Virus vector-mediated genetic modification of brain tumor stromal cells after intravenous delivery

Abstract

The malignant primary brain tumor, glioblastoma (GBM) is generally incurable. New approaches are desperately needed. Adeno-associated virus (AAV) vector-mediated delivery of anti-tumor transgenes is a promising strategy, however direct injection leads to focal transgene spread in tumor and rapid tumor division dilutes out the extra-chromosomal AAV genome, limiting duration of transgene expression. Intravenous (IV) injection gives widespread distribution of AAV in normal brain, however poor transgene expression in tumor, and high expression in non-target cells which may lead to ineffective therapy and high toxicity, respectively. Delivery of transgenes encoding secreted, anti-tumor proteins to tumor stromal cells may provide a more stable and localized reservoir of therapy as they are more differentiated than fast-dividing tumor cells. Reactive astrocytes and tumor-associated macrophage/microglia (TAMs) are stromal cells that comprise a large portion of the tumor mass and are associated with tumorigenesis. In mouse models of GBM, we used IV delivery of exosome-associated AAV vectors driving green fluorescent protein expression by specific promoters (NF-κB-responsive promoter and a truncated glial fibrillary acidic protein promoter), to obtain targeted transduction of TAMs and reactive astrocytes, respectively, while avoiding transgene expression in the periphery. We used our approach to express the potent, yet toxic anti-tumor cytokine, interferon beta, in tumor stroma of a mouse model of GBM, and achieved a modest, yet significant enhancement in survival compared to controls. Noninvasive genetic modification of tumor microenvironment represents a promising approach for therapy against cancers. Additionally, the vectors described here may facilitate basic research in the study of tumor stromal cells in situ.

PSMB8 inhibition decreases tumor angiogenesis in glioblastoma through vascular endothelial growth factor A reduction

“…TMZ is the first‐line chemotherapeutic drug for GBM treatment, but the human glioblastoma cell line LN229 is resistant to TMZ 6,12,13 . IFN‐β increases sensitivity to TMZ and has been shown to improve the therapeutic outcome in GBM patients 14 . In our previous study, the knockdown of PSMB4 expression enhanced the antitumor effect of TMZ on tumor growth in an orthotropic GBM xenograft mouse model 13 .…”

Utilizing minimally purified secreted rAAV for rapid and cost-effective manipulation of gene expression in the CNS

“…One possible explanation is that secreted viral particles may be inherently different than those found intracellularly. Several recent papers have identified secreted rAAV particles inside vesicular bodies such as exosomes [40][41][42][43][44][45][46][47]. The secreted rAAVs utilized in our study may be associated with exosomes and this association could potentially alter the way in which they interact with the target cell.…”

Engineering of Exosomes to Target Cancer Metastasis

“…Viral exosome delivery was used in the treatment of the GL261 mouse glioblastoma (GBM) model to determine its efficacy as a potential treatment for GBM. 117 As described previously, exo-AAV were made by transfecting human 293T cells with an AAV plasmid for 48 h. 71 The role of the vectors was to genetically modify the target GBM cells, causing GFP expression to specifically target Tumor-Associated Macrophages/Microglia (TAMs) and reactive astrocytes. The resulting pathway expressed interferon beta, a cytokine destructive to the brain tumor stromal cells, and was found to increase survival when compared to controls.…”

“…The resulting pathway expressed interferon beta, a cytokine destructive to the brain tumor stromal cells, and was found to increase survival when compared to controls. 117 The ability of exo-AAV to evade neutralizing antibodies is also beneficial in that it allows for reduced dosing for gene therapy applications. 46…”