GPR56 is a member of the adhesion GPCR family. Consistent with their predicted roles as adhesion receptors, several adhesion GPCRs have been implicated in cancer development [8
]. However, all of these studies used xenograft cancer models, and whether those adhesion GPCRs play similar roles in endogenous cancer progression has not been investigated. We report here our analyses on the involvement of GPR56 in endogenous cancer progression using Gpr56−/−
mice and transgenic cancer models. We found that GPR56 plays diverse roles in endogenous cancer progression. GPR56 suppressed prostate cancer development in a background-dependent manner; it may have some effects on the onset of mammary tumors in MMTV-PyMT mice, but did not appear to affect the melanoma progression in Ink4a/Arf −/− tyr-Hras
GPR56 was predicted to inhibit endogenous cancer progression, based our previous findings using xenograft models of melanoma [8
]. Consistent with this prediction, the expression levels of GPR56 were inversely correlated with the metastatic potential of prostate cancer cell lines. In addition, endogenous prostate cancer progression in the TRAMP model with a mixed genetic background was significantly enhanced in Gpr56−/−
mice compared with that in wild-type mice, suggesting that GPR56 suppresses prostate cancer progression in TRAMP mice.
The effects of GPR56 on prostate cancer progression in TRAMP mice appeared to be background-dependent, since effects on cancer development were only observed on a mixed genetic background but not on the pure C57BL/6 background. Several possibilities might account for this difference. First, the FvB/N background may be over-represented in the Gpr56+/− and Gpr56−/− mice (relative to the Gpr56+/+ mice), resulting in an enhanced prostate cancer progression that is independent of GPR56. Alternatively, GPR56 might cooperate with a strain-specific modifier in the mixed genetic background that is not present in the C57BL/6 background to inhibit prostate cancer progression. Finally, the lack of effects on prostate cancer progression in C57BL/6 mice could also be due to developmental defects in Gpr56−/−; C57BL/6 mice. Our preliminary analyses suggest that Gpr56−/−; C57BL/6 mice are defective in testis development (unpublished data). Testis is the major organ that secretes testosterones, which are essential for prostate development and prostate cancer progression. Its dysfunction might therefore impede the otherwise enhanced prostate cancer progression in Gpr56−/− mice.
The mammary tumor onset in Gpr56+/− mice was significantly delayed relative to Gpr56+/+ mice, suggesting that GPR56 might promote mammary tumor onset in this model. However, no significant difference in tumor onsets was observed between Gpr56−/− mice and either Gpr56+/− or Gpr56+/+ mice, and considerable data scattering was present for all three groups, indicating that the effects of GPR56 on mammary tumor onset may be marginal and thus could be readily masked by the complexity of mixed genetic background.
In both TRAMP and MMTV-PyMT models, GPR56 expression was induced in the cancerous lesions relative to the adjacent normal tissues. Increased expression of GPR56 in primary tumors relative to normal tissues in human cancer has been reported [16
] and was frequently shown in analyses from microarray data (www.oncomine.org
). The most straightforward explanation is that GPR56 expression is induced during oncogenesis, although Gpr56
has not been shown to be a direct target of any oncogene. An alternative explanation is that GPR56 is expressed in a minor population of cells in normal prostates and breasts, which are amplified during cancer progression. This minor population might represent cancer-initiating cells, i.e., cancer stem cells. GPR56 has been speculated to play roles in stem cells of normal tissues, since its expression was believed to be up-regulated in both neuronal and hematopoietic stem cells [29
]. It is tempting to connect the potential roles of GPR56 in normal stem cells with its roles in cancer stem cells; however, more careful studies will be needed to address this issue.
The increased expression of GPR56 in poorly differentiated prostate tumors was in contrast to its down-regulation in highly metastatic prostate cancer cell lines. How this occurs needs to be investigated further. It is possible that GPR56 plays distinct roles at different stages of prostate cancer progression: it might promote prostate cancer progression in the primary tumors, but inhibit the establishment of metastases. Or, GPR56 may function differently in prostate xenografts than in endogenous prostate tumors.
In contrast to our conclusions from the xenograft models, GPR56 did not affect melanoma progression in Ink4a/Arf−/− tyr-Hras mice. This might be partly due to the characteristics of the Ink4a/Arf−/− tyr-Hras model. We observed significant variation in tumor onsets and progression among the Ink4a/Arf−/− try-Hras mice. This intrinsic variation might mask the effects of GPR56 on melanoma progression in these mice. In addition, the Ink4a/Arf−/− try-Hras mice die from lymphoma at various ages, making it difficult to assess the roles of GPR56 on melanoma at any fixed time point or at later developmental stages (during metastasis, for example).
Taken together, it is apparent from our study that GPR56 plays varying roles on spontaneous cancer progression. Although it is too early to predict the mechanism of its regulation on cancer, multiple lines of evidence suggest that GPR56 functions as an adhesion receptor and interacts with ECM directly. Its N-terminus binds to TG2, a major crosslinking enzyme in the extracellular matrix [8
], and Gpr56−/−
mice possess defects in basement membrane assembly in both cerebrum and cerebellum [13
]. However, we have not observed any overt differences in ECM deposition between Gpr56−/−
tumors and Gpr56+/−
or wild-type tumors in any of the systems analyzed here (Supplementary Figures 1–3
). Perhaps GPR56 plays additional roles aside from ECM assembly during cancer progression. Its binding partner, TG2, affects fibronectin-mediated cell adhesion through integrins in an enzyme-independent manner [31
]. GPR56 might affect cell adhesion through TG2 and its associated factors. In addition, GPR56 was reported recently to inhibit neuronal cell migration by activating RhoA [33
]. Cell migration and regulation of Rho GTPases are integral processes during cancer progression [34
]. Their regulation by GPR56 might explain the alterations of cancer progression we observed in Gpr56−/−