Established gastric cancer cell lines transplantable into C57BL/6 mice show fibroblast growth factor receptor 4 promotion of tumor growth

Abstract

Previously no mouse gastric cancer cell lines have been available for transplantation into C57BL/6 mice. However, a gastric cancer model in immunocompetent mice would be useful for analyzing putative therapies. N‐Methyl‐N‐nitrosourea (MNU) was given in drinking water to C57BL/6 mice and p53 heterozygous knockout mice. Only 1 tumor from a p53 knockout mouse could be cultured and the cells s.c. transplanted into a C57BL/6 mouse. We cultured this s.c. tumor, and subcloned it. mRNA expression in the most aggressive YTN16 subline was compared to the less aggressive YTN2 subline by microarray analysis, and fibroblast growth factor receptor 4 ( FGFR4) in YTN16 cells was knocked out with a CRISPR/Cas9 system and inhibited by an FGFR4 selective inhibitor, BLU9931. These transplanted cell lines formed s.c. tumors in C57BL/6 mice. Four cell lines (YTN2, YTN3, YTN5, YTN16) were subcloned and established. Their in vitro growth rates were similar. However, s.c. tumor establishment rates, metastatic rates, and peritoneal dissemination rates of YTN2 and YTN3 were lower than for YTN5 and YTN16. YTN16 established 8/8 s.c. tumors, 7/8 with lung metastases, 3/8 with lymph node metastases and 5/5 with peritoneal dissemination. FGFR4 expression by YTN16 was 121‐fold higher than YTN2. FGFR4‐deleted YTN16 cells failed to form s.c. tumors and showed lower rates of peritoneal dissemination. BLU9931 significantly inhibited the growth of peritoneal dissemination of YTN16. These studies present the first transplantable mouse gastric cancer lines. Our results further indicate that FGFR4 is an important growth signal receptor in gastric cancer cells with high FGFR4 expression.

Deep immunophenotyping at the single-cell level identifies a combination of anti-IL-17 and checkpoint blockade as an effective treatment in a preclinical model of data-guided personalized immunotherapy

“…We have previously reported that tumor growth, metastatic rate and peritoneal dissemination rate of YTN16 were higher than YTN2; FGFR4 expression by YTN16 cells was 121-fold higher than YTN2. 8 We demonstrated that FGFR4 disruption by CRISPER-Cas9 system or pharmacological inhibition of FGFR signaling in YTN16 resulted in the reduction of peritoneal dissemination. In this study, we focused on the immunological aspects of these two tumor cell lines.…”

“…YTN2 and YTN16 gastric cancer cell lines are subclones of one line established from a tumor induced by N-methyl-N-nitrosourea treatment of a male heterozygous p53 knockout mouse. 8 YTN2 spontaneously regresses in C57BL/6 mice in a CD8 + T cell-dependent manner, while YTN16 grows progressively ( figure 1 ). These two related cell lines both harbor a similar number of mutation-associated neoantigens.…”

“…To demonstrate the feasibility of an immunological data-guided personalized adaptive approach to immunotherapy, whereby immunomodulatory strategies are tailored to the patient’s specific TME, we used mice-bearing subcutaneous YTN16 gastric cancers. 8 The TME of growing YTN16 tumors was immunologically assessed and the animals were treated based on those results. Using scRNA-Seq, but not bulk RNA-Seq, it was possible to determine that interleukin-17 (IL-17)-producing cells in YTN16 tumors were involved in generating an immunosuppressive microenvironment.…”

Established fibrous peritoneal metastasis in an immunocompetent mouse model similar to clinical immune microenvironment of gastric cancer

“…YTN16 cells were established from subcutaneous tumors by injection of primary cultured cells derived from a mouse gastric adenocarcinoma. Mouse gastric tumors were established in p53 heterozygous knockout C56BL/6 mice by addition of N-Methyl-N-nitrosourea (MNU) to the animals' drinking water [23]. The resulting tumor cells were cultured in high-glucose Dulbecco's modi ed Eagle medium (DMEM, Sigma-Aldrich Japan, Tokyo, Japan) containing 1.0mL/L MITO (Coning Japan, Tokyo), 10mL/L L-Glutamine, 10mL/L Penicillin/Streptomycin and 10% fetal bovine serum (FBS), on plastic dishes coated with Type I collagen solution (0.5% Atelocollagen Acidic Solution IPC-50; Koken Co., Ltd., Japan) at 37°C in 5% CO 2 atmosphere.…”

FGFR4 Inhibitor BLU9931 Attenuates Pancreatic Cancer Cell Proliferation and Invasion While Inducing Senescence: Evidence for Senolytic Therapy Potential in Pancreatic Cancer

“…BLU9931 makes a covalent bond with Cys552 within the ATP-binding pocket of FGFR4, which is not present in FGFR1-3. BLU9931 inhibited gastric cancer cell growth in a transplanted mouse model [ 36 ]. To date, however, there have been no reports about the effects of FGFR4 inhibitors in PDAC.…”