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Src/STAT3-dependent heme oxygenase-1 induction mediates chemoresistance of breast cancer cells to doxorubicin by promoting autophagy.

Tan Q, Wang H, Hu Y, Hu M, Li X - Cancer Sci. (2015)

Bottom Line: In the present study, we found an induction of HO-1 expression in doxorubicin (DOX)-treated MDA-MB-231 human breast adenocarcinoma cells, which showed insensitivity to DOX treatment.Knockdown HO-1 expression dramatically upregulated the incidence of MDA-MB-231 cell death under DOX treatment, indicating that HO-1 functions as a critical contributor to drug resistance in MDA-MB-231 cells.Moreover, upregulation of HO-1 expression required the activation of both signal transducer and activator of transcription (STAT)3 and its upstream regulator, protein kinase Src.

View Article: PubMed Central - PubMed

Affiliation: Department of Stress Medicine, Beijing Institute of Basic Medical Sciences, Beijing, China.

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Src/STAT3/heme oxygenase-1 (HO-1)-dependent autophagy protected MDA-MB-468 cells from doxorubicin (DOX)-induced apoptosis. (a) MDA-MB-468 and MDA-MB-453 cells were left untreated or treated with DOX (0.2 μM) for 48 h and then cell death incidence was determined as described in Figure1(a). (b) MDA-MB-468 and MDA-MB-453 cells were treated with DOX (0.2 μM) for the indicated time periods and then the induction of Src and STAT3 activation and HO-1, Beclin-1 and IC3I/II expression were detected by western-blot assay. (c) MDA-MB-468 and MDA-MB-453 cells were left untreated or treated with DOX (0.2 μM) for 24 h and then autophagy was detected as described in Figure2(a). (d–f) MDA-MB-468 cells were transfected with Src, STAT3, HO-1 siRNA or their control siRNA, respectively. Then the cells were treated with DOX (0.2 μM) 36 h after transfection. The activation of Src, STAT3 and the expressions of HO-1, Beclin-1 and LC3BI/II were determined 12 h after DOX exposure. (g) MDA-MB-468 cells were pretreated with 3-MA followed by exposure to DOX (0.2 μM) and then cell death incidence was detected 48 h after DOX treatment.
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fig06: Src/STAT3/heme oxygenase-1 (HO-1)-dependent autophagy protected MDA-MB-468 cells from doxorubicin (DOX)-induced apoptosis. (a) MDA-MB-468 and MDA-MB-453 cells were left untreated or treated with DOX (0.2 μM) for 48 h and then cell death incidence was determined as described in Figure1(a). (b) MDA-MB-468 and MDA-MB-453 cells were treated with DOX (0.2 μM) for the indicated time periods and then the induction of Src and STAT3 activation and HO-1, Beclin-1 and IC3I/II expression were detected by western-blot assay. (c) MDA-MB-468 and MDA-MB-453 cells were left untreated or treated with DOX (0.2 μM) for 24 h and then autophagy was detected as described in Figure2(a). (d–f) MDA-MB-468 cells were transfected with Src, STAT3, HO-1 siRNA or their control siRNA, respectively. Then the cells were treated with DOX (0.2 μM) 36 h after transfection. The activation of Src, STAT3 and the expressions of HO-1, Beclin-1 and LC3BI/II were determined 12 h after DOX exposure. (g) MDA-MB-468 cells were pretreated with 3-MA followed by exposure to DOX (0.2 μM) and then cell death incidence was detected 48 h after DOX treatment.

Mentions: To determine whether the Src/STAT3/HO-1/autophagy pathway activation plays a general cytoprotective role in breast cancer cells under DOX treatment, we next detected the activation status of this pathway in the MDA-MB-468 cells, which also bear ER, PR and Her2-negative phenotype as MDA-MB-231 cells (data not shown) and are also resistant to DOX-induced apoptosis compared with MDA-MB-453 cells (Fig.6a). We observed that DOX treatment not only induced the activation of Src and STAT3, but also upregulated the expression levels of HO-1, Beclin-1 and LC-3I/II in the MDA-MB-468 cells; in contrast, no signal indicating the activation of these signaling molecules was detected in the MDA-MB-453 cells under the same DOX exposure conditions (Fig.6b). Moreover, an induction of autophagic flux in MDA-MB-468 cells was observed under DOX exposure in the flow cytometry-based analysis (Fig.6c), further confirming the induction of autophagy in the DOX-treated MDA-MB-468 cells. In the following analyses, MDA-MB-468 cells were transfected with Src, STAT3 or HO-1 siRNA, respectively, to test whether signal transduction of Src/STAT3/HO-1 cascade contributed to autophagy. As shown in Figure6(d–f), knockdown Src, STAT3 or HO-1 expression almost totally blocked the activation of their downstream targets as well as the signaling event indicating autophagy induction (Beclin-1 and LC3I/II upregulations). Finally, abrogating autophagy induction by 3-MA obviously increased the sensitivity of MDA-MB-468 cells to DOX-induced cytotoxicity (Fig.6g). Taken together, these data indicate that Src/STAT3/HO-1/autophagy pathway activation also plays a cytoprotective role in MDA-MB-468 cells under DOX treatment.


