Alexandra Colquhoun

Comet assay measures of DNA damage are predictive of bladder cancer cell treatment sensitivity in vitro and outcome in vivo

Bladder cancer patients suffer significant treatment failure, including high rates of recurrence and poor outcomes for advanced disease. If mechanisms to improve tumour cell treatment sensitivity could be identified and/or if tumour response could be predicted, it should be possible to improve local‐control and survival. Previously, we have shown that radiation‐induced DNA damage, measured by alkaline Comet assay (ACA), correlates bladder cancer cell radiosensitivity in vitro. In this study we first show that modified‐ACA measures of cisplatin and mitomycin‐C‐induced damage also correlate bladder cancer cell chemosensitivity in vitro, with essentially the same rank order for chemosensitivity as for radiosensitivity. Furthermore, ACA studies of radiation‐induced damage in different cell‐DNA substrates (nuclei, nucleoids and intact parent cells) suggest that it is a feature retained in the prepared nucleoids that is responsible for the relative damage sensitivity of bladder cancer cells, suggestive of differences in the organisation of DNA within resistant vs. sensitive cells. Second, we show that ACA analysis of biopsies from bladder tumours reveal that reduced DNA damage sensitivity associates with poorer treatment outcomes, notably that tumours with a reduced damage response show a significant association with local recurrence of non‐invasive disease and that reduced damage response was a better predictor of recurrence than the presence of high‐risk histology in this cohort. In conclusion, this study demonstrates that mechanisms governing treatment‐induced DNA damage are both central to and predictive of bladder cancer cell treatment sensitivity and exemplifies a link between DNA damage resistance and both treatment response and tumour aggression.

Dutasteride and Metformin Reduce the Growth of LNCaP Cells and Alter the SREBP-1 Pathway

Prostate cancer (PCa) is the second most prevalent cancer in men after lung cancer. Prostate cancer develop- ment and progression is associated with the dysregulation of a number of molecular pathways; hence, therapeutic strate- gies targeting such pathways bring great promise. Recently we have shown that metformin, the anti-diabetic drug, can in- hibit tumor progression when combined with dutasteride, a 5-alpha-reductase inhibitor (5ARI). Interestingly, both met- formin and dutasteride have been reported to alter the Sterol Regulatory Element Binding Proteins (SREBP) Fatty Acid Synthase (FASN) pathway. The SREBP pathway is involved with lipid and energy homeostasis. In our present study, we investigated if dutasteride in combination with metformin can reduce the proliferation of LNCaP PCa cells, and whether this is mediated through the SREBP-1/FASN pathway. Human PCa cells were treated with either dutasteride (0-100 µM) or metformin (0-50 mM) alone or in combination. The treated cells were then incubated for up to 24 hours, and prolifera- tion assessed using the MTS 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazo- lium) assay. Western blot analysis was performed on cell lysates to assess alterations in key signaling molecules including cleaved SREBP-1, FASN, AR, PSA, pAMPK and apoptotic markers. Results revealed that there was a significant (p<0.05) decrease in cellular proliferation of LNCaP cells treated with a combination of metformin and dutasteride, this effect being greater than either treatment alone. Treating LNCaP cells with both metformin and dutasteride reduced the expression of FASN, cleaved SREBP-1 and pro-caspase-3 with expression of cleaved PARP; suggesting a possible inter- action between FASN and apoptosis. All treatments resulted in reductions in AR, PSA and an upregulation of pAMPK, with the highest expression seen in combination treatment. We report for the first time that metformin and dutasteride in combination can reduce the proliferation of androgen-sensitive cell line through the activation of p-AMPK, and SREBP- 1/FASN pathway and highlight the importance of targeting the SREBP-1 pathway, for improving future therapeutic strate- gies for prostate cancer.

84 THE USE OF MRI-GUIDED TRANSURETHRAL ULTRASOUND THERAPY FOR THE TREATMENT OF LOCALISED PROSTATE CANCER: A PHASE I STUDY

Minimally-invasive treatments for localized prostate cancer, that offer good local control over disease with a low side-effect profile, would have a major impact in improving the management of this disease. MRI-guided transurethral ultrasound therapy is a candidate technology in which planar high-intensity ultrasound energy is delivered to the prostate gland, via a transurethrally inserted device to generate a precise region of thermal coagulation (figure 1). Previous studies in gel models and canines have demonstrated the feasibility of using active MR temperature feedback to control spatial heating to a specified target region. This phase I study was designed to evaluate the safety and feasibility of this technique in humans.