Kinetics and MR-Based Monitoring of AAV9 Vector Delivery into Cerebrospinal Fluid of Nonhuman Primates.

Here we evaluated the utility of MRI to monitor intrathecal infusions in nonhuman primates. Adeno-associated virus (AAV) spiked with gadoteridol, a gadolinium-based MRI contrast agent, enabled real-time visualization of infusions delivered either via cerebromedullary cistern, lumbar, cerebromedullary and lumbar, or intracerebroventricular infusion. The kinetics of vector clearance from the cerebrospinal fluid (CSF) were analyzed. Our results highlight the value of MRI in optimizing the delivery of infusate into CSF. In particular, MRI revealed differential patterns of infusate distribution depending on the route of delivery. Gadoteridol coverage analysis showed that cerebellomedullary cistern delivery was a reliable and effective route of injection, achieving broad infusate distribution in the brain and spinal cord, and was even greater when combined with lumbar injection. In contrast, intracerebroventricular injection resulted in strong cortical coverage but little spinal distribution. Lumbar injection alone led to the distribution of MRI contrast agent mainly in the spinal cord with little cortical coverage, but this delivery route was unreliable. Similarly, vector clearance analysis showed differences between different routes of delivery. Overall, our data support the value of monitoring CSF injections to dissect different patterns of gadoteridol distribution based on the route of intrathecal administration.

Targeted Delivery and Tolerability of MRI-Guided CED Infusion into the Cerebellum of Nonhuman Primates.

This study explored the feasibility of intraparenchymal delivery (gadoteridol and/or Serotype 5 Adeno-Associated Viral Vector-enhanced Green Fluorescent Protein [AAV5-eGFP]) into the cerebellum of nonhuman primates using real-time magnetic resonance imaging-guided convection enhanced delivery (MRI-CED) technology. All animals tolerated the neurosurgical procedure without any clinical sequela. Gene expression was detected within the cerebellar parenchyma at the site of infusion and resulted in transduction of neuronal cell bodies and fibers. Histopathology indicated localized damage along the stem of the cannula tract. These findings demonstrate the potential of real-time MRI-CED to deliver therapeutics into the cerebellum, which has extensive reciprocal connections and may be used as a target for the treatment of neurological disorders.

Intermittent convection-enhanced delivery of GDNF into rhesus monkey putamen: absence of local or cerebellar toxicity.

Glial cell line-derived neurotrophic factor (GDNF) has demonstrated neurorestorative and neuroprotective effects in rodent and nonhuman primate models of Parkinson's disease. However, continuous intraputamenal infusion of GDNF (100 µg/day) resulted in multifocal cerebellar Purkinje cell loss in a 6-month toxicity study in rhesus monkeys. It was hypothesized that continuous leakage of GDNF into the cerebrospinal fluid compartment during the infusions led to down-regulation of GDNF receptors on Purkinje cells, and that subsequent acute withdrawal of GDNF then mediated the observed cerebellar lesions. Here we present the results of a 9-month toxicity study in which rhesus monkeys received intermittent intraputamenal infusions via convection-enhanced delivery. Animals were treated with GDNF (87.1 µg; N = 14) or vehicle (N = 6) once every 4 weeks for a total of 40 weeks (11 treatments). Four of the GDNF-treated animals were utilized in a satellite study assessing the impact of concomitant catheter repositioning prior to treatment. In the main study, eight animals (5 GDNF, 3 control) were euthanized at the end of the treatment period, along with the four satellite study animals, while the remaining eight animals (5 GDNF, 3 control) were euthanized at the end of a 12-week recovery period. There were no GDNF-related adverse effects and in particular, no GDNF-related microscopic findings in the brain, spinal cord, dorsal root ganglia, or trigeminal ganglia. Therefore, 87.1 µg/4 weeks is considered the no observed adverse effect level for GDNF in rhesus monkeys receiving intermittent, convection-enhanced delivery of GDNF for 9 months.

Plasma and CSF oxytocin levels after intranasal and intravenous oxytocin in awake macaques.

Oxytocin (OT) is a neuropeptide that mediates a variety of complex social behaviors in animals and humans. Intranasal OT has been used as an experimental therapeutic for human conditions characterized by deficits in social functioning, especially autism spectrum disorder and schizophrenia. However, it is currently under intense debate whether intranasal delivery of OT reaches the central nervous system. In this study, four female rhesus macaques were implanted with chronic intrathecal catheters and used to investigate the pharmacokinetic profile of OT in the central nervous system and the peripheral vasculature following intravenous (IV) and intranasal (IN) administration of OT. In a randomized, crossover design, OT was given to four awake monkeys at three different doses based on body weight (0.1 IU/kg; 1 IU/kg; 5 IU/kg). A time course of concurrent cerebrospinal fluid (CSF) and plasma samples were taken following administration. We found a dose-dependent effect of IV OT treatment on plasma OT levels, which peaked at 5 min post-dose and gradually returned to baseline by 120 min. In contrast, a change in CSF OT was only observed at the highest IV dose (5 IU/kg) at 15 min post-dose and gradually returned to baseline by 120 min. After IN administration, there was no significant change in plasma OT at any of the three doses. However, at the highest dose level, we found a significant increase in CSF OT at 15-30 min post- dose. The results of this study in light of recent, similar publications highlight the importance of methodological consistency across studies. This study also establishes a non-human primate model that can provide a stable platform for carrying out serial sampling from the central nervous system and peripheral vasculature concurrently.

Microsatellite markers for standardized genetic management of captive colonies of rhesus macaques (Macaca mulatta).

To preserve genetic variability and minimize genetic subdivision among captive Macaca mulatta at each of the U.S. National Institutes of Health (NIH)-sponsored regional research colonies, the genetic structure of each colony must be characterized. To compare population genetic and demographic parameters across colonies and generations, one standard panel of highly informative genetic markers is required. We assembled a core marker set of four multiplex polymerase chain reaction (PCR) panels comprising 15 autosomal short tandem repeat (STR) loci with high information content selected from existing panels of well-characterized markers that are currently used for parentage assessment and genetic management of rhesus macaques. We then assessed the effectiveness of these loci for providing high probabilities of individual identification and parentage resolution, and for estimating population genetic parameters that are useful for genetic management.

Comparative efficacy of a canine distemper-measles and a standard measles vaccine for immunization of rhesus macaques (Macaca mulatta).

Measles virus (MV), a highly infective paramyxovirus, has caused sporadic epizootics characterized by high morbidity and increased mortality in nonhuman primates. Measles vaccines for human use, although effective, are cost prohibitive for use in primate colonies. We compared the efficacy of one or two doses of Vanguard D-M, a canine distemper-measles (CD-M) vaccine, with a single dose of Attenuvax, a human measles vaccine. Compared with 81% of animals inoculated with Attenuvax, all animals inoculated with one or two doses of Vanguard developed detectable MV antibodies. One year after immunization, six juveniles from each vaccine group, along with three unvaccinated controls, were challenged with pathogenic MV and were monitored for clinical signs of disease, viremia, viral shedding, and immune response. All uninoculated controls developed clinical disease and viremia, and shed virus in nasopharangeal secretions. Subclinical viremia without viral shedding was identified in two Attenuvax- and two single-dose Vanguard-inoculated animals. Viremia was not detected in any two-dose Vanguard-inoculated animals. Significantly higher neutralization antibody titers were observed in animals receiving Vanguard. Results of this study indicate that Vanguard is at least as efficacious as Attenuvax for protection of rhesus macaques. The considerably lower cost of Vanguard makes vaccination against measles in large breeding colonies economically feasible.