Role of the bleomycin C-termini in DNA binding and toxicity by metallo-bleomycins

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DESCRIPTION (provided by applicant): There is a fundamental gap in definitely establishing the binding mode characteristic of metallo-bleomycins (MBLMs) to DNA. This gap represents an important problem, since the drugs' C-terminus substituents (tails), which participate on DNA binding by BLM, have also been identified as key factors in the pulmonary toxicity attributed to BLMs. Therefore, this gap hinders the understanding of structure/toxicity correlations for these anticancer drugs. The long-term goal is to better understand the binding of biologically relevant MBLMs to DNA, through an approach that can separate the various factors that affect it, and focusing on the impact of the tails in this binding for BLMs producing different levels of pulmonary fibrosis. The objective in this particular application is to determine how the chemical structures of the BLM tails influence structural interactions between FeIIBLM and DNA, independently of the DNA base sequence, and considering tails with different degrees of pulmonary toxicity. The central hypothesis is that BLM tails with different chemical structures wil correlate with the atoms in DNA in specific ways, and generate different solution structures of the (C-terminus)- DNA regions for the corresponding MBLM-DNA complexes. We have formulated this hypothesis based upon preliminary findings that suggest that the BLM tail can anchor MBLM to DNA, and affect the way in which the rest of the C-terminus of the drug is positioned in the DNA helix. The rationale for the proposed research is that assessing the DNA-binding modes BLMs proposed to cause various levels of pulmonary fibrosis in mice will serve two purposes: 1) to better establish the structural basis for molecular recognition between MBLM and DNA, and 2) to help delineate structure/toxicity correlations for these drugs. Guided by strong preliminary data, this hypothesis will be tested by pursuing two specific aims: 1) Identify interactions between MBLM and DNA in solution using two distinct DNA segments, each containing a specific BLM binding site, and relevant MBLMs with tails with different structures and toxicities; and 2) Characterize the effect of the BLM tail in the structural interactions between MBLM and DNA through molecular modeling of the MBLM-DNA complexes considered. Under the first aim, NMR spectroscopy applied to paramagnetic molecules such as the biologically relevant MBLMs (FeIIBLMs), will be used to identify atom-atom contacts between the MBLMs considered and DNA. Under the second aim, molecular dynamics calculations will be used to find the solution structures of the MBLM-DNA complexes considered. This approach has been established as feasible in the applicant's hands. The proposed research is innovative because it only considers biologically relevant MBLMs, which have no received sufficient attention from the structural point of view due to their paramagnetic nature, and it isolates the factors affecting DNA binding by BLMs. This contribution will be significant because it is expected to vertically advance and expand the understanding of toxicity from the structural point of view, required to guide the development of BLM analogs with milder side effects for cancer patients.