The objective of this work is to understand the nature of the DNA damage leading to MLL translocations in leukemias following anticancer treatment with DNA topoisomerase II inhibitors. The CYP3A4 promoter is polymorphic and CYP3A4 genotype confers susceptibility. CYP3A4 converts etoposide to etoposide catechol; the catechol is readily oxidized to a quinone. These metabolites are genotoxins. MLL joins with one of many partner genes to form the translocations. The genomic breakpoint junction sequences contain evidence of DNA damage and repair. Several genomic breakpoint junction sequences indicate precise or near-precise interchromosomal DNA recombinations, but the cloning of additional breakpoints is essential to discern the damage spectrum. Etoposide and its metabolites induce DNA topoisomerase II cleavage at the translocation breakpoints in MLL and in its partner genes in vitro. We propose that etoposide and its metabolites can stimulate a series of different DNA lesions, which are repaired to form the breakpoint junctions, and that the heterogeneity in genomic breakpoint junction sequences reflects heterogeneity in the damage and its resolution. The DNA lesions to be tested include the direct induction of DNA topoisomerase II cleavage by etoposide parent drug, induction of DNA topoisomerase II cleavage from DNA adduct formation by etoposide quinone or reactive oxygen species, replication fork collisions with DNA topoisomerase II covalent complexes and DNA topoisomerase II-independent damage.
Aim 1 will examine the spectrum and quantify the relative importance of DNA adducts from etoposide metabolites in an MLL bcr DNA substrate using mass spectrometry.
Aim 2 will investigate the induction of functional DNA topoisomerase II covalent complexes in MLL and in the genome by etoposide and etoposide metabolites in human CD34+ hematopoietic progenitor cells using DNA arrays. To answer whether, how often and to what degree precise recombinations, exonucleolytic nibbling, large deletions, insertions, inversions, duplications and nonhomologous end-joining have occurred in creation of the breakpoint junctions, Aim 3 will characterize the genomic sequences of both derivative chromosomes in the leukemias in patients. Solving the mechanism of leukemogenesis of the DNA topoisomerase II inhibitors is highly relevant to the targeted prevention of this usually fatal complication of anticancer treatment.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA077683-05
Application #
6788075
Study Section
Experimental Therapeutics Subcommittee 1 (ET)
Program Officer
Fu, Yali
Project Start
1999-02-16
Project End
2008-07-31
Budget Start
2004-08-01
Budget End
2005-07-31
Support Year
5
Fiscal Year
2004
Total Cost
$281,181
Indirect Cost
Name
Children's Hospital of Philadelphia
Department
Type
DUNS #
073757627
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Yu, Xiang; Davenport, James W; Urtishak, Karen A et al. (2017) Genome-wide TOP2A DNA cleavage is biased toward translocated and highly transcribed loci. Genome Res 27:1238-1249
Mays, Ashley N; Osheroff, Neil; Xiao, Yuanyuan et al. (2010) Evidence for direct involvement of epirubicin in the formation of chromosomal translocations in t(15;17) therapy-related acute promyelocytic leukemia. Blood 115:326-30
Robinson, Blaine W; Felix, Carolyn A (2009) Panhandle PCR approaches to cloning MLL genomic breakpoint junctions and fusion transcript sequences. Methods Mol Biol 538:85-114
Felix, Carolyn A (2009) A safer regimen for high-risk neuroblastoma. Pediatr Blood Cancer 53:3-6
Robinson, Blaine W; Cheung, Nai-Kong V; Kolaris, Christos P et al. (2008) Prospective tracing of MLL-FRYL clone with low MEIS1 expression from emergence during neuroblastoma treatment to diagnosis of myelodysplastic syndrome. Blood 111:3802-12
Hasan, Syed Khizer; Mays, Ashley N; Ottone, Tiziana et al. (2008) Molecular analysis of t(15;17) genomic breakpoints in secondary acute promyelocytic leukemia arising after treatment of multiple sclerosis. Blood 112:3383-90
Felix, Carolyn A; Kolaris, Christos P; Osheroff, Neil (2006) Topoisomerase II and the etiology of chromosomal translocations. DNA Repair (Amst) 5:1093-108
Zheng, Naiyu; Pang, Shaokun; Oe, Tomoyuki et al. (2006) Characterization of an etoposide-glutathione conjugate derived from metabolic activation by human cytochrome p450. Curr Drug Metab 7:897-911
Robinson, Blaine W; Slater, Diana J; Felix, Carolyn A (2006) BglII-based panhandle and reverse panhandle PCR approaches increase capability for cloning der(II) and der(other) genomic breakpoint junctions of MLL translocations. Genes Chromosomes Cancer 45:740-53
Libura, Jolanta; Slater, Diana J; Felix, Carolyn A et al. (2005) Therapy-related acute myeloid leukemia-like MLL rearrangements are induced by etoposide in primary human CD34+ cells and remain stable after clonal expansion. Blood 105:2124-31

Showing the most recent 10 out of 28 publications