Grant Amount: $123,125.4

Dr. Nicola J Mason, BVetMed, PhD, University of Pennsylvania

January 1, 2009 - June 30, 2011

Sponsor(s): Afghan Hound Club of America, American Belgian Tervuren Club, Inc., American Bullmastiff Association, American German Shepherd Dog Charitable Foundation, Inc., American Spaniel Club Foundation, Australian Terrier International, Bernese Mountain Dog Club of America, Briard Club of America Health & Education Trust, Clumber Spaniel Club of America, Collie Health Foundation, English Setter Association of America, Inc., Flat-Coated Retriever Foundation, French Bulldog Club of America, Golden Retriever Foundation, Gordon Setter Club of America, Great Pyrenees Club of America, Leonberger Health Foundation, National Amateur Retriever Club, Poodle Club of America Foundation, Portuguese Water Dog Foundation, Rhodesian Ridgeback Club of the United States, Saluki Health Research, Inc., Samoyed Club of America Education & Research Foundation, Siberian Husky Club of America, Staffordshire Bull Terrier Club of America, TarTan Gordon Setter Club, Versatility in Poodles, Inc.

Breed(s): -All Dogs

Disease(s): Hemangiosarcoma

Project Summary

Canine hemangiosarcoma is a common and highly aggressive tumor of blood vessels that is oftentimes fatal. At diagnosis most dogs have evidence of metastatic disease and despite chemotherapy, survival times rarely exceed 6 months. Novel approaches to the treatment of this disease are needed. Our work supported by CHF and its associated breed clubs aimed to generate a platform technology for generating canine-derived antibody fragments that can be utilized in vivo to specifically target tumor cells or their growth factors that are necessary for tumor cell proliferation and survival. Antibody fragments that target tumor cells can be linked to toxic agents and used to deliver these drugs directly to a cancer cell allowing for increased drug delivery and reduced toxic side effects. Antibody fragments that target growth factors aim to neutralize the effects of these factors on the tumor and so inhibit tumor growth and survival. These antibody fragments do not require linkage to toxic entities to exert their effects. Although antibody targeting approaches have revolutionized the treatment of certain cancers the in human clinics, no such targeting systems are currently available for use in the dog. Antibodies utilized in the human clinics are not used in veterinary medicine because they are not canine in origin so will be rapidly rejected by the dog's immune system. Furthemore, these antibodies may not cross-react with canine tumor molecules so will be ineffective in the canine system. The work performed during the first year lead to our ability to generate libraries of synthetic, canine antibody fragments. Each fragment is specific for a particular molecule. Such molecules may be those expressed on the surface of cancer cells, molecules associated with tumor growth factors or molecules expressed on the surface of infectious agents. Indeed, in theory, any molecule may be recognized by one or more antibody fragments contained within our canine antibody fragment libraries. Having generated these libraries we then aimed to use techniques to isolate fragments that specifically bind to canine hemangiosarcoma cells and canine Vascular Endothelial Growth Factor (VEGF). We were not successful in isolating scFv fragments against tumor associated antigens hypothesized to be expressed by hemangiosarcoma cell lines. Therefore, we opted to focus efforts on selecting antibody fragments that bind the known and bona fide endothelial cell growth factor VEGF. This growth factor plays an important role in ensuring that new blood vessels are generated in response to the presence of the tumor. New blood vessels support tumor growth and agents that inhibit VEGF are important in the treatment of many different malignancies. We have been successful in these efforts and have generated several antibody fragments that bind to canine VEGF. We are currently testing their ability to neutralize VEGF's effects on the growth of hemangiosarcoma cells in vitro. Furthermore, we are now taking steps to modify these fragments and generate canine monoclonal antibodies that are likely to have more favourable pharmacokinetic properties. It is important to note that since the antibody fragments we have generated are replicas of canine antibody fragments they should elicit minimal immune responses when used. As such, these antibody fragments should be able to be administered multiple times if necessary, without loosing their potency. In summary, work supported by this AKC CHF grant has lead to the development of the first canine-derived, antigen-specific targeting approach that may be used for the treatment of many different cancer types including HSA. Furthermore, we have isolated 3 unique canine derived antibody fragments that bind to canine VEGF. Work now continues to further develop these antibodies by evaluating their ability to neutralize canine VEGF and generate mnoclonal antibodies from their sequences - thus moving this work another step closer to the clinic.

Publication(s)

- Braganza, A, Wallace, K, Pell, L, Parrish, Cr, Siegel, Dl and Mason, Nj (2011) Generation and validation of canine single chain variable fragment phage display libraries. Veterinary Immunology and Immunopathology. 139, 27-40. http://www.sciencedirect.com/science/article/B6TD5-50R22V0-4/2/157306239e0cec9cffb66597d2973812