K562 xenograft model
The K562 cell line (human leukemia) is used for the creation of the CDX (Cell Line Derived Xenograft) K562 xenograft mouse model. The K562 xenograft model can be used to investigate the pivotal role of tissue inhibitors of metalloproteinases (TIMPs) for the treatment of leukemia.
Basic study design
1. K562 cells are maintained at exponential phase growth.
2. The cells are prepped for injection by determining the viable number of cells using trypan blue (minimum of 98% viability). Cell counts are adjusted to the appropriate density for injection.
3. Each mouse (10 to 12 weeks old; athymic BALB/C or NOD/SCID) receives single s.c. injections into the flank of a hind leg. Each injection contains one million cells of the K562 + matrigel suspension (vol= 100 microliters).
4. The injections are palpated until tumors are determined. Tumors are measured using calipers (digital) until an average size of 50-150 mm3 is reached.
5. Animals are randomized into appropriate groups . Administration of test material is performed according manufacturer recommendations.
6. Tumors are measured (daily) and mouse weights recorded (3 times weekly).
7. Animals are humanely euthanized when maximum tumor size is reached (or 2,000 sq millimeters).
8. At termination of experiment, tumors are excised and weighed. All tumors are digitally imaged.
10. Necropsies are performed and all tissues collected as requested by client.
11. Tissues can be 1) snap frozen in LN2, 2) prepared for histology, or 3) nucleic acids isolated.
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 K562 xenograft model:
- K562 Tumor Growth Delay (TGD; latency)
- K562 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)
- K562 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