Phenotypic screening using motor neurons derived from the induced pluripotent stem (iPS) cell of a patient with amyotrophic lateral sclerosis led to the identification of Compound X, a novel agent with potent neuroprotective activity that is hypothesized to act by inhibiting hematopoietic progenitor kinase/germinal center kinase-like kinase (HGK). Unlike known HGK inhibitors, including Prostetin and GNE-495, Compound X and related Compounds Y and Z exhibited broader multi-kinase-inhibition profiles, including the strong inhibition of Aurora B kinase and strong-to-moderate inhibition of Src-family kinases in kinase panel screening. Owing to their roles in cytokinesis, we performed in vitro micronucleus tests to assess their genotoxicity. Compounds X, Y, and Z induced the multinucleation of cells at rates of 88.8, 71.5, and 91.8%, respectively, whereas Prostetin and GNE-495 resulted in rates of 0.5 and 1.3%, respectively, which were comparable to the control. A 5-day repeated-dose oral toxicity study of Compound X using mice, at doses of 125, 250, and 500 mg/kg/day, revealed decreased locomotor activity, hypothermia, and mortality at 500 mg/kg/day. Histopathology showed necrosis/hemorrhage in bone marrow cells; single-cell necrosis in the thymus, spleen, and mesenteric lymph nodes; nuclear pleomorphism and single-cell necrosis/necrosis in the gastrointestinal mucosa; and karyomegaly and a reduction in testicular germ cells. These toxicopathological findings suggest that Compound X induces nuclear pleomorphism and necrosis in highly proliferative cells. Although Compound X strongly suppressed HGK, its toxicity may be associated with broader multikinase inhibition, which is consistent with the finding that Prostetin and GNE-495 do not induce multinucleation in vitro.

CByB6F1-Tg (HRAS)2Jic mice (rasH2 mice) are widely used in 26-week short-term carcinogenicity assessments. To ensure phenotypic stability and carcinogenic sensitivity, a 26-week N-methyl-N-nitrosourea (MNU)-induced carcinogenic susceptibility monitoring study was conducted for quality assurance. The study included (i) comprehensive whole-organ monitoring after colony renewal, and (ii) simplified annual monitoring on production colonies focusing on forestomach tumor formation as the most sensitive endpoint. Retrospective analysis of our 26-week historical data indicated a high incidence of forestomach tumors in mice that were moribund or sacrificed before 13 weeks. Therefore, we compared the 13- and 26-week observation periods to evaluate whether 13 weeks was sufficient for simple carcinogenicity monitoring. RasH2 mice were intraperitoneally administered MNU (75 mg/kg) and observed for 13 and 26 weeks. Both groups exhibited small and large forestomach nodules, including hemorrhagic lesions. The corresponding histopathological features were squamous cell papillomas and carcinomas. The incidence of squamous cell papilloma or carcinoma was 100.0% in both sexes at 13 weeks, which was comparable to that at 26 weeks (100.0% in males and 93.3% in females). These results demonstrated that MNU-induced forestomach tumor can be reliably detected within 13 weeks, indicating that a shortened monitoring period of 13 weeks is sufficient to monitor carcinogenic susceptibility for quality assurance using genotoxic carcinogens in rasH2 mice.

An aged female Crl:CD (SD) rat developed a subcutaneous tumor in the left occipital region. Histologically, the tumor was predominantly composed of spindle-shaped cells proliferating in a loosely cohesive fascicular arrangement in a basophilic myxoid matrix with a minor population of epithelioid cells. Metastases were detected in the cervical lymph nodes, kidneys, lungs, and the pleural cavity. Duct-like structures, considered to be entrapped normal salivary ducts, were scattered within the tumor. Immunohistochemically, the tumor cells were diffusely positive for vimentin, focally positive for alpha-smooth muscle actin and p63 in the spindle-shaped population, and positive for cytokeratin AE1/AE3, exclusive to the epithelioid cell population, whereas S100, chromogranin A, and myoglobin were not expressed in any of the tumor cells. Ultrastructurally, spindle-shaped tumor cells exhibited microfilaments, desmosome-like junctions, and cytoplasmic processes. These features were consistent with the diagnosis of malignant myoepithelioma, most likely originating from the parotid gland.

Motugivatrep (SJP-0132), a transient receptor potential cation channel subfamily V member 1 (TRPV1) antagonist, is being developed as an ophthalmic drug for dry eye disease. Since the role of TRPV1 antagonists in skin carcinogenesis remains controversial, this study aimed to investigate whether motugivatrep promotes skin tumorigenesis using an ultra-short-term two-stage skin carcinogenesis model with Tg-rasH2 mice. Following initiation with a single dermal application of 7,12-dimethylbenz[a]anthracene, motugivatrep was applied topically at doses of 0, 150, 300, or 750 µg/50 µL/animal once a day to mouse skin for 7 weeks. The positive control group received 12-O-tetradecanoylphorbol-13-acetate (TPA) once weekly. No skin tumors or preneoplastic lesions were observed in any animals treated with motugivatrep. In contrast, treatment with TPA significantly increased skin tumor formation, confirming the validity of the assay. These results indicated that motugivatrep lacked skin tumor promotion activity in this Tg-rasH2 mice model, providing safety information regarding dermal exposure to motugivatrep.