Invasion and
metastasis are the critical steps in
cancer progression that lead to death from this disease. Intense investigation into the underlying mechanisms of
metastasis has revealed a complex set of signaling pathways that regulate the process. Since the mid-1980s, it has been demonstrated that the Rho family of
proteins plays a major role in these pathways.
Proteins that regulate Rho, including
guanine nucleotide exchange factors,
GTPase-activating proteins, and Rho
GDP dissociation inhibitors (RhoGDIs), have also been shown to contribute to
cancer progression. Among this group of Rho-regulating
proteins is
RhoGDI2 (
RhoGDIbeta/LyGDI/GDID4/RabGDIbeta). Our laboratory initially identified
RhoGDI2 as a
metastasis suppressor due to its differential expression between metastatically capable and poorly metastatic
bladder cancer cell lines. Over the subsequent years, in vivo and in vitro systems have been used to model steps in the metastatic cascade and to test how the expression of
RhoGDI2 affected those processes. This chapter describes several of the more significant methods used to investigate the role of
RhoGDI2 in
bladder cancer invasion and
metastasis. These methods include an in vitro assay for invasion using bladder organ cultures, lung
metastasis assays in immunocompromised murine hosts, polymerase chain reaction-based quantification of metastatic burden, and derivation of increasingly metastatic cell lines.