Transcriptional targeting of tumors with a novel tumor-specific survivin promoter.

It has been demonstrated that survivin, a novel member of the inhibitor of apoptosis (IAP) protein family, is expressed in human cancers but is undetectable in normal differentiated tissues. We employed a recombinant adenoviral vector (reAdGL3BSurvivin) in which a tumor-specific survivin promoter and a luciferase reporter gene were inserted into the E1-deleted region of adenovirus vector. Luciferase activity was measured in both multiple tumor cell lines and two primary melanoma cells infected with reAdGL3BSurvivin.

Mode of transgene expression after fusion to early or late viral genes of a conditionally replicating adenovirus via an optimized internal ribosome entry site in vitro and in vivo.

The expression of therapeutic genes by oncolytic viruses is a promising strategy to improve viral oncolysis, to augment gene transfer compared with a nonreplicating adenoviral vector, or to combine virotherapy and gene therapy. Both the mode of transgene expression and the locale of transgene insertion into the virus genome critically determine the efficacy of this approach. We report here on the properties of oncolytic adenoviruses which contain the luciferase cDNA fused via an optimized internal ribosome entry site (IRES) to the immediate early adenoviral gene E1A (AdDeltaE1AIL), the early gene E2B (AdDeltaE2BIL), or the late fiber gene (AdDeltafiberIL).

Preclinical evaluation of a class of infectivity-enhanced adenoviral vectors in ovarian cancer gene therapy.

Ovarian carcinoma cells are often infected inefficiently by adenoviruses (Ad) due to low expression of coxsackie-adenovirus receptors (CAR), hindering the application of adenovirus-mediated gene therapy in ovarian cancer. In this study, we explored a class of infectivity-enhanced Ad vectors, which contain CAR-independent targeting motifs RGD (Ad5.RGD), polylysine (Ad5.

Adenovirus-mediated gene delivery to dendritic cells.

Dendritic cells (DCs) are "professional" antigen-presenting cells (APCs) that are uniquely capable of activating and instructing a naive immune system to mount a specific cellular and humoral response. Recognition of this crucial function makes the development of technologies for DC-based immuno-therapies a priority for the treatment of a wide variety of diseases. The most immediate impact of this emerging technology will be in the treatment of cancer and the development of third generation vaccines to protect against viral and intracellular pathogens.

Adenovirus-mediated gene delivery to skeletal muscle.

Adenoviral vectors can be employed for gene delivery to skeletal muscle, both ex vivo and in vivo. Although the realization of the full potential of adenoviral vectors awaits the development of methods to allow safe and efficient targeted gene delivery to mature skeletal muscle upon intravenous vector administration (1), the current generation of vectors has nonetheless found utility in preclinical studies of gene therapy and in gene-transfer experiments designed to study muscle biology. Features of adenoviral vectors that have favored their use for gene delivery to skeletal muscle include the ability to infect both actively dividing and terminally differentiated cells, as well as their large insert capacity.

Adenovirus-mediated gene delivery: an overview.

Adenoviruses, which were first isolated in the 1950s, have been developed as gene-delivery vehicles, or vectors, since the early 1980s. The adenoviruses constitute the Adenoviridae family, which is divided into two genera: the Aviadenovirus genus infects only birds, whereas the Mastadenovirus genus contains viruses that infect a range of mammalian species. Human adenoviruses are classified into six subgroups based on the percentage of guanine and cytosine in the DNA molecules and the ability to agglutinate red blood cells.

Retargeting of adenoviral infection to melanoma: combining genetic ablation of native tropism with a recombinant bispecific single-chain diabody (scDb) adapter that binds to fiber knob and HMWMAA.

Gene therapy is an emerging and promising modality for the treatment of malignant melanoma and other neoplasms for which conventional therapies are inadequate. Various therapeutic genes have shown promise for tumor cell killing. However, successful gene therapy depends on the development of efficient and targeted gene transfer vectors.

Engineering regulatory elements for conditionally-replicative adeno-viruses.

Virus-mediated oncolysis is a rapidly growing field with the potential to dramatically alter the future of cancer therapy. Replication-selective viruses are superior to non-replicating vectors in several aspects, such as the amplification of the initial low-dose viral inoculum up to 10(3)-10(3)-fold, lateralization into neighboring cells, introduction of novel cell killing mechanisms, and a potential for a safe profile. However, due to their capacity to replicate, the importance of tumor selectivity is further underscored.

Intravenous delivery of adenovirus-mediated soluble FLT-1 results in liver toxicity.

Purpose: Vascular endothelial growth factor (VEGF) is a potent angiogenic agent and plays a major role in tumor growth and metastases. We have previously reported the locoregional (i.p.

Development of a therapeutic adenoviral vector for cholangiocarcinoma combining tumor-restricted gene expression and infectivity enhancement.

Cholangiocarcinoma is an invasive malignancy that is most often unresectable upon diagnosis and unresponsive to chemotherapy and radiation. While adenoviral gene therapy has shown promise in treating many tumors, systemic toxicity and low tumor transduction efficiency have hampered its application in many gastrointestinal cancers. To overcome these difficulties, we have constructed an adenoviral vector utilizing a tumor-specific promoter (TSP) for selective transgene expression and a vector with an RGD-motif in the fiber-knob region for infectivity enhancement.

Cancer gene therapy.

