Abstract 3374: Fasting chemosensitizes tumor cells by affecting their metabolism
Authors: Bianchi Giovanna , Martella Roberto , Ravera Silvia , Petretto Andrea , Marimpietri Danilo , Emionite Laura , Capitanio Selene , Bottone Gianluca , Orengo Annamaria , Marini Cecilia , Sambuceti Gianmario , Pistoia Vito , Longo Valter D. , Raffaghello Lizzia 
Published: May 2015

Abstract
Background/Aim
The metabolic properties of cancer cells are significantly different from those of normal cells. Energy production in cancer cells is dependent on aerobic glycolysis, fatty acid synthesis and increased rates of glutamine metabolism. Emerging evidences indicate that these features are linked to therapeutic resistance in cancer treatment. Thus, novel therapeutic strategies able to modify tumor metabolism are warranted.
Aim of this study is to investigate the effects and underlying mechanisms of fasting in combination with chemotherapy on tumor metabolism.
Methods
The in vitro citotoxicity of chemodrugs in combination with short-term-starvation (STS) was tested in various colon cancer cell lines by Trypan Blue staining and Annexin V apoptosis assay. Cancer cell proliferation by BrdUdr and CFSE staining was also evaluated. Reactive oxygen species production were evaluated by DCFDA staining. The effects of STS on the regulation of tumor metabolism were evaluated by 2-[(18)F]-Fluoro-2-deoxyglucose uptake. Expression of different proteins involved in metabolic and signaling pathways were studied by western blot and proteomic analyses. In vivo studies were performed using syngeneic BALB/c mice which were subcutaneously injected with CT26 cells. The in vivo STS protocol allowed mice to consume water but not food for 48 hours before chemotherapy. All mice were monitored daily for weight loss and side effects. Tumor growth was monitored every two days by caliper. The effects of STS in combination with chemotherapy (CT), in terms of glucose uptake/consumption and tumor growth inhibition were also evaluated by dynamic micro-Positron Emission Tomography (micro-PET).
Results
In vitro experiments showed that STS+CT maximally reduced cell viability and glucose uptake. A metabolic switch from anaerobic glycolysis to oxidative phosphorilation (OXPHOSPH) and beta-lipidic oxidation was induced by STS+CT. Down-regulation of PI3K/AKT pathway by STS+CT was found to be involved in the reduction of different glycolytic enzymes including hexokinases, phosphofructokinase and piruvate kinase. Furthermore, tumor cells treated with STS+CT exhibited a significant induction of respiratory chain complexes I, III, IV associated to a striking uncoupling between oxygen consumption which was induced and ATP synthesis that was reduced. This latter event was related to significant increase of ROS generation by STS+CT, leading to tumor cell apoptosis. Accordingly, in vivo experiments demonstrated that STS in combination with high-dose CT maximally reduced tumor glucose consumption in parallel with tumor cell growth.
Conclusions
Taken together, these results demonstrated that STS+CT affected tumor metabolism leading to a significant decrease of tumor growth. These findings open a novel scenario in cancer treatment.
Citation Format: Giovanna Bianchi, Roberto Martella, Silvia Ravera, Andrea Petretto, Danilo Marimpietri, Laura Emionite, Selene Capitanio, Gianluca Bottone, Annamaria Orengo, Cecilia Marini, Gianmario Sambuceti, Vito Pistoia, Valter D. Longo, Lizzia Raffaghello. Fasting chemosensitizes tumor cells by affecting their metabolism. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3374. doi:10.1158/1538-7445.AM2014-3374