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Identification of 5-Iodotubercidin as a genotoxic drug with anti-cancer potential.

Zhang X, Jia D, Liu H, Zhu N, Zhang W, Feng J, Yin J, Hao B, Cui D, Deng Y, Xie D, He L, Li B - PLoS ONE (2013)

Bottom Line: We found that 5-Iodotubercidin could cause DNA damage, verified by induction of DNA breaks and nuclear foci positive for γH2AX and TopBP1, activation of Atm and Chk2, and S15 phosphorylation and up-regulation of p53.Moreover, 5-Iodotubercidin showed anti-tumor activity as it could reduce the tumor size in carcinoma xenograft mouse models in p53-dependent and -independent manners.These findings reveal 5-Iodotubercidin as a novel genotoxic drug that has chemotherapeutic potential.

View Article: PubMed Central - PubMed

Affiliation: Bio-X Institutes, Key Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders, Ministry of Education, Shanghai Jiao Tong University, Shanghai, China.

ABSTRACT
Tumor suppressor p53, which is activated by various stress and oncogene activation, is a target for anti-cancer drug development. In this study, by screening panels of protein kinase inhibitors and protein phosphatase inhibitors, we identified 5-Iodotubercidin as a strong p53 activator. 5-Iodotubercidin is purine derivative and is used as an inhibitor for various kinases including adenosine kinase. We found that 5-Iodotubercidin could cause DNA damage, verified by induction of DNA breaks and nuclear foci positive for γH2AX and TopBP1, activation of Atm and Chk2, and S15 phosphorylation and up-regulation of p53. As such, 5-Iodotubercidin induces G2 cell cycle arrest in a p53-dependent manner. Itu also induces cell death in p53-dependent and -independent manners. DNA breaks were likely generated by incorporation of 5-Iodotubercidin metabolite into DNA. Moreover, 5-Iodotubercidin showed anti-tumor activity as it could reduce the tumor size in carcinoma xenograft mouse models in p53-dependent and -independent manners. These findings reveal 5-Iodotubercidin as a novel genotoxic drug that has chemotherapeutic potential.

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Itu generates DNA damage.A. Representative pictures of the karyotypes of HCT116 in the absence or presence of Itu. B. Detection of Itu derivative in genomic DNA in replicating cells. HPLC-MS analysis of Itu metabolites in the genomic DNA of Itu-treated cells. Genomic DNA was isolated from Itu-treated cells and digested into nucleosides, which were analyzed with HPLC (left panel), which was directly linked to mass spectrometer. MS analysis of the molecule with the size of dTU was shown at the right panel. C. The intracellular dATP and dGTP pools were not affected by Itu treatment. A and A1 are derivatives of dATP and G, G1 and G2 are derivatives of dGTP.
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pone-0062527-g002: Itu generates DNA damage.A. Representative pictures of the karyotypes of HCT116 in the absence or presence of Itu. B. Detection of Itu derivative in genomic DNA in replicating cells. HPLC-MS analysis of Itu metabolites in the genomic DNA of Itu-treated cells. Genomic DNA was isolated from Itu-treated cells and digested into nucleosides, which were analyzed with HPLC (left panel), which was directly linked to mass spectrometer. MS analysis of the molecule with the size of dTU was shown at the right panel. C. The intracellular dATP and dGTP pools were not affected by Itu treatment. A and A1 are derivatives of dATP and G, G1 and G2 are derivatives of dGTP.

Mentions: Itu has a purine-like structure with “N-H” being replaced by “C-I” at the 7th position of the purine nucleoside (Fig. 1A). Itu might be incorporated into DNA after being converted to deoxyribose by ribonucleotide reductase [4]. Itu theoretically could form 2 hydrogen bonds with thymidine in double stranded DNA (Fig. S1). However, Itu-T pairing can cause a spacing problem as “C-I” of Itu (position 7) is much bigger than “N-H” of adenosine, and the altered purine ring may not be recognizable by DNA polymerases. Moreover, “C-I” of Itu is rather active and unstable. Itu is thus likely to be metabolized and subsequently incorporated into DNA to cause DNA damage. To confirm this, we first tested whether Itu could alter the karyotypes of HCT116 cells. The cells were first treated with Itu for 36 hrs and then with spindle poison colchicine to condense the chromosomes. The number and appearance of the chromosomes were analyzed under a light microscope after gimsa staining. It is obvious that Itu treatment led to the appearance of broken chromosomes in 32% of Itu-treated cells compared to 0% in untreated cells (Fig. 2A), suggesting that Itu could cause double stranded DNA breaks.


