Prostate stem cell antigen is overexpressed in prostate cancer metastases.

PURPOSE: Prostate stem cell antigen (PSCA) is expressed by a majority of prostate cancers and is a promising therapeutic target. PSCA protein and mRNA expression was examined in prostate cancer bone, lymph node, and visceral metastases to assess the potential of PSCA as an immunotherapeutic target in advanced prostate cancer. EXPERIMENTAL DESIGN: Immunohistochemical analysis of PSCA protein expression and quantitative mRNA expression analysis of PSCA was done on clinical specimens of prostate cancer bone, lymph node, and visceral metastases. PSCA protein and mRNA expression levels were quantified and compared between available matched pairs of bone and lymph node or visceral metastases. RESULTS: Bone metastases stained with higher intensity of PSCA compared with lymph node or liver metastases in seven of eight (87.5%) matched pairs (P = 0.035). PSCA mRNA expression was equal or greater than that of LAPC-9, a PSCA expressing xenograft, in 12 of 24 (50%) cases of prostate cancer metastases and was significantly correlated with PSCA protein expression (sigma = 0.84, P = 0.0019). Overall, PSCA protein expression was detected in 41 of 47 (87.2%), four of six (66.7%), and two of three (66.7%) cases of bone, lymph node, and liver metastases, respectively. Mean PSCA staining intensity was significantly higher in prostate cancer bone metastases compared with lymph node metastases (2.0 +/- 0.02 versus 0.83 +/- 0.31, P = 0.014). CONCLUSIONS: Prostate cancer metastases express PSCA. However, greater PSCA staining intensity and level of PSCA mRNA expression was associated with bone metastases compared with lymph node metastases. This study suggests that PSCA is a promising tumor marker and potential therapeutic target for patients with metastatic prostate cancer.     

FXYD3 is overexpressed in pancreatic ductal adenocarcinoma and influences pancreatic cancer cell growth

The expression and localization of FXYD domain containing ion transport regulator 3 (FXYD3), a transmembrane protein that acts as a chloride channel or chloride channel regulator, was analyzed in pancreatic tissues derived from donors and patients suffering from chronic pancreatitis (CP) or pancreatic ductal adenocarcinoma (PDAC) as well as in pancreatic cancer cells using QRT-PCR, laser-capture microdissection and microarray analysis, in situ hybridization and immunohistochemistry. FXYD3 antisense expressing T3M4 pancreatic cancer cells were generated and compared to control cells using anchorage-dependent and independent growth assays, and xenotransplantation into nude mice. FXYD3 mRNA levels were 3.4-fold increased in PDAC tissues compared to donor specimens (p = 0.006), and 3.9-fold increased in microdissected cancer cells compared to normal pancreatic ductal cells (p = 0.02). FXYD3 was localized in the tubular complexes and PanIN lesions of both CP and PDAC, as well as in pancreatic cancer cells. Downregulation of FXYD3 by stable antisense transfection increased significantly the doubling time of T3M4 pancreatic cancer cells from 44 +/- 2 hr to 55 +/- 12 hr (p = 0.02). Nude mice transplanted with antisense transfected cells displayed a significant increase in tumor doubling time from 3.3 days +/- 1.0 to 4.3 days +/- 0.43 (p = 0.058). Anchorage-independent growth and sensitivity to 5-fluorouracil, gemcitabine and cisplatin as well as to MgCl(2) were not dependent on the level of FXYD3 expression. In conclusion, overexpression of FXYD3 in pancreatic cancer may contribute to the proliferative activity of this malignancy.     

Towards the identification of new markers of pancreatic cancer by gene expression analysis

The poor prognosis of pancreatic cancer has remained unchanged for many years, with a 5-year survival of less than 5 %. Current methods for diagnosing pancreatic cancer are inadequate at identifying small tumors that can be resected by surgery. Characterization of gene expression patterns in pancreatic cancer provided a list of genes that are specifically overexpressed in cancer cells. These genes are putative novel markers for the diagnosis and the prognosis of pancreatic cancer and for the development of targeted therapies. Gene expression analysis should lead to the discovery of molecular markers for early detection of pancreatic cancer that could benefit patients at high risk of developing pancreatic cancer.     

Prostate stem cell antigen is overexpressed in human transitional cell carcinoma.

