Cell Biol Toxicol (2010) 26:1–20 (VSV-G) envelope protein substantially reduces the potential to form wild-type virus (Kafri et al. 1999; Naldini et al. 1996), increases viral stability and increases the host cell tropism, enabling transfection of both dividing and non-dividing cells in vitro (Kafri et al. 1999; Naldini et al. 1996; Reiser et al. 1996) and neurons following direct injection in the brain (Naldini et al. 1996). VSV-G psuedotyping is now widely used, and such vectors recently gained FDA clearance and have entered clinical application (Lu et al. 2004; Manilla et al. 2005). However, VSV-G can be inactivated by serum complement (DePolo et al. 2000) and has been shown to be toxic following long- term infection. Recent studies have therefore investi- gated pseudotyping with alternative glycoproteins (GPs) derived from other viruses, each with specific advantages and disadvantages (Cronin et al. 2005). Examples include pseudotyping with rabies, Ebola Zaire (EboZ), lymphocytic choriomeningitis virus (LCMV), alpha viruses, Sindbis virus and Hantavirus glycoproteins. Both HIV-1 lentiviral and EIAV vec- tors have been successfully pseudotyped with various rabies virus glycoproteins (Conzelmann et al. 1990; Mebatsion et al. 1995; Mitrophanous et al. 1999; Mochizuki et al. 1998). These glycoproteins enable retrograde transport of transgene expression along nerve tracts and into distal brain areas (Wong et al. 2004). MLV-based retroviral vectors pseudotyped with the LCMV GP have been shown to efficiently transduce fibroblasts, epithelial cells, hematopoietic cells, as well as hepatoma, neuroblastoma and glioma cell lines (Miletic et al. 1999). FIV LCMV-WE pseudotypes are potentially useful for targeting neural stem/progenitor cells in vivo. Several reports describ- ing successful pseudotyping of lentiviral vectors with GP from alpha viruses (river virus (RRV), Semliki forest virus and Sindbis virus) have also been described. RRV-pseudotyped FIV vectors (RRV/FIV) predominantly transduced the liver of recipient mice with a 20-fold higher transduction efficiency than that achieved with a VSV-G pseudotype that caused less cytotoxicity. Non-hepatocytes (mainly Kupffer cells) were also transduced, and following injection into the brain, RRV/FIV preferentially transduced neuroglial cells (astrocytes and oligodendrocytes; Kang et al. 2002). Intravenous injection of oncoretroviruses and lentiviruses pseudotyped with a modified chimeric Sindbis virus shows high targeting specificity and low nonspecific infectivity in liver and spleen (Morizono 11 et al. 2001). Successful pseudotyping of HIV-1 vectors using Marburg virus and EboZ virus-derived virion GPs has also been described. Such vectors were more efficient in transducing myocytes, and EboZ pseudotyped vectors injected into muscle in utero were found to transduce muscle satellite or stem cells and lead to transgene expression in newly regenerated muscle after injury (MacKenzie et al. 2005). Direct brain injection of EboZ/HIV pseudo- types in adult mice failed to transduce cells in the brain (Watson et al. 2002). This may be due to the lack of the proper receptor in the CNS. However, EboZ pseudotypes were able to efficiently transduce the apical surface of airway epithelia after apical applica- tion (Kobinger et al. 2001), and therefore, these vectors are now being employed for cystic fibrosis studies (Sanders 2004). Modified lentiviral vector pseudotyped with a strain of Hantavirus (HTNV) has also been used to improve the transduction efficiency into vascular smooth muscle and endothelial cells in vitro and in vivo (Qian et al. 2006). The physical targeting of viral vectors can also be modified by coating the virus surface with polymers such as polyethylene glycol, poly-[N-(2-hydroxy- propyl) methacrylamide] or biodegradable alginate microparticles (Croyle et al. 2000; Kreppel and Kochanek 2008; Sailaja et al. 2002). Such modifica- tions ablate the native tropism of the vector and shield it from the host immune response whilst allowing selec- tive targeting by the attachment of a variety of targeting ligands (peptides, proteins or antibodies) to these polymers (Kreppel and Kochanek 2008; Morrison et al. 2008). As mentioned previously, physical targeting of Ad vectors has been altered by the use of bispecific adaptor molecules (including bispecific antibodies or fusion proteins; Dmitriev et al. 2000; Parrott et al. 2003; Reynolds et al. 2001). In this strategy, however, the two-component nature of bispecific molecules adds complications in manufacturing and in maintaining batch-to-batch homogeneity. Furthermore, these mod- ifications would not be inherited by progeny virions, and therefore, genetic modification of the capsid proteins is a more favoured option at this time. In addition to these viral particle targeting methods, tissue-specific promoters or tissue-specific regulatory elements (Nettelbeck 2008) have been used to limit transgene expression to the tissue of interest. Some tissue-specific promoters investigated are glial fibrillary acidic protein (GFAP), synapsin, muscle creatine