Human coronaviruses cause 15 to 30 percent of common colds, yet there has been little investigation of these human pathogens. Our long term goal is to develop anti-viral drugs to prevent and/or treat human coronavirus infections of the upper respiratory tract. We anticipate that these drugs would be used in combination with drugs to prevent or treat colds due to rhinoviruses which are now being developed in other laboratories. By covering the viruses that cause most common colds, such combination drugs would make it unnecessary to identify the type of common cold virus causing early symptoms before initiating therapy. Our strategy for developing anti-coronavirus drugs is to use information about viral spike protein and receptor structure and function to identify small molecules that may block interactions of the viral spike glycoprotein with its cellular receptor glycoprotein. Our lab has identified receptors for human, murine and feline coronaviruses. We have expressed and purified soluble recombinant receptor glycoproteins, and are testing them for receptor activities. We will use mutagenesis to identify the amino acids and domains of the viral spike proteins and receptor glycoproteins that interact, and test the mutant proteins for binding activity and the ability to induce or undergo conformational changes that lead to membrane fusion and virus entry. We will analyze the structures of the spike proteins and receptor glycoproteins using cryo-electron microscopy and X-ray crystallography. The resulting structural models will be used to develop small molecules that block virus binding and/or receptor induced conformational changes in the spike protein. These molecules will be tested in cell culture model systems for toxicity and for the ability to block virus infection. Non-toxic drugs with anti-viral activity in cultures will be tested in animal model systems for the ability to block coronavirus infections of the respiratory tract. Our experimental models are the interactions of two coronaviruses, mouse hepatitis virus (MHV) and human coronavirus 229E (HCoV-229E), with their receptors, MHVR and hAPN, respectively. The murine model is more advanced than the human model, and will provide information and strategies that will facilitate our studies on human coronavirus-receptor interactions. The MHV model will also provide fundamental information on how an immunoglobulin-related receptor initiates infection with an enveloped virus. The 229E model will elucidate how the enzyme APN acts as a virus receptor. Inhibition of HCoV-229E respiratory infection by candidate drugs will be studied in transgenic mice that express the HCoV-229E receptor, hAPN, in the respiratory epithelium.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Virology Study Section (VR)
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Tseng, Christopher K
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University of Colorado Denver
Schools of Medicine
United States
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Tusell, Sonia M; Schittone, Stephanie A; Holmes, Kathryn V (2007) Mutational analysis of aminopeptidase N, a receptor for several group 1 coronaviruses, identifies key determinants of viral host range. J Virol 81:1261-73
Smith, M K; Tusell, Sonia; Travanty, Emily A et al. (2006) Human angiotensin-converting enzyme 2 (ACE2) is a receptor for human respiratory coronavirus NL63. Adv Exp Med Biol 581:285-8
Tusell, Sonia M; Holmes, Kathryn V (2006) Molecular interactions of group 1 coronaviruses with feline APN. Adv Exp Med Biol 581:289-91
Wentworth, David E; Tresnan, D B; Turner, B C et al. (2005) Cells of human aminopeptidase N (CD13) transgenic mice are infected by human coronavirus-229E in vitro, but not in vivo. Virology 335:185-97
Zelus, Bruce D; Schickli, Jeanne H; Blau, Dianna M et al. (2003) Conformational changes in the spike glycoprotein of murine coronavirus are induced at 37 degrees C either by soluble murine CEACAM1 receptors or by pH 8. J Virol 77:830-40
Breslin, Jamie J; Mork, Irene; Smith, M K et al. (2003) Human coronavirus 229E: receptor binding domain and neutralization by soluble receptor at 37 degrees C. J Virol 77:4435-8
Tan, Kemin; Zelus, Bruce D; Meijers, Rob et al. (2002) Crystal structure of murine sCEACAM1a[1,4]: a coronavirus receptor in the CEA family. EMBO J 21:2076-86
Blau, D M; Holmes, K V (2001) Human coronavirus HCoV-229E enters susceptible cells via the endocytic pathway. Adv Exp Med Biol 494:193-8
Wentworth, D E; Holmes, K V (2001) Molecular determinants of species specificity in the coronavirus receptor aminopeptidase N (CD13): influence of N-linked glycosylation. J Virol 75:9741-52
Wentworth, D E; Holmes, K V (2001) Addition of a single glycosylation site to hAPN blocks human coronavirus-229E receptor activity. Adv Exp Med Biol 494:199-204

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