H2 relaxin overexpression increases in vivo prostate xenograft tumor growth and angiogenesis

Abstract

Our study reports a preliminary investigation into the role of human H2 relaxin in prostate tumor growth. A luciferase-expressing human prostate cancer cell line, PC-3, was generated and termed PC3-Luc. PC3-Luc cells were transduced with lentiviral vectors engineering the expression of either enhanced green fluorescent protein (eGFP) or both H2 relaxin and eGFP in a bicistronic format. These transduced cells were termed PC3-Luc-eGFP and PC3-Luc-H2/eGFP, respectively. To gauge effects, PC3-Luc-H2/eGFP and PC3-Luc-eGFP cells were injected into NOD/SCID mice and monitored over 6 weeks. PC-3 tumor xenografts overexpressing H2 relaxin exhibited greater tumor volumes compared to control tumors. Circulating H2 relaxin levels in sera increased with the relative size of the tumor, with moderately elevated H2 relaxin levels in mice bearing PC3-Luc-H2/eGFP tumors compared to PC3-Luc-eGFP tumors. Zymographic analysis demonstrated that proMMP-9 enzyme activity was significantly downregulated in H2 relaxin-overexpressing tumors. An advanced angiogenic phenotype was observed in H2 relaxin-overexpressing tumors indicated by greater intratumoral vascularization by immunohistochemical staining of endothelial cells with anti-mouse CD31. Moreover, PC3-Luc-H2/eGFP tumors exhibited increased VEGF transcript by reverse-transcription PCR, compared to basal levels in control animals. Taken together, our study provides the first account of a potential role of H2 relaxin in prostate tumor development. ' 2005 Wiley-Liss, Inc.Key words: PC-3; lentivirus; luciferase; VEGF; MMP-9; H2; prorelaxin In recent years, there has been increasing evidence that suggests a role of relaxin in carcinogenesis. Peptide hormones of the relaxin family are reported to be upregulated in a number of neoplasias, including thyroid, 1 breast, 2 gastrointestinal 3 and the reproductive system. 4,5 Many of the molecules characterized as downstream effectors of relaxin's physiological roles in the cardiovascular system and as agents involved in remodeling of connective tissue have also been linked with tumor growth and sustainability. 6 These molecules, including VEGF and NOS, for example, are established angiogenic agents that promote neovascularization and enhanced blood flow carrying oxygen and cellular nutrients to tumors. 7 The regulation of matrix metalloproteinases (MMPs) is the mechanism considered for relaxin's role in connective tissue remodeling during mammary gland involution, softening of reproductive tissues and dilation of the birth canal. 8 However, the regulation of these enzymes by relaxin have also been suggested as a mechanism for its role in facilitating cancer cell migration and invasion. [9][10][11] Although the biological significance of relaxin in the male remains elusive, the prostate gland appears to be relaxin's main source. 8 In the human, there are 3 relaxin alleles encoding 3 hormones, named H1, H2 and H3. While H2 and H3 relaxins are expressed in a number of tissues, in males their predominant expression is found in the prostate and bra...

Relaxin Enhances S100A4 and Promotes Growth of Human Thyroid Carcinoma Cell Xenografts

“…The polypeptide hormone H2-relaxin (RNL2) is increased in human carcinoma and is associated with increased migratory capacity of carcinoma cells of the breast (37,38), prostate (39), and thyroid (10). We showed that RLN2 enhances cell motility and matrix invasion of human thyroid carcinoma cells and this action requires functional RXPF1 signaling (10).…”

Suppression of relaxin receptor RXFP1 decreases prostate cancer growth and metastasis

“…We have studied the tumorigenic effect of RLN, a small peptide hormone produced both in the normal prostate and in prostate cancer (Figueiredo et al 2005, Silvertown et al 2006, Thompson et al 2006, Vinall et al 2006, Feng et al 2007, Liu et al 2008. Previously it has been shown that the RLN expression was increased in aggressive metastatic disease (Figueiredo et al 2005, Thompson et al 2006, Feng et al 2007, and that the stimulation of prostate cancer cells with RLN accelerated their invasiveness, adhesion, survival, and decreased cell apoptosis (Feng et al 2007).…”

“…Despite significant variations in the amino acid sequence of different mammalian RLN peptides, they all efficiently cross-activate human RXFP1 (Sherwood 2004, Bathgate et al 2006. Recently, it was demonstrated that an analog of RLN peptide with a mutated receptor-binding domain has a moderately suppressive effect on prostate tumor when stably or transiently expressed in prostate cancer cells (Silvertown et al 2006). With apparent limitations of an overexpression approach in mind, we decided to investigate the effect of direct RXFP1 knockdown in prostate cancer cells.…”

“…The overall survival was shorter in patients with increased expression of RLN , Feng et al 2007. The in vitro invasiveness of carcinoma cells was significantly increased on incubation with RLN or in cells transfected with RLN expression constructs , Silvertown et al 2006, Feng et al 2007). The stimulating effect on growth and angiogenesis of lentiviral-delivered RLN in the prostate carcinoma cell line PC3 was recently demonstrated in vivo (Silvertown et al 2006).…”

“…The in vitro invasiveness of carcinoma cells was significantly increased on incubation with RLN or in cells transfected with RLN expression constructs , Silvertown et al 2006, Feng et al 2007). The stimulating effect on growth and angiogenesis of lentiviral-delivered RLN in the prostate carcinoma cell line PC3 was recently demonstrated in vivo (Silvertown et al 2006). Furthermore, we have demonstrated that the survival of transgenic adenocarcinoma of mouse prostate (TRAMP) mice with transgenic RLN overexpression was decreased (Feng et al 2007).…”