Limits...
Loss of miR-200c up-regulates CYP1B1 and confers docetaxel resistance in renal cell carcinoma.

Chang I, Mitsui Y, Fukuhara S, Gill A, Wong DK, Yamamura S, Shahryari V, Tabatabai ZL, Dahiya R, Shin DM, Tanaka Y - Oncotarget (2015)

Bottom Line: Additionally, miR-200c, which is significantly down-regulated in RCC regulates CYP1B1 expression and activity.These results suggest that CYP1B1 up-regulation mediated by low miR-200c is one of the mechanisms underlying resistance of RCC cells to docetaxel.Therefore, expression of CYP1B1 and miR-200c in RCC may be useful as a prediction for docetaxel response.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery and Division of Urology, Veterans Affairs Medical Center, San Francisco, California, United States of America.

ABSTRACT
Despite high protein expression and enzymatic activity of cytochrome P450 1B1 (CYP1B1) in renal cell cancer (RCC), its functional significance has not been elucidated. Here we explored the functional role and regulatory mechanism of CYP1B1 in RCC. Reduction of CYP1B1 levels fail to prevent in vitro tumorigenicity such as proliferation, apoptosis, and cell cycle progression of RCC cells. Moreover, the expression levels are not associated with tumor type, stage, Fuhrman grade and 5-year survival probability after surgery. Instead, alteration of CYP1B1 expression regulates the chemosensitivity of RCC cells to docetaxel suggesting its critical contribution to the chemoresistance. Additionally, miR-200c, which is significantly down-regulated in RCC regulates CYP1B1 expression and activity. An inverse association was also observed between the expression levels of miR-200c and CYP1B1 protein in RCC tissues. Finally, alteration of miR-200c levels affects the chemosensitivity of RCC cells. Restoration of docetaxel resistance by exogenous expression of CYP1B1 in miR-200c-over-expressing cells indicates that CYP1B1 is a functional target of miR-200c. These results suggest that CYP1B1 up-regulation mediated by low miR-200c is one of the mechanisms underlying resistance of RCC cells to docetaxel. Therefore, expression of CYP1B1 and miR-200c in RCC may be useful as a prediction for docetaxel response.

No MeSH data available.


Related in: MedlinePlus

MiR-200c regulates docetaxel resistance of RCC cells(A and B) After miR-200c precursor transfection, A498 cells were treated with docetaxel (5 μM) for 72 hrs. Apoptotic cell death was measured with annexin V-FITC/7-AAD staining (A) and cell survival was analyzed by MTS assay (B). **P < 0.01; ***P < 0.001 (C and D) After miR-200c inhibitor transfection, 786-O cells were treated with docetaxel (5 μM) for 72 hrs. Apoptotic cell death was measured with annexin V-FITC/7-AAD staining (C) and cell survival was analyzed by MTS assay (D). *P < 0.05; **P < 0.01 (E-G) After transfection of miR-200c precursor with either control or CYP1B1 plasmid, A498 cells were treated with docetaxel (5 μM) for 72 hrs. CYP1B1 protein expression was determined by Western blot (E), apoptotic cell death was measured with annexin V-FITC/7-AAD staining (F) and cell survival was analyzed by MTS assay (G). *P < 0.05; **P < 0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4480715&req=5

Figure 6: MiR-200c regulates docetaxel resistance of RCC cells(A and B) After miR-200c precursor transfection, A498 cells were treated with docetaxel (5 μM) for 72 hrs. Apoptotic cell death was measured with annexin V-FITC/7-AAD staining (A) and cell survival was analyzed by MTS assay (B). **P < 0.01; ***P < 0.001 (C and D) After miR-200c inhibitor transfection, 786-O cells were treated with docetaxel (5 μM) for 72 hrs. Apoptotic cell death was measured with annexin V-FITC/7-AAD staining (C) and cell survival was analyzed by MTS assay (D). *P < 0.05; **P < 0.01 (E-G) After transfection of miR-200c precursor with either control or CYP1B1 plasmid, A498 cells were treated with docetaxel (5 μM) for 72 hrs. CYP1B1 protein expression was determined by Western blot (E), apoptotic cell death was measured with annexin V-FITC/7-AAD staining (F) and cell survival was analyzed by MTS assay (G). *P < 0.05; **P < 0.01.

Mentions: To determine whether miR-200c affects RCC chemosensitivity, the cytotoxic effect of docetaxel was measured after miR-200c restoration or inhibition. A significant increase in the cytotoxic effect of docetaxel was observed and as a consequence, the cell survival rate was reduced by transfection of miR-200c precursor in A498 cells (Fig. 6A and B). MiR-200c inhibition significantly decreased the cytotoxic effect of docetaxel and enhanced the survival rate of 786-O cells (Fig. 6C and D). To verify the effects of miR-200c via CYP1B1 on the chemosensitivity to docetaxel, we introduced CYP1B1 plasmid into A498 cells after miR-200c overexpression and examined cytotoxicity. Restoration of CYP1B1 protein levels was verified after transfection (Fig. 6E). We observed that the increase in the cytotoxic effect of docetaxel by miR-200c overexpression was reversed by the enforced CYP1B1 expression (Fig. 6F). As a consequence, cell survival was also restored by the exogenous CYP1B1expression (Fig. 6G). These results indicate that CYP1B1 regulation by miR-200c directly influences the chemosensitivity of RCC cells to docetaxel.


