Toxicology Evaluation of Drugs Administered via Uncommon Routes: Intranasal, Intraocular, Intrathecal/Intraspinal, and Intra-Articular.

As the need for nasal, ocular, spinal, and articular therapeutic compounds increases, toxicology assessments of drugs administered via these routes play an important role in human safety. This symposium outlined the local and systemic evaluation to support safety during the development of these drugs in nonclinical models with some case studies. Discussions included selection of appropriate species for the intended route; conducting nonclinical studies that closely mimic the intended use with adequate duration; functional assessment, if deemed necessary; evaluation of local tissues with special histological staining procedure; and evaluations of safety margins based on local and systemic toxicity.

Compilation of international regulatory guidance documents for neuropathology assessment during nonclinical general toxicity and specialized neurotoxicity studies.

Neuropathology analyses as end points during nonclinical efficacy and toxicity studies are challenging and require trained personnel and particular equipment to achieve optimal results. Accordingly, many regulatory agencies have produced explicit guidelines for designing and performing neuropathology assessments for nonclinical studies. This compilation of international regulatory guidance for toxicologic neuropathology end points represents a set of criteria recommended for general toxicity studies and specialized neurotoxicity studies that should facilitate the efforts of individuals who plan, perform, analyze, and report neuropathology evaluations in nonclinical toxicity studies.

Continuing education course #3: current practices and future trends in neuropathology assessment for developmental neurotoxicity testing.

The continuing education course on Developmental Neurotoxicity Testing (DNT) was designed to communicate current practices for DNT neuropathology, describe promising innovations in quantitative analysis and noninvasive imaging, and facilitate a discussion among experienced neuropathologists and regulatory scientists regarding suitable DNT practices. Conventional DNT neuropathology endpoints are qualitative histopathology and morphometric endpoints of particularly vulnerable sites (e.g., cerebral, cerebellar, or hippocampal thickness). Novel imaging and stereology measurements hold promise for automated analysis of factors that cannot be effectively examined in routinely processed specimens (e.g., cell numbers, fiber tract integrity). The panel recommended that dedicated DNT neuropathology data sets be acquired on a minimum of 8 sections (for qualitative assessment) or 3 sections (for quantitative linear and stereological analyses) using a small battery of stains to examine neurons and myelin. Where guidelines permit discretion, immersion fixation is acceptable for younger animals (postnatal day 22 or earlier), and peripheral nerves may be embedded in paraffin. Frequent concerns regarding DNT data sets include false-negative outcomes due to processing difficulties (e.g., lack of concordance among sections from different animals) and insensitive analytical endpoints (e.g., qualitative evaluation) as well as false-positive results arising from overinterpretation or misreading by inexperienced pathologists.

Uniform assessment and ranking of opioid µ receptor binding constants for selected opioid drugs.

The safe disposal of unused opioid drugs is an area of regulatory concern. While toilet flushing is recommended for some drugs to prevent accidental exposure, there is a need for data that can support a more consistent disposal policy based on an assessment of relative risk. For drugs acting at the Mu-opioid receptor (MOR), published measurements of binding affinity (K(i)) are incomplete and inconsistent due to differences in methodology and assay system, leading to a wide range of values for the same drug thus precluding a simple and meaningful relative ranking of drug potency. Experiments were conducted to obtain K(i)'s for 19 approved opioid drugs using a single binding assay in a cell membrane preparation expressing recombinant human MOR. The K(i) values obtained ranged from 0.1380 nM (sufentanil) to 12.486 µM (tramadol). The drugs were separated into three categories based upon their K(i) values: K(i) > 100 nM (tramadol, codeine, meperidine, propoxyphene and pentazocine), K(i)=1-100 nM (hydrocodone, oxycodone, diphenoxylate, alfentanil, methadone, nalbuphine, fentanyl and morphine) and K(i) < 1 nM (butorphanol, levorphanol, oxymorphone, hydromorphone, buprenorphine and sufentanil). These data add to the understanding of the pharmacology of opioid drugs and support the development of a more consistent labeling policies regarding safe disposal.

Can We Panelize Seizure?

Seizure liability remains a significant cause of attrition in drug discovery and development, leading to loss of competitiveness, delays, and increased costs. Current detection methods rely on observations made in in vivo studies intended to support clinical trials, such as tremors or other abnormal movements. These signs could be missed or misinterpreted; thus, definitive confirmation of drug-induced seizure requires a follow-up electroencephalogram study. There has been progress in in vivo detection of seizure using automated video systems that record and analyze animal movements. Nonetheless, it would be preferable to have earlier prediction of seizurogenic risk that could be used to eliminate liabilities early in discovery while there are options for medicinal chemists making potential new drugs. Attrition due to cardiac adverse events has benefited from routine early screening; could we reduce attrition due to seizure using a similar approach? Specifically, microelectrode arrays could be used to detect potential seizurogenic signals in stem-cell-derived neurons. In addition, there is clear evidence implicating neuronal voltage-gated and ligand-gated ion channels, GPCRs and transporters in seizure. Interactions with surrounding glial cells during states of stress or inflammation can also modulate ion channel function in neurons, adding to the challenge of seizure prediction. It is timely to evaluate the opportunity to develop an in vitro assessment of seizure linked to a panel of ion channel assays that predict seizure, with the aim of influencing structure-activity relationship at the design stage and eliminating compounds predicted to be associated with pro-seizurogenic state.