Cancer gene therapy is the transfer of genetic material to the cells of an individual with the goal of eradicating cancer cells, both in the primary tumor and metastases. Cancer gene therapy strategies exploit our expanding knowledge of the genetic basis of cancer, thereby allowing rationally targeted interventions at the molecular level. The successful implementation of cancer gene therapy in the clinic awaits the development of vectors capable of specific and efficient gene delivery to cancer cells.

Generation of an adenoviral vector containing an addition of a heterologous ligand to the serotype 3 fiber knob.

As an initial assessment of the feasibility of employing the adenovirus serotype 3 (Ad3) fiber knob as a locale for introducing a tropism-modifying motif, we generated an adenoviral vector containing a six-histidine tag genetically fused to the carboxy-terminus of the Ad3 fiber knob. The heterologous tag proved to be accessible for binding in the context of the virion and, moreover, had rendered the modified vector capable of mediating gene transfer through an artificial, non-Ad3 receptor.

Transcription initiation activity of adenovirus left-end sequence in adenovirus vectors with e1 deleted.

We analyzed the transcription initiation activity of the left-end sequence (first 342 bp) of the adenovirus genome in the context of an adenovirus vector with E1 deleted in in vitro and in vivo gene transfer models. While nucleotide sequences 1 to 190 and 1 to 342 showed strong activity in three out of three lung cancer cell lines, nucleotide sequence 1 to 103 showed limited activity in H358, cells which show characteristics of type 2 alveolar cells. In vivo, the transcription initiation activities of nucleotide sequence 1 to 103 in the liver and the lung were minimal, while nucleotide sequences 1 to 190 and 1 to 342 showed strong activity comparable to that of the cytomegalovirus promoter.

Dual targeting of adenoviral vectors at the levels of transduction and transcription enhances the specificity of gene expression in cancer cells.

Adenoviral (Ad) vector-mediated strategies for cancer gene therapy mandate a vector that is capable of efficient expression of the therapeutic gene specifically within the target tumor cells. In one approach to the development of cancer cell-specific vectors, Ad vectors have been targeted at the level of transduction to achieve the selective delivery of the therapeutic gene. In an alternative approach to the derivation of cancer cell-specific vectors, Ad vectors have been targeted at the level of transcription by placing the therapeutic gene under the control of transcriptional regulatory sequences that are activated in tumor cells, but not in normal cells, and therefore target expression selectively to the tumor cell.

Conditionally replicative adenoviral vectors for cancer gene therapy.

During the past century, many attempts have been made to exploit the ability of some viruses to infect and destroy cancer cells. Crippled, non-replicative viruses have been used as vectors to transfer genes into tumours. Both strategies have serious limitations.

From molecular changes to customised therapy.

The revolution in molecular methods has allowed the development of approaches whereby cancer-specific changes can be utilised for targeted therapies. Gene therapy strategies include mutation compensation for correction of cancer-associated defects, and molecular chemotherapy for delivering toxic substances locally to tumour cells. Viruses which replicate only in tumour cells represent a powerful novel approach undergoing intensive development, with some exciting clinical results.

Targeting adenovirus to the serotype 3 receptor increases gene transfer efficiency to ovarian cancer cells.

Gene delivery efficiency in clinical cancer gene therapy trials with recombinant adenoviruses (Ads) based on serotype 5 (Ad5) has been limited partly because of variable expression of the primary Ad5 receptor, the coxsackie and adenovirus receptor (CAR), on human primary cancer cells. As a means of circumventing CAR deficiency, Ad vectors have been retargeted by creating chimeric fibers possessing knob domains of alternate Ad serotypes. In this study, we have constructed an Ad5-based vector, Ad5/3luc1, with a chimeric fiber protein featuring a knob domain derived from Ad3.

An adenovirus with enhanced infectivity mediates molecular chemotherapy of ovarian cancer cells and allows imaging of gene expression.

The adenovirus (Ad) is a useful vector for cancer gene therapy due to its unparalleled gene transfer efficiency to dividing and quiescent cells. Primary cancer cells, however, often have highly variable or low levels of the requisite coxsackie-adenovirus receptor (CAR). Also, assessment of gene transfer and vector persistence has been logistically difficult in human clinical trials.

Adenovirus-mediated soluble FLT-1 gene therapy for ovarian carcinoma.

Purpose: We hypothesized that adenovirus-mediated soluble fms-like tyrosine kinase receptor (sFLT-1) gene therapy can inhibit the ovarian tumor growth and increase survival of mice in the context of ovarian carcinoma.

Experimental Design: We constructed an infectivity-enhanced recombinant adenovirus (AdRGDGFPsFLT-1) expressing soluble FLT-1 and green fluorescent protein (GFP). An adenovirus AdRGDGFP expressing GFP alone was used as control.

Genetic targeting of an adenovirus vector via replacement of the fiber protein with the phage T4 fibritin.

The utility of adenovirus (Ad) vectors for gene therapy is restricted by their inability to selectively transduce disease-affected tissues. This limitation may be overcome by the derivation of vectors capable of interacting with receptors specifically expressed in the target tissue. Previous attempts to alter Ad tropism by genetic modification of the Ad fiber have had limited success due to structural conflicts between the fiber and the targeting ligand.

Insertional mutagenesis of AAV2 capsid and the production of recombinant virus.

The structural genes of adeno-associated virus serotype 2 (AAV2) have been altered by linker insertional mutagenesis in order to define critical components of virion assembly and infectivity. An in-frame restriction site linker was inserted across the capsid coding domain of a recombinant plasmid. After complementation in vivo, recombinant AAV2 viruses were generated and assayed for capsid production, packaging, transduction, heparin agarose binding, and morphology.