Prostate stem cell antigen (PSCA), a homologue of the Ly-6/Thy-1 family of cell surface antigens, is expressed by a majority of human prostate cancers and is a promising target for prostate cancer immunotherapy. In addition to its expression in normal and malignant prostate, we recently reported that PSCA is expressed at low levels in the transitional epithelium of normal bladder. In the present study, we compared the expression of PSCA in normal and malignant urothelial tissues to assess its potential as an immunotherapeutic target in transitional cell carcinoma (TCC). Immunohistochemical analysis of PSCA protein expression was performed on tissue sections from 32 normal bladder specimens, as well as 11 cases of low-grade transitional cell dysplasia, 21 cases of carcinoma in situ (CIS), 38 superficial transitional cell tumors (STCC, stages T(a)-T(1)), 65 muscle-invasive TCCs (ITCCs, stages T(2)-T(4)), and 7 bladder cancer metastases. The level of PSCA protein expression was scored semiquantitatively by assessing both the intensity and frequency (i.e., percentage of positive tumor cells) of staining. We also examined PSCA mRNA expression in a representative sample of normal and malignant human transitional cell tissues. In normal bladder, PSCA immunostaining was weak and confined almost exclusively to the superficial umbrella cell layer. Staining in CIS and STCC was more intense and uniform than that seen in normal bladder epithelium (P < 0.001), with staining detected in 21 (100%) of 21 cases of CIS and 37 (97%) of 38 superficial tumors. PSCA protein was also detected in 42 (65%) of 65 of muscle-invasive and 4 (57%) of 7 metastatic cancers, with the highest levels of PSCA expression (i.e., moderate-strong staining in >50% of tumor cells) seen in 32% of invasive and 43% of metastatic samples. Higher levels of PSCA expression correlated with increasing tumor grade for both STCCs and ITCCs (P < 0.001). Northern blot analysis confirmed the immunohistochemical data, showing a dramatic increase in PSCA mRNA expression in two of five muscle-invasive transitional cell tumors when compared with normal samples. Confocal microscopy demonstrated that PSCA expression in TCC is confined to the cell surface. These data demonstrate that PSCA is overexpressed in a majority of human TCCs, particularly CIS and superficial tumors, and may be a useful target for bladder cancer diagnosis and therapy.     

Differentially expressed genes in pancreatic ductal adenocarcinomas identified through serial analysis of gene expression

Serial analysis of gene expression (SAGE) is a powerful tool for the discovery of novel tumor markers. The publicly available online SAGE libraries of normal and neoplastic tissues (http://www.ncbi.nlm.nih.gov/SAGE/) have recently been expanded; in addition, a more complete annotation of the human genome and better biocomputational techniques have substantially improved the assignment of differentially expressed SAGE "tags" to human genes. These improvements have provided us with an opportunity to re-evaluate global gene expression in pancreatic cancer using existing SAGE libraries. SAGE libraries generated from six pancreatic cancers were compared to SAGE libraries generated from 11 non-neoplastic tissues. Compared to normal tissue libraries, we identified 453 SAGE tags as differentially expressed in pancreatic cancer, including 395 that mapped to known genes and 58 "uncharacterized" tags. Of the 395 SAGE tags assigned to known genes, 223 were overexpressed in pancreatic cancer, and 172 were underexpressed. In order to map the 58 uncharacterized differentially expressed SAGE tags to genes, we used a newly developed resource called TAGmapper (http://tagmapper.ibioinformatics.org), to identify 16 additional differentially expressed genes. The differential expression of seven genes, involved in multiple cellular processes such as signal transduction (MIC-1), differentiation (DMBT1 and Neugrin), immune response (CD74), inflammation (CXCL2), cell cycle (CEB1) and enzymatic activity (Kallikrein 6), was confirmed by either immunohistochemical labeling of tissue microarrays (Kallikrein 6, CD74 and DMBT1) or by RT-PCR (CEB1, Neugrin, MIC1 and CXCL2). Of note, Neugrin was one of the genes whose previously uncharacterized SAGE tag was correctly assigned using TAGmapper, validating the utility of this program. Novel differentially expressed genes in a cancer type can be identified by revisiting updated and expanded SAGE databases. TAGmapper should prove to be a powerful tool for the discovery of novel tumor markers through assignment of uncharacterized SAGE tags.     

