Adeno-Associated Virus-Mediated Dorsal Root Ganglion Toxicity in the New Zealand White Rabbit.

Recombinant adeno-associated virus (AAV)-mediated degeneration of sensory neurons in the dorsal root ganglia (DRG) and trigeminal ganglia (TG) has been observed in non-human primates (NHPs) following intravenous (IV) and intrathecal (IT) delivery. Administration of recombinant AAV encoding a human protein transgene via a single intra-cisterna magna (ICM) injection in New Zealand white rabbits resulted in histopathology changes very similar to NHPs: mononuclear cell infiltration, degeneration/necrosis of sensory neurons, and nerve fiber degeneration of sensory tracts in the spinal cord and of multiple nerves. AAV-associated clinical signs and incidence/severity of histologic findings indicated that rabbits were equally or more sensitive than NHPs to sensory neuron damage.

Scientific and Regulatory Policy Committee Technical Review: Biology and Pathology of Ganglia in Animal Species Used for Nonclinical Safety Testing.

Dorsal root ganglia (DRG), trigeminal ganglia (TG), other sensory ganglia, and autonomic ganglia may be injured by some test article classes, including anti-neoplastic chemotherapeutics, adeno-associated virus-based gene therapies, antisense oligonucleotides, nerve growth factor inhibitors, and aminoglycoside antibiotics. This article reviews ganglion anatomy, cytology, and pathology (emphasizing sensory ganglia) among common nonclinical species used in assessing product safety for such test articles (TAs). Principal histopathologic findings associated with sensory ganglion injury include neuron degeneration, necrosis, and/or loss; increased satellite glial cell and/or Schwann cell numbers; and leukocyte infiltration and/or inflammation.

Efficacy and safety of a SOD1-targeting artificial miRNA delivered by AAV9 in mice are impacted by miRNA scaffold selection.

Toxic gain-of-function mutations in superoxide dismutase 1 (SOD1) contribute to approximately 2%-3% of all amyotrophic lateral sclerosis (ALS) cases. Artificial microRNAs (amiRs) delivered by adeno-associated virus (AAV) have been proposed as a potential treatment option to silence SOD1 expression and mitigate disease progression. Primary microRNA (pri-miRNA) scaffolds are used in amiRs to shuttle a hairpin RNA into the endogenous miRNA pathway, but it is unclear whether different primary miRNA (pri-miRNA) scaffolds impact the potency and safety profile of the expressed amiR .

Scientific and Regulatory Policy Committee Points to Consider: Sampling, Processing, Evaluation, Interpretation, and Reporting of Test Article-Related Ganglion Pathology for Nonclinical Toxicity Studies.

Certain biopharmaceutical products consistently affect dorsal root ganglia, trigeminal ganglia, and/or autonomic ganglia. Product classes targeting ganglia include antineoplastic chemotherapeutics, adeno-associated virus-based gene therapies, antisense oligonucleotides, and anti-nerve growth factor agents. This article outlines "points to consider" for sample collection, processing, evaluation, interpretation, and reporting of ganglion findings; these points are consistent with published best practices for peripheral nervous system evaluation in nonclinical toxicity studies.

Antisense Oligonucleotide-Related Macrovesicular Vacuolation of Hippocampal Neurons in Nonhuman Primates.

2'-methoxyethyl (MOE) antisense oligonucleotides (ASOs) tested in multidose intrathecal nonhuman primate (NHP) toxicity studies have consistently revealed the presence of single large vacuoles in pyramidal neurons of the hippocampus in the absence of any cellular response. Termed "macrovesicular," these vacuoles were characterized by immunohistochemistry and transmission electron microscopy which showed that these vacuoles are dilated lysosomes in neurons containing accumulated ASO. Additionally, two NHP studies were conducted to investigate the role of tissue fixation on their histogenesis.

