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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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cDNA microarray analysis of LoVo and isogenic L2 and L3 cells overexpressing c-Myc. (A) The number of genes identified as differentially expressed is a function of the cutoff used. Selection of a stringent cutoff value of four-fold reduces the number of false positives expected when assessing differences in gene expression between LoVo and L2/L3 cells. (B) TreeView image showing the expression profile of the 34 genes with over four-fold expression difference between LoVo and L2/L3 that characterise camptothecin-resistant and -sensitive cells. Both replicas are shown. Red and green indicate genes that are over-represented and under-represented, respectively, relative to the reference RNA used (see Materials and Methods).
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fig8: cDNA microarray analysis of LoVo and isogenic L2 and L3 cells overexpressing c-Myc. (A) The number of genes identified as differentially expressed is a function of the cutoff used. Selection of a stringent cutoff value of four-fold reduces the number of false positives expected when assessing differences in gene expression between LoVo and L2/L3 cells. (B) TreeView image showing the expression profile of the 34 genes with over four-fold expression difference between LoVo and L2/L3 that characterise camptothecin-resistant and -sensitive cells. Both replicas are shown. Red and green indicate genes that are over-represented and under-represented, respectively, relative to the reference RNA used (see Materials and Methods).

Mentions: To identify additional genes that could serve as markers predicting apoptotic response to camptothecin, we measured the relative expression of 9216 sequences, in duplicate, in resistant LoVo cells, as well as in sensitive c-Myc-overexpressing L2 and L3 isogenic cells, using cDNA microarray analysis. The complete databases are available at http://sequence.aecom.yu.edu/bioinf/Augenlicht/default.html. First, to assess the background variability due to methodological and biological factors, we averaged the values for the two LoVo replicas and compared them to the average of another two different replicates of LoVo cells, and quantified the number of genes identified as differentially expressed as a function of selected cutoff values (Figure 8AFigure 8


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)

cDNA microarray analysis of LoVo and isogenic L2 and L3 cells overexpressing c-Myc. (A) The number of genes identified as differentially expressed is a function of the cutoff used. Selection of a stringent cutoff value of four-fold reduces the number of false positives expected when assessing differences in gene expression between LoVo and L2/L3 cells. (B) TreeView image showing the expression profile of the 34 genes with over four-fold expression difference between LoVo and L2/L3 that characterise camptothecin-resistant and -sensitive cells. Both replicas are shown. Red and green indicate genes that are over-represented and under-represented, respectively, relative to the reference RNA used (see Materials and Methods).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2394410&req=5

fig8: cDNA microarray analysis of LoVo and isogenic L2 and L3 cells overexpressing c-Myc. (A) The number of genes identified as differentially expressed is a function of the cutoff used. Selection of a stringent cutoff value of four-fold reduces the number of false positives expected when assessing differences in gene expression between LoVo and L2/L3 cells. (B) TreeView image showing the expression profile of the 34 genes with over four-fold expression difference between LoVo and L2/L3 that characterise camptothecin-resistant and -sensitive cells. Both replicas are shown. Red and green indicate genes that are over-represented and under-represented, respectively, relative to the reference RNA used (see Materials and Methods).
Mentions: To identify additional genes that could serve as markers predicting apoptotic response to camptothecin, we measured the relative expression of 9216 sequences, in duplicate, in resistant LoVo cells, as well as in sensitive c-Myc-overexpressing L2 and L3 isogenic cells, using cDNA microarray analysis. The complete databases are available at http://sequence.aecom.yu.edu/bioinf/Augenlicht/default.html. First, to assess the background variability due to methodological and biological factors, we averaged the values for the two LoVo replicas and compared them to the average of another two different replicates of LoVo cells, and quantified the number of genes identified as differentially expressed as a function of selected cutoff values (Figure 8AFigure 8

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