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Adenovirus has strong therapeutic potential as an oncolytic virus and gene therapy vector. However, injecting human species C serotype 5 adenovirus, HAdv-C5, into the bloodstream leads to numerous interactions with plasma proteins that affect viral tropism and biodistribution, and can lead to potent immune responses and viral neutralization. The HAdv/factor X (FX) interaction facilitates highly efficient liver transduction and protects virus particles from complement-mediated neutralization after intravenous delivery. Ablating the FX interaction site on the HAdv-C5 capsid leaves the virus susceptible to neutralization by natural IgM followed by activation of the complement cascade and covalent binding of complement components C4b and C3b to the viral capsid. Here we present structural models for IgM and complement components C1, C4b, and C3b in complex with HAdv-C5. Molecular dynamics simulations indicate that when C3b binds near the vertex, multiple stabilizing interactions can be formed between C3b, penton base, and fiber. These interactions may stabilize the vertex region of the capsid and prevent release of the virally encoded membrane lytic factor, protein VI, which is packaged inside of the viral capsid, thus effectively neutralizing the virus. In a situation where FX and IgM are competing for binding to the capsid, IgM may not be able to form a bent conformation in which most of its Fab arms interact with the capsid. Our structural modeling of the competitive interaction of FX and IgM with HAdv-C5 allows us to propose a mechanistic model for FX inhibition of IgM-mediated virus neutralization. According to this model, although IgM may bind to the capsid, in the presence of FX it will likely retain a planar conformation and thus be unable to promote activation of the complement cascade at the virus surface. [ABSTRACT FROM AUTHOR]

The expression of cytokines and chemokines in response to adenovirus infection is tightly regulated by the innate immune system. Cytokine-mediated toxicity and cytokine storm are known clinical phenomena observed following naturally disseminated adenovirus infection in immunocompromised hosts as well as when extremely high doses of adenovirus vectors are injected intravenously. This dose-dependent, cytokine-mediated toxicity compromises the safety of adenovirus-based vectors and represents a critical problem, limiting their utility for gene therapy applications and the therapy of disseminated cancer, where intravenous injection of adenovirus vectors may provide therapeutic benefits. The mechanisms triggering severe cytokine response are not sufficiently understood, prompting efforts to further investigate this phenomenon, especially in clinically relevant settings. In this review, we summarize the current knowledge on cytokine and chemokine activation in response to adenovirus- and adenovirus-based vectors and discuss the underlying mechanisms that may trigger acute cytokine storm syndrome. First, we review profiles of cytokines and chemokines that are activated in response to adenovirus infection initiated via different routes. Second, we discuss the molecular mechanisms that lead to cytokine and chemokine transcriptional activation. We further highlight how immune cell types in different organs contribute to synthesis and systemic release of cytokines and chemokines in response to adenovirus sensing. Finally, we review host factors that can limit cytokine and chemokine expression and discuss currently available and potential future interventional approaches that allow for the mitigation of the severity of the cytokine storm syndrome. Effective cytokine-targeted interventional approaches may improve the safety of systemic adenovirus delivery and thus broaden the potential clinical utility of adenovirus-based therapeutic vectors. [ABSTRACT FROM AUTHOR]

