HepG2 Xenograft Model

HepG2 xenograft model

The HepG2 cell line (human liver) is used to create the CDX (Cell Line Derived Xenograft) HepG2 xenograft mouse model. The HepG2 xenograft mouse model of human hepatocellular carcinoma (HCC) allows for studies consisting of antiangiogenesis targeting (i.e. bevacizumab, rapamycin, etc.) or as a tumor growth inhibition model (e.g. sorafenib).

*NOTE: All IP rights to the HepG2 cell line belongs solely to the Wistar Institute. The client needs to obtain a license from the Wistar Institute prior to initiation of a xenograft study using the HepG2 cell line.

Basic study design

1.  Exponential growth of HepG2 cells are maintained prior to collection.
2. The cells are collected by trypsinizing the cells.  Cell count concentration and viability is determined with trypan blue (min 98% viability).  Cell suspensions are then adjusted to the required concentration for inoculation.
3. One million cells, in a volume of 100 uL (matrigel + HepG2 suspension), is inoculated subcutaneously (s.c) via a single injection into the hind leg (NOD/SCID or athymic BALB/C, 10-12 wks old).
4. The injection areas are monitored until tumors are palpatable.  Calipers are utilized for tumor measurement until average sizes of tumors are 50-150 mm3.
5. After randomization (sorting) into appropriate cohorts, test compounds are injected according to the agreed upon treatment schedule.
6. Tumor measurements are recorded (daily) and mouse weights are documented (3 times weekly).
7. As tumor size limit is reached (or maximum of 2,000 mm2), the animals are euthanized.
8. Predetermined for the study, a necropsy is performed at the end of the study.
9. Tumors are removed, their weights logged and the tumor is documented (digital imaging).
10. Tissues are collected from the animals (submersed in RNAlater, snap frozen, nucleic acids isolated or tissues prepared for histological analysis) by performing standard gross necropsies.

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HepG2 Xenograft Model

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 HepG2 xenograft model:

  • HepG2 Tumor Growth Delay (TGD; latency)
  • HepG2 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)
  • HepG2 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

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HepG2 Xenograft Model