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PARP inhibitor ABT-888 affects response of MDA-MB-231 cells to doxorubicin treatment, targeting Snail expression.

Mariano G, Ricciardi MR, Trisciuoglio D, Zampieri M, Ciccarone F, Guastafierro T, Calabrese R, Valentini E, Tafuri A, Del Bufalo D, Caiafa P, Reale A - Oncotarget (2015)

Bottom Line: MDA-MB-231 is a Snail-expressing metastatic breast cancer cell line, which exhibits chemoresistance properties when treated with damaging agents.In this study, we show that the PARP inhibitor ABT-888 was capable to modulate the MDA-MB-231 cell response to doxorubicin, leading to an increase in the rate of apoptosis.Our further results indicate that PARP-1 controlled Snail expression at transcriptional level in cells exposed to doxorubicin.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular Biotechnologies and Haematology, "Sapienza" University of Rome, Italy.

ABSTRACT
To overcome cancer cells resistance to pharmacological therapy, the development of new therapeutic approaches becomes urgent. For this purpose, the use of poly(ADP-ribose) polymerase (PARP) inhibitors in combination with other cytotoxic agents could represent an efficacious strategy. Poly(ADP-ribosyl)ation (PARylation) is a post-translational modification that plays a well characterized role in the cellular decisions of life and death. Recent findings indicate that PARP-1 may control the expression of Snail, the master gene of epithelial-mesenchymal transition (EMT). Snail is highly represented in different resistant tumors, functioning as a factor regulating anti-apoptotic programmes. MDA-MB-231 is a Snail-expressing metastatic breast cancer cell line, which exhibits chemoresistance properties when treated with damaging agents. In this study, we show that the PARP inhibitor ABT-888 was capable to modulate the MDA-MB-231 cell response to doxorubicin, leading to an increase in the rate of apoptosis. Our further results indicate that PARP-1 controlled Snail expression at transcriptional level in cells exposed to doxorubicin. Given the increasing interest in the employment of PARP inhibitors as chemotherapeutic adjuvants, our in vitro results suggest that one of the mechanisms through which PARP inhibition can chemosensitize cancer cells in vivo, is targeting Snail expression thus promoting apoptosis.

No MeSH data available.


Related in: MedlinePlus

Snail knockdown sensitizes MDA-MB-231 cells to doxo-induced apoptosis and allows recovery of PTEN expressionA. Levels of Snail protein were measured by Western blot analyses in shCT and shSnail MDA-MB-231 cells treated for 24 h with 1 μM doxo B. Annexin V positive cells were counted in the right upper and lower squares. The diagram reports the percentage of Annexin V positive cells in untreated shCT and shSnail cells (black bars) and after treatment of shCT (white bars) and shSnail cells (light gray bars) with 1 μM doxo at the indicated times in relation to total cells. Data represented are the mean+SEM of three independent experiments performed in duplicates. Comparisons were made with ANOVA/Turkey's test. *P < 0.05, **P < 0.01 compared to cells treated with doxo at 24 h, 48 h respectively. C. Expression levels of PTEN after treatment of shCT and shSnail MDA-MB-231 cells with 1 μM doxo for 7 h (light gray bars) or 24 h (dark gray bars) in relation to untreated shCT and shSnail cells (white bars), considered as 1.0. Data represent mean+SEM of three independent experiments performed in triplicates. *P < 0.05 by Student's t-test.
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Figure 5: Snail knockdown sensitizes MDA-MB-231 cells to doxo-induced apoptosis and allows recovery of PTEN expressionA. Levels of Snail protein were measured by Western blot analyses in shCT and shSnail MDA-MB-231 cells treated for 24 h with 1 μM doxo B. Annexin V positive cells were counted in the right upper and lower squares. The diagram reports the percentage of Annexin V positive cells in untreated shCT and shSnail cells (black bars) and after treatment of shCT (white bars) and shSnail cells (light gray bars) with 1 μM doxo at the indicated times in relation to total cells. Data represented are the mean+SEM of three independent experiments performed in duplicates. Comparisons were made with ANOVA/Turkey's test. *P < 0.05, **P < 0.01 compared to cells treated with doxo at 24 h, 48 h respectively. C. Expression levels of PTEN after treatment of shCT and shSnail MDA-MB-231 cells with 1 μM doxo for 7 h (light gray bars) or 24 h (dark gray bars) in relation to untreated shCT and shSnail cells (white bars), considered as 1.0. Data represent mean+SEM of three independent experiments performed in triplicates. *P < 0.05 by Student's t-test.

Mentions: To confirm that the rate of apoptosis in MDA-MB-231 cells depended on Snail levels, MDA-MB-231 cells were transfected with shRNA-Snail or shRNA-CT plasmids. Figure 5A shows that depletion of endogenous Snail was efficient as Snail signal resulted low after doxo treatment too. A significant increase in the number of Annexin V positive cells after treatment with doxo at 24–48 h was observed in Snail-depleted cells with respect to control cells (around 3 fold) (Figure 5B). These data demonstrate that Snail deficiency is required for efficient apoptosis in doxo-treated cells and support the idea that PARP-1 inhibition/depletion sensitizes MDA-MB-231 cells by limiting Snail upregulation. Interestingly, Real-Time PCR analysis (Figure 5C) showed that, upon doxo treatment, the clear PTEN downregulation of control cells was not detectable in Snail-depleted cells. This result is in agreement with the presence of high Snail levels and their repressive function on PTEN expression in control cells exposed to doxo.


