Mutations in TGFBR2, a component of the transforming growth factor (TGF)-β signaling pathway, occur in high-frequency microsatellite instability (MSI-H) colorectal cancer (CRC). In mouse models, ...Tgfbr2 inactivation in the intestinal epithelium accelerates the development of malignant intestinal tumors in combination with disruption of the Wnt-β-catenin pathway. However, no studies have further identified the genes influenced by TGFBR2 inactivation following disruption of the Wnt-β-catenin pathway. We previously described CDX2P-G19Cre;Apcflox/flox mice, which is stochastically null for Apc in the colon epithelium. In this study, we generated CDX2P-G19Cre;Apcflox/flox;Tgfbr2flox/flox mice, with simultaneous loss of Apc and Tgfbr2. These mice developed tumors, including adenocarcinoma in the proximal colon. We compared gene expression profiles between tumors of the two types of mice using microarray analysis. Our results showed that the expression of the murine homolog of GSDMC was significantly upregulated by 9.25-fold in tumors of CDX2P-G19Cre;Apcflox/flox;Tgfbr2flox/flox mice compared with those of CDX2P-G19Cre;Apcflox/flox mice. We then investigated the role of GSDMC in regulating CRC tumorigenesis. The silencing of GSDMC led to a significant reduction in the proliferation and tumorigenesis of CRC cell lines, whereas the overexpression of GSDMC enhanced cell proliferation. These results suggested that GSDMC functioned as an oncogene, promoting cell proliferation in colorectal carcinogenesis. In conclusion, combined inactivation of both Apc and Tgfbr2 in the colon epithelium of a CRC mouse model promoted development of adenocarcinoma in the proximal colon. Moreover, GSDMC was upregulated by TGFBR2 mutation in CRC and promoted tumor cell proliferation in CRC carcinogenesis, suggesting that GSDMC may be a promising therapeutic target.
Recently, mesenchymal stem cells (MSCs) were reported to migrate to tumor stroma as well as injured tissue. We examined the role of human MSCs in tumor stroma using an orthotopic nude mice model of ...KM12SM colon cancer. In in vivo experiments, systemically injected MSCs migrated to the stroma of orthotopic colon tumors and metastatic liver tumors. Orthotopic transplantation of KM12SM cells mixed with MSCs resulted in greater tumor weight than did transplantation of KM12SM cells alone. The survival rate was significantly lower in the mixed‐cell group, and liver metastasis was seen only in this group. Moreover, tumors resulting from transplantation of mixed cells had a significantly higher proliferating cell nuclear antigen labeling index, significantly greater microvessel area and significantly lower apoptotic index. Splenic injection of KM12SM cells mixed with MSCs, in comparison to splenic injection of KM12SM cells alone, resulted in a significantly greater number of liver metastases. MSCs incorporated into the stroma of primary and metastatic tumors expressed α‐smooth muscle actin and platelet‐derived growth factor receptor‐β as carcinoma‐associated fibroblast (CAF) markers. In in vitro experiments, KM12SM cells recruited MSCs, and MSCs stimulated migration and invasion of tumor cells through the release of soluble factors. Collectively, MSCs migrate and differentiate into CAFs in tumor stroma, and they promote growth and metastasis of colon cancer by enhancing angiogenesis, migration and invasion and by inhibiting apoptosis of tumor cells.
Gastric cancer (GC) develops through deregulation of gene expression and accumulation of epigenetic abnormalities, leading to tumor cell acquisition of malignant features. MicroRNAs (miRNAs) play a ...critical role in cancer development where they can act as oncogenes or oncosuppressors. To identify miRNAs that are associated with some clinicopathologic features of GC and/or participate in tumor progression, miRNA expression in 20 GC tissues and five corresponding non‐neoplastic gastric mucosa was examined by miRNA microarray. Oligonucleotide array analysis was carried out for miRNA target prediction. The functions of candidate miRNAs and their target genes were also analyzed by quantitative RT‐PCR, Western blotting, reporter gene assay, and cell invasion assay. Comparison of miRNA expression profiles revealed that downregulation of miR‐148a was identified in most of the GC tissues. Downregulation of miR‐148a was significantly correlated with an advanced clinical stage, lymph node metastasis, and poor clinical outcome. Custom oligonucleotide array analysis revealed that MMP7 expression was markedly downregulated in miR‐148a‐overexpressing GC cells; MMP7 was found to be a direct and functional target of miR‐148a, participating in cell invasion. These results suggest that miR‐148a contributes to the maintenance of homeostasis in normal stomach tissue and plays an important role in GC invasion by regulating MMP7 expression.
miR‐148a expression was significantly downregulated in gastric cancer, and its downregulation was significantly correlated with an worse clinico‐pathologic characters. MMP7 was found to be a direct and functional target of miR‐148a, participating in cell invasion.
