Vlahos, N. S., Futscher, B. W., Hora, N. K., Trent, J. M., & Erickson, L. C. (1990). Gene amplification affecting O6-alkylguanine-DNA alkyltransferase activity is not detected in nitrosourea resistant or sensitive human cell lines. Carcinogenesis, 11(3), 479-483.
PMID: 2311191;Abstract:
An attempt was made to characterize the genetic regulation of the human DNA repair protein O6-alkylguanine-DNA alkyltransferase (AGT) in the absence of the cloned gene. Four human cell lines, differing in AGT activity from very proficient to essentially absent, were assayed for gene amplification as a possible mediator of the methylation repair phenotype (Mer+, AGT activity and MER-, no AGT activity) using in-gel DNA renaturation and G-banded karyotype analysis. The former technique allows subsequent analysis of amplification units and cloning of observed amplified DNA fragments, a hopeful approach to the isolation of the human AGT gene. Within the sensitivities of the techniques, no correlation between AGT activity and gene amplification was observed in the four cell lines tested.
Pieper, R. O., Fuscher, B. W., Dong, Q., & Th.Erickson, M. E. (1990). Comaparison of 0-6-methylguanine DNA methyltransferase (MGMT) mRNA levels in Mer+ and Mer- human tumor cell lines containing the MGMT gene by the polymerase chain reaction technique. Cancer Communications, 2(1), 13-20.
Danks, M. K., Warmoth, M. R., Friche, E., Granzen, B., Bugg, B. Y., Harker, W. G., Zwelling, L. A., Futscher, B. W., Suttle, D. P., & Beck, W. T. (1993). Single-strand conformational polymorphism analysis of the Mr 170,000 isozyme of DNA topoisomerase II in human tumor cells. Cancer Research, 53(6), 1373-1379.
PMID: 8383009;Abstract:
Five cell lines selected for resistance to the cytotoxicity of inhibitors of DNA topoisomerase II have point mutations in the gene that codes for the Mr 170,000 form of this enzyme. In each case, the mutation results in an amino acid change in or near an ATP binding sequence of the Mr 170,000 isozyme of topoisomerase II. We used single-strand conformational polymorphism analysis to screen for similar mutations in other drug-resistant cell lines or in leukemic cells from patients previously treated with etoposide or teniposide. We also analyzed the region of the gene that codes for amino acids adjacent to the tyrosine at position 804 of topoisomerase II which binds covalently to DNA. CEM/VM-1, CEM/VM-1-5, and HL-60/AMSA human leukemic cell lines were used as controls; 3 of 3 known mutations were detected by migration differences of polymerase chain reaction products from the RNA extracted from these three lines. A previously unknown mutation was found in the tyrosine 804 region of the Mr 170,000 topoisomerase II expressed by CEM/VM-1 and CEM/VM-1-5 cells. Sequence analysis showed that substitution of a T for a C at nucleolide 2404 resulted in an amino acid change of a serine for a proline at amino acid 802. No mutations in any of the ATP binding sequences or in the tyrosine 804 region were detected in polymerase chain reaction products from RNA extracted from human leukemia HL-60/MX2 or CEM/MX1 cells (both cell lines selected for resistance to mitoxantrone) or in human myeloma 8226/Dox1V cells (selected for resistance by simultaneous exposure to doxorubicin and verapamil). No mutations were detected in polymerase chain reaction products from RNA extracted from blasts of 15 patients with relapsed acute lymphocytic leukemia, previously treated with etoposide or teniposide. We conclude that: (a) single-strand conformational polymorphism analysis is useful for screening for mutations in topoisomerase II; (b) resistance to the cytotoxicity of inhibitors of DNA topoisomerase II is not always associated with mutations in ATP binding sequences or the active site tyrosine region of Mr 170,000 topoisomerase II; and (c) mutations similar to those detected in drug resistant cells selected in culture have not been identified in blast cells from patients with relapsed acute lymphocytic leukemia, previously treated with etoposide or teniposide.
Futscher, B. W., O'Meara, M. M., Kim, C. J., Rennels, M. A., Di, L. u., Gruman, L. M., E., R., J., M., & Domann, F. E. (2004). Aberrant methylation of the maspin promoter is an early event in human breast cancer. Neoplasia, 6(4), 380-389.
PMID: 15256060;PMCID: PMC1502109;Abstract:
The maspin gene functions as a tumor suppressor in human breasts, and its expression is frequently lost during breast cancer progression. In vitro models of human breast cancer indicate that the loss of maspin expression is closely linked to aberrant methylation of the maspin promoter. We conducted a study on 30 archival ductal carcinoma in situ (DCIS) specimens to determine if aberrant methylation of the maspin promoter occurred in vivo, and whether it occurred early in breast cancer evolution. Healthy tissue obtained from reduction mammoplasty was used as normal control. Results from immunohistochemical analysis indicate that maspin expression is lost in a substantial fraction of DCIS specimens (57%). Bisulfite sequencing of DNA isolated from laser capture-microdissected normal and neoplastic ducts showed that loss of maspin expression was often, but not always, linked to aberrant methylation of the maspin promoter, suggesting that other mechanisms, in addition to aberrant methylation, participate and/or cooperate to silence maspin gene expression. Taken together, these results indicate that aberrant methylation of the maspin promoter is an early event in human breast cancer.
Futscher, B. W. (2016). Epigenetic silencing of MORT is an early lesion in cancer and is associated with luminal, receptor positive, breast tumor subtypes. Journal of Breast Cancer.