Identification of new drugs and the evaluation of vaccines for treating and preventing canine distemper in dogs and raccoons

Canine distemper virus (CDV) is a morbillivirus that is closely related to measles virus (MeV). These viruses are highly contagious and infect lymphocytes, airway epithelial cells, and cells of the neurological system. There have been recent outbreaks of CDV in urban populations of raccoons in Halifax and Toronto. In addition, there are global concerns originating in China, where CDV was observed to jump from dogs into macaques at various primate centres. Our laboratory discovered the cellular receptors for morbilliviruses (SLAMF1, Nectin-4, and CD46). This application represents a new venture into antiviral drugs and the evaluation of vaccines directed against CDV. We propose to use ferret and a SLAMF1 transgenic model to evaluate drugs and vaccines. Ferrets are extremely susceptible to CDV and die within 2 weeks of contracting disease. We also have found that a single mutation in the CDV-H receptor binding protein permits the virus to replicate in our human SLAMF1 transgenic mouse model. In Aim 1 we propose to test the same mouse with wild type CDV strains including neurotropic Snyder Hill virus and local raccoon isolates. Infected mice will be dissected and tissue will be cryo-sectioned and analyzed for virus using immune fluorescence microscopy and RT-PCR. We will use the ferret and mouse models to evaluate new antiviral drugs directed against CDV in Aim 2. There are no therapeutic drugs for treating morbillivirus infections, but I previously discovered a small peptide Z-D-Phe-L-Phe Gly which blocked MeV-mediated membrane fusion and entry into the host cell [Richardson et al. (1980) Virology 105: 205-222]. A small molecular analogue to this peptide (AS48) has been synthesized and both molecules target a similar hydrophobic cavity in the MeV fusion (F) protein [Ha et al. (2017) J. Virol. 91(23):e01026-17]. Similar molecules which inhibit the F protein of CDV will be identified using a chemical library at the Li Ka Shing Virology Institute, Edmonton. Another target for antiviral drugs directed against CDV is the viral RNA polymerase (L). One lead compound (ERDRP-0519) appears effective against MeV and we propose to identify compounds that target the CDV polymerase. Aim 3 addresses a perennial problem with CDV vaccines characterized by vaccine-related encephalitis. The Rockborn vaccine is particularly prone to this debility. The safer Onderstepoort vaccine is attenuated exclusively in eggs, while the problematic Rockborn strain is propagated in mammalian cell culture. We propose to assess receptor usage of various strains of CDV using CHO, CHO-CD46, CHO-dogSLAMF1, CHO-dogNectin-4, CHO-chickNectin-4, and CHO-PVRL2 cell lines. We will study the poorly understood process of virus attenuation by studying vaccine and wild type CDV infections in canine peripheral blood lymphocytes and primary canine tracheal/bronchial epithelial cells. Infectious particle formation, viral protein synthesis, and interferon production by these cells will be evaluated.