The objective of this work is to better understand the molecular genetic changes in leukemia in infants. Translocations of the MLL gene at chromosome band 11q23 are present in most cases and are considered primary alterations. The translocations frequently occur in utero, suggesting that prenatal exposures to DNA topoisomerase II inhibitors may be relevant etiologic factors, as leukemias in infants resemble leukemias linked to chemotherapy that targets DNA topoisomerase II. Alternatively, abasic sites are the most common form of spontaneous DNA damage and abasic sites may affect the DNA topoisomerase II cleavage- religation equilibrium. We hypothesize that the MLL genomic translocation mechanism involves chromosomal breakage by DNA topoisomerase II followed by recombination of DNA free ends through DNA repair. We have shown a correspondence between DNA topoisomerase II cleavage sites and some MLL genomic translocation breakpoints. As signs of DNA repair, we have recognized untemplated nucleotides at the breakpoint junctions and sequence similarities between MLL and partner genes that suggest homologous end-joining. Latency to the onset of leukemia has implied that secondary genetic changes in addition to the translocations may be of importance, but the candidate gene approach to investigations of the secondary changes has been inconclusive. Although MLL has many different partner genes, the influence of various translocation partners on sufficiency of MLL gene translocations for full leukemogenesis has not been assessed. We hypothesize that there are secondary changes and that the necessity for such changes depends, in part, on the translocation partner.
The first aim uses a new approach called panhandle variant PCR to clone MLL genomic breakpoints, characterize the partner genes, and examine breakpoint junctions.
The second aim addresses whether the translocation breakpoints are functional DNA topoisomerse II cleavage sites and examines the effects on cleavage of introduction of abasic sites at the translocation breakpoints. If the first hypothesis is correct, the translocation breakpoints should coincide with DNA topoisomerase II cleavage sites and abasic sites should enhance the cleavage.
The third aim uses cDNA microarray technology to identify secondary genetic changes in leukemias with MLL gene rearrangements involving different translocation partners. If the second hypothesis is correct, there will be modulations of gene expression that reflect the secondary changes, and the kinds of secondary changes will vary with the partner gene. This work will increase our understanding of the etiology of MLL gene translocations and identify secondary genetic changes in leukemia in infants.
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