Biomedical subjects

K Binley

Publications and source records attributed to K Binley.

7 recordsLinked to original sources

PubMed2006

EIAV vector-mediated delivery of endostatin or angiostatin inhibits angiogenesis and vascular hyperpermeability in experimental CNV.

We evaluated the efficacy of equine infectious anaemia virus (EIAV)-based lentiviral vectors encoding endostatin (EIAV.endostatin) or angiostatin (EIAV.angiostatin) in inhibiting angiogenesis and vascular hyperpermeability in the laser-induced model of choroidal neovascularisation (CNV). Equine infectious anaemia virus.endostatin, EIAV.angiostatin or control (EIAV.null) vectors were administered into the subretinal space of C57Bl/6J mice. Two weeks after laser injury CNV areas and the degree of vascular hyperpermeability were measured by image analysis of in vivo fluorescein angiograms. Compared with EIAV.null-injected eyes, EIAV.endostatin resulted in a 59.5% (P<0.001) reduction in CNV area and a reduction in hyperpermeability of 25.6% (P<0.05). Equine infectious anaemia virus.angiostatin resulted in a 50.0% (P<0.05) reduction in CNV area and a 23.9% (P<0.05) reduction in hyperpermeability. Equine infectious anaemia virus.endostatin, but not EIAV.angiostatin significantly augmented the frequency of apoptosis within the induced CNV as compared with injected controls. TdT-dUTP terminal nick end labeling analysis 5 weeks post-injection, and histological and retinal flatmount analysis 12 months post-injection revealed no evidence of vector- or transgene expression-related deleterious effects on neurosensory retinal cells, or mature retinal vasculature in non-lasered eyes. Highly expressing EIAV-based vectors encoding endostatin or angiostatin effectively control angiogenesis and hyperpermeability in experimental CNV without long-term deleterious effects, supporting the use of such a strategy in the management of patients with exudative age-related macular degeneration.

Angiogenesis Inhibitors↗

PubMed2006

Stable and efficient intraocular gene transfer using pseudotyped EIAV lentiviral vectors.

BACKGROUND: We have developed minimal non-primate lentiviral vectors based on the equine infectious anaemia virus (EIAV). We evaluated the in vivo expression profiles of these vectors delivered regionally to ocular tissues to define their potential utility in ocular gene therapy. METHODS: EIAV vectors pseudotyped with VSV-G or rabies-G envelope proteins were delivered subretinally, intravitreally or into the anterior chambers (intracameral administration) in mice. Reporter gene (eGFP) expression was analysed using in vivo retinal imaging or histological examination of eyes and brains at intervals between 3 days and 16 months. We investigated the effects of vector titre, pseudotype, genome configuration, site of intraocular administration, intentional retinal trauma and the degree of retinal maturation on the spatial and temporal expression profiles of these vectors. RESULTS: Subretinal vector delivery resulted in efficient and stable transduction of retinal pigment epithelial (RPE) cells and variable transduction of photoreceptors up to 16 months post-injection. Retinal trauma facilitated the local transduction of neurosensory retinal cells. Intracameral administration of VSV-G- but not rabies-G-pseudotyped vectors produced stable eGFP expression in corneal endothelial cells and trabecular meshwork. CONCLUSIONS: The cellular tropism and expression kinetics of optimised EIAV vectors after intraocular administration make them attractive vehicles for delivering therapeutic genes in the management of inherited and acquired retinal and anterior segment disorders.

Animals↗

PubMed2003

Hypoxia-mediated tumour targeting.

Hypoxia is a common physiological feature of tumours. It activates a signalling cascade that culminates in the stabilization of the HIF-1 transcription factor and activation of genes that possess a hypoxia response element (HRE). We have used an optimized hypoxia responsive promoter (OBHRE) to investigate hypoxia-targeted gene expression in vivo in the context of an adenovirus vector. The OBHRE promoter showed limited activity in the liver or spleen such that expression was 1000-fold lower than that driven by the strong CMV/IE promoter. However, in the context of the tumour microenvironment, the OBHRE promoter achieved expression levels comparable to that of the CMV/IE promoter. Next, we showed that an adenovirus expressing the human cytochrome P450 (CYP2B6) regulated by the OBHRE promoter delays tumour growth in response to the prodrug cyclophosphamide (CPA). Finally, we exploited the hepatotropism of adenovirus to investigate whether the OBHRE promoter could mitigate the hepatotoxicity of a recombinant adenovirus expressing thymidine kinase (TK) in the context of the prodrug ganciclovir (GCV). High-dose Ad.CMVTK/GCV treatment caused significant liver necrosis whereas the same dose of Ad.HRETK was well tolerated. These in vivo data demonstrate that hypoxia-targeted gene expression via the OBHRE promoter can be used to increase the therapeutic window of cytotoxic cancer gene therapy.

