Previous stud ies have shown that when MCF10DCIS cells are injected into the mammary fat pad of immunodeficient nude mice, tumors develop within 2 3 weeks. These tumors faithfully http://www.selleckchem.com/products/BI6727-Volasertib.html recapitulate the human comedo DCIS condition, with the basement membrane limiting duct like structure being comprised of an outer myoepithelial layer, an inner layer of luminal epithelial cells, and a cen tral necrotic lumen. We chose to use sub cutaneous injections instead of orthotopic or intraductal methods, as previous work by Hu et al. showed that the progression and phenotype of the MCF10DCIS tumors grown subcutaneously in the mammary fat pad were highly similar to human high grade comedo DCIS tumors.

In our study, we found that PADI2 protein expression was restricted to the luminal epithelium of the duct like structures in the MCF10DCIS xenografts, and was not observed in the stromal tissue or the necrotic core. At the subcellu lar level, PADI2 appears to be expressed in both the cytoplasmic and nuclear compartments of luminal epi thelial cells. This observation sup ports our recent findings that PADI2 can be targeted to the nucleus of both human normal mammary tissue and breast cancer cells and regulate gene activity via citrullination. Next, we examined whether the observed correlation between PADI2 and HER2 ERBB2 expression also occurred in vivo. We found that both HER2 ERBB2 and PADI2 were expressed within the luminal epithelium of MCF10DCIS tumors. Inter estingly, a previous report by Behbod et. al. found low levels of HER2 ERBB2 in MCF10DCIS tumors that were grown intraductally.

The disparity between this data and our data may be due to differences in the microenviron ment. We then quantified PADI2 mRNA in the MCF10DCIS xenografts by qRT PCR, and found that PADI2 levels were significantly higher in the tumors when compared to monolayer cultures. We also car ried out immunofluorescence analysis of these tumors to examine PADI2 intratumoral localization, and found that PADI2 protein expression appears entirely limited to cytokeratin positive luminal epithelial cells, while no detect able PADI2 signal was observed in the p63 positive myoe pithelial cells. Treatment of MCF10DCIS xenografts with Cl amidine suppresses tumor growth Anacetrapib Given the inhibitory effects of Cl amidine on MCF10 DCIS monolayer and spheroid growth, we next tested whether the treatment of mice with this inhibitor would suppress the growth of MCF10DCIS derived tu mors. For this study, mouse fat pads were injected with MCF10DCIS cells and the tumors were al lowed to establish and grow for 2 weeks as described previously. Mice were randomly assigned into treatment or control groups and administered daily intra peritoneal injections of either Cl amidine or vehicle.