20 or anti p53. Statistical analysis All data were expressed as mean SD. The means were compared between groups with one way analysis of vari ance and the Student t test. p value 0. 05 was considered significant. till Results High levels of A20 and low levels of p53 in colon cancer Immune deregulation plays a role in the pathogenesis of cancer, recent reports indicate that A20 contributes to immune regulation. Whether A20 is involved in the pathogenesis of colon cancer is unclear. Thus, we collected 88 colon cancer tissue from the clinic. As shown by qRT PCR and Western blotting, the A20 levels were higher in colon cancer tissue than that in the IBS colon tissue. The expression of p53 was also assessed in all the samples. The results showed that the expres sion of p53 was significantly suppressed in colon cancer tissue as compared with controls.

When reviewed the disease history, we noted that 32 88 cases also suffered from colon polyp. A correl ation assay was performed with the A20 levels of the colon cancer tissue and their polyp history. The results showed that the A20 levels were positively correlated with the polyp history. By immunohistochemistry, we observed the expression of A20 was mainly localized in the epithelial cells, which was much stronger in cancerous tissue than that from the IBS colon mucosa. Levels of A20 and p53 are correlated with recurrence of colon polyp Colon polyps have tendency to develop into colon cancer. We then recruited 136 patients with non cancer colon polyp at the first diagnosis. The colon polyps were removed under colonoscopy.

The levels of A20 in the polyp epithelium were assessed by ELISA. The results showed that the levels of A20 in the polyp epi thelium were higher, p53 levels were lower, as com pared to controls. All the patients were followed up three times a year. The follow up data showed that the polyp recurred in 59 136 pa tients. Their colon polyps were removed under colonoscopy again. The polyp epithelium was also examined by ELISA for the levels of A20 and p53. The results showed the levels of A20 were higher, p53 were lower, in the recurred polyp than the levels in the original polyp. We then performed a correlation assay with the levels of A20 and p53 in the polyp and the recurrence of the original 136 patients. A significant positive correlation was identified between the levels of A20 or p53 and the recurrence of colon polyps.

We further analyzed the relation between the phenotypes of the colon Carfilzomib polyps and the levels of A20 U0126 MAPK and p53. As shown by ELISA data, the levels of A20 were higher, p53 were lower, in adenomas and hyperplastic polyps than the inflammatory polyps. Colon polyps with high levels of A20 show tumorigenic tendency The 136 patients with colon polyp were followed up for 3 6 years. During this period, 45 patients were diagnosed colon cancer. We analyzed the cancerous rate of the pathological phenotypes of the colon polyps. The results showed 4 79 inflammatory colon polyps, 14 21 hyperpl

treated with do ycycline in drinking water. Within three weeks, all the 31 mice injected with control cells gave rise to tumors with a mean diameter of considering 8 mm. In contrast, 38% of mice injected with p130Cas silenced cells did not give rise to detectable tumors and the remaining 45 mice developed small tumors, with a mean diameter of 2 mm. Interestingly, p130Cas silencing was sufficient to halt tumor growth in mice that have already developed tumors with a diameter of 3 to 4 mm. Indeed, by adding do ycycline to drinking water two weeks after cell injection, p130Cas silenced tumors regressed, becoming undetectable by palpation within two to three weeks, while control tumors contin ued to grow. Consistently, after do ycycline withdrawal p130Cas silenced tumors resumed growing.

These data strengthen the in vivo rele vance of p130Cas as a major regulator of the tumorigenic properties of mesenchymal breast cancer cells. We have previously shown that intranipple injection of p130Cas siRNAs in the mammary gland of Balb c NeuT mice sig nificantly decreases the number of cancer lesions com pared to glands injected with control siRNAs, with a significant downregulation of proliferative and survival pathways. Overall these data indicate that tight modula tion of p130Cas levels can affect in vivo tumor properties of distinct breast cancer subtypes, implying the compel ling need of studying its transcriptional regulation in nor mal mammary epithelial cells and in tumors in the near future.

Hemato ylin and eosin staining of tumor sections showed that tumors derived from p130Cas silenced cells consisted of cells with an epithelial like shape, while the control tumors presented elongated, mesenchymal cells. Moreover, immunohistochemis try analysis indicated that tumors from p130Cas silenced cells were characterized by decreased vascularization and proliferation, and increased apoptosis. Western blot analysis of p130Cas silenced tumors showed a significant in vivo p130Cas silencing together with Co 2 downregulation, compromised activation of c Src and JNK kinases and decreased e pression of Cyclin D1. A parallel downregu lation of Snail, Cilengitide Slug and Twist e pression was also detected, indicating that p130Cas silencing compromises tumor growth through inhibition of cell signaling controlling cell cycle progres sion and the acquirement of epithelial like features.

In parallel, syngeneic mice were subcutaneously injected with 105 Co 2 silenced or control A17 cells and treated with do ycycline in drinking water. As shown in Figure 3D, while mice injected with control cells gave rise to tumors with a mean diameter of 10 mm within si weeks, mice injected with Co 2 silenced cells give rise to barely detectable Bosutinib cost tumors. Taken together these data show that p130Cas Co 2 a is controls in vivo survival and proliferative pathways of mesenchymal breast can cer cells and silencing of either p130Cas or Co 2 is suf ficient for switching cells to an epithelial state leading to impaired