Adenoviridae↗

PubMed2003

Hypoxia-regulated transgene expression in experimental retinal and choroidal neovascularization.

Recombinant AAV vectors mediate efficient and sustained transgene expression in retinal tissues and offer a powerful approach to the local, sustained delivery of angiostatic proteins for the treatment of ocular neovascular disorders. The application of such strategies may also require regulated gene expression to minimize the potential for unwanted adverse effects. In this study, we have evaluated the effect of a hypoxia-responsive element (HRE) on the kinetics of recombinant adeno-associated (rAAV)-mediated reporter gene expression in murine models of retinal and choroidal neovascularization. In murine ischaemia-induced retinal neovascularization, intravitreal delivery of rAAV.HRE.GFP results in reporter gene expression specifically at sites of vascular closure during the period of active neovascularization and not after vector delivery in normal controls. In murine laser-induced choroidal neovascularization, subretinal delivery of rAAV.HRE.GFP results in reporter gene expression at sites of active neovascularization but not elsewhere or after vector delivery in normal controls. HRE-driven gene expression offers an attractive strategy for the targeted and regulated delivery of angiostatic proteins to the retina in the management of neovascular disorders.

Animals↗

PubMed2000

The macrophage - a novel system to deliver gene therapy to pathological hypoxia.

The use of activated macrophages in the treatment of cancer has been largely ineffectual. By 'arming' these cells with the ability to express a therapeutic gene we demonstrate significant advances in the efficacy of this approach. We have used a hypoxia-regulated adenoviral vector to transduce human macrophages with either a reporter or a therapeutic gene encoding human cytochrome P4502B6 (CYP2B6). Infiltration of transduced macrophages into a tumour spheroid results in induction of gene expression. We demonstrate significant tumour cell killing only in the presence of cyclophosphamide via activation by P4502B6 and show that this can be further targeted to tumours through hypoxia regulated gene expression. Gene Therapy (2000) 7, 255-262.

Adenoviridae↗

PubMed1999

Characterization of physiologically regulated vectors for the treatment of ischemic disease.

A high therapeutic index is as important for gene-based therapies as it is for chemotherapy or radiotherapy. One approach has been transcriptional targeting through the use of tissue-specific regulatory elements. A more versatile approach would be to use a regulatory element that is controlled via a parameter common to a broad range of diseases. Ischemia is characteristic of a number of pathologies that range from vascular occlusion through to cancer. The state of low oxygen, hypoxia, triggers a transcriptional signaling pathway that is mediated by transcription factors binding to a specific enhancer, the hypoxia response element (HRE). These observations have therefore led to the use of HREs to drive gene expression in a number of target tissues from tumors to cardiac muscle. To translate these observations into a clinically useful vector system we have now assessed the potency of a number of naturally derived HREs in various configurations combined with minimal promoters. The optimal HRE has been introduced into a single transcription unit retroviral vector that can deliver regulated gene expression in response to hypoxia. An important feature of this new physiologically regulated vector is the combination of low basal expression and high-level activated expression that is on a par with that obtained with the cytomegalovirus immediate-early (CMV IE) promoter. The role of elements that stabilize mRNA in the presence of hypoxia has also been assessed. These hypoxia-regulated vectors may have utility for restricting the delivery of therapeutic proteins to tumors and ischemic sites.

Cell Line↗

PubMed1999

An adenoviral vector regulated by hypoxia for the treatment of ischaemic disease and cancer.

Recombinant adenoviral vectors have a number of advantages for gene therapy, including transduction of a range of dividing and non-dividing cell types. However, this broad range may be a disadvantage if non-target cells are transduced and are adversely affected by expression of the transferred gene. Here we describe a novel adenoviral vector in which transcription of the transgene is restricted to the patho-physiological condition of low oxygen tension (hypoxia). Hypoxia activates the expression of a number of genes, principally via the stabilisation of members of the bHLH/PAS family of transcription factors that bind to a con- sensus DNA sequence, the hypoxia response element (HRE). We have configured an optimised HRE expression cassette into an adenoviral vector, AdOBHRE. A range of cell types, including primary human skeletal muscle, when transduced with AdOBHRE display a low basal level of transgene expression that is highly induced in hypoxia to levels equivalent to that obtained from the CMV promoter. The AdOBHRE vector could be exploited for transcriptionally targeted gene therapy for the treatment of diseases such as cancer, peripheral arterial disease, arthritis and anaemia where tissue hypoxia is a cardinal feature.

Adenoviridae↗