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c-Myc overexpression sensitises colon cancer cells to camptothecin-induced apoptosis.

Arango D, Mariadason JM, Wilson AJ, Yang W, Corner GA, Nicholas C, Aranes MJ, Augenlicht LH - Br. J. Cancer (2003)

Bottom Line: Both camptothecin exposure and c-Myc overexpression in LoVo cells resulted in elevation of p53 protein levels, suggesting a role of p53 in the c-Myc-imposed sensitisation to the apoptotic effects of camptothecin.Targeted inactivation of p21(Waf1/Cip1) in HCT116 colon cancer cells resulted in significantly increased levels of apoptosis following treatment with camptothecin, demonstrating the importance of p21(Waf1/Cip1) in the response to this agent.Moreover, the expression of 10 of these genes was confirmed to be significantly correlated with response to camptothecin in a panel of 30 colorectal cancer cell lines.

View Article: PubMed Central - PubMed

Affiliation: Albert Einstein Cancer Center, Montefiore Medical Center, Oncology Department, 111 East 210th St, Bronx, NY 10467, USA. diego.arango@helsinki.fi

ABSTRACT
The proto-oncogene c-Myc is overexpressed in 70% of colorectal tumours and can modulate proliferation and apoptosis after cytotoxic insult. Using an isogenic cell system, we demonstrate that c-Myc overexpression in colon carcinoma LoVo cells resulted in sensitisation to camptothecin-induced apoptosis, thus identifying c-Myc as a potential marker predicting response of colorectal tumour cells to camptothecin. Both camptothecin exposure and c-Myc overexpression in LoVo cells resulted in elevation of p53 protein levels, suggesting a role of p53 in the c-Myc-imposed sensitisation to the apoptotic effects of camptothecin. This was confirmed by the ability of PFT-alpha, a specific inhibitor of p53, to attenuate camptothecin-induced apoptosis. p53 can induce the expression of p21(Waf1/Cip1), an antiproliferative protein that can facilitate DNA repair and drug resistance. Importantly, although camptothecin treatment markedly increased p21(Waf1/Cip1) levels in parental LoVo cells, this effect was abrogated in c-Myc-overexpressing derivatives. Targeted inactivation of p21(Waf1/Cip1) in HCT116 colon cancer cells resulted in significantly increased levels of apoptosis following treatment with camptothecin, demonstrating the importance of p21(Waf1/Cip1) in the response to this agent. Finally, cDNA microarray analysis was used to identify genes that are modulated in expression by c-Myc upregulation that could serve as additional markers predicting response to camptothecin. Thirty-four sequences were altered in expression over four-fold in two isogenic c-Myc-overexpressing clones compared to parental LoVo cells. Moreover, the expression of 10 of these genes was confirmed to be significantly correlated with response to camptothecin in a panel of 30 colorectal cancer cell lines.

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Role of p21 in the apoptotic response to camptothecin. Targeted inactivation of both alleles of p21Waf1/Cip1 in HCT116 colon cancer cells resulted in significant sensitisation to camptothecin-induced apoptosis. The mean of four different experiments±standard error of the mean is shown. Asterisks indicate significant differences (Student's t-test, P<0.004) between apoptotic levels in HCT116 p21+/+ and −/− for a given dose.
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fig7: Role of p21 in the apoptotic response to camptothecin. Targeted inactivation of both alleles of p21Waf1/Cip1 in HCT116 colon cancer cells resulted in significant sensitisation to camptothecin-induced apoptosis. The mean of four different experiments±standard error of the mean is shown. Asterisks indicate significant differences (Student's t-test, P<0.004) between apoptotic levels in HCT116 p21+/+ and −/− for a given dose.

Mentions: Importantly, although exposure of parental LoVo cells to 0.1 or 0.5 μM camptothecin for 24 h resulted in upregulation of p53 and the p53 target gene p21Waf1/Cip1, c-Myc-overexpressing L2 and L3 cells failed to upregulate p21Waf1/Cip1 protein levels in response to camptothecin treatment (Figure 4), strongly suggesting that p21Waf1/Cip1 levels could modulate apoptosis induced by camptothecin. To directly investigate this possibility, we used an engineered in vitro system where both alleles of the p21Waf1/Cip1 gene have been inactivated by targeted deletion in HCT116 cells, a colon cancer cell line with a wild-type p53 gene (Bunz et al, 1998). Inactivation of p21Waf1/Cip1 in these cells resulted in significantly (P<0.004) increased sensitivity to apoptosis induced by camptothecin compared to parental wild-type p21Waf1/Cip1 HCT116 cells (Figure 7Figure 7


c-Myc overexpression sensitises colon cancer cells to camptothecin-induced apoptosis.

