In Situ Evaluation of the Role of the Small GTPase Rac3 in Breast Cancer

摘要

To test the hypothesis that the signaling protein Rac3 is critical for the initiation of breast cancer metastasis, we created stable fluorescently- tagged breast cancer cell lines that express active and inactive forms of Rac3 and its close homolog Rac1 in a highly metastatic variant of the MDA-ME-435 metastatic breast cancer cell line. Dominant active forms of Rac3 and Racl were stably expressed in a low metastatic variant of MDA-MB-435. High metastatic cell lines expressing dominant negative Racl or Rac3 demonstrated similar reduction in invasion and migration in vitro compared to controls. Cell lines expressing a dominant negative Rac1 or Rac3 did not show differences in proliferation; however, when implanted in nude mice, both cell lines produced smaller primary tumors that demonstrated reduced metastasis compared to controls. Expression of dominant active Racl or Rac3 in a low metastatic breast cancer cell variant resulted in enhanced migratory and invasive properties. The metastatic efficiency of the Rac1 or Rac3 mutant cells was analyzed in the nude mouse model of experimental metastasis. Mammary tumors from cells expressing Rac1 or Rac3 dominant active mutations were more metastatic while the mammary tumors from cells expressing dominant negative Rac1 or Rac3 were less metastatic when compared to controls. To provide a direct assessment of these mutant cell lines, we developed a fluorescence illumination system to specifically image fluorescence breast cancer progression in live mice.