Hypoxia-regulated therapeutic gene as a preemptive treatment strategy against ischemia/reperfusion tissue injury

Abstract

Ischemia and reperfusion represent major mechanisms of tissue injury and organ failure. The timing of administration and the duration of action limit current treatment approaches using pharmacological agents. In this study, we have successfully developed a preemptive strategy for tissue protection using an adenoassociated vector system containing erythropoietin hypoxia response elements for ischemia-regulated expression of the therapeutic gene human heme-oxygenase-1 (hHO-1). We demonstrate that a single administration of this vector several weeks in advance of ischemia͞reperfusion injury to multiple tissues such as heart, liver, and skeletal muscle yields rapid and timely induction of hHO-1 during ischemia that resulted in dramatic reduction in tissue damage. In addition, overexpression of therapeutic transgene prevented long-term pathological tissue remodeling and normalized tissue function. Application of this regulatable system using an endogenous physiological stimulus for expression of a therapeutic gene may be a feasible strategy for protecting tissues at risk of ischemia͞reperfusion injury.

Oxidative stress-inducible lentiviral vectors for gene therapy

“…This induction is of the same magnitude as the B8-fold hypoxic induction in the luciferase activity of the AAV construct driven by four tandem repeats of the erythropoietin HRE element subsequently shown in vivo to provide significant protection against I/R injury when driving the HO-1 transgene. 10 Likewise, HRE-regulated AAV-vector having nine copies of erythropoietin HRE elements yielded similar magnitude of induction of LacZ marker gene in vitro (10-to 30-fold depending on the cell type) in hypoxic conditions. 38 This vector was also shown to effectively increase the expression of vascular endothelial growth factor-A (VEGF-A) transgene in hypoxic mouse myocardium, despite significant induction of the endogenous mouse VEGF-A.…”

“…3,26 Regulatable vectors offer a solution to this problem, especially if they can be regulated by endogenous stimuli relevant to the pathology. Hypoxiaactivated adeno-associated virus (AAV) vector-expressing HO-1 has been developed and successfully used for treatment of cardiac I/R injury, 10 and HO-1 gene transfer also offers protection against chronic recurrent I/R injury and prevents post-infarction heart failure. 10,27,28 AREregulated gene transcription has been shown to be a central mechanism providing protection against oxidative stress in the cardiovascular system.…”

“…5 Nevertheless, HO-1 has been used for gene therapy in several experimental animal models of CVD. 10,11,27,28 In the study by Pachori et al 10 Oxidative stress-inducible vectors H Hurttila et al significant induction of endogenous HO-1 was observed, yet HRE-regulated HO-1 overexpression provided an additional benefit. In our study, both in controls as well as 2 Â GCLM-HO-1-transduced HUVECs, 15d-PGJ 2 significantly attenuated TNF-a-induced NF-kB activation and VCAM-1 expression, but the effect was greater in HO-1-transduced cells (Figure 6).…”

“…Hypoxiaregulated gene therapy vectors using tandem repeats of hypoxia response elements (HREs), enhancer regions in the promoters of genes activated during hypoxia, have been developed and used to regulate expression of the HO-1 transgene in vectors used for cardioprotection against I/R injury. 10,11 By analogy, vectors that would physiologically respond to oxidative stress could be useful for gene therapy in pathologies in which ROS are important.…”

Heme-oxygenase-1-induced protection against hypoxia/reoxygenation is dependent on biliverdin reductase and its interaction with PI3K/Akt pathway

“…Heme oxygenase-1 (HO-1) is an inducible protein whose expression is increased several fold in response to a variety of cellular stresses and stimuli including ischemia [3] hypoxia [4], oxidative stress [5] and inflammatory cytokines [6], suggesting an important role for this enzyme in tissue protection. Several studies have shown that HO-1 over-expression protects against ischemia/reperfusion injury in a variety of tissues including the myocardium [7][8][9][10][11]. Conversely, reduced HO-1 levels increase susceptibility to injury in a variety of stress conditions.…”