Src/STAT3-dependent heme oxygenase-1 induction mediates chemoresistance of breast cancer cells to doxorubicin by promoting autophagy.

Tan Q, Wang H, Hu Y, Hu M, Li X - Cancer Sci. (2015)

Src/STAT3/heme oxygenase-1 (HO-1)-dependent autophagy protected MDA-MB-468 cells from doxorubicin (DOX)-induced apoptosis. (a) MDA-MB-468 and MDA-MB-453 cells were left untreated or treated with DOX (0.2 μM) for 48 h and then cell death incidence was determined as described in Figure1(a). (b) MDA-MB-468 and MDA-MB-453 cells were treated with DOX (0.2 μM) for the indicated time periods and then the induction of Src and STAT3 activation and HO-1, Beclin-1 and IC3I/II expression were detected by western-blot assay. (c) MDA-MB-468 and MDA-MB-453 cells were left untreated or treated with DOX (0.2 μM) for 24 h and then autophagy was detected as described in Figure2(a). (d–f) MDA-MB-468 cells were transfected with Src, STAT3, HO-1 siRNA or their control siRNA, respectively. Then the cells were treated with DOX (0.2 μM) 36 h after transfection. The activation of Src, STAT3 and the expressions of HO-1, Beclin-1 and LC3BI/II were determined 12 h after DOX exposure. (g) MDA-MB-468 cells were pretreated with 3-MA followed by exposure to DOX (0.2 μM) and then cell death incidence was detected 48 h after DOX treatment.
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fig06: Src/STAT3/heme oxygenase-1 (HO-1)-dependent autophagy protected MDA-MB-468 cells from doxorubicin (DOX)-induced apoptosis. (a) MDA-MB-468 and MDA-MB-453 cells were left untreated or treated with DOX (0.2 μM) for 48 h and then cell death incidence was determined as described in Figure1(a). (b) MDA-MB-468 and MDA-MB-453 cells were treated with DOX (0.2 μM) for the indicated time periods and then the induction of Src and STAT3 activation and HO-1, Beclin-1 and IC3I/II expression were detected by western-blot assay. (c) MDA-MB-468 and MDA-MB-453 cells were left untreated or treated with DOX (0.2 μM) for 24 h and then autophagy was detected as described in Figure2(a). (d–f) MDA-MB-468 cells were transfected with Src, STAT3, HO-1 siRNA or their control siRNA, respectively. Then the cells were treated with DOX (0.2 μM) 36 h after transfection. The activation of Src, STAT3 and the expressions of HO-1, Beclin-1 and LC3BI/II were determined 12 h after DOX exposure. (g) MDA-MB-468 cells were pretreated with 3-MA followed by exposure to DOX (0.2 μM) and then cell death incidence was detected 48 h after DOX treatment.
Mentions: To determine whether the Src/STAT3/HO-1/autophagy pathway activation plays a general cytoprotective role in breast cancer cells under DOX treatment, we next detected the activation status of this pathway in the MDA-MB-468 cells, which also bear ER, PR and Her2-negative phenotype as MDA-MB-231 cells (data not shown) and are also resistant to DOX-induced apoptosis compared with MDA-MB-453 cells (Fig.6a). We observed that DOX treatment not only induced the activation of Src and STAT3, but also upregulated the expression levels of HO-1, Beclin-1 and LC-3I/II in the MDA-MB-468 cells; in contrast, no signal indicating the activation of these signaling molecules was detected in the MDA-MB-453 cells under the same DOX exposure conditions (Fig.6b). Moreover, an induction of autophagic flux in MDA-MB-468 cells was observed under DOX exposure in the flow cytometry-based analysis (Fig.6c), further confirming the induction of autophagy in the DOX-treated MDA-MB-468 cells. In the following analyses, MDA-MB-468 cells were transfected with Src, STAT3 or HO-1 siRNA, respectively, to test whether signal transduction of Src/STAT3/HO-1 cascade contributed to autophagy. As shown in Figure6(d–f), knockdown Src, STAT3 or HO-1 expression almost totally blocked the activation of their downstream targets as well as the signaling event indicating autophagy induction (Beclin-1 and LC3I/II upregulations). Finally, abrogating autophagy induction by 3-MA obviously increased the sensitivity of MDA-MB-468 cells to DOX-induced cytotoxicity (Fig.6g). Taken together, these data indicate that Src/STAT3/HO-1/autophagy pathway activation also plays a cytoprotective role in MDA-MB-468 cells under DOX treatment.

Bottom Line: In the present study, we found an induction of HO-1 expression in doxorubicin (DOX)-treated MDA-MB-231 human breast adenocarcinoma cells, which showed insensitivity to DOX treatment.Knockdown HO-1 expression dramatically upregulated the incidence of MDA-MB-231 cell death under DOX treatment, indicating that HO-1 functions as a critical contributor to drug resistance in MDA-MB-231 cells.Moreover, upregulation of HO-1 expression required the activation of both signal transducer and activator of transcription (STAT)3 and its upstream regulator, protein kinase Src.

View Article: PubMed Central - PubMed

Affiliation: Department of Stress Medicine, Beijing Institute of Basic Medical Sciences, Beijing, China.

Show MeSH
Related in: MedlinePlus