Practical Considerations in Clinical Pathology Data Interpretation and Description.

Although interpretation and description of clinical pathology test results for any preclinical safety assessment study should employ a consistent standard approach, companies differ regarding that approach and the appearance of the end product. Some rely heavily on statistical analysis, others do not. Some believe reference intervals are important, most do not. Some prefer severity of effects be described by percentage differences from, or multiples of, baseline or control, others prefer only word modifiers. Some expect a definitive decision for every potential effect, others accept uncertainty. This commentary addresses these differences and underscores the need for flexibility in a "consistent standard approach" because the conditions of every study are unique. This article constitutes an overview of material originally presented at Session 2 of the 2016 Society of Toxicologic Pathology Annual Symposium.

Deciphering Sources of Variability in Clinical Pathology.

The objectives of this session were to explore causes of variability in clinical pathology data due to preanalytical and analytical variables as well as study design and other procedures that occur in toxicity testing studies. The presenters highlighted challenges associated with such variability in differentiating test article-related effects from the effects of experimental procedures and its impact on overall data interpretation. These presentations focused on preanalytical and analytical variables and study design-related factors and their influence on clinical pathology data, and the importance of various factors that influence data interpretation including statistical analysis and reference intervals. Overall, these presentations touched upon potential effect of many variables on clinical pathology parameters, including animal physiology, sample collection process, specimen handling and analysis, study design, and some discussion points on how to manage those variables to ensure accurate interpretation of clinical pathology data in toxicity studies. This article is a brief synopsis of presentations given in a session entitled "Deciphering Sources of Variability in Clinical Pathology-It's Not Just about the Numbers" that occurred at the 35th Annual Symposium of the Society of Toxicologic Pathology in San Diego, California.

Naphthoquine-induced Central Nervous System and Hepatic Vasculocentric Toxicity in the Beagle Dog.

Naphthoquine phosphate (NP) was considered as a partner drug with a promising antimalarial drug candidate. Here we report unexpected adverse clinical signs and microscopic findings in a canine pilot toxicology study with NP. Male and female dogs were dosed daily by oral gavage with NP at 2, 10, or 50 mg/kg/day for a maximum of 14 days. NP was not tolerated at ≥10 mg/kg/day; several animals were sacrificed in moribund condition and marked neurological clinical signs were noted at 50 mg/kg/day. The main microscopic observation was central nervous system vasculocentric inflammation (mainly lymphocytes and macrophages) in the white and gray matter of various regions of the brain at ≥2 mg/kg/day and at lower incidence in the spinal cord at ≥10 mg/kg/day. Vasculocentric microscopic changes predominantly centered on the centrilobular vein were also observed in the liver at ≥2 mg/kg/day. Females were more sensitive than males with comparable NP plasma exposure. In conclusion, under the conditions of this study, the administration of NP to dogs via daily oral gavage for up to 2 weeks was not tolerated causing moribundity, marked neurological clinical signs, and vasculocentric microscopic changes in the central nervous system and the liver.

Factors affecting urine reagent strip blood results in dogs and nonhuman primates and interpretation of urinalysis in preclinical toxicology studies: a Multi-Institution Contract Research Organization and BioPharmaceutical Company Perspective.

BACKGROUND: Urinalysis data in preclinical toxicology studies can be influenced by preanalytic and analytic factors which have the potential to confound interpretation. There is a paucity of information regarding positive reagent strip urinary blood reactions in healthy nonhuman primates (NHP) and Beagle dogs used in preclinical toxicology studies. OBJECTIVES: The objectives were (1) to establish historical control data for reagent strip urinary blood reactions in healthy NHP and Beagle dogs, (2) to determine the incidence of positive urinary blood reactions during predose and dosing phases, and (3) to determine if collection practice was a relevant parameter. METHODS: Historical control data from 2 institutions in the biopharmaceutical industry were retrospectively analyzed for reagent strip urinary blood reactions in healthy NHP and Beagles. The incidence of positive results between the 2 institutions with different urine collection practices and between males and females was compared. RESULTS: The incidence of positive urinary blood reactions in NHP was comparable between institutions (≤ 14% in males; ≤ 33% in females), while the incidence of positive urinary blood reactions in Beagles was more variable (≤ 77% in males; ≤ 69% in females), and higher in females during the dosing phase. CONCLUSIONS: Positive urinary blood results that could potentially be misinterpreted as toxicologically relevant were identified in healthy NHP and Beagles during predose and dosing phases. Different incidences of positive results between the 2 institutions were likely related to collection practices. Strategies to reduce feces and food contamination of collected urine samples should help minimize false-positive urinary blood reactions.

Best practices for clinical pathology testing in carcinogenicity studies.

The Society of Toxicologic Pathology (STP) and American Society for Veterinary Clinical Pathology (ASCVP) convened a Clinical Pathology in Carcinogenicity Studies Working Group to recommend best practices for inclusion of clinical pathology testing in carcinogenicity studies. Regulatory guidance documents and literature were reviewed, and veterinary pathologists from North America, Japan, and Europe were surveyed regarding current practices, perceived value, and recommendations for clinical pathology testing in carcinogenicity studies. For two-year rodent carcinogenicity studies, the Working Group recommends that clinical pathology testing be limited to collection of blood smears at scheduled and unscheduled sacrifices to be examined only if indicated to aid in the diagnosis of possible hematopoietic neoplasia following histopathologic evaluation. Additional clinical pathology testing is most appropriately used to address specific issues from prior toxicity studies or known test article-related class effects. Inadequate data were available to make a recommendation concerning clinical pathology testing for alternative six-month carcinogenicity assays using genetically modified mice, although the Working Group suggests that it may be appropriate to use the same approach as for two-year carcinogenicity studies since the study goal is the same.

Refinement of the urine concentration test in rats.

The urine concentration test is a potentially stressful procedure used to assess renal function. Historically, animals have been deprived of water for 24 h or longer during this test, creating the potential for distress. Refinement of the technique to lessen distress may involve decreasing the water-deprivation period. To determine the feasibility of reduced water-deprivation time, 10 male and 10 female rats were food- and water-deprived for 22 h. Clinical condition and body weights were recorded, and urine was collected every 2 h, beginning 16 h after the onset of food and water deprivation. All rats lost weight (P < 0.001). All rats were clinically normal after 16 h, but 90% of the males and 30% of the females appeared clinically dehydrated after 22 h. After 16 h, mean urine specific gravities were 1.040 and 1.054 for males and females, respectively, and mean urine osmolalities were 1,362 and 2,080 mOsm/kg, respectively, indicating the rats were adequately concentrating urine. The rats in this study tolerated water deprivation relatively well for 16 h but showed clinical signs of dehydration after 22 h. Based on this study, it was concluded that the urine concentration test can be refined such that rats are not deprived of water for more than 16 h without jeopardizing test results.

Factors affecting the interpretation of canine and nonhuman primate clinical pathology.

Interpreting canine and nonhuman primate clinical pathology data from preclinical studies can be challenging. Relatively few animals are tested (typically beagles and macaques), and they often undergo study-related procedures (eg, sample collection for pharmacokinetic analysis) that can affect clinical pathology test results. Data interpretation requires an understanding of the significance of each test, species differences for each test, normal interanimal and intraanimal variability, the effects of study design variables, and supporting data from other disciplines. Interpretation of hematology, coagulation, clinical chemistry, and urinalysis parameters are discussed, with emphasis on species peculiarities and study design variables that may affect clinical pathology test results.