Prostate stem cell antigen (PSCA) expression in human prostate cancer tissues: implications for prostate carcinogenesis and progression of prostate cancer

OBJECTIVE: Prostate stem cell antigen (PSCA) is a recently defined homolog of the Thy-1/Ly-6 family of glycosylphosphatidylinositol (GPI)-anchored cell surface antigens. The objective of the present study was to examine the expression status of PSCA protein and mRNA in clinical specimens of human prostate cancer (PCa) and to validate it as a potential molecular target for diagnosis and treatment of PCa. METHODS: Immunohistochemical (IHC) and in situ hybridization (ISH) analyses of PSCA expression were simultaneously performed on paraffin-embedded sections of 20 benign prostatic hyperplasia (BPH), 20 prostatic intraepithelial neoplasm (PIN) and 48 prostate cancer (PCa) tissues, including 9 androgen-independent prostate cancers. The level of PSCA expression was semiquantitatively scored by assessing both the percentage and intensity of PSCA-positive staining cells in the specimens. We then compared the PSCA expression between BPH, PIN and PCa tissues and analyzed the correlations of PSCA expression level with pathological grade, clinical stage and progression to androgen-independence in PCa. RESULTS: In BPH and low grade PIN, PSCA protein and mRNA staining were weak or negative and less intense and uniform than that observed in high grade PIN (HGPIN) and PCa. Moderate to strong PSCA protein and mRNA expression were noted in 8 of 11 (72.7%) HGPIN and in 40 of 48 (83.4%) PCa specimens examined by IHC and ISH analyses, and their statistical significance was compared with BPH (20%) and low-grade PIN (22.2%) specimens (P < 0.05). The expression level of PSCA increased with a higher Gleason grade, advanced stage and progression to androgen-independence (P < 0.05). In addition, IHC and ISH staining revealed a high degree of correlation between PSCA protein and mRNA overexpression. CONCLUSIONS: Our data demonstrate that PSCA as a new cell surface marker is overexpressed in a majority of cases of human PCa. PSCA expression correlates positively with adverse tumor characteristics, such as increasing pathological grade (poor cell differentiation), worsening clinical stage and androgen-independence and speculatively with prostate carcinogenesis. PSCA may possess prognostic utility and may be a promising molecular target for diagnosis and treatment of PCa.     

EphA4 receptor, overexpressed in pancreatic ductal adenocarcinoma, promotes cancer cell growth

To isolate novel diagnostic markers and drug targets for pancreatic ductal adenocarcinoma (PDAC), we previously performed expression profile analysis of PDAC cells using a genome-wide cDNA microarray combined with laser microdissection. Among dozens of up-regulated genes identified in PDAC cells, we herein focused on one tyrosine kinase receptor, Eph receptor A4 (EphA4), as a molecular target for PDAC therapy. Immunohistochemical analysis validated EphA4 overexpression in approximately half of the PDAC tissues. To investigate its biological function in PDAC cells, we knocked down EphA4 expression by siRNA, which drastically attenuated PDAC cell viability. In concordance with the siRNA experiment, PDAC-derivative cells that were designed to constitutively express exogenous EphA4 showed a more rapid growth rate than cells transfected with mock vector, suggesting a growth-promoting effect of EphA4 on PDAC cells. Furthermore, the expression analysis for ephrin ligand family members indicated the coexistence of ephrinA3 ligand in PDAC cells with EphA4 receptor, and knockdown of ephrinA3 by siRNA also attenuated PDAC cell viability. These results suggest that the EphA4-ephrinA3 pathway is likely to be a promising molecular target for pancreatic cancer therapy.     

SPAG9 is overexpressed in human prostate cancer and promotes cancer cell proliferation

Sperm-associated antigen 9 (SPAG9) was recently reported to be overexpressed in several cancers and associated with the malignant behavior of cancer cells. However, the expression pattern of SPAG9 and its clinical significance in human prostate cancer have not been reported. In the present study, we analyzed SPAG9 expression in human prostate cancer tissues by immunohistochemistry and found that SPAG9 was overexpressed in 36.5 % of prostate cancer specimens. There was a significant association between SPAG9 overexpression and tumor stage (p?=?0.0020) and Gleason score (p?=?0.0377). Transfection of SPAG9 plasmid was performed in PC-3 cell line and siRNA knockdown was carried out in DU145 cells. Colony formation and MTT showed that SPAG9 overexpression promoted while siRNA knockdown inhibited prostate cancer cell proliferation. In addition, we found that SPAG9 could regulate cyclin D1 and cyclin E protein expression. In conclusion, SPAG9 is overexpressed in human prostate cancers and contributes to prostate cancer cell growth, possibly through cyclin protein regulation.     

