Selective Cytotoxicity of the NAMPT Inhibitor FK866 Toward Gastric Cancer Cells With Markers of the Epithelial-Mesenchymal Transition, Due to Loss of NAPRT

Abstract

Background & aims: Markers of the epithelial-to-mesenchymal transition (EMT) in gastric tumor tissues are associated with poor patient outcomes. We performed a screen to identify pharmacologic compounds that kill gastric cancer cells with EMT-associated gene expression patterns and investigate their mechanisms.

Methods: We identified 29 gastric cancer cell lines with a gene expression signature previously associated with an EMT subtype, based on data from RNA sequence analyses, and confirmed the mesenchymal phenotypes of 7 lines (Hs746T, SNU1750, MKN1, SK4, SNU484, SNU668, and YCC11), based on invasive activity and protein markers. We screened 1,345 compounds for their ability to kill cells with the EMT signature compared with cell lines without this pattern. We tested the effects of identified compounds in BALB/c nude mice bearing GA077 tumors; mice were given intraperitoneal injections of the compound or vehicle (control) twice daily for 24 days and tumor growth was monitored. Proteins associated with the toxicity of the compounds were overexpressed in MKN1 and SNU484 cells or knocked down in MKN45 and SNU719 using small interfering RNAs. We performed immunohistochemical analyses of 942 gastric cancer tissues and investigated associations between EMT markers and protein expression patterns.

Results: The nicotinamide phosphoribosyltransferase inhibitor FK866 killed 6 of 7 gastric cancer cell lines with EMT-associated gene expression signatures but not gastric cancer cells without this signature. The 6 EMT-subtype gastric cell lines expressed significantly low levels of nicotinic acid phosphoribosyltransferase (NAPRT), which makes the cells hypersensitive to nicotinamide phosphoribosyltransferase inhibition. Gastric cell lines that expressed higher levels of NAPRT, regardless of EMT markers, were sensitized to FK866 after knockdown of NAPRT, whereas overexpression of NAPRT in deficient EMT cell lines protected them from FK866-mediated toxicity. Administration of FK866 to nude mice with tumors grown from GA077 cells (human gastric cancer tumors of the EMT subtype) led to tumor regression in 2 weeks; FK866 did not affect tumors grown from MKN45 cells without the EMT expression signature. Loss of NAPRT might promote the EMT, because it stabilizes β-catenin. We correlated the EMT gene expression signature with lower levels of NAPRT in 942 gastric tumors from patients; we also found lower levels of NAPRT mRNA in colorectal, pancreatic, and lung adenocarcinoma tissues with the EMT gene expression signature.

Conclusions: FK866 selectively kills gastric cancer cells with an EMT gene expression signature by inhibiting nicotinamide phosphoribosyltransferase in cells with NAPRT deficiency. Loss of NAPRT expression, frequently through promoter hypermethylation, is observed in many gastric tumors of the EMT subtype. FK866 might be used to treat patients with tumors of this subtype.

Keywords: Drug Screen; Partial Epithelial-to-Mesenchymal Transition; Stomach Cancer; Synthetic Lethal.