De-livering an oncolytic virus: Oncolytic viruses, optimized for killing tumor cells, are a promising modality in cancer immunotherapy. However, there are some challenges that these viruses must overcome, including antiviral immune responses that can affect both the efficacy and the safety of these viruses. For example, naturally occurring immunoglobulin M (IgM) antibodies bind to human species C adenovirus, resulting in trapping of the virus in the liver. To overcome this, Atasheva et al. engineered a modified version of this adenovirus, with mutations in the binding site for IgM. This modified virus was not trapped in the liver and effectively suppressed tumor growth in mouse models without safety concerns. Oncolytic virus therapy is a cancer treatment modality that has the potential to improve outcomes for patients with currently incurable malignancies. Although intravascular delivery of therapeutic viruses provides access to disseminated tumors, this delivery route exposes the virus to opsonizing and inactivating factors in the blood, which limit the effective therapeutic virus dose and contribute to activation of systemic toxicities. When human species C adenovirus HAdv-C5 is delivered intravenously, natural immunoglobulin M (IgM) antibodies and coagulation factor X rapidly opsonize HAdv-C5, leading to virus sequestration in tissue macrophages and promoting infection of liver cells, triggering hepatotoxicity. Here, we showed that natural IgM antibody binds to the hypervariable region 1 (HVR1) of the main HAdv-C5 capsid protein hexon. Using compound targeted mutagenesis of hexon HVR1 loop and other functional sites that mediate virus-host interactions, we engineered and obtained a high-resolution cryo–electron microscopy structure of an adenovirus vector, Ad5-3M, which resisted inactivation by blood factors, avoided sequestration in liver macrophages, and failed to trigger hepatotoxicity after intravenous delivery. Systemic delivery of Ad5-3M to mice with localized or disseminated lung cancer led to viral replication in tumor cells, suppression of tumor growth, and prolonged survival. Thus, compound targeted mutagenesis of functional sites in the virus capsid represents a generalizable approach to tailor virus interactions with the humoral and cellular arms of the immune system, enabling generation of "designer" viruses with improved therapeutic properties. [ABSTRACT FROM AUTHOR]

Adenovirus is a highly evolutionary successful pathogen, as it is widely prevalent across the animal kingdom, infecting hosts ranging from lizards and frogs to dolphins, birds, and humans. Although natural adenovirus infections in humans rarely cause severe pathology, intravenous injection of high doses of adenovirus‐based vectors triggers rapid activation of the innate immune system, leading to cytokine storm syndrome, disseminated intravascular coagulation, thrombocytopenia, and hepatotoxicity, which individually or in combination may cause morbidity and mortality. Much of the information on exactly how adenovirus activates the innate immune system has been gathered from mouse experimental systems. Intravenous administration of adenovirus to mice revealed mechanistic insights into cellular and molecular components of the innate immunity that detect adenovirus particles, activate pro‐inflammatory signaling pathways and cytokine production, sequester adenovirus particles from the bloodstream, and eliminate adenovirus‐infected cells. Collectively, this information greatly improved our understanding of mechanisms of activation of innate immunity to adenovirus and may pave the way for designing safer adenovirus‐based vectors for therapy of genetic and acquired human diseases. [ABSTRACT FROM AUTHOR]

Inflammation is a highly coordinated host response to infection, injury, or cell stress. In most instances, the inflammatory response is pro-survival and is aimed at restoring physiological tissue homeostasis and eliminating invading pathogens, although exuberant inflammation can lead to tissue damage and death. Intravascular injection of adenovirus (Ad) results in virus accumulation in resident tissue macrophages that trigger activation of CXCL1 and CXCL2 chemokines via the IL-1α-IL-1RI signaling pathway. However, the mechanistic role and functional significance of this pathway in orchestrating cellular inflammatory responses to the virus in vivo remain unclear. Resident metallophilic macrophages expressing macrophage receptor with collagenous structure (MARCO+) in the splenic marginal zone (MZ) play the principal role in trapping Ad from the blood. Here we show that intravascular Ad administration leads to the rapid recruitment of Ly-6G+7/4+ polymorphonuclear leukocytes (PMNs) in the splenic MZ, the anatomical compartment that remains free of PMNs when these cells are purged from the bone marrow via a non-inflammatory stimulus. Furthermore, PMN recruitment in the splenic MZ resulted in elimination of virus-containing cells. IL-1α-IL-1RI signaling is only partially responsible for PMN recruitment in the MZ and requires CXCR2, but not CXCR1 signaling. We further found reduced recruitment of PMNs in the splenic MZ in complement C3-deficient mice, and that pre-treatment of IL-1α-deficient, but not wild-type mice, with complement inhibitor CR2-Crry (inhibits all complement pathways at C3 activation) or CR2-fH (inhibits only the alternative complement activation pathway) prior to Ad infection, abrogates PMN recruitment to the MZ and prevents elimination of MARCO+ macrophages from the spleen. Collectively, our study reveals a non-redundant role of the molecular factors of innate immunity – the chemokine-activating IL-1α-IL-1RI-CXCR2 axis and complement – in orchestrating local inflammation and functional cooperation of PMNs and resident macrophages in the splenic MZ, which collectively contribute to limiting disseminated pathogen spread via elimination of virus-containing cells. [ABSTRACT FROM AUTHOR]