Murine six-transmembrane epithelial antigen of the prostate, prostate stem cell antigen, and prostate-specific membrane antigen: prostate-specific cell-surface antigens highly expressed in prostate cancer of transgenic adenocarcinoma mouse prostate mice

To identify genes that are differentially up-regulated in prostate cancer of transgenic adenocarcinoma mouse prostate (TRAMP) mice, we subtracted cDNA isolated from mouse kidney and spleen from cDNA isolated from TRAMP-C1 cells, a prostate tumor cell line derived from a TRAMP mouse. Using this strategy, cDNA clones that were homologous to human six-transmembrane epithelial antigen of the prostate (STEAP) and prostate stem cell antigen (PSCA) were isolated. Mouse STEAP (mSteap) is 80% homologous to human STEAP at both the nucleotide and amino acid levels and contains six potential membrane-spanning regions similar to human STEAP. Mouse PSCA (mPsca) shares 65% homology with human PSCA at the nucleotide and amino acid levels. mRNA expression of mSteap and mPsca is largely prostate-specific and highly detected in primary prostate tumors and metastases of TRAMP mice. Both mSteap and mPsca map to chromosome 5. Another known gene coding for mouse prostate-specific membrane antigen (mPsma) is also highly expressed in both primary and metastatic lesions of TRAMP mice. These results indicate that the TRAMP mouse model can be used to effectively identify genes homologous to human prostate-specific genes, thereby allowing for the investigation of their functional roles in prostate cancer. mSteap, mPsca, and mPsma constitute new tools for preventative and/or therapeutic vaccine construction and immune monitoring in the TRAMP mouse model that may provide insights into the treatment of human prostate cancer.     

Search for new biomarkers of gastric cancer through serial analysis of gene expression and its clinical implications

Gastric cancer is one of the most common human cancers and is the second most frequent cause of cancer-related death in the world. Serial analysis of gene expression (SAGE) is a powerful technique to allow genome-wide analysis of gene expression in a quantitative manner without prior knowledge of the gene sequences. SAGE on 5 samples of gastric cancer with different histology and clinical stages have created large SAGE libraries of gastric cancer that enable us to identify new cancer biomarkers. Commonly up-regulated genes in gastric cancer in comparison with normal gastric epithelia included CEACAM6, APOC1 and YF13H12. By comparing gene expression profiles of gastric cancers at early and advanced stages, several genes differentially expressed by tumor stage were also identified, including FUS, CDH17, COL1A1 and COL1A2, which should be novel genetic markers for high-grade malignancy. Regenerating gene type IV (REGIV) is one of the most up-regulated genes in a SAGE library of a scirrhous-type gastric cancer. In vitro studies using RegIV-transfected cells revealed that RegIV is secreted by cancer cells and inhibits apoptosis, suggesting that RegIV may serve as a novel biomarker and therapeutic target for gastric cancer. Production of RNA aptamers could be a useful approach to establish a detection system in blood. A custom-made array, named Ex-STOMACHIP, consisting of 395 genes, including highly differentially expressed genes identified by our SAGE and other known genes related to carcinogenesis and chemosensitivity, is useful to study the molecular pathogenesis of gastric cancer and to obtain information about biological behavior and sensitivity to therapy in the clinical setting. Combined analyses of gene expression profile, genetic polymorphism and genetic instability will aid not only cancer detection, but also characterization of individual cancers and patients, leading to personalized medicine and cancer prevention.     

