Pre-clinical delivery of gene therapy products to the cerebrospinal fluid: challenges and considerations for clinical translation

While the majority of gene therapy studies in neurological indications have focused on direct gene transfer to the central nervous system (CNS), there is growing interest in the delivery of therapeutics using the cerebrospinal fluid (CSF) as a conduit. Historically, direct CNS routes-of-administration (RoAs) have relied on tissue dynamics, displacement of interstitial fluid, and regional specificity to achieve focal delivery into regions of interest, such as the brain. While intraparenchymal delivery minimizes peripheral organ exposure, one perceived drawback is the relative invasiveness of this approach to drug delivery. In this mini review, we examine the CSF as an alternative RoA to target CNS tissue and discuss considerations associated with the safety of performing such procedures, biodistribution of therapeutics following single administration, and translation of findings given differences between small and large animals. These factors will help delineate key considerations for translating data obtained from animal studies into clinical settings that may be useful in the treatment of neurological conditions

Compressive Mechanical Characterization of Non-Human Primate Spinal Cord White Matter

The goal of developing computational models of spinal cord injury (SCI) is to better understand the human injury condition. However, finite element models of human SCI have used rodent spinal cord tissue properties due to a lack of experimental data. Central nervous system tissues in non human primates (NHP) closely resemble that of humans and therefore, it is expected that material constitutive models obtained from NHPs will increase the fidelity and the accuracy of human SCI models. Human SCI most often results from compressive loading and spinal cord white matter properties affect FE predicted patterns of injury; therefore, the objectives of this study were to characterize the unconfined compressive response of NHP spinal cord white matter and present an experimentally derived, finite element tractable constitutive model for the tissue. Cervical spinal cords were harvested from nine male adult NHPs (Macaca mulatta). White matter biopsy samples (3 mm in diameter) were taken from both lateral columns of the spinal cord and were divided into four strain rate groups for unconfined dynamic compression and stress relaxation (post-mortem <1-hour). The NHP spinal cord white matter compressive response was sensitive to strain rate and showed substantial stress relaxation confirming the viscoelastic behavior of the material. An Ogden 1st order model best captured the non-linear behavior of NHP white matter in a quasi-linear viscoelastic material model with 4-term Prony series. This study is the first to characterize NHP spinal cord white matter at high (>10/sec) strain rates typical of traumatic injury. The finite element derived material constitutive model of this study will increase the fidelity of SCI computational models and provide important insights for transferring pre-clinical findings to clinical treatments

A novel inhibitor of p75-neurotrophin receptor improves functional outcomes in two models of traumatic brain injury.

The p75 neurotrophin receptor is important in multiple physiological actions including neuronal survival and neurite outgrowth during development, and after central nervous system injury. We have discovered a novel piperazine-derived compound, EVT901, which interferes with p75 neurotrophin receptor oligomerization through direct interaction with the first cysteine-rich domain of the extracellular region. Using ligand binding assays with cysteine-rich domains-fused p75 neurotrophin receptor, we confirmed that EVT901 interferes with oligomerization of full-length p75 neurotrophin receptor in a dose-dependent manner. Here we report that EVT901 reduces binding of pro-nerve growth factor to p75 neurotrophin receptor, blocks pro-nerve growth factor induced apoptosis in cells expressing p75 neurotrophin receptor, and enhances neurite outgrowth in vitro Furthermore, we demonstrate that EVT901 abrogates p75 neurotrophin receptor signalling by other ligands, such as prion peptide and amyloid-β. To test the efficacy of EVT901 in vivo, we evaluated the outcome in two models of traumatic brain injury. We generated controlled cortical impacts in adult rats. Using unbiased stereological analysis, we found that EVT901 delivered intravenously daily for 1 week after injury, reduced lesion size, protected cortical neurons and oligodendrocytes, and had a positive effect on neurological function. After lateral fluid percussion injury in adult rats, oral treatment with EVT901 reduced neuronal death in the hippocampus and thalamus, reduced long-term cognitive deficits, and reduced the occurrence of post-traumatic seizure activity. Together, these studies provide a new reagent for altering p75 neurotrophin receptor actions after injury and suggest that EVT901 may be useful in treatment of central nervous system trauma and other neurological disorders where p75 neurotrophin receptor signalling is affected

Macrophage presence is essential for the regeneration of ascending afferent fibres following a conditioning sciatic nerve lesion in adult rats

Abstract Background Injury to the peripheral branch of dorsal root ganglia (DRG) neurons prior to injury to the central nervous system (CNS) DRG branch results in the regeneration of the central branch. The exact mechanism mediating this regenerative trigger is not fully understood. It has been proposed that following peripheral injury, the intraganglionic inflammatory response by macrophage cells plays an important role in the pre-conditioning of injured CNS neurons to regenerate. In this study, we investigated whether the presence of macrophage cells is crucial for this type of regeneration to occur. We used a clodronate liposome technique to selectively and temporarily deplete these cells during the conditioning phase of DRG neurons. Results Retrograde and anterograde tracing results indicated that in macrophage-depleted animals, the regenerative trigger characteristic of pre-conditioned DRG neurons was abolished as compared to injury matched-control animals. In addition, depletion of macrophage cells led to: (i) a reduction in macrophage infiltration into the CNS compartment even after cellular repopulation, (ii) astrocyte up-regulation at rostral regions and down-regulation in brain derived neurotrophic factor (BDNF) concentration in the serum. Conclusion Activation of macrophage cells in response to the peripheral nerve injury is essential for the enhanced regeneration of ascending sensory neurons.</p