Numerous human genetic and acquired diseases could be corrected or ameliorated if viruses are harnessed to safely and effectively deliver therapeutic genes to diseased cells and tissues in vivo. Innate immune and inflammatory response represents one of the key stumbling blocks during the development of viral-based therapies. In this review, current data on the early innate immune responses to viruses and to the most commonly used gene therapy vectors (using adenovirus and adeno-associated virus) will be discussed. Recent findings in the field may help develop new approaches to moderate these innate immune anti-viral responses and thus improve the safety of viral vectors for human gene therapy applications. [ABSTRACT FROM AUTHOR]

Gene therapy for hemoglobinopathies requires efficient gene transfer into hematopoietic stem cells and high-level erythroid-specific gene expression. Toward this goal, we constructed a helper-dependent adenovirus vector carrying the β-globin locus control region (LCR) to drive green fluorescent protein (GFP) expression, whereby the LCR-GFP cassette is flanked by adeno-associated virus (AAV) inverted terminal repeats (Ad.LCR-β-GFP). This vector possesses the adenovirus type 35 fiber knob that allows efficient infection of hematopoietic cells. Transduction and vector integration studies were performed in MO7e cells, a growth factor-dependent CD34+ erythroleukemic cell line, and in cord blood-derived human CD34+ cells. Stable transduction of MO7e cells with Ad.LCR-β-GFP was more efficient and less subject to position effects and silencing than transduction with a vector that did not contain the β-globin LCR. Analysis of integration sites indicated that Ad.LCR-β-GFP integration in MO7e cells was not random but tethered to chromosome 11, specifically to the globin LCR. More than 10% of analyzed integration sites were within the chromosomal β-globin LCR. None of the Ad.LCR-β-GFP integrations occurred in exons. The integration pattern of a helper-dependent vector that contained X-chromosomal stuffer DNA was different from that of the β-globin LCR-containing vector. Infection of primary CD34+ cells with Ad.LCR-β-GFP did not affect the clonogenic capacity of CD34+ cells. Transduction of CD34+ cells with Ad.LCR-β-GFP resulted in vector integration and erythroid lineage-specific GFP expression. [ABSTRACT FROM AUTHOR]

Adenoviruses (Ad) are efficient vehicles for gene delivery in vitro and in vivo. Therefore, they are a promising tool in gene therapy, particularly in the treatment of cancer and cardiovascular diseases. However, preclinical and clinical studies undertaken during the last decade have revealed a series of problems that limit both the safety and efficacy of Ad vectors, specifically after intravenous application. Major obstacles to clinical use include innate toxicity and Ad sequestration by nontarget tissues. The factors and mechanisms underlying these processes are poorly understood. The majority of intravenously injected Ad particles are sequestered by the liver, which in turn causes an inflammatory response characterized by acute transaminitis and vascular damage. Here, we describe a novel pathway that is used by Ad for infection of hepatocytes and Kupffer cells upon intravenous virus application in mice. We found that blood factors play a major role in targeting Ad vectors to hepatic cells. We demonstrated that coagulation factor IX and complement component C4-binding protein can bind the Ad fiber knob domain and provide a bridge for virus uptake through cell surface heparan sulfate proteoglycans and low-density lipoprotein receptor-related protein. An Ad vector, Ad5mut, which contained mutations in the fiber knob domain ablating blood factor binding, demonstrated significantly reduced infection of liver cells and liver toxicity in vivo. This study contributes to a better understanding of adenovirus-host interactions for intravenously applied vectors. It also provides a rationale for novel strategies to target adenovirus vector to specific tissues and to reduce virus-associated toxicity after systemic application. [ABSTRACT FROM AUTHOR]

