ABSTRACT
BACKGROUND: Fibulins, encoded by FBLN genes, are extracellular matrix proteins influencing cell adhesion and migration. Altered expression of fibulins is associated with progression of several cancer types, but has not been studied in prostate cancer. METHODS: Expression of FBLN1 (major splice forms C and D), FBLN4, FBLN5, SPOCK1, and TENC was compared between 47 prostate cancer samples and 13 benign prostatic tissues by quantitative RT-PCR. Fibulin-1 and fibulin-5 expression was studied by immunohistochemistry. Effects of androgens and the DNA methylation inhibitor 5-aza-2'-deoxycytidine on fibulin expression were investigated in different prostate cancer cell lines. RESULTS: Our recent microarray analysis suggested downregulation of three fibulins, FBLN1, FBLN4, and FBLN5, in prostate cancer, while two further ECM genes, SPOCK1 (testican) and TENC (tenascin C), appeared upregulated or unchanged. These observations were corroborated by quantitative RT-PCR. Accordingly, FBLN1 and FBLN4 were weakly expressed in carcinoma lines compared to normal prostate epithelial cells (PrECs). Only FBLN4 was induced by 5-aza-2'-deoxycytidine, but its promoter was unmethylated. Androgen did not affect expression of FBLN genes. The FBLN1C and FBLN1D splice forms were coordinately expressed. Fibulin-1 protein was weakly detectable in benign PrECs, but tended to accumulate in cancer cells. Fibulin-5 was predominantly located in the stroma with a strong gradient from the periurethral to the peripheral zone, and lost in cancers. CONCLUSIONS: Three FBLN genes are significantly downregulated in prostate cancer, whereas SPOCK1 is often upregulated. FBLN5 downregulation fits its postulated anticancerous function, whereas FBLN1 and FBLN4 behave different than in certain other cancers.
Subject(s)
Extracellular Matrix Proteins/genetics , Gene Expression Regulation, Neoplastic , Prostatic Neoplasms/genetics , Aged , Azacitidine/analogs & derivatives , Azacitidine/pharmacology , Calcium-Binding Proteins/biosynthesis , Calcium-Binding Proteins/genetics , Calcium-Binding Proteins/metabolism , Cell Line, Tumor , DNA Methylation/drug effects , Decitabine , Down-Regulation , Extracellular Matrix Proteins/biosynthesis , Extracellular Matrix Proteins/metabolism , Gene Dosage , Gene Expression Profiling/methods , Humans , Immunohistochemistry , Male , Middle Aged , Prostatic Neoplasms/metabolism , Proteoglycans/biosynthesis , Proteoglycans/genetics , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
BACKGROUND: Alterations of chromosome 8 and hypomethylation of LINE-1 retrotransposons are common alterations in advanced prostate carcinoma. In a former study including many metastatic cases, they strongly correlated with each other. To elucidate a possible interaction between the two alterations, we investigated their relationship in less advanced prostate cancers. RESULTS: In 50 primary tumor tissues, no correlation was observed between chromosome 8 alterations determined by comparative genomic hybridization and LINE-1 hypomethylation measured by Southern blot hybridization. The discrepancy towards the former study, which had been dominated by advanced stage cases, suggests that both alterations converge and interact during prostate cancer progression. Therefore, interaction analysis was performed on microarray-based expression profiles of cancers harboring both alterations, only one, or none. Application of a novel bioinformatic method identified Gene Ontology (GO) groups related to innate immunity, cytoskeletal organization and cell adhesion as common targets of both alterations. Many genes targeted by their interaction were involved in type I and II interferon signaling and several were functionally related to hereditary prostate cancer genes. In addition, the interaction appeared to influence a switch in the expression pattern of EPB41L genes encoding 4.1 cytoskeleton proteins. Real-time RT-PCR revealed GADD45A, MX1, EPB41L3/DAL1, and FBLN1 as generally downregulated in prostate cancer, whereas HOXB13 and EPB41L4B were upregulated. TLR3 was downregulated in a subset of the cases and associated with recurrence. Downregulation of EPB41L3, but not of GADD45A, was associated with promoter hypermethylation, which was detected in 79% of carcinoma samples. CONCLUSION: Alterations of chromosome 8 and DNA hypomethylation in prostate cancer probably do not cause each other, but converge during progression. The present analysis implicates their interaction in innate immune response suppression and cytoskeletal changes during prostate cancer progression. The study thus highlights novel mechanisms in prostate cancer progression and identifies novel candidate genes for diagnostic and therapeutic purposes. In particular, TLR3 expression might be useful for prostate cancer prognosis and EPB41L3 hypermethylation for its detection.