Use of anesthetic agents in neonates and young children.

BACKGROUND: Some drugs used for sedation and anesthesia produce histopathologic central nervous system changes in juvenile animal models. These observations have raised concerns regarding the use of these drugs in pediatric patients. We summarized the findings in developing animals and describe the steps that the Food and Drug Administration (FDA) and others are taking to assess potential risks in pediatric patients. The FDA views this communication as opening a dialog with the anesthesia community to address this issue. METHODS: We reviewed the available animal studies literature examining the potential neurotoxic effects of commonly used anesthetic drugs on the developing brain. The search strategy involved crossing the keywords neurotoxic and neuroapoptosis with the following general and specific terms: anesthetic, N-methyl-d-aspartate (NMDA), ketamine, midazolam, lorazepam, fentanyl, methadone, morphine, meperidine, isoflurane, nitrous oxide, sevoflurane, halothane, enflurane, desflurane, propofol, etomidate, barbiturate, methoxyflurane, and chloral hydrate. We summarized several studies sponsored by the FDA in rats and monkeys, initially examining the potential for ketamine, as a prototypical agent, to induce neurodegeneration in the developing brain. RESULTS: Numerous animal studies in rodents indicate that NMDA receptor antagonists, including ketamine, induce neurodegeneration in the developing brain. The effects of ketamine are dose dependent. The data suggest that limiting exposure limits the potential for neurodegeneration. There is also evidence that other general anesthetics, such as isoflurane, can induce neurodegeneration in rodent models, which may be exacerbated by concurrent administration of midazolam or nitrous oxide. There are very few studies that have examined the potential functional consequences of the neurodegeneration noted in the animal models. However, the studies that have been reported suggest subtle, but prolonged, behavioral changes in rodents. Although the doses and durations of ketamine exposure that resulted in neurodegeneration were slightly larger than those used in the clinical setting, those associated with isoflurane were not. There are insufficient human data to either support or refute the clinical applicability of these findings. CONCLUSIONS: Animal studies suggest that neurodegeneration, with possible cognitive sequelae, is a potential long-term risk of anesthetics in neonatal and young pediatric patients. The existing nonclinical data implicate not only NMDA-receptor antagonists, but also drugs that potentiate gamma-aminobutyric acid signal transduction, as potentially neurotoxic to the developing brain. The potential for the combination of drugs that have activity at both receptor systems or that can induce more or less neurotoxicity is not clear; however, recent nonclinical data suggest that some combinations may be more neurotoxic than the individual components. The lack of information to date precludes the ability to designate any one anesthetic agent or regimen as safer than any other. Ongoing studies in juvenile animals should provide additional information regarding the risks. The FDA anticipates working with the anesthesia community and pharmaceutical industry to develop strategies for further assessing the safety of anesthetics in neonates and young children, and for providing data to guide clinicians in making the most informed decisions possible when choosing anesthetic regimens for their pediatric patients.

Immunomodulation and lymphoma in humans.

Observational and clinical studies have associated increased cancer risks with primary or acquired immunodeficiencies, autoimmunity, and use of immunotherapies to treat chronic inflammation (e.g. autoimmunity) or support organ engraftment. Understanding of the relationship between immune status and cancer risk is generally grounded in two juxtaposing paradigms: that the immune system protects the host via surveillance of tumors and oncogenic viruses (e.g. immunosurveillance model) and that chronic inflammation can augment tumor growth and metastasis (inflammation model). Whereas these models support a role of immune status in many cancers, they are insufficient to explain the disproportionate increase in B-cell lymphoma risk observed across patient populations with either chronic immunosuppression or inflammation. Evaluation for the presence of Epstein-Barr virus (EBV) in lymphomas obtained from various populations demonstrates a variable role for the virus in lymphomagenesis across patient populations. An evaluation of the DNA alterations found in lymphomas and an understanding of B-cell ontogeny help to provide insight into the unique susceptibility of lymphocytes, primarily B-cells, to oncogenic transformation. EBV-independent B-cell oncogenic transformation is driven by chronic antigenic stimulation due to either inflammation (as seen in patients with autoimmune disease or a tissue allograft) or to unresolved infection (as seen in immunosuppressed patients), and the transformation arises as a result of DNA damage from genomic recombination and mutation during class switching and somatic hypermutation. This model explains the increased background rate of lymphoma in some patients with autoimmunity, and highlights the challenge of resolving the confounding that occurs between disease severity and use of targeted immunotherapies to treat chronic inflammation. The ability to distinguish between disease- and treatment-related risk of lymphoma and an appreciation of the etiology of B-cell transformation is central to an improved risk assessment by scientists, clinicians and regulators, including the approval, labeling, and chronic use of immunotherapies.