Identification of 5-Iodotubercidin as a genotoxic drug with anti-cancer potential.

Zhang X, Jia D, Liu H, Zhu N, Zhang W, Feng J, Yin J, Hao B, Cui D, Deng Y, Xie D, He L, Li B - PLoS ONE (2013)

Itu generates DNA damage.A. Representative pictures of the karyotypes of HCT116 in the absence or presence of Itu. B. Detection of Itu derivative in genomic DNA in replicating cells. HPLC-MS analysis of Itu metabolites in the genomic DNA of Itu-treated cells. Genomic DNA was isolated from Itu-treated cells and digested into nucleosides, which were analyzed with HPLC (left panel), which was directly linked to mass spectrometer. MS analysis of the molecule with the size of dTU was shown at the right panel. C. The intracellular dATP and dGTP pools were not affected by Itu treatment. A and A1 are derivatives of dATP and G, G1 and G2 are derivatives of dGTP.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3646850&req=5

pone-0062527-g002: Itu generates DNA damage.A. Representative pictures of the karyotypes of HCT116 in the absence or presence of Itu. B. Detection of Itu derivative in genomic DNA in replicating cells. HPLC-MS analysis of Itu metabolites in the genomic DNA of Itu-treated cells. Genomic DNA was isolated from Itu-treated cells and digested into nucleosides, which were analyzed with HPLC (left panel), which was directly linked to mass spectrometer. MS analysis of the molecule with the size of dTU was shown at the right panel. C. The intracellular dATP and dGTP pools were not affected by Itu treatment. A and A1 are derivatives of dATP and G, G1 and G2 are derivatives of dGTP.
Mentions: Itu has a purine-like structure with “N-H” being replaced by “C-I” at the 7th position of the purine nucleoside (Fig. 1A). Itu might be incorporated into DNA after being converted to deoxyribose by ribonucleotide reductase [4]. Itu theoretically could form 2 hydrogen bonds with thymidine in double stranded DNA (Fig. S1). However, Itu-T pairing can cause a spacing problem as “C-I” of Itu (position 7) is much bigger than “N-H” of adenosine, and the altered purine ring may not be recognizable by DNA polymerases. Moreover, “C-I” of Itu is rather active and unstable. Itu is thus likely to be metabolized and subsequently incorporated into DNA to cause DNA damage. To confirm this, we first tested whether Itu could alter the karyotypes of HCT116 cells. The cells were first treated with Itu for 36 hrs and then with spindle poison colchicine to condense the chromosomes. The number and appearance of the chromosomes were analyzed under a light microscope after gimsa staining. It is obvious that Itu treatment led to the appearance of broken chromosomes in 32% of Itu-treated cells compared to 0% in untreated cells (Fig. 2A), suggesting that Itu could cause double stranded DNA breaks.

Bottom Line: We found that 5-Iodotubercidin could cause DNA damage, verified by induction of DNA breaks and nuclear foci positive for γH2AX and TopBP1, activation of Atm and Chk2, and S15 phosphorylation and up-regulation of p53.Moreover, 5-Iodotubercidin showed anti-tumor activity as it could reduce the tumor size in carcinoma xenograft mouse models in p53-dependent and -independent manners.These findings reveal 5-Iodotubercidin as a novel genotoxic drug that has chemotherapeutic potential.

View Article: PubMed Central - PubMed

Affiliation: Bio-X Institutes, Key Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders, Ministry of Education, Shanghai Jiao Tong University, Shanghai, China.

ABSTRACT
Tumor suppressor p53, which is activated by various stress and oncogene activation, is a target for anti-cancer drug development. In this study, by screening panels of protein kinase inhibitors and protein phosphatase inhibitors, we identified 5-Iodotubercidin as a strong p53 activator. 5-Iodotubercidin is purine derivative and is used as an inhibitor for various kinases including adenosine kinase. We found that 5-Iodotubercidin could cause DNA damage, verified by induction of DNA breaks and nuclear foci positive for γH2AX and TopBP1, activation of Atm and Chk2, and S15 phosphorylation and up-regulation of p53. As such, 5-Iodotubercidin induces G2 cell cycle arrest in a p53-dependent manner. Itu also induces cell death in p53-dependent and -independent manners. DNA breaks were likely generated by incorporation of 5-Iodotubercidin metabolite into DNA. Moreover, 5-Iodotubercidin showed anti-tumor activity as it could reduce the tumor size in carcinoma xenograft mouse models in p53-dependent and -independent manners. These findings reveal 5-Iodotubercidin as a novel genotoxic drug that has chemotherapeutic potential.

Show MeSH
Related in: MedlinePlus