Adenovirus (Ad) vectors are widely used for gene delivery in vitro and in vivo. A solid understanding of the biology of this virus is imperative for the development of novel, effective, and safe vectors. For the group C adenovirus serotypes 2 and 5 that use CAR as a primary attachment receptor, it is known that the penton base RGD motifs interact with cellular integrins and that this interaction promotes virus internalization. However, the RGD motif's impact on the efficiency of postinternalization steps, such as the escape of the virus particle from the endosome, is less defined. Furthermore, the role of penton-integrin interactions remains unknown for new vectors possessing group B Ad fiber knobs that use CD46 as a primary virus attachment receptor. In this study, we used vectors with the RGD motif deleted that contained Ad5 and B-group Ad35 fiber knobs and long fiber shafts and studied the role of RGD-integrin interactions in virus internalization and endosome escape. The deletion of the RGD motif in the penton base did not affect virus attachment, regardless of the type of cellular receptor used for attachment. RGD motif deletion, however, significantly reduced the rate of virus internalization for both the Ad5 and Ad35 fiber knob-containing vectors. This study also demonstrates the role of penton RGD motifs in facilitating the endosome escape step of virus infection and indicates that penton-integrin interactions are involved in internalization of capsid-chimeric CD46-interacting Ads with long fiber shafts. [ABSTRACT FROM AUTHOR]

Adenovirus serotype 5 (Ad5) vectors containing Ad B-group fibers have become increasingly popular as gene transfer vectors because they efficiently transduce human cell types that are relatively refractory to Ad5 infection. So far, most B-group fiber-containing vectors have been first-generation vectors, deleted of E1 and/or E3 genes. Transduction with these vectors, however, results in viral gene expression and is associated with cytotoxicity and immune responses against transduced cells. To circumvent these problems, we developed fiber-chimeric Ad vectors devoid of all viral genes that were produced either by the homologous recombination of first-generation vectors or by using the Cre/lox-based helper virus system. In this study we compared early steps of infection between first-generation (35-kb genome) and Ad vectors devoid of all viral genes with genome sizes of 28 kb and 12.6 kb. All vectors possessed an Ad35-derived fiber knob domain, which uses CD46 as a primary attachment receptor. Using immortalized human hematopoietic cell lines and primary human CD34-positive hematopoietic cells, we found that the Ad genome size did not affect the efficiency of virus attachment to and internalization into cells. Furthermore, independently of the genome length and structure, all vectors migrated to the nucleus through late endosomal and lysosomal cellular compartments. However, the vector containing the short 12.6-kb genome was unable to efficiently escape from endosomes and deliver its DNA into the nucleus. Moreover, compared to other vectors, these Ad particles were less stable and had an abnormal capsid protein composition, including a lack of capsid-stabilizing protein IX. Our data indicate that the size and structure of the packaged viral genomes can affect the integrity of Ad particles, which in turn results in lower infectivity of Ad vectors. [ABSTRACT FROM AUTHOR]

After intravenous administration, adenovirus (Ad) vectors are predominantly sequestered by the liver. Delineating the mechanisms for Ad accumulation in the liver is crucial for a better understanding of Ad clearance and Ad-associated innate toxicity. To help address these issues, in this study, we used Ad vectors with different fiber shaft lengths and either coxsackievirus-Ad receptor (CAR)-interacting Ad serotype 9 (Ad9) or non-CAR-interacting Ad35 fiber knob domains. We analyzed the kinetics of Ad vector accumulation in the liver, uptake into hepatocytes and Kupffer cells, and induction of cytokine expression and release in response to systemic vector application. Immediately after intravenous injection, all Ad vectors accumulated equally efficiently in the liver; however, only genomes of long-shafted Ads were maintained in the liver tissue over time. We found that Kupffer cell uptake of long-shafted Ads was mediated by the fiber knob domain and was CAR independent. The short-shafted Ads were unable to efficiently interact with hepatocellular receptors and were not taken up by Kupffer cells. Moreover, our studies indicated that Kupffer cells were not the major reservoir for the observed accumulation of Ads (used in this study) in the liver within the first 30 min after virus infusion. The lower level of liver cell transduction by short-shafted Ads correlated with a significantly reduced inflammatory anti-Ad response as well as liver damage induced by the systemic administration of these vectors. This study contributes to a better understanding of the biology of systemically applied Ad and will help in designing safer vectors that can efficiently transduce target tissues. [ABSTRACT FROM AUTHOR]