Scientific and Regulatory Policy Committee Points to Consider: Nonclinical Research and Development of In Vivo Gene Therapy Products, Emphasizing Adeno-Associated Virus Vectors.

Sequencing of the human genome and numerous advances in molecular techniques have launched the era of genetic medicine. Increasingly precise technologies for genetic modification, manufacturing, and administration of pharmaceutical-grade biologics have proved the viability of in vivo gene therapy (GTx) as a therapeutic modality as shown in several thousand clinical trials and recent approval of several GTx products for treating rare diseases and cancers. In recognition of the rapidly advancing knowledge in this field, the regulatory landscape has evolved considerably to maintain appropriate monitoring of safety concerns associated with this modality.

The role of the toxicologic pathologist in the biopharmaceutical industry.

Toxicologic pathologists contribute significantly to the development of new biopharmaceuticals, yet there is often a lack of awareness of this specialized role. As the members of multidisciplinary teams, toxicologic pathologists participate in all aspects of the drug development process. This review is part of an initiative by the Society of Toxicologic Pathology to educate scientists about toxicologic pathology and to attract junior scientists, veterinary students, and veterinarians into the field.

Effects of nicotine on pulmonary surfactant proteins A and D in ovine lung epithelia.

Maternal smoking during pregnancy increases the incidence and severity of respiratory infections in neonates. Surfactant proteins A and D (SP-A and SP-D, respectively) are components of pulmonary innate immunity and have an important role in defense against inhaled pathogens. The purpose of this study was to determine if nicotine exposure during the third trimester of pregnancy alters the expression of SP-A and SP-D of fetal lung epithelia.

Maternal alcohol ingestion reduces surfactant protein A expression by preterm fetal lung epithelia.

In addition to neurodevelopmental effects, alcohol consumption at high levels during pregnancy is associated with immunomodulation and premature birth. Premature birth, in turn, is associated with increased susceptibility to various infectious agents such as respiratory syncytial virus (RSV). The initial line of pulmonary innate defense includes the mucociliary apparatus, which expels microorganisms trapped within the airway secretions.

Differential expression of ovine innate immune genes by preterm and neonatal lung epithelia infected with respiratory syncytial virus.

Preterm infants have increased susceptibility to severe manifestations of respiratory syncytial virus (RSV) infection. The cause(s) for this age-dependent vulnerability is/are not well-defined, but alterations in innate immune products have been implicated. In sheep, RSV disease severity has similar age-dependent characteristics and sheep have several related innate molecules for study during pulmonary infection including surfactant protein A (SP-A), surfactant protein D (SP-D), sheep beta defensin 1 (SBD1), monocyte chemotactic protein 1 (MCP1), and Toll-like receptor 4 (TLR4).

Expression of select immune genes (surfactant proteins A and D, sheep beta defensin 1, and toll-like receptor 4) by respiratory epithelia is developmentally regulated in the preterm neonatal lamb.

Preterm infants experience enhanced susceptibility and severity to respiratory syncytial virus (RSV) infection. Terminal airway epithelium is an important site of RSV infection and the extent of local innate immune gene expression is poorly understood. In this study, expression of surfactant proteins A and D (SP-AD), sheep beta defensin 1 (SBD1), and toll-like receptor 4 (TLR4) mRNA were determined in whole lung homogenates from lambs.

Reduced clearance of respiratory syncytial virus infection in a preterm lamb model.

Respiratory syncytial virus (RSV) causes significant respiratory disease in children worldwide. For the study of severe RSV disease seen in preterm infants, a suitable animal model is lacking. The novel hypothesis of this study was that preterm lambs are susceptible to bovine RSV (bRSV) infection, an analogous pneumovirus with ruminant host specificity, and that there would be age-dependent differences in select RSV disease parameters.

Adenovirus-mediated gene therapy enhances parainfluenza virus 3 infection in neonatal lambs.

Parainfluenza viruses are a common cause of seasonal respiratory disease, but in high-risk individuals (e.g., young children) these viruses can cause severe clinical manifestations that require hospitalization.

