Evaluation for synergistic suppression of breast cancer proliferation by Chloroquine in combination with the rapamycin derivative, Rad001

Fragestellung: Breast cancer remains the most prevalent cancer among women in the Western world. Interactions between the p53 and the IGF-1/PI3K/Akt/mTOR pathway are present in many cancer cells. Therapeutical modification of each of these two pathways has been proven to exert anti-tumoral effects. The macrolide derivative of rapamycin Rad001 (everolimus) is an inhibitor of the serine-threonine kinase mTOR, and has been developed as an anti-proliferative drug with applications as an immunosuppressant and anticancer agent.

The anti-malarial Chloroquine (CQ) is a 4-alkylamino substituted quinoline and up-regulates the tumor-suppressor p53, its downstream target gene p21, and causes G1 cell cycle arrest in breast cancer cells (Loehberg et al, 2007). Methodik: Cell culture (MCF7 breast cancer cell line), Cell growth assay, Flow cytometry, Western Blot Analysis. Ergebnisse: In MCF7 cells CQ and Rad001 showed synergistic reduction of cell proliferation. FACS analysis revealed that CQ and Rad001 induce increased G1 cell cycle arrest also in a synergistic manner. Western Blot analysis showed reduced activation of p70 S6 kinase, which is a mitogen activated kinase that is required for cell growth and G1 cell cycle progression and is an mTOR downstream target gene, by CQ and Rad001 treatment. We further sought to determine CQ's effects on the Rad001-induced activation of the oncogene Akt. Treatment of MCF7 cells with CQ resulted in a reduction of Rad001-induced increase of Akt phosphorylation. Schlussfolgerung: Currently, there are very few agents that have a proven ability to prevent or treat breast cancer and their mechanism of action remains undetermined.

We reasoned that possible agents would include those that augment p53-dependent pathways. Our data suggest that the p53-activator CQ might exert at least parts of its anti-cancer effects through modifications of the IGF-1/PI3K/Akt/mTOR pathway and presents itself as an interesting combination partner for the mTOR-inhibitor Rad001. Reducing the Rad001-induced Akt phosphorylation, which has been suspected to be a key mechanism of Rad001-resistence in breast cancer, might play a central. CQ is part of the promising quinoline chemical family, like e.g. the pan-PI3K-Inhibitor NVP-BEZ235, and our results, even though not completely transferable, might show an interesting proof-of-principle in designing new breast cancer therapy strategies. In further experiments we intend to examine to what extent Rad001 and CQ add to each other's effect on growth of breast cancer in an animal model.