Group B adenoviruses, a subgenus of human Adenoviridae, are associated with a variety of often-fatal illnesses in immunocompromised individuals, including bone marrow transplant recipients and cancer and AIDS patients. Recently, group B adenovirus derivatives have gained interest as attractive gene therapy vectors because they can transduce target tissues, such as hematopoietic stem cells, dendritic cells and malignant tumor cells, that are refractory to infection by commonly used adenoviral vectors. Whereas many adenoviruses infect cells through the coxsackievirus and adenovirus receptor (CAR), group B adenoviruses use an alternate, as-yet-unidentified cellular attachment receptor. Using mass spectrometric analysis of proteins interacting with a group B fiber, we identified human CD46 as a cellular attachment receptor for most group B adenoviruses. We show that ectopic expression of human CD46 rendered nonhuman cells susceptible to infection with group B viruses in vitro and in vivo. In addition, both siRNA-mediated knockdown of CD46 and a soluble form of CD46 blocked infection of human cell lines and primary human cells. The discovery that group B adenoviruses use CD46, a ubiquitously expressed complement regulatory protein, as a cellular attachment receptor elucidates the diverse clinical manifestation of group B virus infections, and bears directly on the application of these vectors for gene therapy. [ABSTRACT FROM AUTHOR]

Most of the presently used adenovirus (Ad) vectors are based on serotype 5. However, the application of these vectors is limited by the native tropism of Ad5. To address this problem, a series of fiber chimeric vectors were produced to take advantage of the different cellular receptors used by Ad of different subgroups. In this study we utilize an Ad5-based chimeric vector containing sequences encoding the Ad35 fiber knob domain instead of the Ad5 knob (Ad5/35L) to analyze factors responsible for selection of intracellular trafficking routes by Ads. By competition analysis with recombinant Ad5 and Ad35 knobs we showed that the Ad5/35L vector infected cells through a receptor different from the Ad5 receptor. Intracellular trafficking of Ad5 and Ad5/35L viruses was analyzed in HeLa cells by tracking fluorophore-conjugated Ad particles, by immunostaining for capsid hexon protein, by electron microscopy, and by Southern blotting for viral DNA. These studies showed that the interaction with the Ad35 receptor(s) predestines Ad5/35L vector to intracellular trafficking pathways different from those of Ad5. Ad5 efficiently escaped from the endosomes early after infection. In contrast, Ad5/35L remained longer in late endosomal/lysosomal compartments and used them to achieve localization to the nucleus. However, a significant portion of Ad5/35L particles appeared to be recycled back to the cell surface. This phenomenon resulted in significantly less efficient Ad5/35L-mediated gene transfer compared to that of Ad5. We also demonstrated that the selection of intracellular trafficking routes was determined by the fiber knob domain and did not depend on the length of the fiber shaft. This study contributes to a better understanding of the mechanisms that govern the infection of retargeted, capsid-modified vectors which have potential application for hematopoietic stem cell and tumor gene therapy. [ABSTRACT FROM AUTHOR]

The construction and amplification of adenoviral (Ad) vectors expressing biologically active transgenes that are cytotoxic or inhibit Ad replication can be extremely difficult, if not impossible. In this study, we harnessed the ability of Ad genomes to undergo efficient homologous recombination to reconstitute the adeno-associated virus (AAV) rep78 gene, a cytotoxic gene that strongly inhibits Ad replication, which was divided between two parental, first-generation Ad vectors. A functional open reading frame was generated by recombination only upon co-infection of both parental vectors and after the onset of viral DNA replication. We were able to amplify both parental rep78 vectors to normal titers without any signs of inhibition or toxicity and could use them to generate progeny vectors containing a functional rep78 gene without any Ad genes. Using this vector recombination system in AAV rescue assays demonstrated that no Ad protein was essential for Rep78 mediated rescue of AAV ITR flanked DNA from plasmid or Ad backbones; the amount of rescue product generated was substantially greater in the presence of Ad infection; neither cellular nor viral DNA replication was necessary for rescue to occur; and progeny vector genomes were efficiently co-replicated along with conventional, first-generation Ad vectors. [Copyright &y& Elsevier]

