Syngeneic mouse models for the testing of immunotherapeutics

Syngeneic Mouse Models: Impact on Immunotherapeutics Testing 

Immunotherapies in oncology clinical practice have rapidly changed the therapeutic landscape. In particular checkpoint inhibitors have rapidly expanded their clinical use and can provide durable responses in a significant fraction of patients in select diseases. Unfortunately, many patients derive no clinical benefit because of primary resistance to these agents and develop acquired resistance compromising long term survival.  There is a significant need for the development of new immunotherapeutics that can target novel molecules and mechanisms outside of PD1/PDL1 and CTLA4, and to develop agents that work in combination with approved therapeutics to enhance response to new and existing patients.  

Although checkpoint inhibitors like nivolumab (PD1) and ipilimumab (CTLA4) have had multiple successes in clinical use, the broad use of these inhibitors has exposed characteristic weaknesses that prevent successful response in patients.  Tumors develop multiple mechanisms to exclude the immune system from recognizing and infiltrating tumor tissue. These non-inflamed or “cold tumors” are not responsive to checkpoint inhibition and require methods to bypass the resistance mechanisms to be successfully treated.  An important focus of current preclinical research is to convert the tumor environment from being non-inflamed to inflamed in order to facilitate the subsequent response to checkpoint inhibitors. 

Preclinical animal models provide a system to evaluate these new immuno-oncology agents as both single agents and in combination with checkpoint inhibitors in a controlled and reproducible setting. The most common test systems are mouse models implanted with syngeneic tumor cell lines. These syngeneic murine tumor models have become the primary method for the initial evaluation of these agents. They provide a large diversity of tumor cell types from a variety of tissues of origin and have a diversity of mechanisms represented for sensitivity and resistance to known agents.  Syngeneic mouse models can provide a reproducible and robust system to test a wide variety of immuno-oncology agents.   

Strengths of the murine syngeneic tumor models include:   

• The use of immortalized tumor cell lines provides a homogenous cell source with the ability to incorporate a tracking label (luciferase) or genetically manipulate cells for mechanistic studies. 

• The ability to use an animal with an intact immune system and generate a complete reactive immune response.  In addition, for many of the mouse strains, availability of genetically engineered mice (such as T-cell deficiency) enables the evaluation of host interactions. 

• Availability of a robust set of reagents allows in depth characterization of the murine immune system, and murine versions of checkpoint inhibitors provide representative activity in mouse models.  

• Unlike humanized mice, syngeneic mouse models can be used for long term studies to perform vaccine and rechallenge studies.  

Although a variety of syngeneic tumor models from a multitude of tissues are now available there are three most popular models that provide many of the characteristics needed to evaluate immunotherapeutics. These include: 

1. 4T1 Breast carcinoma Model: Tumors grow well in both subcutaneous and orthotopic sites and serve as a good metastatic model. 4T1 is refractory to checkpoint inhibition and provides a good model for a “cold tumor” 
2. B16F10 Melanoma Model:  Aggressive growth in subcutaneous and metastatic models. Poorly responsive to checkpoint inhibitors but provides a robust model for testing model antigens and for mechanistic studies.   
3. CT26 Colon Cancer Model: Provides a reliable subcutaneous growth and drug response. This model is resistant to PD1 inhibitors but responsive to CTLA4 inhibitors.  

Figure 1 below shows an example tumor efficacy study. Figure 2 below shows an example of the utilization of labelled tumor cell line to detect primary, orthotopic and metastatic tumors

In conclusion, syngeneic mouse models provide valuable systems for the reproducible and robust testing of immuno-oncology agents. In addition, these models can be effectively used to test novel modulators of the immune system and combinations with known immuno-oncology agents.