Surfactant protein D expression in normal and pneumonic ovine lung.

Surfactant protein D (SP-D) is a collagenous calcium-dependent lectin constitutively expressed by alveolar type II pneumocytes and non-ciliated bronchiolar epithelial (Clara) cells. It binds to surface glycoconjugates expressed by a wide variety of microorganisms such as Gram-negative bacteria, influenza A virus, and various fungi, leading to pathogen inactivation or enhanced neutrophil and macrophage activity. Since a hallmark of bronchopneumonia is the initiation of inflammation in the bronchi and bronchoalveolar junction, we chose a classic ruminant model of bronchopneumonia caused by Mannheimia haemolytica to study the expression of SP-D within the bronchioles of infected lambs.

Differential expression of sheep beta-defensin-1 and -2 and interleukin 8 during acute Mannheimia haemolytica pneumonia.

Beta-defensins are antimicrobial peptides produced by several cell types, including respiratory epithelia and leukocytes. Expression of some beta-defensins is increased by bacterial-induced inflammatory responses whereas expression of other beta-defensins is constitutive. Two beta-defensins are expressed in lungs of sheep (sheep beta-defensin-1 and -2; SBD-1/-2) and expression of SBD-1 is increased during parainfluenza virus type 3 (PI-3) infection.

Enhanced surfactant protein and defensin mRNA levels and reduced viral replication during parainfluenza virus type 3 pneumonia in neonatal lambs.

Defensins and surfactant protein A (SP-A) and SP-D are antimicrobial components of the pulmonary innate immune system. The purpose of this study was to determine the extent to which parainfluenza type 3 virus infection in neonatal lambs alters expression of sheep beta-defensin 1 (SBD-1), SP-A, and SP-D, all of which are constitutively transcribed by respiratory epithelia. Parainfluenza type 3 viral antigen was detected by immunohistochemistry (IHC) in the bronchioles of all infected lambs 3 days postinoculation and at diminished levels 6 days postinoculation, but it was absent 17 days postinoculation.

Distribution of substance P receptor (neurokinin-1 receptor) in normal ovine lung and during the progression of bronchopneumonia in sheep.

Substance P contributes to the physiological homeostasis of pulmonary airways and vasculature. During pneumonia, alterations in substance P production and receptor expression can influence bronchoconstriction and vascular perfusion. The distribution of substance P receptor [neurokinin-1 receptor (NK-1R)] in lungs of normal sheep and sheep with acute (1 day), subacute (15 days), and chronic (45 days) bronchopneumonia caused by Mannheimia haemolytica was determined by immunohistochemistry (IHC).

Developmental expression and distribution of sheep beta-defensin-2.

The aim of this study was to define the ontogeny of sheep beta-defensin-2 (SBD-2) mRNA and peptide in selected tissues of fetal, neonatal and adult sheep by real-time PCR and immunohistochemistry, respectively. Fetal and neonatal lambs had significantly greater SBD-2 tissue distribution than adult sheep. For all ages, the intestines had consistent SBD-2 mRNA expression while extra intestinal expression was sporadic and weak.

Repeated intravenous doses of all-trans-retinoyl beta-D-glucuronide is not effective in the treatment of bacterial bronchopneumonia in lambs but is devoid of gross and acute toxicity.

Background: All-trans-retinoyl beta-D-glucuronide, a water-soluble glucuronic acid conjugate of all-trans-retinoic acid, has demonstrated high biological activity and low toxicity in most in vitro and in vivo models. Since the reparative effects of retinoids on epithelium are well-known, our aim was to study the effect(s) of intravenously-administered all-trans-retinoyl beta-D-glucuronide in lambs with chronic bacterial bronchopneumonia.

Material/methods: Two groups of lambs were inoculated intrabronchially with either pyrogen-free saline or Mannheimia haemolytica.