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Abstract. Human adenovirus type 2 (Ad2) enters host cells by receptor-mediated endocytosis, an event mediated by the virus penton base binding to cell surface integrins otv/33 and otv/35. While both o~v integrins promote virus internalization, olv/35 is involved in the subsequent event of membrane permeabilization. Cells transfected with the r5 or/33 subunit, expressing either otvfl5 and otv/33, respectively, were capable of supporting Ad2 infection to varying degrees. In this case, cells expressing otvfl5 were significantly more susceptible to Ad2-induced membrane permeabilization, as well as to Ad2 infection, than cells expressing avfl3. Adenovirus-mediated gene delivery was also more efficient in cells expressing txvfl5. These results suggest that the interaction of oLv/35 with Ad2 penton base facilitates the subsequent step of virus penetration into the cell. These studies provide evidence for the involvement of a cellular receptor in virusmediated membrane permeabilization and suggest a novel biological role for integrin avfl5 in the infectious pathway of a human adenovirus. crucial step in virus infection of host cells is penetration/permeabilization of the cell plasma membrane, a past-internalization event required for delivery of the viral genome into the cytoplasm. Although a substantial amount of knowledge exists on cell entry by enveloped viruses, the mechanism(s) by which nonenveloped viruses penetrate cells is not well understood. Adenovirus, a nonenveloped DNA virus that is a major cause of respiratory and gastrointestinal infections of children (3,14), has proved useful for studying cell entry by nonenveloped viruses.Of the over 40 different serotypes of human adenovirus, the majority of cell interaction studies have been performed with serotype 2 (human adenovirus type 2; Ad2) t. Initial attachment of Ad2 to host cells is mediated by the fiber protein (13, 17), an elongated 62-kD protein that is present on each of the 12 vertices of the virion capsid (28). The fiber receptor, which is broadly distributed on a variety of cells, has not yet been identified. After Ad2 attachment to the fiber receptor, virus particles are rapidly internalized into clathrin-coated vesicles by receptor-mediated endocytosis (4, 33). The fiber protein is dissociated from the virion particle early in the entry pathway (12). Ad2 internalization is mediated by either of two secondary host cell receptors, integrins olv/33 and o~v/35 (38).

The human adenovirus serotype 5 (Ad5) is used widely for applications in human gene therapy. Cellular attachment of Ad5 is mediated by binding of the carboxyl-terminal knob of its fiber coat protein to the Coxsackie adenovirus receptor (CAR) protein. However, Ad5 binding to CAR hampers the development of adenovirus vectors capable of specifically targeting (diseased) tissues or organs. Through sequence analysis and mutagenesis, a conserved receptor-binding region was identified on the side of three divergent CAR-binding knobs. The feasibility of simultaneous CAR ablation and redirection of an adenovirus to a new receptor is demonstrated.

Purpose: Therapeutic nanoparticles are designed to deliver their drug payloads through enhanced permeability and retention (EPR) in solid tumors. The extent of EPR and its variability in human tumors is highly debated and has been proposed as an explanation for variable responses to therapeutic nanoparticles in clinical studies. Experimental Design: We assessed the EPR effect in patients using a 64Cu-labeled nanoparticle, 64Cu-MM-302 (64Cu-labeled HER2-targeted PEGylated liposomal doxorubicin), and imaging by Positron Emission Tomography/Computed Tomography (PET/CT). Nineteen patients with HER2-positive metastatic breast cancer underwent 2–3 PET/CT scans post-administration of 64Cu-MM-302 as part of a clinical trial of MM-302 plus trastuzumab with and without cyclophosphamide (). Results: Significant background uptake of 64Cu-MM-302 was observed in liver and spleen. Tumor accumulation of 64Cu-MM-302 at 24–48 h varied 35-fold (0.52 to 18.5 %ID/kg) including deposition in bone and brain lesions, and was independent of systemic plasma exposure. Computational analysis quantified rates of deposition and washout, indicating peak liposome deposition at 24–48 h. Patients were classified based on 64Cu-MM-302 lesion deposition using a cut-point that is comparable to a response threshold in preclinical studies. In a retrospective exploratory analysis of patient outcomes relating to drug levels in tumor lesions, high 64Cu-MM-302 deposition was associated with more favorable treatment outcomes (hazard ratio = 0.42). Conclusions: These findings provide important evidence and quantification of the EPR effect in human metastatic tumors, and support imaging nanoparticle deposition in tumors as a potential means to identify patients well-suited for treatment with therapeutic nanoparticles.

Adenovirus (Ad) is used as a vector for gene delivery in therapies involving genetic disease, vascular disease, and cancer. The first step for efficient gene transfer is effective virus binding to the target cells. We have found that Ad-mediated gene delivery to multiple cell types is much less efficient compared to epithelial-derived cells. Low gene delivery to nonepithelial cell types was directly correlated to a deficiency of the cellular receptor which mediates Ad binding. To overcome this inefficiency we constructed a new virus, AdPK, that contains a heparin-binding domain that targets the virus to broadly expressed, heparan-containing cellular receptors. AdPK delivers genes to multiple cell types at markedly higher efficiencies than unmodified Ad. Viruses with enhanced attachment characteristics significantly improve gene transfer efficiency and may expand the tissues amenable to efficient Ad-mediated gene therapy.