Balb c nude mice

Antibodies used in this study are listed in Table S1. MCF-7, SK-BR-3, SK-OV-3, Daudi and K562 were obtained from ATCC. These cell lines were cultured according to ATCC guidelines. oNK cells were cultured as described previously [27 (link)]. Female peripheral blood mononuclear cells (PBMC) were bought from PPA Research Group and cultured under the manufacturer’s instructions. Female NSG (NOD.Cg-Prkdc^scid Il2rg^tm1Wjl/SzJ, 6–10 weeks old) and BALB/c nude mice were respectively purchased from The Jackson Laboratory (Stock No.: 005557) and BioLASCO Taiwan Co. Ltd. (Taipei City, Taiwan), and housed under the regulation of the Institutional Animal Care and Use Committee (IACUC) of the contract research organizations.

LN229 cells (1 × 10⁶) were injected subcutaneously into the dorsal flank of athymic BALB/c nude mice (4-week-old, female) from Jackson laboratory. Once the tumor volume reached 100 mm³, animals were divided into five subgroups: control 1 (TMZ vehicle), control 2 (volasertib vehicle), TMZ (Selleckchem, Cat: S1237) volasertib (Selleckchem, Cat: S2235), combination (TMZ + volasertib) (n = 3 mice). TMZ (25 mg/kg formulated in 10% DMSO in sterile PBS) and administrated intravenously every 3 days. TMZ (25 mg/kg formulated in 10% DMSO in sterile PBS) and administrated intravenously every 3 days. volasertib was formulated in hydrochloric acid (0.1 N) and diluted with 0.9% NaCl at 10 mg/kg concentration and delivered intravenously every 3 days. Each mouse’s tumor size was measured every 3 days after the first injection by a Vernier caliper. The volume of the tumor was calculated with the formula: volume = (length × width²)/2. In this case, 30 days after the injection, the mice were euthanized and the tumors were dissected for histological analysis.

Four-week-old female BALB/c nude mice were purchased from The Jackson Laboratory Japan (Yokohama, Japan). All animal experiments were approved by the Institutional Animal Care and Use Committee of the University of Tokyo, and carried out under the University of Tokyo guidelines for animal experiments, the Japanese government’s Law 105 Notification 6, regarding human cell culture, and the ARRIVE guidelines.

The BALB/c nude mice (Jackson Immuno Research Laboratories, West Grove, PA) were inoculated subcutaneously with GBM8401 cells (5 × 10⁷ cells) mixed with 100 μl Matrigel ™ (5 mg/mL, BD Biosciences), to enhance tumor growth ability in nude mice [55 (link)]. After 1 day of the implantation, VPA (400 mg/kg) [40 (link), 41 (link)] or PBS was injected intraperitoneally every two days for 60 days, and PBS group were used as control. Once palpable tumors were established after the inoculation, the tumor xenografts were measured with two dimensions by caliper every two days. These nude mice were sacrificed at 60 days after tumor inoculation. Tumor volume was calculated by the following equation: length × height × width (mm³). The Institutional Animal Care and Use Committee of the Mackay Memorial Hospital approved the use of animals in this study.

TRAMP mouse were bought from Jackson lab and castrated at 12-wk-old. Tumor samples were collected and analyzed at 16wks, 20wks, 24wks and 28wks. Single cell suspensions of LNCap cell lines (5×10⁶ cells) in Matrigel (BD Biosciences) (1/1) were injected subcutaneously in the flank of 8-wk-old BALB/C nude mice with androgen recruitment (Jackson Laboratories). Tumor incidence and growth was monitored during different time periods post injection. Tumors grew up to 1.0 cm in diameter, at which point animals were euthanized. After the diameter of LNCaP xenograft tumors reached to 0.8-1.0 cm, the tumor samples were collected and named androgen dependent xenograft tumor. We removed androgen supplementary from the nude mice with diameter-0.8cm LNCaP xenograft tumors and castrated them for one week, then sacrificed the mice and collected tumors named ADT tumor. When the diameter of LNCaP xenograft tumor re-grew to 0.8cm in the castrated mice, we sacrificed the mice and collected the tumors named castration resistant LNCaP xenograft tumor. Each tumor was dissected, fixed in formalin, and processed for histopathology examination.

The mouse experiments were conducted and approved in accordance with the Animal Care Committee of the University of UCSD (S11020). Five-week-old male Balb/c nude mice (NU/J-002019) were obtained from The Jackson Laboratory (JAX). After anesthesia, the mice were made ~1 cm incision in the left abdominal flank. GIST-T1 (GFP-conjugated; 5 × 10⁶ cells), GIST-T1 (5 × 10⁶ cells) with GF (1 × 10⁶ cells), and GIST-T1 with GF pretreated with TGF-β1 for 24 h were injected into the spleen. After 3 weeks, livers collected from each group were analyzed using the IVIS imaging system, and the GFP signals produced from livers were graphed by total photon flux (p/s).