Arango D, Mariadason JM, Wilson AJ, Yang W, Corner GA, Nicholas C, Aranes MJ, Augenlicht LH - Br. J. Cancer (2003)

Role of p21 in the apoptotic response to camptothecin. Targeted inactivation of both alleles of p21Waf1/Cip1 in HCT116 colon cancer cells resulted in significant sensitisation to camptothecin-induced apoptosis. The mean of four different experiments±standard error of the mean is shown. Asterisks indicate significant differences (Student's t-test, P<0.004) between apoptotic levels in HCT116 p21+/+ and −/− for a given dose.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2394410&req=5

fig7: Role of p21 in the apoptotic response to camptothecin. Targeted inactivation of both alleles of p21Waf1/Cip1 in HCT116 colon cancer cells resulted in significant sensitisation to camptothecin-induced apoptosis. The mean of four different experiments±standard error of the mean is shown. Asterisks indicate significant differences (Student's t-test, P<0.004) between apoptotic levels in HCT116 p21+/+ and −/− for a given dose.
Mentions: Importantly, although exposure of parental LoVo cells to 0.1 or 0.5 μM camptothecin for 24 h resulted in upregulation of p53 and the p53 target gene p21Waf1/Cip1, c-Myc-overexpressing L2 and L3 cells failed to upregulate p21Waf1/Cip1 protein levels in response to camptothecin treatment (Figure 4), strongly suggesting that p21Waf1/Cip1 levels could modulate apoptosis induced by camptothecin. To directly investigate this possibility, we used an engineered in vitro system where both alleles of the p21Waf1/Cip1 gene have been inactivated by targeted deletion in HCT116 cells, a colon cancer cell line with a wild-type p53 gene (Bunz et al, 1998). Inactivation of p21Waf1/Cip1 in these cells resulted in significantly (P<0.004) increased sensitivity to apoptosis induced by camptothecin compared to parental wild-type p21Waf1/Cip1 HCT116 cells (Figure 7Figure 7

Bottom Line: Both camptothecin exposure and c-Myc overexpression in LoVo cells resulted in elevation of p53 protein levels, suggesting a role of p53 in the c-Myc-imposed sensitisation to the apoptotic effects of camptothecin.Targeted inactivation of p21(Waf1/Cip1) in HCT116 colon cancer cells resulted in significantly increased levels of apoptosis following treatment with camptothecin, demonstrating the importance of p21(Waf1/Cip1) in the response to this agent.Moreover, the expression of 10 of these genes was confirmed to be significantly correlated with response to camptothecin in a panel of 30 colorectal cancer cell lines.

View Article: PubMed Central - PubMed

Affiliation: Albert Einstein Cancer Center, Montefiore Medical Center, Oncology Department, 111 East 210th St, Bronx, NY 10467, USA. diego.arango@helsinki.fi

ABSTRACT
The proto-oncogene c-Myc is overexpressed in 70% of colorectal tumours and can modulate proliferation and apoptosis after cytotoxic insult. Using an isogenic cell system, we demonstrate that c-Myc overexpression in colon carcinoma LoVo cells resulted in sensitisation to camptothecin-induced apoptosis, thus identifying c-Myc as a potential marker predicting response of colorectal tumour cells to camptothecin. Both camptothecin exposure and c-Myc overexpression in LoVo cells resulted in elevation of p53 protein levels, suggesting a role of p53 in the c-Myc-imposed sensitisation to the apoptotic effects of camptothecin. This was confirmed by the ability of PFT-alpha, a specific inhibitor of p53, to attenuate camptothecin-induced apoptosis. p53 can induce the expression of p21(Waf1/Cip1), an antiproliferative protein that can facilitate DNA repair and drug resistance. Importantly, although camptothecin treatment markedly increased p21(Waf1/Cip1) levels in parental LoVo cells, this effect was abrogated in c-Myc-overexpressing derivatives. Targeted inactivation of p21(Waf1/Cip1) in HCT116 colon cancer cells resulted in significantly increased levels of apoptosis following treatment with camptothecin, demonstrating the importance of p21(Waf1/Cip1) in the response to this agent. Finally, cDNA microarray analysis was used to identify genes that are modulated in expression by c-Myc upregulation that could serve as additional markers predicting response to camptothecin. Thirty-four sequences were altered in expression over four-fold in two isogenic c-Myc-overexpressing clones compared to parental LoVo cells. Moreover, the expression of 10 of these genes was confirmed to be significantly correlated with response to camptothecin in a panel of 30 colorectal cancer cell lines.

Show MeSH
Related in: MedlinePlus