2014;74:3306\3316. demonstrates that it is possible to target FGF19 for anti\HCC therapies without adversely affecting its normal bile acid regulatory function, and highlights the exciting promise of G1A8 or HS29 as potential therapy for HCC. Keywords: antibody therapy, bile acid, drug development, FGF19, HCC Fibroblast growth factor?19 (FGF19) is a driver in hepatocellular carcinoma (HCC); however, it exerts important physiological function in regulation of bile acid synthesis. Our study, for the first time, shows that selectively targeting a defined region of FGF19 by our newly developed Abs could provide an effective and safe novel treatment option for HCC. 1.?INTRODUCTION Liver cancer ranks as the third most common cause of cancer\related deaths.1 Among all liver cancers, hepatocellular carcinoma (HCC) is the most common type; it accounts for approximately 90% of all liver cancer cases.2 Currently, two tyrosine kinase inhibitors (TKIs), sorafenib and lenvatinib, are the only systemic brokers approved by the FDA as first\line treatment of HCC. They can only extend patient survival by approximately 3?months.3, 4 Other drugs in second\line therapy, including several TKIs and immune checkpoint inhibitors, show slight survival advantages and antitumor activities, have limited therapeutic efficacy.5 Thus, there is obviously a need for more effective therapeutic agents for HCC patients. Several studies have identified that HCC tumors feature highly focal amplification of fibroblast growth factor?19 (FGF19),6, 7, 8, 9 a bile acid\induced and ileum\derived peptide growth factor that functions to regulate bile acid metabolism. FGF19 binds to its receptor, hepatocyte\expressed FGF receptor?4 (FGFR4), and its co\receptor, \klotho (KLB), to repress the hepatic transcription of a gene encoding cholesterol\7\alpha\hydroxylase 1 (CYP7A1), an essential enzyme for bile acid biosynthesis.10, 11, 12, LP-935509 13, 14 In addition to LP-935509 its bile\acid\regulatory function, both FGF19 and its cognate receptor FGFR4 are highly expressed in tumors compared to adjacent non\tumorous tissues.15, 16 The high expression of these proteins promotes tumor progression; moreover, it is also associated with poor prognosis in HCC patients.16, 17 In transgenic mice, the overexpression of FGF19 caused hepatocellular dysplasia, neoplasia, and ultimately HCC,18 yet these outcomes were abolished in FGFR4 knockout mice,19 thus mechanistically confirming the tumorigenic activity of aberrant FGF19/FGFR4 signaling. Although several selective FGFR4 small molecule inhibitors are under development for the treatment of HCC, each of these elevates bile acid synthesis and causes liver toxicity (either in preclinical animal models or in early human clinical trials).20, 21, 22, 23, 24 Another drug development effort directly targeting FGF19 for treating HCC was based on a neutralizing anti\FGF19 Ab, 1A6. Treatment with 1A6 prevented transgenic mice overexpressing FGF19 from developing HCC, and such treatment also suppressed the growth of HCC xenografts in mice.6, 15 Unfortunately, however, in a toxicology study, treatment with the humanized 1A6 Ab to cynomolgus monkeys increased hepatic transcription of and elevated bile acid synthesis, thus dramatically altering bile acid metabolism and causing severe dose\limiting side?effects.25 Considering both FGF19s physiological function in regulating bile acid metabolism and its tumorigenic activity in driving the pathogenesis of HCC, it is unclear if a strategy that targets FGF19 can effectively treat HCC while being safe for patients. Previous studies exploring the function of FGF19s N\terminus (NT) have established that a variant (M70) with NT substitutions and deletions, as well as a chimeric variant substituted with the 20?N\terminal residues from FGF21 exhibit reduced ability to induce hepatocyte proliferation but retained their ability to suppress hepatic expression.26, 27, 28 Building on these insights, we surmised that this NT of FGF19 may be essential for its tumorigenic activity but may not be required for its physiological bile\acid\regulatory function. We further hypothesized that selectively targeting the NT of FGF19 with an Ab instead of a small molecule drug may be both effective and safe. In this study, we first identified Abs that specifically bind to FGF19 in an NT\dependent manner. We then exhibited that Rabbit Polyclonal to FANCG (phospho-Ser383) one high\affinity NT\dependent Ab, G1A8, and its close variant Ab, HS29, effectively inhibit FGF19\induced HCC cell proliferation in vitro and significantly suppress HCC tumor growth in cell line\derived xenograft and patient\derived xenograft (PDX) mouse models. Importantly, G1A8 did not affect FGF19\mediated repression of mouse hepatic transcription. Moreover, G1A8 did not cause bile\acid\related side?effects in cynomolgus monkeys. Collectively, our study demonstrates that selectively targeting the NT of FGF19 with an Ab can be both effective and safe, and the Abs we developed, G1A8 and HS29, show strong promise to be further LP-935509 developed into a safe and therapeutic agent for treating FGF19\driven HCC. 2.?MATERIALS AND METHODS 2.1. Cell lines FreeStyle 293\F cells were cultured according to the manufacturers instructions (Thermo Fisher Scientific). The.