Male athymic nude mice

All animal studies were conducted in full compliance with a protocol approved by the Johns Hopkins University Animal Care and Use Committee. Five young adult male athymic nude mice (Taconic Biosciences, Hudson, NY) were prepared as described previously [33 (link),34 (link)] to carry a single subcutaneous xenograft each of PSMA-positive PC-3 PIP cells and PSMA-negative PC-3 flu cells (a gift from Warren B. Heston, the Cleveland Clinic). Near IR dye-labeled intact IgG or 5D3 Fab, as indicated, was injected via the tail vein when tumor xenografts had reached 4–6 mm in diameter. Imaging was performed on a Pearl Impulse imager (LI-COR Biosciences). All images were displayed using the manufacturer’s software and normalized to the same acquisition time to facilitate direct comparison between mice and over time. Image acquisition began 45 min after fluorescent antibody injection and concluded 72 hr post-injection. Following the 72 hr image acquisition, each mouse was euthanized by 3% v/v isoflurane-anesthetized cervical dislocation and dissected to allow imaging of the tumors without attenuation from the skin.

Male athymic nude mice, age 6–8 wk, were obtained from Taconic Farms. Mice were housed and monitored at the Division of Laboratory Animal Resources at Stony Brook University. All experimental procedures and protocols were approved by the Stony Brook University institutional animal care and use committee (IACUC). Tumors were established by resuspending 1 × 10⁶ tumor cells in 100 μl PBS and injecting the cells into the mid-flanks of mice using a 26-gauge needle. For each tumor, the tumor length (l) and width (w) was measured every 4–5 d with an electronic caliper. Tumor volume (v) was calculated using the formula v = (l × w²)/2 and plotted in mm³.

Male athymic nude mice were purchased from Taconic Farms (Hudson, NY). All animals were 6–8 weeks old at the time of injection. Metastatic liver models of colorectal cancer were developed as described previously ^{29 (link)}. For all animal experiments, HCT116 cells (labelled with a stable expression luciferase gene) were used. For all therapeutic experiments, the dose of FLANC small interfering RNA (siRNA) was 200 μg/kg, as described previously ^{30 (link)}. The oligos were incorporated into neutral 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine (DOPC) nanoliposomes. These were administered via intravenous injection twice weekly beginning two weeks after tumor cell injection and continued for 4 weeks. All mouse studies were approved and supervised by the MD Anderson Cancer Center (MDACC) Institutional Animal Care and Use Committee.
Additional methods are presented in thesupplementary methods section.