and BxPC-3 cells were found to be more resistant to gemcitabine than those in the fibroblast-poor co-culture. Immunohistochemistry revealed that the expression of mammalian target of rapamycin was increased at the invasive front of fibroblast-rich co-cultures. In addition, in mouse xenografts of fibroblast-rich co-cultures, tumors were larger and had a higher Ki-67 index compared with that of the fibroblast-poor co-culture xenografts. These results indicate that fibroblast-rich co-cultures may promote the malignant potential of the pancreatic cancer cell line BxPC-3, both in vitro and in vivo.

Significant intratumoral heterogeneity of human epidermal growth factor receptor 2 status in gastric cancer: A comparative study of immunohistochemistry, FISH, and dual-color in situ hybridization.

The assessment of human epidermal growth factor receptor 2 (HER2) status is crucial for selecting patients with gastric cancer who may benefit from HER2-targeted therapy. Accurate assessment using biopsy specimens is important for patients with advanced-stage cancer. Intratumoral heterogeneity of HER2, however, is a major challenge in HER2 testing. Here, we aimed to examine whether assessment of HER2 status could be accurately carried out with currently used methods, namely, immunohistochemistry (IHC), FISH, and dual-color in situ hybridization (DISH). Human epidermal growth factor receptor 2 status was evaluated in 108 biopsy tissues from patients with gastric adenocarcinoma and 70 matched surgical specimens by IHC, FISH, and DISH; HER2 amplification was detected in 11 (10.2%) out of 108 biopsy specimens. The IHC and FISH results were well correlated, and FISH and DISH results were consistent for all cases. The overall concordance rate of HER2 status between biopsy tissues and surgical specimens was 91.4%. All six discordant cases were false negative on biopsy; of these cases, five showed HER2 heterogeneity on surgical resection. Assessment of the HER2 status of biopsy tissues could predict the status of the whole tumor; however, a proportion of these cases may be discordant because of intratumoral heterogeneity.

Inflammatory suppressive effect of prostate cancer cells with prolonged exposure to transforming growth factor ß on macrophage-differentiated cells via downregulation of prostaglandin E2.

Transforming growth factor ß1 (TGFß1) regulates a variety of cellular functions, including cell growth, apoptosis and differentiation. The aim of the current study was to investigate the alterations of phenotypic events in the long-term exposure of prostate cancer (PCa) cells to TGFß1 and its effect on macrophage-differentiated cells. The PCa cell line, PC-3, and the subclone, M1, were exposed to TGFß1 for short- or long-term periods. TGFß1 signaling was assessed by Smad3 phosphorylation, and non-canonical signaling was analyzed by quantitative polymerase chain reaction-based regulatory gene expression profiles. TGFß1-exposed PCa cells were also co-cultured with phorbol 12-myristate 13-acetate (PMA)-treated THP-1 macrophages as a model of the tumor microenvironment. The phosphorylation of Smad3 in the PCa cells with long-term exposure was lower than that in the PCa cells with short-term exposure. Interleukin-6 mRNA expression in the PMA-treated THP-1 macrophages was significantly downregulated by co-culture with the PCa cells with long-term exposure. Cyclooxygenase-2 expression in the long-term TGFß1-exposed PCa cells was lower than that in the control PCa cells, and the production of prostaglandin E2 (PGE2) in the long-term TGFß1-exposed PCa cells was also significantly lower. The results of the current study demonstrated that the long-term TGFß1 exposure of PCa cells induces phenotypic changes, including the downregulation of PGE2 production. This indicates that prolonged TGFß-exposed PCa cells may change the cytokine production of macrophages in the tumor microenvironment.

RhoB enhances migration and MMP1 expression of prostate cancer DU145.

Rho family protein regulates variety of cellular functions as cytoskeletal organization, cell proliferation and apoptosis. In the present study, we demonstrate that RhoB-overexpressed prostate cancer cells showed an enhanced cell motility and the administration of the GSK-3 inhibitors inhibited this increase in migration. Among the extracellular matrix and adhesion-related molecules, MMP1 RNA expression was increased in RhoB-overexpressed cells, administration of MMP inhibitor suppressed the collagen gel invasion in these cells. This is the first report evaluating RhoB function and the downstream signaling events in prostate cancer cell. Our results indicate that RhoB promotes cell motility and invasion in a metastatic prostate cancer cell.

High level expression of STAG1/PMEPA1 in an androgen-independent prostate cancer PC3 subclone.

In this paper, we describe the isolation and characterization of two PC3 subclones. One subclone, mr, showed an epithelial phenotype, the other, M1, showed a sarcomatous morphology. Transplanted into nude mice, mr developed tumors at a dramatically faster rate than M1. Comparing the two subclones, differentially expressed genes were identified, including E-cadherin, IL-8 and STAG1/PMEPA1. These genes were expressed at higher levels in mr than in M1.

Reduced heterochromatin protein 1-beta (HP1beta) expression is correlated with increased invasive activity in human melanoma cells.

Heterochromatin protein 1 (HP1) is associated with heterochromatin formation and the regulation of gene expression. In this study, we demonstrated that decreased HP1beta, but not HPla, mRNA and protein expression, correlates with invasive potential in five human melanoma cell lines, and we used immunohistochemistry to confirm that HP1beta expression is suppressed during melanoma progression. HPIP levels are decreased in (V600E)B-RAF-transformed mouse melanocytes, suggesting that HP1beta-mediated suppressive mechanisms correlate with melanoma oncogenesis. Expression of microphthalmia associated-transcription factor (MITF), an important melanocyte differentiation factor, is reduced in melanoma, which is correlated with poor prognosis. In CRL1579, SK-MEL-28 and HMV-II human melanoma cells in which HP1beta expression is reduced by RNAi, MITF RNA levels and invasiveness activities are differentially altered and are not correlated with each other. Our findings indicate that the (V600E)B-RAF mutation induces HPIbeta down-regulation, which causes epigenetic gene regulation associated with melanoma progression.

The 3'UTR of stem cell factor suppresses protein expression from a cotransfected vector.

One of the isoforms of stem cell factor (SCF) was isolated from an embryonic kidney library with the ability to suppress the expression of cotransfected gene. The responsible region resides in the SCF 3' untranslated region (UTR). The SCF 5'UTR also has repressive element with a smaller effect than 3'UTR. The mRNA expression level of the cotransfected gene was not markedly changed. These results suggest that the SCF 3'UTR should negatively regulate gene expression at a posttranscriptional level.