Research Article Enhanced delivery and expression of a nanoencapsulated DNA vaccine vector for respiratory syncytial virus Seyhan Boyoglu, MS a , Komal Vig, PhD a , Shreekumar Pillai, PhD a , Vijay Rangari, PhD b , Vida A. Dennis, PhD c , Fayaz Khazi, PhD d , Shree R. Singh, PhD a, ⁎ a Center for NanoBiotechnology Research, Alabama State University, Montgomery, Alabama, USA b Tuskegee University, Tuskegee, Alabama, USA c Tulane University Health Sciences Center, Covington, Louisiana, USA d Intrexon Corporation, Norristown, Pennsylvania, USA Received 19 November 2008; accepted 13 February 2009 Abstract This study evaluated the efficiency of chitosan-encapsulated DNA-based respiratory syncytial virus (RSV) vaccine. Antigenic regions of RSV F, M2, and G genes were cloned into the human cytomegalovirus promoter–based constitutive expression vector, resulting in a DNA vaccine vector named DR-FM2G. This vector was used to formulate DNA-chitosan nanoparticles (DCNPs) using a complex coacervation process that yielded an encapsulation efficiency of 94.7%. The DCNP sizes ranged from 80 to 150 nm with uniform size distribution and spherical shape. DNA release was between 50% and 60% when DCNPs were incubated with similar gastrointestinal fluid (pH 2), whereas 21% to 25% of DNA was released from DCNPs in 30 minutes at pH 10. Differential scanning calorimetry showed DCNPs to be more stable than naked DNA or chitosan, offering protection from DNA degradation by nucleases. DCNPs were not toxic to cells when used at concentrations ≤400 μg/mL. Immunohistochemical and real-time polymerase chain reaction results showed a higher level of RSV protein expression in mouse tissues given when DCNPs were injected intravenously as compared with naked DNA. From the Clinical Editor: This study evaluated the efficiency of chitosan-encapsulated DNA-based respiratory syncytial virus (RSV) vaccine, showing a higher level of RSV protein expression in mouse tissues given when DCNPs were injected intravenously as compared with naked DNA. © 2009 Elsevier Inc. All rights reserved. Key words: DNA vaccine; Respiratory syncytial virus; Chitosan; Nanoencapsulation Human respiratory syncytial virus (RSV) is classified in the order Mononegavirales, family Paramyxoviridae, subfamily Pneumovirinae, genus Pneumovirus. RSV causes severe lower respiratory tract infections in infants and can lead to pneumonia. RSV accounts for approximately 4500 deaths and 90,000 hospitalizations in infants and children in the United States each year. 1 In the United States RSV is also responsible for almost 3.3 million cases of respiratory tract diseases in elderly patients. 2 The RSV genome is a negative-sense RNA strand consisting of 15,200 nucleotides, which encode 11 proteins. 3 Among them, the fusion (F) protein, attachment glycoprotein (G), and matrix protein (M2) are the leading candidates for vaccine development. However, a safe and effective vaccine against this virus is not available for many reasons, among them the causation of exacerbated disease by vaccines, residual virulence, overattenuation, or failure to induce an adequate immune response. 4 Development of an effective RSV vaccine requires the induction of mucosal immunity. The nasal administration of proteins, attenuated RSV, and DNA induces humoral and cellular immune responses. However, the major hurdle is the efficient induction of mucosal immune responses when immunogens are administered via the intranasal route. In particular, DNA administration does not induce high titers of mucosal antibodies because of poor cellular uptake and a rapid in vivo degradation of DNA. Therefore, effective delivery systems are required to deliver DNA to antigen-presenting cells and to specific organs. Traditionally, DNA delivery systems have been classified as viral vector-mediated systems and nonviral vector-mediated systems. However, the application of viral carriers has several disadvantages, such as undesired Available online at www.sciencedirect.com Nanomedicine: Nanotechnology, Biology, and Medicine 5 (2009) 463 – 472 www.nanomedjournal.com This work was supported by National Science Foundation-CREST (HRD-0734232) and NSF-HBCU-UP (HRD-0505872) grants. ⁎ Corresponding author. E-mail address: ssingh@alasu.edu (S.R. Singh). Please cite this article as: S. Boyoglu, et al, Nanomedicines for ocular NSAIDs: safety on drug delivery, Nanomedicine: NBM 2009;5:463-472, doi:10.1016/j.nano.2009.02.004 1549-9634/$ – see front matter © 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.nano.2009.02.004