About Preclinical Therapeutics
The UCSF Preclinical Therapeutics Core (PTC) facility generates tumor-bearing animals and conducts preclinical oncology trials for UCSF Helen Diller Family Comprehensive Cancer Center investigators. Offering a variety of xenograft-based cancer models, the core provides a complete set of services that include consultation regarding experimental design, tumor cell growth in culture and generation of tumors in mice, preparation and administration of experimental agents, monitoring of tumor burden and response to therapy, and interpretation of study results.
The PTC maintains a cryorepository of commonly used human cancer cell lines derived from multiple tumor types along with data regarding their in vivo growth characteristics. The PTC is also a resource of expertise in small animal survival surgery, and is available to provide training in these techniques on a recharge basis. Although the core primarily functions to test experimental anti-cancer agents in vivo, it also provides animal models of human cancer for use in novel diagnostic, tumor imaging, and basic mechanistic research. In addition, the core oversees, maintains and provides as a service a number of small animal imaging technologies housed within the barrier facility.
The availability of centralized cell and animal resources together with personnel with expertise in conducting preclinical studies ensures appropriate experimental design and reproducibility, compliance with local and federal regulatory guidelines for tumor-bearing animals, and maximum resource utilization through coordinated animal purchasing and housing.
The use of rodents as a model for human cancer has been an important tool in the development and translation of innovative anti-cancer therapies. The service that we provide makes use of xenograft tumor models in immune deficient (nu/nu) mice, as well as transgenic and conditional/knock-in based engineered models that target known tumor promoting pathways. Genetic and phenotypic characterization of both xenograft tissue and genetically engineered animals allows the correlation of drug-target interaction, as well as the ability to test predictions of pathway-targeted anti-tumor efficacy. To test synergistic activity, novel therapeutics—generated both from within UCSF as well as externally—are tested alone and in rational combinations and dosing sequences. Tumor response to therapeutics is detected by a combination of direct tumor measurement and molecular imaging methods.
Cell/tissue culture and engraftment
The PTC has expertise in high volume cell culture, and maintains a large cryorepository of commonly used tumor cell lines and tissues. In certain cases, tumor tissue is maintained and passaged in vivo. Core personnel generate xenografts via cell/tissue implantation by a variety of methodologies. In addition to subcutaneous, sites of tumor cell implantation include intra-venous, intra-cardiac and orthotopic engraftment directly into mammary fat pad, brain, pancreas, liver, spleen, bladder, and kidney capsule. When needed, matrigel is injected in combination with the tumor cells. Some tumors require hormone supplementation for growth in vivo typically provided by a subcutaneous implanted slow-release pellet.
Compound formulation and delivery
The Preclinical Therapeutics Core formulates agents for delivery using a variety of commonly used methods and delivery vehicles and has expertise in pharmacology and toxicology. Core personnel deliver therapeutic agent by a number of routes including oral gavage, intra-peritoneal, intra-venous, intra-tumoral, subcutaneous, and by food and/or drinking water. The core also implants Alzet osmotic minipumps for continuous drug delivery (up to 28 days).
Tumor and animal monitoring
Animal body weight is measured twice weekly and weight loss is used as an index of toxicity. Tumor volume is measured twice weekly to calculate tumor volumes. Electronic balances and calipers link directly to a server-based data management system. In some studies in vivo imaging is used to assess tumor size and location in disseminated and orthotopic tumor models (see below).
Body fluids, such as blood, urine, feces, organs, and cells are collected as needed per experimental design. At the close of a study, animals are euthanized and tissues and tumors are collected and processed in a myriad of ways depending on experimental design. For example, tumors are placed in 10% buffered formalin for transfer to the Mouse Pathology Core for histopathology and immunohistology analyses, in Trizol for RNA analyses, or snap-frozen for other types of studies (i.e., protein analysis, etc). In some studies, blood (50-100 ul) for serum or plasma is collected by saphenous vein bleeding at weekly intervals per standard CAR procedures. All animal procedures are performed according to the UCSF-approved CAR Protocols.
Pharmacokinetic (PK) and pharmacodynamic (PD) analysis
The Core performs PD and PK studies in a variety of ways depending on study goals and requirements. For PK analysis, a relatively low dose of experimental agent, typically 5 mg/kg, is administered by the route of choice and plasma/serum is collected at multiple time points over a 24-48 hr period. In a typical PD study, a group of tumor bearing mice are treated with an experimental agent for a period of time before tissue is collected for molecular analysis such as target modulation or compound distribution.
Patient derived tumor xenografting
The Core also engrafts primary human tumor tissue into mice mice for research purposes.. Tissues derived from either cancer patient biopsy or tumor removal surgery are implanted into a variety of immunodeficient mouse backgrounds including nu/nu, NOD SCID and NSG, and mice are monitored for evidence of successful tumor engraftment. Tumor tissue that demonstrates growth in vivo is maintained and propagated by surgical removal, dissection into small fragments, and reimplantation into additional host mice as well as cryopreservation for subsequent in vivo implantation and growth. All tissues are engrafted with patient consent, and no patient identifiers are in any way associated with these tumor tissues lines.
Models of metastatic disease
The PTC provides as a service a number of animal models of metastatic disease. In most cases, molecular imaging methods allow rapid evaluation of tumor location and overall tumor burden, including metastases. We use a combination of fluorescent, bioluminescent and ultrasound imaging methodologies to monitor and quantify tumor status. Viral transduction is commonly used to generate reporter cell lines to enable non-invasive imaging. The measure of tumor burden provides a basis for study enrollment, tumor localization and response to therapy. Finally, at the conclusion of the study, post-euthanasia autopsy under direct fluorescent visualization allows sensitive detection of gross and microscopic metastatic disease. Core personnel are skilled in application of imaging technologies and data analysis of bioluminescent images. Reporter cell lines are maintained for use in studies requiring imaging technology.
Genetically engineered models
UCSF develops and validates many genetically engineered mouse models of human cancer. These have certain advantages over xenograft models resulting from de novo tumor development and outgrowth in situ, including the complex multi-component tissue architecture of the tumor. The Preclinical Therapeutics Core therefore offers services that support preclinical trials in these models such as drug dosing and PK studies, animal weight and tumor measurements, whole animal imaging, tissue collection and necropsy.
The Preclinical Therapeutics Core works closely with project investigators and UCSF statisticians in order to design studies that yield adequate statistical power while minimizing the number of animals required. The number of animals required to achieve statistical significance is based upon historical data generated in the PTC, as well as relevant published data.
In support of in vivo imaging, the PTC maintains and oversees several molecular imaging devices at multiple UCSF campus locations. Housed within the animal barrier facility, these instruments are made available to the UCSF investigators for an hourly fee. The PTC offers a number of instruments that enable bioluminescent and/or fluorescent imnaging capabilities. These include a Xenogen IVIS 100 bioluminescent imaging system located at Mt Zion, a Xenogen IVIS Spectrum bioluminescent and fluorescent imaging system located at Mission Bay, and an INDEC Fluorvivo 300 fluorescent imaging system located at Mission Bay. In addition, the PTC offers a high-resolution ultrasound imaging device, the VisualSonics Vevo 770, located at the Mission Bay campus. Finally, as part of its optical imaging resources, the Core also offers a Leica MZ FLIII, fluorescent stereoscope for ex vivo applications. This includes filters for both GFP and Texas Red, and 8X zoom lens, and associated digital color camera. All of these are available for booking using UCSF’s CoreAdmin system.
A searchable database of core facilities at all UCSF campus locations, provided by the Clinical and Translational Science Institute at UCSF, is available here.