Background and purpose Hypoxia is a hallmark of solid cancers and

Background and purpose Hypoxia is a hallmark of solid cancers and associated with metastases and treatment failure. lost cell-cell contacts. Expression of epithelial markers such as E-cadherin decreased whereas mesenchymal markers such as vimentin and N-cadherin increased. Combining hypoxia with TGFβ or EGFRvIII expression lead to more rapid and pronounced EMT-like phenotype. Interestingly E-cadherin expression and the mesenchymal appearance were reversible upon reoxygenation. Mesenchymal conversion and E-cadherin loss were associated with radioresistance. Conclusions Our findings describe a mechanism by which the tumor microenvironment may contribute to tumor radioresistance via E-cadherin loss and EMT. < 0.05 two-way Anova) than cells seeded at high density at all doses tested (Fig. 4a). Fig. 4 Cells with mesenchymal phenotype show increased radioresistance (a) Clonogenic survival of MCF7 cells seeded at different densities. (b) Western blot for E-cadherin Omecamtiv mecarbil of MCF7 lysates made form cell cultures grown at different densities. (c) Immunofluorescent ... We reasoned that if cells with low cell Rabbit Polyclonal to BAIAP2L1. surface E-cadherin are more resistant to radiation reversion to the epithelial phenotype by reintroduction of E-cadherin may sensitize cells. To test this hypothesis we compared the radiation response of the mesenchy-mal MDA-MB 231 metastatic breast cancer cells with an Omecamtiv mecarbil epithelial revertant generated by expressing E-cadherin in MDA-MB 231 (RevCdh1) cells. Parental MDA-MB 231 cells show undetectable E-cadherin expression (Fig. 4e) and appear with a mesenchymal morphology. Expression of E-cadherin induced a marked conversion to a more epithelial morphology (compare left and right panels in Fig. 4f). Next we tested the radiation sensitivity of two independently generated MDA-MD 231 cell lines in which we reconstituted E-cadherin expression. MDA-MB 231 cells expressing E-cadherin were significantly more sensitive than their parental E-cadherin negative MDA-MD 231 cells as determined by their clonogenic survival after irradiation at increasing doses of γ-irradiation (< 0.001 two-way Anova) (Fig. 4d). Discussion In the present study we have shown that various stimuli emanating from tumor cells or their microenvironment can induce Omecamtiv mecarbil mesenchymal conversion of normal and cancerous epithelial cells. Interestingly our data indicate that Omecamtiv mecarbil combining these signals results in a more pronounced EMT-like induction. Moreover EMT-like transformation by the microenvironment is reversible upon reoxygenation. We have correlated these findings with changes in the expression of E-cadherin a cell adhesion protein implicated in EMT and associated with tumor progression in many human cancers. We found that breast cancer cells that express E-cadherin were more sensitive to radiation than their counterparts without E-cadherin. Thus changes in E-cadherin expression in tumors induced by changes in the microenvironment such as hypoxia and reoxygenation may contribute to the intrinsic sensitivity of tumor cells to radiotherapy. Our data are in line with previous studies that highlight the contribution of hypoxia in inducing phenotypic changes in cells. This induction has been linked to various mechanisms including the involvement of uPAR [24] activation of the PI3K/Akt pathway [7] inactivation of GSK3β or the production of ROS [25]. Other reports have indicated a role for Snail [4] and we and others have shown Twist1 to be induced upon hypoxia [13 23 Here we confirm that hypoxia attenuates E-cadherin expression in epithelial cells as previously observed Omecamtiv mecarbil by others [11 26 Since the tumor microenvironment is highly complex [10] we reasoned that other factors inherently present in solid tumors would also contribute to this phenotypic switch. Some of these signals might function as autocrine loops in malignancy cells such as manifestation of EGFRvIII shown to be indicated in malignant Omecamtiv mecarbil mind lung and breast tumors [27 28 We have previously demonstrated that EGFRvIII contributes to improved malignancy and survival under hypoxic conditions [22]. Others have reported that EGFRvIII manifestation disrupts adherens junctions [3] and that silencing EGFRvIII reduces manifestation of factors involved in EMT [29]. Our present data are consistent with both these observations and display additionally that the effect is definitely even more.