DU-145 xenograft model
Prostate cancer is the most prevalent cancer in older males and is the second common malignancy in the Western world. Xenograft preclinical studies can be conducted to find more complex treatment for patients with advanced stage disease. The DU-145 epithelial cell line was isolated from prostate carcinoma cells of a 69-year-old Caucasian male. DU-145 has microvilli, tonofilaments, and desmosomes. The DU-145 cell line is not hormone sensitive and fails to express prostate-specific antigen. Concurrent chemoradiotherapy (CCRT) has proven to be a potent tool used during the past decades. A 2014 study by Wang et al. published in Clinical and Translational Oncology, examined the efficacy of combining radiation with low-dose docetaxel in the DU-145 prostate cancer xenograft model. The article indicates that CCRT with low-dose docetaxel blocks the growth of DU-145 prostate cancer xenografts in vivo, by enhancing apoptosis and suppressing angiogenesis. Therefore, CCRT with low-dose docetaxel could be a new treatment strategy for prostate cancer patients. The DU-145 cell line (human prostate) is used to create the CDX (Cell Line Derived Xenograft) DU-145 xenograft mouse model. The DU-145 xenograft model enables studies of tumor growth inhibition by systemic viral agents, small molecules (farnesol, darinaparsin, taxotere) or siRNA/miRNA.
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Basic study design
- Cells are maintained under conditions of exponential growth.
- The DU-145 cells are then collected from the flasks by trypsinization followed by determining viable cell counts using a trypan blue exclusion assay (minimum of 98% cell viability). Cell suspension concentration is adjusted to the appropriate density.
- Each mouse (athymic BALB/c (nu/nu), 10-12 weeks old) receives a single subcutaneous (s.c) injection into the flank of the hind leg. Each injection contains one million cells (in 100 uL) of the matrigel-DU-145 cell suspension.
- Injection sites are palpated up to three times a week until tumors are established. Tumors are measured via digital calipers until an average size of 50-150 mm3 is reached.
- According to the treatment schedule, animals are randomized into the treatment cohorts. Administration of the test compound is performed.
- Tumors are continually measured daily while mouse weights are recorded 3 times a week.
- Animals are humanely euthanized as tumor size reaches 2,000 cu millimeters, or the predetermined size limit of the study.
- A necropsy and tissue collection is performed at the termination of the experiment. Tumors are excised and weighed, and then documented by digital imaging.
- Gross necropsies are performed to collect tissues for downstream analysis.
- The tumors/tissues can be submersed in RNAlater, snap frozen in LN2, prepared for histology or nucleic acid isolated for genetic analysis.
Xenograft animal models are used to assess the effectiveness of drugs against specific types of cancer. New medicines are tested on staged tumor growths that have been engrafted via subcutaneous or orthotopic inoculation in an immunocompromised mouse or rat model. All clinically approved anti-cancer agents have been evaluated with conventional preclinical in vivo models. Xenograft studies can be highly complex, starting with the selection of the appropriate animal model, choice of tumorigenic cell line, administration method, dosing, analysis of tumor growth rates and tumor analysis (histology, mRNA and protein expression levels).
Altogen Labs provides an array of laboratory services using over 30 standard Cell Line Derived Xenograft (CDX) models and over 20 PDX models. Researchers investigating the role of specific proteins or gene products in regulating tumor growth can benefit from development of protein overexpression (genetically engineered to ectopically express proteins, tumor suppressors, or oncogenes) and RNAi cell lines with long term gene silencing. Altogen Labs provides quantitative gene expression analysis of mRNA expression (RT-PCR) and protein expression analysis using the WES system (ProteinSimple).
The dosing of the experimental compound of interest is initiated, for a staged study, when the mean tumor size reaches a specified volume (typically 50-100 mm3). In an unstaged study, the dosing of the compound of interest is initiated immediately after xenografting. Mice are dosed once or twice a day for 28 days (or other desired study duration) via the chosen route of administration. Tumor volume (mm3) is calculated via the “(W x W x L) / 2” formula, where W is tumor width and L is tumor length.
Animal handling and maintenance at the Altogen Labs facility is IACUC-regulated and GLP-compliant. Following acclimatization to the vivarium environment, mice are sorted according to body mass. The animals are examined daily for tumor appearance and clinical signs. We provide detailed experimental procedures, health reports and data (all-inclusive report is provided to the client that includes methods, results, discussion and raw data along with statistical analysis). Additional services available include collection of tissue, histology, isolation of total protein or RNA and analysis of gene expression. Our animal facilities have the flexibility to use specialized food or water systems for inducible gene expression systems.
Following options are available for the DU-145 xenograft model:
- DU-145 Tumor Growth Delay (TGD; latency)
- DU-145 Tumor Growth Inhibition (TGI)
- Dosing frequency and duration of dose administration
- Dosing route (intravenous, intratracheal, continuous infusion, intraperitoneal, intratumoral, oral gavage, topical, intramuscular, subcutaneous, intranasal, using cutting-edge micro-injection techniques and pump-controlled IV injection)
- DU-145 tumor immunohistochemistry
- Alternative cell engraftment sites (orthotopic transplantation, tail vein injection and left ventricular injection for metastasis studies, injection into the mammary fat pad, intraperitoneal injection)
- Blood chemistry analysis
- Toxicity and survival (optional: performing a broad health observation program)
- Gross necropsies and histopathology
- Positive control group employing cyclophosphamide, at a dosage of 50 mg/kg administered by intramuscular injection to the control group daily for the study duration
- Lipid distribution and metabolic assays
- Imaging studies: Fluorescence-based whole body imaging, MRI