Colorectal cancer (CRC) is the second leading cause of cancer‐related mortality worldwide. Kinesin Family Member C1 (KIFC1) has been proposed as a promising therapeutic target due to its pivotal role ...in centrosome clustering to mediate cancer cell progression. This study aimed to analyze the expression and biological function of KIFC1 in CRC. Immunohistochemically, 67 (52%) of 129 CRC cases were positive for KIFC1 and statistically associated with poorer overall survival. KIFC1 small interfering RNA (siRNA)‐transfected cells demonstrated lower cell proliferation as compared to the negative control cells. A specific KIFC1 inhibitor, kolavenic acid analog (KAA) drastically inhibited CRC cell proliferation. Microarray analysis revealed that KAA‐treated CRC cells presented reduced ZW10 interacting kinetochore protein (ZWINT) expression as compared to control cells. Immunohistochemical analysis demonstrated that 61 (47%) of 129 CRC cases were positive for ZWINT and ZWINT expression was significantly correlated with KIFC1 expression. ZWINT‐positive cases exhibited significantly worse overall survival. KIFC1 siRNA‐transfected cells showed reduced ZWINT expression while ZWINT siRNA‐transfected cells decreased cell proliferation. Both KIFC1 and ZWINT knockdown cells attenuated spheroid formation ability. This study provides new insights into KIFC1 regulating ZWINT in CRC progression and its potential as a therapeutic target.
Gastric cancer (GC) is one of the most common malignancies worldwide. In particular, scirrhous type GC is highly metastatic and is characterized clinically by rapid disease progression and poor ...prognosis. MicroRNAs (miRNAs) play crucial roles in cancer development and progression. In the present study, we identified several miRNAs that are expressed at higher levels in scirrhous type GC than in non‐scirrhous type GC by miRNA microarray analysis. Among these, microRNA‐143 (miR‐143) expression was higher in scirrhous type GC than in non‐scirrhous types of GC. In situ hybridization and quantitative RT‐PCR analysis showed that miR‐143 is expressed by stromal fibroblasts but not by cancer cells. In stromal cells, miR‐143 enhanced collagen type III expression in normal gastric fibroblasts and cancer‐associated fibroblasts through activation of transforming growth factor‐β)/SMAD signaling. Furthermore, high miR‐143 expression in GC was associated with worse cancer‐specific mortality (P = 0.0141). Multivariate analysis revealed that miR‐143 was an independent prognostic factor. Treatment of GC cell lines with 5‐aza‐2′‐deoxycytidine restored the expression of miR‐143, and precursor miR‐143 caused the inhibition of cancer cell invasion. These data suggest that miR‐143 regulates fibrosis of scirrhous type GC through induction of collagen expression in stromal fibroblasts and that miR‐143 expression serves as a prognostic marker of GC.
MicroRNA‐143 regulates the collagen type III expression through activation of transforming growth factor‐β/SMAD pathway in stromal fibroblasts of scirrhous type gastric cancer.
We previously reported that claspin is a key regulator in the progression of gastric cancer and renal cell carcinoma. However, the clinicopathological significance of claspin in urothelial carcinoma ...(UC) has not been investigated. We analyzed the expression and distribution of claspin in UC cases by immunohistochemistry. In the non-neoplastic urothelium, the expression of claspin was either weak or absent, whereas UC tissues showed nuclear staining. The expression of claspin was detected in 58 (42%) of a total of 138 upper tract UC cases treated by radical nephroureterectomy without neoadjuvant chemotherapy. Claspin-positive UC cases were associated with nodular/flat morphology, variant histology, high tumor grade, high pathological T grade, and lymphatic and venous invasion. The expression of claspin was significantly associated with decreased progression-free survival and cancer-specific survival. In addition, claspin was co-expressed with Ki-67, PD-L1, HER2, EGFR, and p53 in consecutive tumor sections of UC. An immunohistochemical analysis of claspin in biopsy specimens revealed that strong to moderate claspin staining was more frequently observed in carcinoma in situ in comparison to dysplasia or the benign urothelium. Furthermore, immunocytochemistry for claspin on urine cytology slides demonstrated that the proportion of claspin-positive cells was significantly greater in high-grade UC than in benign cases. These results suggest that claspin may be a novel prognostic marker and a possible therapeutic target molecule for UC. Moreover, claspin could be a useful diagnostic biomarker of urothelial neoplasia.
Abstract We previously reported that in an orthotopic nude mouse model of human colon cancer, bone marrow–derived mesenchymal stem cells (MSCs) migrated to the tumor stroma and promoted tumor growth ...and metastasis. Here, we evaluated the proliferation and migration ability of cancer cells cocultured with MSCs to elucidate the mechanism of interaction between cancer cells and MSCs. Proliferation and migration of cancer cells increased following direct coculture with MSCs but not following indirect coculture. Thus, we hypothesized that direct contact between cancer cells and MSCs was important. We performed a microarray analysis of gene expression in KM12SM colon cancer cells directly cocultured with MSCs. Expression of epithelial-mesenchymal transition (EMT)–related genes such as fibronectin ( FN ), SPARC , and galectin 1 was increased by direct coculture with MSCs. We also confirmed the upregulation of these genes with real-time polymerase chain reaction. Gene expression was not elevated in cancer cells indirectly cocultured with MSCs. Among the EMT-related genes upregulated by direct coculture with MSCs, we examined the immune localization of FN, a well-known EMT marker. In coculture assay in chamber slides, expression of FN was seen only at the edges of cancer clusters where cancer cells directly contacted MSCs. FN expression in cancer cells increased at the tumor periphery and invasive edge in orthotopic nude mouse tumors and human colon cancer tissues. These results suggest that MSCs induce EMT in colon cancer cells via direct cell-to-cell contact and may play an important role in colon cancer metastasis.
