Antiviral Research 68 (2005) 163–172 A novel inhibitor of respiratory syncytial virus isolated from ethnobotanicals Joshua O. Ojwang a,∗ , Yan-Hong Wang b , Philip R. Wyde c , Nikolaus H. Fischer b , Wolfgang Schuehly b , James R. Appleman a,1 , Soreeta Hinds a , Craig D. Shimasaki d a ZymeTx, Inc., 655 Research Parkway Suite 554, Oklahoma City, OK 73104, USA b University of Mississippi, Department of Pharmacognosy and Natural Center for Natural Products Research, RIPS, University, MS 38677, USA c Baylor College of Medicine, Department of Molecular Virology & Microbiology, One Baylor Plaza, Houston, TX 77030, USA d InterGenetics, Inc., 655 Research Parkway Suite 300, Oklahoma City, OK 73104, USA Received 30 June 2005; accepted 21 September 2005 Abstract A novel low molecular weight compound, CJ 4-16-4, isolated from ethnobotanicals using bioassay-guided fractionation, was found to be a potent inhibitor of respiratory syncytial virus (RSV) in vitro and in vivo. In vitro, a very low micromolar efficacious dose was obtained against at least four of subtype A (RSV-Long, RSV A2, and RSV A6 57754) and one of subtype B (Washington) RSV strains without seeing any significant cytotoxicity to Hep-2, MDCK or Vero cell lines. The drug inhibits growth of RSV in Hep-2 cells maintained in tissue culture at a very low concentration (∼0.07 M) with cell toxicity >400 M (TI > 5880). In a cotton rat model of RSV infection, the drug was able to reduce viral titers by ∼1 log at dose 12.5 and 25 mg/kg/day, and by >2 log at 100 mg/kg/day. This antiviral activity was specific as influenza A and B and herpes simplex 1 and 2 viruses were not inhibited. The results obtained indicate that CJ 4-16-4 warrants clinical development. © 2005 Elsevier B.V. All rights reserved. Keywords: Inhibitors; Antivirals; Respiratory syncytial virus; Natural product; Ethnobotanicals 1. Introduction Respiratory syncytial virus (RSV) belongs to Pneumovirus genus of the Paramyxoviridae family. RSV is a negative stranded RNA virus comprised of two antigenic subgroups, A and B. The two subgroups are different in their G surface glycoprotein but remain antigenitically similar in their F or fusion surface glyco- protein (Chanock and McIntosh, 1990; Grosfeld et al., 1995). RSV is the leading cause of severe lower respiratory tract infections in infants and children under 2 years of age (Parrott et al., 1973; Glezen et al., 1982; Chanock and McIntosh, 1990). It has been reported to be responsible for 40–50% of hospitaliza- tions for bronchiolitis in the United States and 25% of pediatric hospitalizations for pneumonia (Dowell et al., 1996). Reinfec- tion with RSV is common, with about 76% of children being reinfected during the second year of life. RSV reinfection also ∗ Corresponding author. Tel.: +1 405 809 1314; fax: +1 405 809 1944. E-mail address: Ojwangj@zymetx.com (J.O. Ojwang). 1 Present Address: Anadys Pharmaceutical, Inc., San Diego, CA, USA. occurs in adults and is a common cause of hospitalization of adult patients with chronic lung conditions such as asthma and chronic obstructive pulmonary disease. It can also impact healthy adults. It was reported that 43% of adults with confirmed RSV missed work for periods up to 2 weeks (Dowell et al., 1996; Falsey and Walsh, 2000; Murry and Dowell, 1997). This virus has also been reported to be an important factor in the development of otitis media infections and hyper-reactive airway disease in later life (La Via et al., 1992). RSV has also been shown to be significant pathogen in the elderly (Falsey et al., 1992,1995; Mills, 1996; Treanor and Falsey, 1999). In this population, RSV infection often mani- fests as a flu-like illness that can be misdiagnosed as influenza (Mathur et al., 1980; Osterweil and Norman, 1990). The impact of RSV-related mortality was recently illustrated in an epidemi- ology study, which estimated that in the United States more than 17,000 deaths per year are caused by RSV infection, with greater than 80% of these deaths occurring in people over 65 years of age (Thompson et al., 2003). Nursing residents are at greater risk, with outbreak rates measured as high as 40% (Mathur et al., 1980; Sorvillo et al., 1984). RSV is also responsible for serious 0166-3542/$ – see front matter © 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.antiviral.2005.09.003