Distribution of nanoparticles throughout the cerebral cortex of rodents and non-human primates: Implications for gene and drug therapy

When nanoparticles/proteins are infused into the brain, they are often transported to distal sites in a manner that is dependent both on the characteristics of the infusate and the region targeted. We have previously shown that adeno-associated virus (AAV

Correlating Tissue Mechanics and Spinal Cord Injury: Patient-Specific Finite Element Models of Unilateral Cervical Contusion Spinal Cord Injury in Non-Human Primates

Non-human primate (NHP) models are the closest approximation of human spinal cord injury (SCI) available for pre-clinical trials. The NHP models, however, include broader morphological variability that can confound experimental outcomes. We developed subject-specific finite element (FE) models to quantify the relationship between impact mechanics and SCI, including the correlations between FE outcomes and tissue damage. Subject-specific models of cervical unilateral contusion SCI were generated from pre-injury MRIs of six NHPs. Stress and strain outcomes were compared with lesion histology using logit analysis. A parallel generic model was constructed to compare the outcomes of subject-specific and generic models. The FE outcomes were correlated more strongly with gray matter damage (0.29 &lt; R2 &lt; 0.76) than white matter (0.18 &lt; R2 &lt; 0.58). Maximum/minimum principal strain, Von-Mises and Tresca stresses showed the strongest correlations (0.31 &lt; R2 &lt; 0.76) with tissue damage in the gray matter while minimum principal strain, Von-Mises stress, and Tresca stress best predicted white matter damage (0.23 &lt; R2 &lt; 0.58). Tissue damage thresholds varied for each subject. The generic FE model captured the impact biomechanics in two of the four models; however, the correlations between FE outcomes and tissue damage were weaker than the subject-specific models (gray matter [0.25 &lt; R2 &lt; 0.69] and white matter [R2 &lt; 0.06] except for one subject [0.26 &lt; R2 &lt; 0.48]). The FE mechanical outputs correlated with tissue damage in spinal cord white and gray matters, and the subject-specific models accurately mimicked the biomechanics of NHP cervical contusion impacts

Mechanical Design and Analysis of a Unilateral Cervical Spinal Cord Contusion Injury Model in Non-Human Primates.

Non-human primate (NHP) models of spinal cord injury better reflect human injury and provide a better foundation to evaluate potential treatments and functional outcomes. We combined finite element (FE) and surrogate models with impact data derived from in vivo experiments to define the impact mechanics needed to generate a moderate severity unilateral cervical contusion injury in NHPs (Macaca mulatta). Three independent variables (impactor displacement, alignment, and pre-load) were examined to determine their effects on tissue level stresses and strains. Mechanical measures of peak force, peak displacement, peak energy, and tissue stiffness were analyzed as potential determinants of injury severity. Data generated from FE simulations predicted a lateral shift of the spinal cord at high levels of compression (&gt;64%) during impact. Submillimeter changes in mediolateral impactor position over the midline increased peak impact forces (&gt;50%). Surrogate cords established a 0.5 N pre-load protocol for positioning the impactor tip onto the dural surface to define a consistent dorsoventral baseline position before impact, which corresponded with cerebrospinal fluid displacement and entrapment of the spinal cord against the vertebral canal. Based on our simulations, impactor alignment and pre-load were strong contributors to the variable mechanical and functional outcomes observed in in vivo experiments. Peak displacement of 4 mm after a 0.5N pre-load aligned 0.5-1.0 mm over the midline should result in a moderate severity injury; however, the observed peak force and calculated peak energy and tissue stiffness are required to properly characterize the severity and variability of in vivo NHP contusion injuries

Strong Cortical and Spinal Cord Transduction After AAV7 and AAV9 Delivery into the Cerebrospinal Fluid of Nonhuman Primates

The present study builds on previous work showing that infusion of adeno-associated virus type 9 (AAV9) into the cisterna magna (CM) of nonhuman primates resulted in widespread transduction throughout cortex and spinal cord. Transduction efficiency was severely limited, however, by the presence of circulating anti-AAV antibodies. Accordingly, we compared AAV9 to a related serotype, AAV7, which has a high capsid homology. CM infusion of either AAV7 or AAV9 directed high level of cell transduction with similar patterns of distribution throughout brain cortex and along the spinal cord. Dorsal root ganglia and corticospinal tracts were also transduced. Both astrocytes and neurons were transduced. Interestingly, little transduction was observed in peripheral organs. Our results indicate that intrathecal delivery of either AAV7 or AAV9 directs a robust and widespread cellular transduction in the central nervous system and other peripheral neural structures