Abstract
To detect muscle-invasive upper tract urothelial carcinoma, we evaluated the internal texture of the tumor using texture analysis of computed tomography images in 86 cases of upper tract ...urothelial carcinoma. The internal texture of the tumor was evaluated as the value of computed tomography attenuation number of the unenhanced image, and the median, standard deviation, skewness and kurtosis were calculated. Each parameter was compared with clinicopathological factors, and their associations with postoperative prognosis were investigated. Immunohistochemistry was performed to investigate the histological and molecular mechanisms of the inflammatory tumor microenvironment. The histogram of computed tomography attenuation number in non-muscle invasive tumor was single-peaked, whereas muscle invasive tumor showed a multi-peaked shape. In the parameters obtained by texture analysis, standard deviation was significantly associated with pathological stage (
p
< 0.0001), tumor grade (
p
= 0.0053), lymphovascular invasion (
p
= 0.0078) and concomitant carcinoma in situ (
p
= 0.0177) along with recurrence-free (
p
= 0.0191) and overall survival (
p
= 0.0184). The standard deviation value correlated with the amount of stromal components (
p
< 0.0001) and number of tumor-infiltrating macrophages (
p
< 0.0001). In addition, higher expression of high mobility group box 1 was found in heterogeneous tumor. Tumor heterogeneity evaluated by texture analysis was associated with muscle-invasive upper tract urothelial carcinoma and represented an inflammatory tumor microenvironment and useful as the clinical assessment to differentiate muscle invasive tumor.
Colorectal cancer (CRC) is one of the leading causes of cancer‐related death worldwide. In order to identify novel prognostic markers or therapeutic targets for CRC, we searched for candidate genes ...in our comprehensive gene expression libraries, and focused on SEC11A, which encodes the SPC18 protein. SPC18 plays a key role in the endoplasmic reticulum‐Golgi secretory pathway and presumably regulates the secretion of various secretory proteins. An immunohistochemical analysis of SPC18 in 137 CRC tissue samples demonstrated that 79 (58%) CRC cases were positive for SPC18. SPC18‐positive CRC cases were more advanced in terms of N classification (P = 0.0315) and tumor stage (P = 0.0240) than SPC18‐negative CRC cases. Furthermore, the expression of SPC18 was an independent prognostic classifier for CRC patients. The cell growth and invasiveness of SPC18 siRNA‐transfected CRC cell lines was less than that of the negative control siRNA‐transfected cell lines. The levels of phosphorylated epidermal growth factor receptor, Erk and Akt were lower in SPC18 siRNA‐transfected CRC cells than in control cells. The expression of SPC18 was colocalized with β‐catenin nuclear localization and MMP7 at the invasive front. An immunohistochemical analysis of human colorectal polyp specimens revealed a sequential increase in the expression of SPC18 through the conventional adenoma‐carcinoma pathway, while SPC18 was not expressed or was expressed to a lesser extent in serrated pathway‐related tumors. These results suggest that SPC18 is involved in tumor progression, and is an independent prognostic classifier in patients with CRC.
SPC18 plays a key role in the endoplasmic reticulum‐Golgi secretory pathway and presumably regulates the secretion of various secretory proteins.Our results suggest that SPC18 is involved in tumor progression, and is an independent prognostic classifier in patients with CRC.
Intelectin‐1 (ITLN1) is an adipokine with an anti‐inflammatory function that is involved in neoplastic diseases such as pleural mesothelioma and gastric and prostate cancers. However, the expression ...and function of ITLN1 in colorectal cancer (CRC) remain unknown. To identify novel prognostic markers or therapeutic targets for CRC, we focused on ITLN1 protein. By immunohistochemistry, 87 (59%) of 148 CRC cases showed reduced expression of ITLN1. ITLN1‐reduced CRC cases were associated with higher M grades (P = 0.0017) than ITLN1‐retained CRC cases. Furthermore, the cases with ITLN1 retained expression tended toward a more favorable prognosis than those with reduced expression. Cell growth of the CRC cell lines transfected with ITLN1 siRNA were greater than those of the negative control cell lines transfected with siRNA. Levels of phosphorylated epidermal growth factor receptor, Erk and Akt were higher in the CRC cells transfected with ITLN1 siRNA than in control cells. Immunohistochemical analysis of human colorectal polyp specimens also revealed a sequential decrease in the expression of ITLN1 through both the conventional adenoma–carcinoma pathway and the serrated pathway. These results indicated that ITLN1 might play an important role in regulating colorectal tumorigenesis.
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