PARP inhibitor ABT-888 affects response of MDA-MB-231 cells to doxorubicin treatment, targeting Snail expression.

Mariano G, Ricciardi MR, Trisciuoglio D, Zampieri M, Ciccarone F, Guastafierro T, Calabrese R, Valentini E, Tafuri A, Del Bufalo D, Caiafa P, Reale A - Oncotarget (2015)

Snail knockdown sensitizes MDA-MB-231 cells to doxo-induced apoptosis and allows recovery of PTEN expressionA. Levels of Snail protein were measured by Western blot analyses in shCT and shSnail MDA-MB-231 cells treated for 24 h with 1 μM doxo B. Annexin V positive cells were counted in the right upper and lower squares. The diagram reports the percentage of Annexin V positive cells in untreated shCT and shSnail cells (black bars) and after treatment of shCT (white bars) and shSnail cells (light gray bars) with 1 μM doxo at the indicated times in relation to total cells. Data represented are the mean+SEM of three independent experiments performed in duplicates. Comparisons were made with ANOVA/Turkey's test. *P < 0.05, **P < 0.01 compared to cells treated with doxo at 24 h, 48 h respectively. C. Expression levels of PTEN after treatment of shCT and shSnail MDA-MB-231 cells with 1 μM doxo for 7 h (light gray bars) or 24 h (dark gray bars) in relation to untreated shCT and shSnail cells (white bars), considered as 1.0. Data represent mean+SEM of three independent experiments performed in triplicates. *P < 0.05 by Student's t-test.
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Related In: Results  -  Collection

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Figure 5: Snail knockdown sensitizes MDA-MB-231 cells to doxo-induced apoptosis and allows recovery of PTEN expressionA. Levels of Snail protein were measured by Western blot analyses in shCT and shSnail MDA-MB-231 cells treated for 24 h with 1 μM doxo B. Annexin V positive cells were counted in the right upper and lower squares. The diagram reports the percentage of Annexin V positive cells in untreated shCT and shSnail cells (black bars) and after treatment of shCT (white bars) and shSnail cells (light gray bars) with 1 μM doxo at the indicated times in relation to total cells. Data represented are the mean+SEM of three independent experiments performed in duplicates. Comparisons were made with ANOVA/Turkey's test. *P < 0.05, **P < 0.01 compared to cells treated with doxo at 24 h, 48 h respectively. C. Expression levels of PTEN after treatment of shCT and shSnail MDA-MB-231 cells with 1 μM doxo for 7 h (light gray bars) or 24 h (dark gray bars) in relation to untreated shCT and shSnail cells (white bars), considered as 1.0. Data represent mean+SEM of three independent experiments performed in triplicates. *P < 0.05 by Student's t-test.
Mentions: To confirm that the rate of apoptosis in MDA-MB-231 cells depended on Snail levels, MDA-MB-231 cells were transfected with shRNA-Snail or shRNA-CT plasmids. Figure 5A shows that depletion of endogenous Snail was efficient as Snail signal resulted low after doxo treatment too. A significant increase in the number of Annexin V positive cells after treatment with doxo at 24–48 h was observed in Snail-depleted cells with respect to control cells (around 3 fold) (Figure 5B). These data demonstrate that Snail deficiency is required for efficient apoptosis in doxo-treated cells and support the idea that PARP-1 inhibition/depletion sensitizes MDA-MB-231 cells by limiting Snail upregulation. Interestingly, Real-Time PCR analysis (Figure 5C) showed that, upon doxo treatment, the clear PTEN downregulation of control cells was not detectable in Snail-depleted cells. This result is in agreement with the presence of high Snail levels and their repressive function on PTEN expression in control cells exposed to doxo.

Bottom Line: MDA-MB-231 is a Snail-expressing metastatic breast cancer cell line, which exhibits chemoresistance properties when treated with damaging agents.In this study, we show that the PARP inhibitor ABT-888 was capable to modulate the MDA-MB-231 cell response to doxorubicin, leading to an increase in the rate of apoptosis.Our further results indicate that PARP-1 controlled Snail expression at transcriptional level in cells exposed to doxorubicin.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular Biotechnologies and Haematology, "Sapienza" University of Rome, Italy.

ABSTRACT
To overcome cancer cells resistance to pharmacological therapy, the development of new therapeutic approaches becomes urgent. For this purpose, the use of poly(ADP-ribose) polymerase (PARP) inhibitors in combination with other cytotoxic agents could represent an efficacious strategy. Poly(ADP-ribosyl)ation (PARylation) is a post-translational modification that plays a well characterized role in the cellular decisions of life and death. Recent findings indicate that PARP-1 may control the expression of Snail, the master gene of epithelial-mesenchymal transition (EMT). Snail is highly represented in different resistant tumors, functioning as a factor regulating anti-apoptotic programmes. MDA-MB-231 is a Snail-expressing metastatic breast cancer cell line, which exhibits chemoresistance properties when treated with damaging agents. In this study, we show that the PARP inhibitor ABT-888 was capable to modulate the MDA-MB-231 cell response to doxorubicin, leading to an increase in the rate of apoptosis. Our further results indicate that PARP-1 controlled Snail expression at transcriptional level in cells exposed to doxorubicin. Given the increasing interest in the employment of PARP inhibitors as chemotherapeutic adjuvants, our in vitro results suggest that one of the mechanisms through which PARP inhibition can chemosensitize cancer cells in vivo, is targeting Snail expression thus promoting apoptosis.

No MeSH data available.


Related in: MedlinePlus