Low levels of ATM in breast cancer patients with clinical radiosensitivity.

BACKGROUND AND PURPOSE: Adjuvant radiotherapy for cancer can result in severe adverse side effects for normal tissues. In this respect, individuals with anomalies of the ATM (ataxia telangiectasia) protein/gene are of particular interest as they may be at risk of both breast cancer and clinical radiosensitivity. The association of specific ATM gene mutations with these pathologies has been well documented, however, there is uncertainty regarding pathological thresholds for the ATM protein. RESULTS: Semi-quantitative immuno-blotting provided a reliable and reproducible method to compare levels of the ATM protein for a rare cohort of 20 cancer patients selected on the basis of their severe adverse normal tissue reactions to radiotherapy. We found that 4/12 (33%) of the breast cancer patients with severe adverse normal tissue reactions following radiotherapy had ATM protein levels < 55% compared to the mean for non-reactor controls. CONCLUSIONS: ATM mutations are generally considered low risk alleles for breast cancer and clinical radiosensitivity. From results reported here we propose a tentative ATM protein threshold of ~55% for high-risk of clinical radiosensitivity for breast cancer patients.

Potent inhibition of tumoral hypoxia-inducible factor 1alpha by albendazole.

BACKGROUND: Emerging reports suggest resistance, increased tumor invasiveness and metastasis arising from treatment with drugs targeting vascular endothelial growth factor (VEGF). It is believed that increased tumoral hypoxia plays a prominent role in the development of these phenomena. Inhibition of tumoral hypoxia inducible factor (HIF-1alpha) is thus becoming an increasingly attractive therapeutic target in the treatment of cancer. We hypothesized that the anti-VEGF effect of albendazole (ABZ) could be mediated through inhibition of tumoral HIF-1alpha. METHOD: In vitro, the effects of ABZ on HIF-1alpha levels in human ovarian cancer cells (OVCAR-3) were investigated using hypoxic chamber or desferrioxamine (DFO) induced-hypoxia. In vivo, the effects of ABZ (150 mg/kg, i.p., single dose) on the tumor levels of HIF-1alpha and VEGF protein and mRNA were investigated by western blotting, RT-PCR and real time-PCR. RESULTS: In vitro, ABZ inhibited cellular HIF-1alpha protein accumulation resulting from placement of cells under hypoxic chamber or exposure to DFO. In vivo, tumors excised from vehicle treated mice showed high levels of both HIF-1alpha and VEGF. Whereas, tumoral HIF-1alpha and VEGF protein levels were highly suppressed in ABZ treated mice. Tumoral VEGFmRNA (but not HIF-1alphamRNA) was also found to be highly suppressed by ABZ. CONCLUSION: These results demonstrate for the first time the effects of an acute dose of ABZ in profoundly suppressing both HIF-1alpha and VEGF within the tumor. This dual inhibition may provide additional value in inhibiting angiogenesis and be at least partially effective in inhibiting tumoral HIF-1alpha surge, tumor invasiveness and metastasis.

Albendazole-cyclodextrin complex: enhanced cytotoxicity in ovarian cancer cells.

BACKGROUND: Over recent years, we have identified a potentially new indication for albendazole (ABZ) namely that of an anticancer agent. Our recent data indicate that besides regional use, the drug is quite likely to be useful as a systemic anticancer agent. However, with extremely low solubility, ABZ has to be prepared in a biocompatible solubilized form before any systemic evaluation is possible. The present study aimed at preparing soluble ABZ and evaluating its in vitro antiproliferative efficacy and toxicity. EXPERIMENTAL DESIGN: Using beta-cyclodextrins (CDs), various formulations of ABZ were prepared and tested in cell culture for antiproliferative efficacy, cell integrity and cell toxicity against human ovarian cancer cell lines 1A9, OVCAR-3 and SKOV-3. Hepatocytes isolated from patients undergoing liver tumor resection were used for toxicity evaluations. RESULTS: Treatment of tumor cells with ABZ-CD + citric acid (CA) solution led to dose-dependent inhibition of cell proliferation. Compared to an ethanolic solution of ABZ, ABZ-CD + CA increased the antiproliferative efficacy of ABZ. Furthermore, in contrast to the ethanolic solution, ABZ-CD-CA complex profoundly (p<0.001) reduced the number of OVCAR-3 colonies formed. Fresh human hepatocytes exposed for 3 days to the highest ABZ concentration used in the study (1 microM), revealed no drug toxicity. CONCLUSION: Complexation of ABZ with beta-cyclodextrin leads to the formation of an ABZ solution with potent antiproliferative effects. This solution may find clinical value as an intravenous anticancer agent.

Epothilone-paclitaxel resistant leukemic cells CEM/dEpoB300 are sensitive to albendazole: Involvement of apoptotic pathways.

Altered or deficient activation of apoptosis signalling pathways may contribute to drug resistance. Here, we assess the role of apoptotic mediators in eliciting an anti-proliferative response to paclitaxel (PTX) in a T cell acute lymphoblastic leukemia (ALL) cell line CEM and its epothilone-paclitaxel resistant sub-line CEM/dEpoB300. Furthermore, the cellular response to PTX was compared to those elicited by cells in response to treatment with albendazole (ABZ; a microtubule depolymerizing agent). In cell proliferation studies, CEM cells were sensitive to both PTX and ABZ, while the CEM/dEpoB300 cells were highly resistant to PTX (IC(50) 2.86 nM versus 30.26 nM, respectively). In contrast, the resistant cells showed a 2-fold increase in sensitivity to ABZ (0.32 microM in CEM compared to 0.16 microM in CEM/dEpoB300). Analysis of caspase-3 activity and cytochrome c release in response to PTX or ABZ treatment (24, 48 and 72 h) revealed that, compared to the parent cells, the resistant cells have diminished response to PTX and enhanced response to ABZ. A similar pattern was observed for the pro-apoptotic protein Bax. Levels of the anti-apoptotic protein Bcl-2 was highly elevated in CEM/dEpoB300 cells and in these cells, ABZ was more effective in lowering the Bcl-2 levels than PTX. Similarly, ABZ treatment led to profound down regulation of the Mcl-1 protein. These results reveal for the first time, the changes in apoptotic mediators following development of resistance to PTX in an ALL cell and the significantly increased sensitivity of these PTX resistant cells to ABZ.

The chaperone function of cyclophilin 40 maps to a cleft between the prolyl isomerase and tetratricopeptide repeat domains.

Cyclophilin 40 (CyP40), an immunophilin cochaperone present in steroid receptor-Hsp90 complexes, contains an N-terminal peptidylprolyl isomerase (PPIase) domain separated from a C-terminal Hsp90-binding tetratricopeptide repeat (TPR) domain by a 30-residue linker. To map CyP40 chaperone function, CyP40 deletion mutants were prepared and analysed for chaperone activity. CyP40 fragments containing the PPIase domain plus linker or the linker region and the adjoining TPR domain retained chaperone activity, whilst individually, the catalytic and TPR domains were devoid of chaperoning ability. CyP40 chaperone function then, is localized within the linker that forms a binding cleft with potential to accommodate non-native substrates.