Generation of a Liver Orthotopic Human Uveal Melanoma Xenograft Platform in Immunodeficient Mice
November 6th, 2019
Orthotopic human liver metastatic uveal melanoma xenograft mouse models were created using surgical orthotopic implantation techniques with patient-derived tumor chunk and needle injection techniques with cultured human uveal melanoma cell lines.
Most patient-derived tumor animal models are subcutaneously implanted, preventing the evaluation of important features of tumors and of the drug efficacy. Housing the implanted tumors within the liver facilitated this analysis without the risk of too much deviations occurring within the liver. Before beginning the procedure, spray all of the materials and instruments with 70%ethyl alcohol.
Confirm a lack of response to toe pinch in the anesthetized animal. Place the mouse in the supine position on a heating pad, and wet the fur with additional 70%ethyl alcohol. Spread the fur to visualize the skin below the left subcostal area.
Remove hair, and apply iodine to be absorbed into the skin. Place a sterile surgical drape with a two-centimeter hole over the mouse, and use curved ultra fine forceps to lift the abdominal skin. Make a one-centimeter transverse left subcostal incision into the tented skin, and insert curved scissor tips into the incision to separate the peritoneum from the skin.
Retract the scissors without opening the blades, and locate the liver under the peritoneum by its dark reddish color. Use the curved scissors to make a one-centimeter transverse incision in the peritoneum. Using the other hand, insert the edge of a cotton swab beneath the left liver lobe, and roll the swab downward to push up the liver.
Lift the swab to transfer the liver onto a nonwoven, absorbent fabric sheet. Holding a sterile number 11 scalpel blade parallel to the surface, press the blade horizontally into the liver to make a five-millimeter deep by five-millimeter wide incision in the parenchyma while softly pressing the incision site with a cotton swab. Roll the cotton swab upward to open the incision site.
Use curved ultra fine forceps to implant a one-millimeter cube of tumor tissue into the pocket. Retract the forceps while rolling the cotton swab in the reverse direction while pressing down, and remove the swab. Immediately place an absorbable hemostat on the incision site, and confirm hemostasis.
Next use blunt ended forceps to peel the liver from the fabric sheet, and return the liver to the abdominal cavity. Then close the peritoneum with a 5-0 absorbable suture and a double ligature, and close the skin with a 5-0 absorbable suture and a triple ligature. The xenograft tumor engrafts in the liver as a solitary tumor without daughter nodules.
Surgical orthotopic injection into the liver using microneedles can result in a successful engraftment of a cluster of cultured human liver metastatic uveal melanoma cells within the liver. Using this liver orthotopic patient-derived tumor model, the efficacy of the drugs injected directly into the liver can be tested with similar expected outcomes to those observed in clinical patients.
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Overview
This study presents the creation of orthotopic human liver metastatic uveal melanoma xenograft mouse models. These models were developed using surgical orthotopic implantation techniques with patient-derived tumor chunks and needle injection techniques with cultured human uveal melanoma cell lines.
Key Study Components
Area of Science
- Oncology
- Neuroscience
- Animal Models
Background
- Traditional patient-derived tumor models are often subcutaneously implanted.
- Subcutaneous models limit the evaluation of tumor features and drug efficacy.
- Orthotopic models allow for a more accurate representation of tumor behavior.
- Housing tumors within the liver minimizes deviations in analysis.
Purpose of Study
- To develop a more accurate model for studying uveal melanoma.
- To facilitate the evaluation of tumor characteristics and treatment responses.
- To improve the understanding of metastatic behavior in liver environments.
Methods Used
- Surgical orthotopic implantation techniques.
- Needle injection of cultured human uveal melanoma cell lines.
- Use of patient-derived tumor chunks.
- Preparation of materials with 70% ethyl alcohol for sterility.
Main Results
- Successful establishment of orthotopic models in mice.
- Enhanced ability to study tumor behavior in the liver.
- Improved assessment of drug efficacy against uveal melanoma.
- Demonstrated reduced variability in tumor responses.
Conclusions
- Orthotopic models provide a better platform for research on uveal melanoma.
- These models can lead to more effective treatment strategies.
- Further studies are needed to explore the full potential of these models.
Frequently Asked Questions
Executive Industry Relevance
Establishing orthotopic liver xenograft models for metastatic uveal melanoma addresses a critical gap in preclinical oncology by enabling physiologically relevant tumor microenvironment studies. This platform supports mechanistic de-risking of therapeutic candidates through improved predictive confidence in drug efficacy assessments. It enhances translational continuity from discovery to preclinical evaluation by reducing biological variability inherent in subcutaneous models.
Strategic Applications in Biopharma R&D
Early Discovery & Target Validation
- Scientific Value: Enables interrogation of tumor-host interactions in the liver microenvironment to validate therapeutic targets.
- Operational Value: Provides a reproducible system for assessing target modulation in metastatic disease contexts.
Screening & Assay Development
- Scientific Value: Supports preparation of validated biological systems for downstream compound screening in organ-specific settings.
- Operational Value: Facilitates assay standardization through consistent tumor engraftment using surgical or injection techniques.
Translational & Preclinical Research
- Scientific Value: Maintains disease-relevant system fidelity for studying liver metastatic progression and treatment response.
- Operational Value: Enables continuity from discovery through preclinical validation via re-engraftment of cryopreserved tumors.
Pipeline & Workflow Integration
The method integrates into the discovery continuum by supporting hypothesis testing in Early Discovery, assay readiness in Screening, and quantitative tumor growth measurements in Preclinical work.
- Discovery Biology: Supports pathway clarification and biological de-risking through orthotopic tumor engraftment in immunocompromised hosts.
- Screening: Describes assay readiness via reproducible tumor implantation using either surgical chunks or cultured cell line injection.
- Analytics: Highlights CT scanning as a quantitative readout for detecting interior liver tumors and monitoring engraftment success.
- Translational Research: Connects to preclinical continuity through re-implantation protocols using cryopreserved tumors to maintain model fidelity.
- Enterprise Reuse: Positions the platform as a reusable capability for studying liver-metastatic cancers across multiple therapeutic programs.
Operational & Enterprise Impact
- Scientific Value: Predictive confidence in target validation through physiologically relevant tumor modeling.
- Operational Value: Standardization and reproducibility via defined surgical and injection protocols with high technical success rates.
- Strategic Value: Improved go/no-go decisions by reducing late-stage biological risk in metastasis-focused programs.
- Portfolio Impact: Risk-adjusted prioritization of candidates based on liver-specific efficacy data.
Implementation Considerations
- Requires expertise in murine surgical techniques and aseptic procedures.
- Dependent on sterile instrumentation, absorbent fabric sheets, and absorbable sutures.
- Necessitates cross-team standardization between surgery, imaging, and pharmacology groups.
- Involves adaptation considerations when applying to different tumor types or model systems.
- Practical limitations include the technical complexity of liver incision and tumor pocket formation.