In this study, we examined the ability of adenoviral (Ad) vectors to undergo homologous recombination. The lacZ gene was divided between two parental, first-generation vectors such that neither encoded a functional product but both shared 494 bp in common. The open reading frame could only be restored by homologous recombination. We observed β-galactosidase activity only upon co-infection of both parental vectors and after the onset of viral DNA replication, creating a delay in expression of 24–36 hours in HeLa cells. At peak efficiency, this recombination vector system resulted in β-galactosidase activity levels 100× above background and just 18× less than a conventional, first-generation vector in HeLa cells. After recombination, the resultant progeny vector genomes containing reconstituted expression cassettes were devoid of all viral genes and contained two packaging signals. These progeny genomes were efficiently packaged, could be separated from their parental vectors based on their lighter buoyant densities in CsCl gradients, and were subsequently used as functional gene transfer vectors. This novel recombination vector system should be useful for transferring large transgenes (because the carrying capacity of two Ad vectors can be exploited) or expressing any cytotoxic or Ad replication inhibitory protein (because the parental vectors exhibit no background expression). [Copyright &y& Elsevier]

An abstract of the article "Blood Factors Affect Adenovirus Infectivity and Bio-Distribution In Vivo," by Dmitry M. Shayakhmetov is presented.

An abstract of the article "Localization of Regions in CD46 That Interact with Adenovirus," by Anuj Gaggar, Dmitry M. Shayakhmetov, M. Kathryn Liszewski, John P. Atkinson, and Andre Lieber is presented.

Human adenovirus (Ad) is a ubiquitous pathogen causing a wide range of diseases. Although the interactions of human Ad serotype 5 (Ad5) with susceptible cells in vitro are known in great detail, host factors controlling the tissue specificity of Ad5 infection in vivo remain poorly understood. Here, we analyzed the mechanisms of sequestration by the liver for blood-born human Ads and Ad5-based vectors. Our data suggest that several known mechanisms that lead to Ad5 sequestration by the liver become engaged in a redundant, sequential, and synergistic manner to ensure the rapid clearance of circulating virus particles from the blood. These mechanisms include (i) trapping of the virus by liver residential macrophages, Kupffer cells; (ii) Ad5 hepatocyte infection via blood factor–hexon interactions; and (iii) Ad5 penton RGD motif–mediated interactions with liver endothelial cells and hepatocytes, mediating virus retention in the space of Disse. More important, we show that when all of these mechanisms are simultaneously inactivated via mutations of Ad5 capsid proteins and pharmacological interventions, virus sequestration by the liver is markedly reduced. Therefore, our study is the first demonstration of the principal possibility of ablating the sequestration of blood-born Ad in the liver via specific inactivation of a defined set of mechanisms that control this process.Molecular Therapy (2009) 17 4, 675–684 doi:10.1038/mt.2008.307 [ABSTRACT FROM AUTHOR]

The article discusses research being done on adenovirus-mediated evasion against host-derived natural antibodies. It references the study "Coagulation Factor X Shields Adenovirus Type 5 From Attack by Natural Antibodies and Complement," by Z. Xu et al. in the 2013 issue of "Nature Medicine." The researcher observed that host-derived coagulation factor X (FX) has linked to the adenovirus capsid and entered hepatocytes in mice without natural immunoglobulin M (IgM) antibodies.

Adeno-associated virus 2 (AAV2) and adenovirus 5 (Ad5) are promising gene therapy vectors. Both display liver tropism and are currently thought to enter hepatocytes in vivo through cell surface heparan sulfate proteoglycans (HSPGs). To test directly this hypothesis, we created mice that lack Ext1, an enzyme required for heparan sulfate biosynthesis, in hepatocytes. Ext1HEP mutant mice exhibit an 8-fold reduction of heparan sulfate in primary hepatocytes and a 5-fold reduction of heparan sulfate in whole liver tissue. Conditional hepatocyte Ext1 gene deletion greatly reduced AAV2 liver transduction following intravenous injection. Ad5 transduction requires blood coagulation factor X (FX); FX binds to the Ad5 capsid hexon protein and bridges the virus to HSPGs on the cell surface. Ad5.FX transduction was abrogated in primary hepatocytes from Ext1HEP mice. However, in contrast to the case with AAV2, Ad5 transduction was not significantly reduced in the livers of Ext1HEP mice. FX remained essential for Ad5 transduction in vivo in Ext1HEP mice. We conclude that while AAV2 requires HSPGs for entry into mouse hepatocytes, HSPGs are dispensable for Ad5 hepatocyte transduction in vivo. This study reopens the question of how adenovirus enters cells in vivo. [ABSTRACT FROM AUTHOR]

