Abstract 964: Nicotinamide phosphoribosyltransferase (Nampt) regulates growth of non-small cell lung cancer cells with c-Met overexpression
Authors: Okumura Shunsuke , Minami Yoshinori , Endoh Satoshi , Sasaki Takaaki , Satoh Kazuhiro , Kitada Masahiro , Ohsaki Yoshinobu 
Published: June 2012

Background: Non-small cell lung cancer (NSCLC) often has an oncogene addiction, including epidermal growth factor receptor (EGFR) signaling or hepatocyte growth factor (HGF)/c-Met signaling. NSCLC cells with these addictions require a large amount of intracellular ATP to activate their receptor tyrosine kinases (RTKs). Nicotinamide phosphoribosyltransferase (Nampt) is an enzyme to synthesize Nicotinamide adenine dinucleotide (NAD), which regulates cellular redox reactions as well as intracellular ATP biosynthesis. We have reported that the inhibition of intracellular Nampt function suppressed tumor growth via intracellular ATP reduction in EGFR-gene mutated NSCLC. We have developed a novel Nampt inhibitor, which is encapsulated within poly (lactic-co-glycolic acid) (PLGA) microspheres, to enhance the tumoricidal activity in vivo. We hypothesized that the Nampt inhibitor has a high anti-tumor activity in NSCLC with c-MET overexpression, and we evaluated the role of Nampt in growth of NSCLC with c-Met overexpression.

Methods: In vitro study, expression of Nampt protein and mRNA in NSCLC cell lines (H1975, H1648, H820, H1993, from ATCC) were studied using western blotting and real-time RT-PCR (Lightcycler 2.0, Roche). NSCLC cells were treated with Nampt-siRNA (Invitrogen) or a Nampt inhibitor, FK866 (Axon medchem), and cell proliferation was examined using BrdU assay (Roche). Cell apoptosis in the NSCLC was detected by Flow cytometry (BD FACSAria II, BD Bioscience). Intracellular ATP levels in the treated NSCLC cells were also evaluated using bioluminescence assay (Roche). In western blotting, expression of c-MET signal proteins in the treated NSCLC was also studied. In vivo study, NSCLC cells were transplanted into 7-10 weeks-old female nude mice (CD-1Foxn1, Charles River), and the mice were treated with or without FK866. We used PLGA-FK866 in the tumor-bearing mice model. In immunohistochemistry, c-Met and MAPK signal proteins in xenograft tumors was evaluated using immunofluorescence microscopy (FV1000-D, OLYMPUS).

Results: Nampt mRNA was overexpressed in all NSCLC cell lines. BrdU assay showed that treatment of Nampt-siRNA and FK866 suppressed cell proliferation and induced apoptosis in NSCLC cells. The expression of c-MET and MAPK signals were suppressed in FK866-treated NSCLC. ATP bioluminescence assay exhibited that ATP levels were decreased in FK866-treated NSCLC (p&lt;0.05). In the tumor-bearing mice model, PLGA-FK866 suppressed growth of xenograft tumors (p&lt;0.05), and achieved downregulation of MAPK signal proteins in xenografts.

Conclusion: In the present study, we founded that the Nampt inhibitor suppressed NSCLC growth via suppression of intracellular ATP levels, and the expression of c-Met and MAPK signal proteins. These results suggest that Nampt has a potential to become a novel therapeutic target in NSCLC with c-MET overexpression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 964. doi:10.1158/1538-7445.AM2011-964