Modified adenoviral vectors ablated for coxsackievirus-adenovirus receptor, alphav integrin, and heparan sulfate binding reduce in vivo tissue transduction and toxicity.
Coxsackievirus and adenovirus receptor (CAR), alphav integrins, and heparan sulfate glycosaminoglycans (HSGs) are the tropism determinants of adenoviral (Ad) vectors in vivo. For the development of a targeted Ad vector, its broad tropism needs to be blocked (or reduced). We have previously developed Ad vectors with ablation of CAR, alphav integrin, and HSG binding by mutation of the FG loop in the fiber knob (deletion of T489, A490, Y491, and T492 of the fiber protein), deletion of the RGD motif of the penton base, and substitution of the fiber shaft domain for that derived from Ad type 35, respectively, and have shown that this triple-mutant Ad vector [Ad/deltaF(FG)deltaP-S35-L2] exhibits significantly lower transduction in mouse liver compared with the conventional Ad vector [Koizumi, N., Mizuguchi, H., Sakurai, F., Yamaguchi, T., Watanabe, Y., and Hayakawa, T. (2003). J. Virol. 77, 13062-13072]. In the present study, we optimized the fiber knob mutation for further reduced in vivo transduction and examined toxicity of the modified Ad vectors. Ad/deltaF(AB)deltaPS35- L2, a triple-mutant Ad vector containing a mutation of the AB loop in the fiber knob (R412S, A415G, E416G, and K417G), mediated approximately 15,000- and 500-fold lower mouse liver transduction by intravenous and intraperitoneal administration, respectively, than the conventional Ad vector, and mediated 10- fold lower mouse liver transduction than did Ad/deltaF(FG)deltaP-S35-L2. Ad/deltaF(AB)deltaP-S35-L2 also exhibited lower transduction of other organs compared with Ad/deltaF(FG)deltaP-S35-L2 and the conventional Ad vector. Levels of both liver serum enzymes (aspartate transferase [AST] and alanine transferase (ALT)] and interleukin (IL)-6 in mouse serum after intravenous administration of Ad/deltaF(AB)deltaP-S35-L2 were similar to those in the nontreatment mouse serum, whereas the conventional Ad vector led to high levels of AST, ALT, and IL-6. We therefore succeeded in further improving the mutant Ad vector, abolishing both viral natural tropism and toxicity. This new Ad vector appears to be a fundamental vector for targeted gene delivery.
National Institute of Biomedical Innovation, Osaka 567-0085, Japan.
Naoya
N
Koizumi
Kenji
K
Kawabata
Fuminori
F
Sakurai
Yoshiteru
Y
Watanabe
Takao
T
Hayakawa
Hiroyuki
H
Mizuguchi
D000260:Adenoviruses, Human / Q000235:genetics* / Q000378:metabolism; D000410:Alanine Transaminase / Q000378:metabolism; D000818:Animals; D001219:Aspartate Aminotransferases / Q000378:metabolism; D001665:Binding Sites; D062727:Coxsackie and Adenovirus Receptor-Like Membrane Protein; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006497:Heparan Sulfate / Q000378:metabolism*; D006801:Humans; D039564:Integrin alphaV / Q000378:metabolism*; D015850:Interleukin-6 / Q000378:metabolism; D008099:Liver / Q000201:enzymology / Q000378:metabolism / Q000821:virology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009842:Oligopeptides / Q000235:genetics / Q000378:metabolism; D011991:Receptors, Virus / Q000378:metabolism*; D017384:Sequence Deletion; D014161:Transduction, Genetic
D000410:Alanine Transaminase; D001219:Aspartate Aminotransferases; D062727:Coxsackie and Adenovirus Receptor-Like Membrane Protein; D006497:Heparan Sulfate; D039564:Integrin alphaV; D015850:Interleukin-6; D009842:Oligopeptides; D011991:Receptors, Virus; C568771:CLMP protein, human; C568772:CLMP protein, mouse; C047981:arginyl-glycyl-aspartic acid