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Bcl-2/adenovirus E1B 19 kDa-interacting protein 3 (BNIP3) expression is epigenetically regulated by one-carbon metabolism in invasive duct cell carcinoma of breast

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Abstract

In view of recent studies highlighting the prognostic relevance of expression and CpG island methylator phenotype (CIMP) of Bcl-2/adenovirus E1B 19 kDa-interacting protein 3 (BNIP3) in invasive duct cell carcinoma (IDC), we hypothesized in this article that impaired one-carbon metabolism might influence CIMP phenotype of BNIP3. In order to substantiate the prognostic relevance of BNIP3, we explored its association with 8-oxo-2′deoxyguanosine (8-oxodG), a marker of oxidative stress with prognostic relevance. BNIP3 expression and CIMP phenotype were studied using semi-quantitative RT-PCR and combined bisulfite restriction analysis (COBRA), respectively, in 56 IDC tumors. Eight polymorphisms in one-carbon metabolism were studied using PCR–RFLP and PCR–AFLP approaches. 8-oxodG was measured using competitive ELISA kit. BNIP3 was found to be upregulated in IDC (cases vs. controls: 0.94 ± 0.05 vs. 0.18 ± 0.08, P < 0.0001). COBRA analysis confirmed hypomethylation of BNIP3 promoter CpG island in these cases. CIMP phenotype of BNIP3 showed positive association with tubule formation (P = 0.034) and methionine synthase reductase (MTRR) A66G (P = 0.002); inverse association with cytosolic serine hydroxyl methyltransferase (cSHMT) C1420T (P < 0.005) and 8-oxodG (<10% vs. >10% methylation: 7.24 ± 2.77 ng/ml vs. 4.42 ± 2.93 ng/ml, P < 0.0005); and no association with nuclear pleomorphism or mitotic index or ER, PR, and HER statuses. Synergistic effect of MTR A2756G and MTRR A66G variants on BNIP3 hypermethylator phenotype was clearly evident (P < 0.0007). MTRR A66G and cSHMT C1420T polymorphisms influence CIMP phenotype of BNIP3, thus epigenetically regulating BNIP3 in breast cancer. The linear association between BNIP3 and 8-oxodG substantiates the role of BNIP3 as redox sensor as well as prognostic marker in breast cancer.

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Abbreviations

8-oxodG:

8-Oxo-2′-deoxyguanosine

AFLP:

Amplified fragment length polymorphism

BNIP3:

Bcl-2/adenovirus E1B 19 kDa-interacting protein 3

CIMP:

CpG island methylator phenotype

COBRA:

Combined bisulfite restriction analysis

cSHMT:

Cytosolic serine hydroxymethyltransferase

DCIS:

Duct cell in situ

DNMT:

DNA methyltransferases

HIF-1:

Hypoxia inducible factor-1

IDC:

Invasive duct cell carcinoma

MTHFR:

Methylene tetrahydrofolate reductase

RFLP:

Restriction fragment length polymorphism

ROS:

Reactive oxygen species

RT-PCR:

Reverse transcriptase PCR

TYMS:

Thymidylate synthase

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Acknowledgment

This study was supported by the grant funded by Indian Council of Medical Research (ICMR), New Delhi (Ref No. 5/13/32/2007). Vijay Kumar Kutala is a recipient of Ramanujan Fellowship awarded by Department of Science and Technology, Government of India.

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Authors and Affiliations

  1. Department of Clinical Pharmacology & Therapeutics, Nizam’s Institute of Medical Sciences, Panjagutta, Hyderabad, 500082, Andhra Pradesh, India

    Shaik Mohammad Naushad & Vijay Kumar Kutala

  2. Department of Pathology, Nizam’s Institute of Medical Sciences, Panjagutta, Hyderabad, 500082, Andhra Pradesh, India

    Aruna Prayaga

  3. Department of Medical Oncology, Nizam’s Institute of Medical Sciences, Panjagutta, Hyderabad, 500082, Andhra Pradesh, India

    Raghunadha Rao Digumarti

  4. Department of Surgical Oncology, Nizam’s Institute of Medical Sciences, Panjagutta, Hyderabad, 500082, Andhra Pradesh, India

    Suryanarayana Raju Gottumukkala

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  1. Shaik Mohammad Naushad
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  2. Aruna Prayaga
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  3. Raghunadha Rao Digumarti
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  4. Suryanarayana Raju Gottumukkala
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  5. Vijay Kumar Kutala
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Correspondence to Vijay Kumar Kutala.

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Naushad, S.M., Prayaga, A., Digumarti, R.R. et al. Bcl-2/adenovirus E1B 19 kDa-interacting protein 3 (BNIP3) expression is epigenetically regulated by one-carbon metabolism in invasive duct cell carcinoma of breast. Mol Cell Biochem 361, 189–195 (2012). https://doi.org/10.1007/s11010-011-1103-z

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  • DOI: https://doi.org/10.1007/s11010-011-1103-z

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