Abstract 1149: Consistency between genomic and proteomic profiles reveals novel molecular mechanisms of fasting antitumor activity
Authors: Raffaghello Lizzia , Bianchi Giovanna , Martella Roberto , Marimpietri Danilo , Petretto Andrea , Inglese Elvira , Amaro Adriana , Pistoia Vito , Pfeffer Ulrich , Longo Valter 
Published: August 2015

Abstract
Background/Aim
Fasting or Short term Starvation (STS) represents a novel therapeutic strategy which appears to: i) protect normal but not tumor cells against the chemotherapy-mediated cytotoxicity, ii) induce a potent chemosensitizing effect in a wide range of experimental tumor model, and iii) be feasible, safe and able to reduce common side effects induced by chemotherapy in cancer patients. In addition, fasting alone has been shown to retard in vitro and in vivo tumor growth. However, the molecular mechanisms coupling STS with antitumor activity remain only partially understood. Thus, aim of this study is to investigate the modulation of the genomic and proteomic profiles by fasting in cancer cells with particular attention to elements involved in cell growth and metabolism signaling pathways.
Methods
Tumor cell lines were cultured under control (glucose: 1 g/L + 10% fetal bovine serum) or starved (glucose: 0.5 g/L + 1% fetal bovine serum) conditions for 48 hours. The in vitro viability of STS was tested by Trypan Blue staining and Annexin V apoptosis assay. Tumor cell proliferation was evaluated by 5-bromo-2-deoxyuridine (BrdU) and Carboxyfluorescein Succinimidyl ester (CFSE) staining. Gene expression was tested by microarray analysis. Protein expression were studied by western blot and Label Free Quantitation (LFQ) on High Resolution/Mass Accuracy Liquid Chromatography Tandem Mass Spectrometry (HR/MA LC MS/MS). Proteomic data were evaluated by network analysis using Cytoscape.
Results
In vitro experiments showed that STS significantly reduced cell proliferation and promoted apoptosis. The latter effect was mediated by a fine regulation of genes and proteins involved in cell growth and metabolic signaling pathways. In particular, STS significantly reduced the expression of elements of PI3K/AKT pathway and glycolytic enzymes. This latter modulation likely occurred at the transcriptional level since the expression profiles of STS treated cells showed a clear down-regulation of the genes encoding these enzymes as compared to untreated controls. STS also down-regulated glutaminase both at the mRNA and protein level. In contrast, proteomic and genomic analyses of the expression of oxidative phosphorylation enzymes did not show a clear up-regulation effect by STS. Finally, preliminary data revealed that STS modulates genes and proteins involved in Jak/STAT, MAPK pathways and ubiquitin mediated proteolysis. Noteworthy, for 90% of genes down- or up-regulated by STS for which a protein was identified by HR/MA LC MS/MS, a corresponding mRNA level was modulated by STS with a similar trend.
Conclusions
Taken together, the genomic and proteomic profiles demonstrated that STS caused an anti-Warburg effect and downregulated proliferation and inflammatory pathways. These findings open a novel scenario in cancer treatment.
Citation Format: Lizzia Raffaghello, Giovanna Bianchi, Roberto Martella, Danilo Marimpietri, Andrea Petretto, Elvira Inglese, Adriana Amaro, Vito Pistoia, Ulrich Pfeffer, Valter Longo. Consistency between genomic and proteomic profiles reveals novel molecular mechanisms of fasting antitumor activity. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1149. doi:10.1158/1538-7445.AM2015-1149