Loss of miR-200c up-regulates CYP1B1 and confers docetaxel resistance in renal cell carcinoma.

Chang I, Mitsui Y, Fukuhara S, Gill A, Wong DK, Yamamura S, Shahryari V, Tabatabai ZL, Dahiya R, Shin DM, Tanaka Y - Oncotarget (2015)

MiR-200c regulates docetaxel resistance of RCC cells(A and B) After miR-200c precursor transfection, A498 cells were treated with docetaxel (5 μM) for 72 hrs. Apoptotic cell death was measured with annexin V-FITC/7-AAD staining (A) and cell survival was analyzed by MTS assay (B). **P < 0.01; ***P < 0.001 (C and D) After miR-200c inhibitor transfection, 786-O cells were treated with docetaxel (5 μM) for 72 hrs. Apoptotic cell death was measured with annexin V-FITC/7-AAD staining (C) and cell survival was analyzed by MTS assay (D). *P < 0.05; **P < 0.01 (E-G) After transfection of miR-200c precursor with either control or CYP1B1 plasmid, A498 cells were treated with docetaxel (5 μM) for 72 hrs. CYP1B1 protein expression was determined by Western blot (E), apoptotic cell death was measured with annexin V-FITC/7-AAD staining (F) and cell survival was analyzed by MTS assay (G). *P < 0.05; **P < 0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4480715&req=5

Figure 6: MiR-200c regulates docetaxel resistance of RCC cells(A and B) After miR-200c precursor transfection, A498 cells were treated with docetaxel (5 μM) for 72 hrs. Apoptotic cell death was measured with annexin V-FITC/7-AAD staining (A) and cell survival was analyzed by MTS assay (B). **P < 0.01; ***P < 0.001 (C and D) After miR-200c inhibitor transfection, 786-O cells were treated with docetaxel (5 μM) for 72 hrs. Apoptotic cell death was measured with annexin V-FITC/7-AAD staining (C) and cell survival was analyzed by MTS assay (D). *P < 0.05; **P < 0.01 (E-G) After transfection of miR-200c precursor with either control or CYP1B1 plasmid, A498 cells were treated with docetaxel (5 μM) for 72 hrs. CYP1B1 protein expression was determined by Western blot (E), apoptotic cell death was measured with annexin V-FITC/7-AAD staining (F) and cell survival was analyzed by MTS assay (G). *P < 0.05; **P < 0.01.
Mentions: To determine whether miR-200c affects RCC chemosensitivity, the cytotoxic effect of docetaxel was measured after miR-200c restoration or inhibition. A significant increase in the cytotoxic effect of docetaxel was observed and as a consequence, the cell survival rate was reduced by transfection of miR-200c precursor in A498 cells (Fig. 6A and B). MiR-200c inhibition significantly decreased the cytotoxic effect of docetaxel and enhanced the survival rate of 786-O cells (Fig. 6C and D). To verify the effects of miR-200c via CYP1B1 on the chemosensitivity to docetaxel, we introduced CYP1B1 plasmid into A498 cells after miR-200c overexpression and examined cytotoxicity. Restoration of CYP1B1 protein levels was verified after transfection (Fig. 6E). We observed that the increase in the cytotoxic effect of docetaxel by miR-200c overexpression was reversed by the enforced CYP1B1 expression (Fig. 6F). As a consequence, cell survival was also restored by the exogenous CYP1B1expression (Fig. 6G). These results indicate that CYP1B1 regulation by miR-200c directly influences the chemosensitivity of RCC cells to docetaxel.

Bottom Line: Additionally, miR-200c, which is significantly down-regulated in RCC regulates CYP1B1 expression and activity.These results suggest that CYP1B1 up-regulation mediated by low miR-200c is one of the mechanisms underlying resistance of RCC cells to docetaxel.Therefore, expression of CYP1B1 and miR-200c in RCC may be useful as a prediction for docetaxel response.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery and Division of Urology, Veterans Affairs Medical Center, San Francisco, California, United States of America.

ABSTRACT
Despite high protein expression and enzymatic activity of cytochrome P450 1B1 (CYP1B1) in renal cell cancer (RCC), its functional significance has not been elucidated. Here we explored the functional role and regulatory mechanism of CYP1B1 in RCC. Reduction of CYP1B1 levels fail to prevent in vitro tumorigenicity such as proliferation, apoptosis, and cell cycle progression of RCC cells. Moreover, the expression levels are not associated with tumor type, stage, Fuhrman grade and 5-year survival probability after surgery. Instead, alteration of CYP1B1 expression regulates the chemosensitivity of RCC cells to docetaxel suggesting its critical contribution to the chemoresistance. Additionally, miR-200c, which is significantly down-regulated in RCC regulates CYP1B1 expression and activity. An inverse association was also observed between the expression levels of miR-200c and CYP1B1 protein in RCC tissues. Finally, alteration of miR-200c levels affects the chemosensitivity of RCC cells. Restoration of docetaxel resistance by exogenous expression of CYP1B1 in miR-200c-over-expressing cells indicates that CYP1B1 is a functional target of miR-200c. These results suggest that CYP1B1 up-regulation mediated by low miR-200c is one of the mechanisms underlying resistance of RCC cells to docetaxel. Therefore, expression of CYP1B1 and miR-200c in RCC may be useful as a prediction for docetaxel response.

No MeSH data available.


Related in: MedlinePlus