Multiply deleted [E1, polymerase-, and pTP-] adenovirus vector persists despite deletion of the preterminal protein

Background The inherent Limitations of [E1-]Ad vectors as gene therapy vehi cles suggest that further modifications may improve their overall performan ce profiles. However, Ad vector modifications can have untoward effects on their basic biology, e.g., some helper-virus dependent Ad vectors have been found to be unstable without the presence of preterminal protein (pTP) act ivities. Despite this concern, we generated a new class of helper-virus ind ependent Ad vector that was multiply deleted for the E1, polymerase, and pT P genes, and investigated the ramifications of these deletions upon several vector performance parameters. Methods The construction and propagation of an [E1-, polymerase-, pTP-]Ad v ector was achieved with the use of trans-complementing cells co-expressing the Ad E1, polymerase and pTP genes. Results High titer production of the [E1-, polymerase-, pTP-]Ad vector was successfully accomplished via conventional Ad purification techniques. This unique class of Ad vector was capable of long-term gene transfer in vivo ( despite lacking pTP functions) that was concomitant with a significantly de creased hepatic toxicity. Conclusions Previous studies had suggested that Ad genome persistence in vi vo may be dependent upon the presence of low level vector genome replicatio n and/or pTP functions. Our results suggest that [E1-, polymerase-, pTP-]Ad vectors can overcome these barriers. The further benefits afforded by the use of this class of Ad vector (increased cloning capacity, high level grow th, decreased propensity to generate replication competent Ad (RCA), decrea sed toxicity) suggests that they will be highly beneficial for use in sever al aspects of human gene therapy. Copyright (C) 2000 John Wiley & Sons, Ltd .