10,11,12 Innate responses can prime for a humoral immune response generating neutralizing antibodies to adenoviral capsid antigens, which are augmented by T-cell help.1,2 Visualization and measurements of transgene expression are needed to refine and optimize gene therapy strategies. We have recently set up in nonhuman primates a semiquantitative selleck chemicals Pacritinib transgene-specific positron emission tomography (PET) technique,13 which allows detection and imaging of the reporter gene herpes simplex virus type 1 thymidine kinase (HSV1-tk). HSV1-tk as a reporter transgene can be traced by means of PET imaging in nonhuman primates13,14 and in humans.15,16 This imaging technique is based on measurements of retention of [18F]9-(4-[18F]-fluoro-3-hydroxymethylbutyl)-guanine ([18F]FHBG) in the liver parenchyma once phosphorylated by HSV1-tk.

This noninvasive imaging technique is useful for assessing the possibility of repeated gene transfer. We reasoned that if first-generation recombinant adenovirus could be readministered, it would be easier to repeatedly use less immunogenic adenoviral vector generations. An additional advantage of our adenoviral system is the transient time course of transgene expression.15 Once transgene expression is extinguished in about 2 weeks, the visualization of the efficiency of subsequent gene transfers is experimentally feasible. Transgene expression extinction is explained by immune elimination of transduced cells and gene silencing affecting the cytomegalovirus (CMV) promoter.17 Many approved drugs exist5,18,19,20,21,22 that thwart the immune response and might allow for viral vector readministration.

A previous report23 has shown the feasibility and safety of pharmacological immunosuppression of three macaques in an attempt to block T-cell responses to AAV encoding factor IX coagulation factor. In this study, we have demonstrated that pharmacological immunosuppression can achieve repeated liver gene transfer with the same first-generation adenoviral vector in spite of the high degree of immunogenicity attributed to this viral vector. B-cell depletion with anti-CD20 monoclonal antibody19,24 and concomitant T-cell inhibition with clinically available drugs permits repeated liver gene Cilengitide transfer in macaques. Results Rituximab and FK506 treatment reduce the antiadenoviral humoral and cellular immunity that prevents gene-transfer repetition with the same vector To test whether immunosuppression could allow repeated administration of an adenoviral vector, a group of macaques treated i.v. with the clinical-grade first-generation recombinant adenovirus AdCMVHSV1.tk25 (Figure 1a) received two courses of Rituximab to deplete B cells26 plus a conventional daily oral regimen with FK506 (ref. 18) to repress T cells (Figure 1a).