Viral vector gene therapy approaches for regeneration and repair in spinal cord injury

List of abbreviations

SCI

spinal cord injury

CNS

central nervous system

AdV

adenovirus

HSV

herpes simplex virus

LV

lentivirus

AAV

adeno-associated virus

IM

intramuscular

IN

intraneural

IV

intravenous

IT

intrathecal

NGF

nerve growth factor

BDNF

brain-derived neurotrophic factor

NT

neurotrophin

BBB

blood-brain barrier

CSPG

chondroitin sulfate proteoglycan

ChABC

chondroitinase ABC

ADAMTS4

a disintegrin and metalloproteinase with thrombospondin motifs 4

Introduction

Traumatic spinal cord injury (SCI) results from a physical insult to the spinal cord. This leads to disruption of the blood supply and loss of tissue near the site of impact. Following this, there is ongoing detrimental change to the tissue, including inflammation, formation of an inhibitory scar, and inhibition of axonal regeneration and remyelination ( Fig. 1 ). A number of approaches have been tested in pre-clinical models including increasing the intrinsic regenerative ability of neurons or removing the inhibitory molecules present after injury. Approaches include the delivery of neurotrophic factors ( ), strategies to block inhibitory receptor signaling ( ) or using enzymes to degrade inhibitory molecules ( ). However, conventional methods of delivery of such factors have limitations, such as poor stability in vivo, poor tissue penetration, and the potential for off-target effects ( ). Viral vector gene therapy can be applied to a number of these approaches to help address these issues. This chapter describes different viral vectors that have been used to target the spinal cord, different delivery methods, and examples of how viral gene therapy has been used to effectively target therapeutic agents to the spinal cord.

Fig. 1, Overview of traumatic spinal cord injury mechanisms and potential targets for gene-based therapies. Mechanism : (A) Mechanical trauma impairs axonal function leading to synaptic dysfunction. (B) ECM deposition by reactive astrocytes at the lesion border and tissue cavitation significantly impairs axonal regrowth through the injury site. (C) Vascular hemorrhage and leakage disrupts nervous tissue homeostasis whilst a large infiltrating immune cell population responds aggressively to inflammatory signals resulting from tissue trauma. (D) Myelin debris resulting from oligodendrocyte death inhibits remyelination and axon regrowth. Overwhelming of lipid metabolism within phagocytes leads to prolonged inflammatory activation. Treatment targets : (E) Growth cone reformation is achieved by the re-establishment of anterograde axonal transport. Neurite extension can be stimulated by the presence of transgenic growth factor cues. (F) The digestion of extracellular matrix (ECM) molecules such as chondroitin sulfate proteoglycans (CSPGs) reduces the inhibition of axonal regrowth exerted by the astroglial scar. (G) The mechanisms of many gene therapy approaches have beneficial effects upon multiple injury processes. For example, stimulation of remyelination using current treatment approaches likely results from the direct effects of transgene products upon endogenous oligodendrocyte precursors (OPCs), i.e., neurotropic factors or removal of CSPGs. However, removal of CSPGs can also regulate phagocytic phenotype and function, which in turn promotes myelin repair. Further work should take consideration the interactions of these factors and how combinational viral gene therapy could be used to optimize this beneficial synergy.

Viral vector approaches

Viral vectors allow for long-term, stable, and targeted gene expression. A number of viral vectors have been trialed as a gene therapy approach to SCI repair. Each of these approaches has advantages and shortcomings ( Fig. 2 ).

Fig. 2, Overview of viral vector systems commonly used in the treatment and study of spinal cord injury. A number of viral vectors can be used for gene delivery to the spinal cord, each of which has advantages (+) and disadvantages (−).

Adenovirus

Adenoviruses (AdVs) are encapsulated double-stranded DNA viruses with a 36 kb genome and therefore have the capability of holding large transgenes. They can transduce both dividing and non-dividing cells and exhibit high transduction efficiency in a wide variety of cell types. The genome remains extra chromosomal in the nucleus, thereby the transgene is not integrated into the host cell’s genome, avoiding risk of tumorigenesis. However, transgene expression is only transient and expression is lost during division as the non-integrated genome is received by only one daughter cell. Also, an immune response is elicited against AdV viral antigens leading to inflammation and limiting the period of transgene expression ( ).

Poliovirus

Polioviruses are small, non-enveloped RNA viruses and recombinant poliovirus replicons can be used for gene therapy ( ). As infection is restricted to motor neurons in the hind brain and spinal cord these have been suggested as a potential vector for use in the spinal cord ( ). Advantages of poliovirus include that its RNA genome is replicated in the cytoplasm, leading to very high levels of gene expression ( ). Multiple inoculations of poliovirus replicons in the spinal cord has been shown to be safe, give good levels of gene expression with no functional deficits or inflammatory response. However, studies show that gene expression is not sustained ( ), although this could be of benefit if transient gene expression is required.

Herpes simplex virus

Herpes simplex virus (HSV) is an enveloped virus that contains a large (150 kb) double-stranded genome, allowing for an almost unlimited packaging capacity for transgenes. They demonstrate an exceptional capability to provide long-lived infections, with the genome being maintained extra-chromosomally in the nucleus of host cells ( ). Advantages of HSV are it is neurotrophic and has a tendency to remain latent while permanently transducing target cells. The large size of the vector allows for expression of multiple genes which has been suggested as advantageous in development of effective treatments ( ). However, following transduction a number of viral proteins are expressed, leading to cytotoxicity and immune responses against targeted cells ( ).

Lentivirus

Lentiviral vectors (LV) are a sub-class of retroviruses derived from human immunodeficiency viruses. Due to their ability to transduce dividing and non-dividing cells, LV vectors can transduce post-mitotic neurons, making them a powerful tool for targeting the CNS ( ) and as such have been used extensively for gene therapy targeting of the spinal cord ( ). Because LV genomes are integrated into the host genome they can provide long-term stable gene expression. While integration poses a risk of oncogenic transformation it has been suggested that this is reduced by the virus integrating into gene ends rather than promoter regions ( ).

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