An aggressive RhoC phenotype is associated with relapse after external beam radiation therapy of a prostate cancer xenograft model.

Abstract

e14748 Background: Relapsed treatment resistant prostate cancer remains the second most common cause of male mortality in the western world. New therapies for therapy resistant prostate cancer depend on defining tumor cell sub-populations capable of surviving, reinitiating, and sustaining net growth of the cancer. In this study we focused on RhoC expression in a prostate cancer model after external beam radiation therapy (EBRT). RhoC has earlier been implicated in metastatic processes in most solid tumors. Methods: In mice, established s.c. LNCaP xenografts (n = 12) were irradiated, the beam collimated with lead to restrict it to the xenograft, with X-rays (200 kV, 1 Gymin-1, Gulmay Medical) for an absorbed dose of 15 Gy. Tumor tissue was collected after seven days or after 3-4 weeks when recurrent growth was seen. RhoC, AR, CD166 and Ki67 immunohistochemistry in tumor sections was visualized by using DAB or Peroxidase Green and nuclear red or haematoxylin blue as counter stain. Sections stained for RhoC and Ki67 were quantified with the Halo software (v2.3.2089.34) using the Indica Labs – cytonuclear v1.6 algorithm. The intensity of the RhoC staining was also quantified in a separate set of sections only stained for RhoC. Results: The RhoC staining intensity was significantly higher in relapsed xenografts (P = 0.003). Also, a significant difference in percentage of RhoC expressing cells in the two populations (P = 0.045) with 40.6 ± 5.8% (mean ± SEM) in relapsed tumors as compared to 15.0 ± 8.1% after 7d. The percentage of cells expressing Ki67 was not significantly different (P = 0.477). However, the percentage of double positive cells (Ki67, RhoC) was significantly higher in treated relapsed xenografts (P = 0.042). Further, a subpopulation of RhoC-positive cells were positive for AR and the stem cell marker CD166. Conclusions: Here we show that a subpopulation of proliferating RhoC-positive cells are more prevalent in relapsing PCa xenografts after EBRT, suggesting an aggressive phenotype that is associated with both proliferation and invasiveness. Another subpopulation of RhoC cells was found to express the prostate stem cell marker CD166, suggesting that pathways and properties involved in cellular self-renewal and invasiveness may be responsible for survival and recurrent growth in relapsed prostate tumors.