studies

HEPATOLOGY, VOLUME 62, NUMBER 1 (SUPPL) AASLD ABSTRACTS 1175A
The following authors have nothing to disclose: Yasutoshi Nozaki, Satoshi
Tanaka, Sadatsugu Sakane, Yugo Kai, Yuki Makino, Tasuku Nakabori, Yoshinobu
Saito, Ryotaro Sakamori, Naoki Hiramatsu, Tomohide Tatsumi
1986
FGF15 and FGF19 Induce Disparate FGFR4-Mediated
Hepatocarcinogenicity In-Vitro and In Two Murine Models:
Implications for Drug-Associated Carcinogenicity
Risk Assessments
Lei Ling, Mei Zhou, Darrin Lindhout, Jian Luo, Michael Chen,
Hong Yang, Mark Humphrey, Van Phung, Kalyani Mondal, Hugo
Matern, Marc Learned, Stephen Rossi, Alex M. DePaoli, Hui Tian;
NGM Biopharmaceuticals, S. San Francisco, CA
Background and Aims: FGF19 is a regulator of bile acid (BA)
synthesis in humans. However, FGF19 produces hepatocellular
carcinoma (HCC) in transgenic mice and is linked to increased
risk of HCC post-resection recurrence in humans. FGF15 is the
rodent ortholog of human FGF19 with 50% amino acid (AA)
homology and used in rodents to assess the pharmacologic and
caricongenic activity of FGF19. Both FGF15 and FGF19 are
potent inhibitors of Cyp7a1-mediated BA synthesis in rodents
but the carcinogenicity risk of FGF15 is not well characterized.
The hepatocarcinogenicity of FGF15 and FGF19 was
evaluated in leptin receptor-deficient (db/db) mice after 24
wks or diet-induced obese (DIO) mice after 52 weeks of treatment.
In vitro receptor interaction and activation studies were
performed. Methods: FGF15, FGF19 or control was dosed
by long-term transgene expression using 1 dose of adeno-associated
virus (AAV)-mediated gene delivery. Liver tissue was
examined for tumors and liver weight at 24wks post-dose (db/
db) or 52wks post-dose (DIO). Gene expression of HCC-related
and cell proliferation markers were measured in liver
tissue. FGFR4-Klothoβ (FGFR4-KLB) receptor complex binding
was assessed with SPR assays and transfected rat L6 cells were
used to assess receptor activation. Results: FGF19 induced
liver tumors in both models post-treatment at concentrations
as low as 1 ng/ml (Figures 1a, 1b). In contrast, FGF15 failed
to induce liver tumors at supraphysiologic concentrations and
maintained normal liver weight and liver-body weight ratios.
Liver tissue expression of Ki-67 (Figure 1c), α-fetoprotein, glypican-3,
cyclin-a2, Ccnb1 and Ccnb2 were induced with FGF19
but not FGF15. Reduced activation of FGFR4-KLB receptor complex
was only seen with FGF15, as measured by decreased
expression of STAT-3 target genes. These differences may be
mediated by AA sequences unique to FGF15. Conclusions:
Significant differences in hepatocellular proliferation and tumorigenesis
were observed with FGF15 vs FGF19. These data
demonstrate the potential challenges of assessing the risks of
increased FGF19 levels in humans using FGF15 in rodent carcinogenicity
models.
Disclosures:
Lei Ling - Employment: NGM Biopharmaceuticals, Inc.
Mei Zhou - Stock Shareholder: NGM Biopharmaceuticals
Darrin Lindhout - Patent Held/Filed: NGM Biopharmaceuticals; Stock Shareholder:
NGM Biopharmaceuticals
Jian Luo - Employment: NGM Biopharmaceuticals
Hong Yang - Employment: NGM Biopharmaceuticals
Mark Humphrey - Employment: NGM Biopharmaceuticals; Stock Shareholder:
NGM Biopharmaceuticals
Van Phung - Employment: NGM Biopharmaceuticals; Stock Shareholder: NGM
Biopharmaceuticals
Kalyani Mondal - Stock Shareholder: NGM BioPharmaceuticals
Hugo Matern - Employment: NGM; Stock Shareholder: NGM
Marc Learned - Employment: NGM Biopharmaceuticals, Inc.; Stock Shareholder:
NGM Biopharmaceuticals, Inc.
Stephen Rossi - Employment: NGM Biopharmaceuticals, Inc; Stock Shareholder:
NGM Biopharmaceuticals, Gilead Sciences
Alex M. DePaoli - Employment: NGM Biopharmaceuticals
Hui Tian - Management Position: NGM Biopharma
The following authors have nothing to disclose: Michael Chen
1987
Lymphotoxin–β regulated by NF–κB is a useful biomarker
related to poor prognosis in hepatocellular carcinoma
Takehisa Watanabe, Satomi Fujie, Youko Yoshimaru, Katsuya
Nagaoka, Hiroko Setoyama, Kotaro Fukubayashi, Masakuni Tateyama,
Hideaki Naoe, Jiro Fujimoto, Motohiko Tanaka, Yutaka
Sasaki; Hepatology, Kumamoto Univ., Kunamoto, Japan
The Lymphotoxin-β (LTβ) is a major pro-inflammatory cytokine
that belongs to the TNF superfamily and mediates local
inflammatory responses as a membrane-bound cytokine. The
LTβ pathway is considered to be an important pathway in
the hepato-carcinogenesis. We previously reported that the
CCCTC-binding factor (CTCF) mediated higher order chromatin
conformation of TNF/LT locus, including LTβ gene, is deeply
involved in the regulation of LTβ expression in human HCC,
by the global analysis with ChIP-chip and ChIP-seq. In this
study, we attempted to reveal the molecular mechanism of LTβ
regulation and the clinical characteristics of the patients with
HCC expressing LTβ. Because chronic inflammation is one of
the most crucial factors for hepato-carcinogenesis, we firstly
focused on NF-κB, a major pro-inflammatory transcription factor,
and investigated the influence of NF-κB on LTβ expression
and higher order chromatin conformation, by using cell lines
differed on the activity of NF-κB. As a result, LTβ was expressed
continuously in the cells with constitutive active NF-κB in
response to the stimulation with TNF. On the other hand, LTβ
degraded rapidly in the cells without constitutive active NF-κB.
These cells differ on the higher order chromatin conformation
to control the interaction between the LTβ promoter and appropriate
NF-κB responsive enhancer. These results indicated that
NF-κB was involved in the higher order chromatin conformation
and regulation of LTβ. Additionally, we found that LTβ
could be quantified in the exosomes extracted from serum of
the patient, whereas liver biopsy had been considered to be
indispensable to describe how LTβ is expressed in hepatic lobules.
To examine whether exosomal LTβ reflects LTβ expression
in HCC tissue, we performed immunohistochemistry with LTβ
antibodies in 24 samples from HCC patients and compare with
the amount of exosomal LTβ. Consequently, high level exosomal
LTβ reflected the high expression LTβ in the tumor tissues,
not in the adjacent non-tumor tissues. Moreover, the patients
with the high level exosomal LTβ exhibited significantly shorter
survival time. Therefore, these results suggest that exosomal LTβ
might be a useful biomarker related to poor prognosis in HCC.
Disclosures:
Yutaka Sasaki - Grant/Research Support: Chugai Pharmaceutical Co. JAPAN,
MSD Co. JAPAN