Triple-Gene Therapy for Stroke: A Proof-of-Concept in Vivo Study in Rats

Abstract

Natural brain repair after stroke is extremely limited, and current therapeutic options are even more scarce with no clinical break-through in sight. Despite restricted regeneration in the central nervous system, we have previously proved that human umbilical cord blood mono-nuclear cells (UCB-MC) transduced with adenoviral vectors carrying genes encoding vascular endothelial growth factor (VEGF), glial cell-derived neurotrophic factor (GDNF), and neural cell adhesion molecule (NCAM) successfully rescued neurons in amyotrophic lateral sclerosis and spinal cord injury. This proof-of-principle project was aimed at evaluating the beneficial effects of the same triple-gene approach in stroke. Rats subjected to distal occlusion of the middle cerebral artery were treated intrathecally with a combination of these genes either directly or using our cell-based (UCB-MC) approach. Various techniques and markers were employed to evaluate brain injury and subsequent recovery after treatment. Brain repair was most prominent when therapeutic genes were delivered via adenoviral vector- or UCB-MC-mediated approach. Remodeling of brain cortex in the stroke area was confirmed by reduction of infarct volume and attenuated neural cell death, depletion of astrocytes and microglial cells, and increase in the number of oligodendroglial cells and synaptic proteins expression. These results imply that intrathecal injection of genetically engineered UCB-MC over-expressing therapeutic molecules (VEGF, GDNF, and NCAM) following cerebral blood vessel occlusion might represent a novel avenue for future research into treating stroke.

Preventive Triple Gene Therapy Reduces the Negative Consequences of Ischemia-Induced Brain Injury after Modelling Stroke in a Rat

“…An important factor is the absence of legal, ethical, and religious prohibitions related to blood cell transplantation. In our previous study, we showed that intrathecal injection of adenoviral vectors carrying vegf, gdnf, and ncam, or genetically-modified UCB-MC+Ad5-VEGF-GDNF-NCAM, 4 h after stroke modelling in rats, had a positive effect on the morpho-functional recovery of the post-ischemic brain [25]. Adenoviral vectors and genetically-modified UCB-MC with cerebrospinal fluid reached the ischemic area and delivered the production of recombinant VEGF, GDNF, and NCAM, lasting up to 21 days in the experiment.…”

“…The neuroprotective effect of these factors has been experimentally proven, but there is no unequivocal, let alone exhaustive, answer to the question of which of these factors may be recommended as neuroprotective factors in practical medicine. It has been established that combinations of several neurotrophic factors may have a more pronounced effect on nerve cell survival [25,28].…”

Genetic Aspects of Inflammation and Immune Response in Stroke

“…Results obtained by Sokolov et al imply that intrathecal injection of genetically engineered umbilical cord blood mononuclear cells (UCB-MC) over-expressing therapeutic molecules vascular endothelial growth factor (VEGF), glial cell line-derived neurotrophic factor (GDNF), neural cell adhesion molecule (NCAM) might represent a novel avenue for future research into treating stroke. Remodeling of the brain cortex in the stroke area was confirmed by the reduction of infarct volume and attenuated neural cell death, depletion of astrocytes and microglial cells, and an increase in the number of oligodendroglial cells and synaptic proteins expression [ 94 ].…”

Regenerative Medicine Therapies for Targeting Neuroinflammation After Stroke

“…The interactions between implanted hydrogel and endogenous brain cells have the potential to induce many different reparative and anti-inflammatory cellular pathways, through binding of CAPs (including RGD, IKVAV, and YISGR motifs) to specific cell surface receptors ( 101 ). Anti-inflammatory targets of CAPs include; cell adhesion molecules (CAMs), which are involved in the recruitment and trafficking of leukocytes ( 51 , 116 ); integrin receptors, which in addition to having anti-inflammatory effects can have proangiogenic properties ( 115 , 117 , 118 ), reduce reactive gliosis ( 43 , 119 ) and promote the infiltration of neural progenitor cells to the site of injury ( 43 , 111 ); and growth factor receptors that can initiate similar anti-inflammatory effects ( 120 ). CAPs can also be used to mimic growth factors and initiate preferential cellular pathways.…”

“…The use of gene therapies offers the potential to alter cellular and molecular processes that are important to recovery from ischemic stroke, reducing the inflammatory response and initiating regeneration of damaged tissue. This approach has been used to deliver anti-inflammatory gene therapies that promote production of VEGF ( 116 , 173 ), anti-inflammatory neural cell adhesion molecule (NCAM) ( 116 ), or IL-1Ra ( 50 ). These therapies were administered in rodents by intrathecal injection, but could be improved through encapsulation within a hydrogel for injection or implantation as this would control the release of gene-edited cells over time to increase the therapeutic effect.…”