Phillips Laboratory
Department of Biochemistry
Research - Anti-cancer drugs
The primary interest of my laboratory is to elucidate the molecular and cellular responses to clinical and experimental DNA-acting anticancer agents and to use that knowledge to devise strategies to:
1) enhance the anticancer activity of these drugs.
2) overcome resistance to these agents.
3) define the downstream molecular targets and signalling processes.
4) develop tumour-targetting procedures.
This work is carried out in collaboration with Dr Suzanne Cutts.
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We now know that the anthracycines doxorubicin (Adriamycin), epirubicin, idarubicin, daunomycin, as well as mitoxantrone, can be activated by formaldehyde to yield anthracyline-DNA covalent adducts which form preferentially at 5'-GC-3' sequences.
The formaldehyde can be readily released in cells from prodrugs such as AN-9.
The formaldehyde activation of clinical agents such as doxorubicin leads to enhanced levels of doxorubicin-DNA adducts, and this results in an enhanced cytotoxic response.
Mitoxantrone is also activated by formaldehyde and this results in the formation of unstable mitoxantrone-DNA adducts. |
Confocal, Comet and light
microscopy analysis of
drug-treated cells in culture. |
These adducts form preferentially at 5'-CG-3' sequences, and even more so when the cytosine residues are methylated. Key questions that we are now addressing include: |
1. What are the cellar responses to these drug-DNA adducts?
2. What forms of DNA damage are induced by the adducts, and how are they repaired?
3. Does the preference of mitoxantrone for methylated-CG sequences trigger gene-specific responses?
4. Can the synergistic combination of active drugs and formaldehyde release from prodrugs be harnessed to yield clinical advantages?
Techniques used to address these questions include growth inhibition and colony survival cell culture assays, flow cytometry, scintillation counting of 14C adducts, Comet assay, micronucleus assay, gene-specific crosslinking assays, quantitative PCR, real-time PCR, in vitro transcription. |
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Structures of Adriamycin and AN-9, and the pathway of Adriamycin activation by formaldehyde released by esterase-mediated hydrolysis of AN-9. Click figure to enlarge. |
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