The molecular mechanisms governing the infectivity of adenovirus (Ad) toward specific cell and tissue types in vivo remain poorly understood. The direct Ad binding to hepatic heparan sulfate proteoglycans via the KKTK motif within the fiber shaft domain was suggested to be the major mechanism of Ad liver cell infection in vivo. Here, we describe the generation and in vitro and in vivo infectivity studies of Ad5-based vectors possessing long Ad31- or Ad41-derived fiber shaft domains, which lack the KKTK motif. We found that all the critical early steps of Ad infection, including attachment to the cellular receptor, internalization, and virus genome transfer into the nucleus, occurred with similar levels of efficiency for fiber shaft-chimeric vectors and unmodified Ad5. Upon intravenous delivery into mice, fiber shaft-chimeric vectors accumulated in liver tissue, transduced liver cells, and induced the production of proinflammatory cytokines (tumor necrosis factor alpha and interleukin-6) and the chemokine monocyte chemoattractant protein 1 at levels indistinguishable from those observed for Ad5. Thus, our data provide evidence that the Ad5 fiber shaft amino acid sequence does not play any substantial role in determining adenovirus infectivity toward hepatic cells in vivo. The data obtained contribute to improving our understanding of the molecular mechanisms determining Ad infectivity and biodistribution in vivo and may aid in designing novel Ad-based vectors for gene therapy applications. [ABSTRACT FROM AUTHOR]

Intravascular delivery of adenovirus (Ad) vectors is being developed for liver-directed gene therapy for targeting disseminated disease in cancer therapeutics and for targeting non-hepatic tissues and organs through vector engineering strategies. The utility of Ad vectors is not limited to serotype 5 (Ad5), and many alternate human serotypes and non-human serotypes of Ad are currently being investigated. Critical to intravascular delivery of Ad is the interaction of the virus with host blood cells and plasma proteins, because immediate contact is observed following injection. Although incompletely understood, recent studies suggest that these interactions are critical in dictating the particle bio-distribution and resulting transduction properties of Ad in vivo. For example, plasma proteins—in particular, vitamin K–dependent coagulation zymogens—are able to directly bind to Ad, and “bridge” the virus to receptors in the liver. Unraveling and characterizing these mechanisms will be of fundamental importance both for understanding basic Ad biology in vivo and for refinement and optimization of Ad vectors for human gene therapy.Molecular Therapy (2007) 15 8, 1410–1416. doi:10.1038/sj.mt.6300206 [ABSTRACT FROM AUTHOR]

Avoiding transduction of normal tissue after intravenous application of oncolytic adenoviruses (Ad) is an important strategy to improve the safety and efficacy of these vectors in gene therapy. As a model for a targeted vector, we used Ad vectors with type 35 fibers (Ad5/35), which efficiently transduce human cervical carcinoma cells but not liver cells. In an in vitro model of liver metastases, in which small nests of HeLa cells were surrounded by mouse hepatocytes, we showed that an Ad5/35-based conditionally replicating vector regulated by DNA replication-dependent recombination conferred increased gene transfer to tumor cells and enhanced viral replication and tumor cell lysis compared to the nontargeted Ad5 vector. Intravenous injection of Ad5/35 vectors into mice bearing liver metastases derived from HeLa cells caused markedly less hepatotoxicity than Ad5 vectors; however, it did not result in enhanced tumor cell transduction, viral replication, or oncolysis. Apparently, other factors, including the stability of virus in the blood, trapping within the liver sinusoids, transendothelial transfer, and/or vector diffusion of viral particles to tumor cells, limit tumor transduction, even if the vector is not taken up by liver cells. [Copyright &y& Elsevier]

An abstract of the article "Identifying Functional Adenovirus-Host Interactions Using Tandem Mass Spectrometry," by Anuj Gaggar, Martin Sadilek, Andre Lieber and Dmitry M. Shayakhmetov is presented.

An abstract of the article "Trapping of Adenovirus Vectors in Blood," by Daniel Stone, Shaoheng Ni, Zong-Yi Li, Dmitry M. Shayakhmetov and Andre Lieber is presented.