Insig2 is overexpressed in pancreatic cancer and its expression is induced by hypoxia

A hypoxic microenvironment is a characteristic feature of pancreatic cancer, and induces the expressions of various genes involved in malignant behaviors. Insulin‐induced gene 2 (Insig2) has recently been shown to be correlated with cellular invasion in colon cancer. However, there have been no reports regarding its expression in pancreatic cancer. In this study, we evaluated Insig2 mRNA expression and the biological function of Insig2 in pancreatic cancer. We measured Insig2 mRNA expression in cultured pancreatic cancer cell lines and invasive ductal carcinoma (IDC) cells, normal pancreatic epithelial cells, and pancreatic intraepithelial neoplasia cells obtained by laser‐capture microdissection. We also investigated the effects of Insig2‐targeting siRNAs on the cell proliferation and cell invasion of pancreatic cancer cell lines. All pancreatic cancer cell lines expressed Insig2 mRNA. The PANC‐1 and MIA PaCa‐2 pancreatic cancer cell lines showed >2‐fold higher Insig2 mRNA expression levels under hypoxic conditions (1% O₂) than under normoxic conditions (21% O₂). Cell proliferation was significantly decreased in SUIT‐2 cells and cell invasion was significantly decreased in SUIT‐2, Capan‐2, and CFPAC‐1 cells after transfection of the Insig2‐targeting siRNAs. In analyses of microdissected cells, cells from IDC tissues expressed significantly higher levels of Insig2 mRNA than normal pancreatic cells (P < 0.001) and pancreatic intraepithelial neoplasia cells (P = 0.082). In analyses of IDC cells, the levels of Insig2 mRNA expression were significantly higher in late‐stage patients than in early‐stage patients. The present data suggest that Insig2 is associated with the malignant potential of pancreatic cancer under hypoxic conditions. (Cancer Sci 2011; 102: 1137–1143)     

前列腺干细胞抗原在人胰腺癌神经浸润转移中作用的研究

目的探讨前列腺干细胞抗原(PSCA)在人胰腺癌神经浸润中的作用.方法80例胰腺癌手术标本切片行HE染色,观察记数小血管、小淋巴管、神经纤维浸润数.用鼠抗人PSCA单抗行免疫组化(Elivison二步法)染色,观察肿瘤细胞的阳性情况.结果肿瘤细胞神经浸润阳性与小血管、小淋巴管的浸润呈正相关(P<0.01);肿瘤细胞PSCA表达阳性(53例)与神经浸润阳性(66例)呈正相关(P<0.01);肿瘤的恶性程度与PSCA表达呈正相关;PSCA表达与小血管、小淋巴管浸润无关.结论PSCA可能是胰腺癌的特征性分子之一,可引导胰腺癌细胞向神经纤维趋化和黏附,即起"导航"和"停泊"作用.     

前列腺干细胞抗原在人胰腺癌神经浸润转移中作用的研究

目的探讨前列腺干细胞抗原(PSCA)在人胰腺癌神经浸润中的作用.方法80例胰腺癌手术标本切片行HE染色,观察记数小血管、小淋巴管、神经纤维浸润数.用鼠抗人PSCA单抗行免疫组化(Elivison二步法)染色,观察肿瘤细胞的阳性情况.结果肿瘤细胞神经浸润阳性与小血管、小淋巴管的浸润呈正相关(P<0.01);肿瘤细胞PSCA表达阳性(53例)与神经浸润阳性(66例)呈正相关(P<0.01);肿瘤的恶性程度与PSCA表达呈正相关;PSCA表达与小血管、小淋巴管浸润无关.结论PSCA可能是胰腺癌的特征性分子之一,可引导胰腺癌细胞向神经纤维趋化和黏附,即起导航和停泊作用.     

前列腺干细胞抗原在人胰腺癌神经浸润转移中作用的研究

目的探讨前列腺干细胞抗原(PSCA)在人胰腺癌神经浸润中的作用.方法80例胰腺癌手术标本切片行HE染色,观察记数小血管、小淋巴管、神经纤维浸润数.用鼠抗人PSCA单抗行免疫组化(Elivison二步法)染色,观察肿瘤细胞的阳性情况.结果肿瘤细胞神经浸润阳性与小血管、小淋巴管的浸润呈正相关(P<0.01);肿瘤细胞PSCA表达阳性(53例)与神经浸润阳性(66例)呈正相关(P<0.01);肿瘤的恶性程度与PSCA表达呈正相关;PSCA表达与小血管、小淋巴管浸润无关.结论PSCA可能是胰腺癌的特征性分子之一,可引导胰腺癌细胞向神经纤维趋化和黏附,即起"导航"和"停泊"作用.