Nod scid mice

Animal procedures were approved by the Administrative Panel on Laboratory Animal Care of the VA Palo Alto Health Care System. Young C57BL/6J mice (strain 000664) and R26R^YFP mice on the C57BL/6J background (strain 006148) were purchased from The Jackson Laboratory. NOD-SCID mice were purchased from Taconic Biosciences. Old C57BL/6N mice were obtained from Charles River Laboratories through the National Institute on Aging. Pax7^CreER mice on the C57BL/6 x 129/SvJ background were kindly provided by Dr. Charles Keller (Oregon Health & Science University). Ccnd1^flox mice on the C57BL/6J x 129/Sv background were kindly provided by Dr. Peter Sicinski (Dana-Farber Cancer Institute). TRE-Ccnd1^WT and TRE-Ccnd1^K112E mice on the FVB background were kindly provided by Dr. Richard Pestell (Thomas Jefferson University). Pax7^rtTA mice were generated by genOway on the C57BL/6J background (details below). Mice were housed in specific pathogen-free conditions in barrier-protected rooms under a twelve-hour light-dark cycle and were fed ad libitum. All mice used for analysis were male. Young adult mice, Pax7^CreER mice and Ccnd1^flox mice were 3-4 months old, R26R^YFP mice were either 4 or 20-22 months old, NOD-SCID mice were 4-5 months old, and old mice were 18-22 months old. When possible, littermate controls were used.

Ctx^R H226 cell line xenografts were established as previously described (76 (link), 77 (link)). Briefly, Ctx^R H226 xenografts were established in athymic nude mice by injecting cells (2×10⁶) subcutaneously in the lower left flank. Tumors were allowed to grow to 100 mm³ prior to randomization and treatment with either cetuximab (1 mg/mouse) or IgG by intraperitoneal injection twice weekly. Tumors were monitored for cetuximab resistance, which was defined as marked tumor growth in the presence of continued cetuximab therapy. Ctx^R tumors were harvested when they reached approximately 2000 mm³. NSCLC PDXs were established and evaluated for cetuximab response as previously described (50 (link)). Briefly, surgical tumor samples were cut into 3–4 mm pieces and transplanted subcutaneously into 3–6 immunodeficient NOD/SCID mice (Taconic, Lille Skensved, Denmark). After three successful passages, cetuximab dose response studies were initiated. The treatment schedule for cetuximab (Erbitux, Merck KgaA, Darmstadt, Germany) was 50mg/kg/d, qd 1–5, by intraperitoneal injection.

All procedures were approved by the Institutional Animal Care and Use Committee at the University of Kentucky College of Medicine and conform to the legal mandates and federal guidelines for the care and maintenance of laboratory animals. Animals were maintained and treated under pathogen-free conditions. Female NOD-SCID mice (6–8 weeks old; Taconic, Germantown, NY) were injected with breast cancer SUM1315 (2×10⁶ cells/mouse) cells via mammary fat pad, and mice had three groups: vector control and Wnt5a-knockdown stable clones. Tumor growth was monitored with caliper measurements. When tumors were approximately 1.0 cm in size, mice were euthanized and tumors excised. Data were analyzed by Student's t test; p < 0.05 was considered significant.

Animal studies were performed in accordance with NIH guidelines and with protocols approved by the Fred Hutchinson Cancer Research Center and the University of Washington Institutional Animal Care and Use Committees. All surgeries were performed under isoflurane anesthesia, and all efforts were made to minimize animal suffering. LNCaP, VCaP, and 22Rv1 human PC mouse xenograft tumors were propagated as previously described (7 ). Briefly, two million PC cells were first aggregated with matrigel (BD Biosciences) and then injected subcutaneously into flanks of male NOD Scid mice (Taconic Farms). The tumor’s size was measured daily and tumors were harvested after eight to ten weeks in biological triplicates. Total RNA was then extracted using an RNA isolation kit (Qiagen) according to the manufacturer’s protocol, and used for human microarray and quantitative real time-PCR analysis.