Two weeks dose range-finding and four weeks repeated dose oral toxicity study of a novel reversible monoamine oxidase B inhibitor KDS2010 in cynomolgus monkeys.

A novel reversible monoamine oxidase B inhibitor, KDS2010, has been developed as a therapeutic candidate for neurodegenerative diseases. This study investigated its potential toxicity in non-human primates before human clinical trials. Daily KDS2010 doses (25, 50, or 100 mg/kg) were orally administered to cynomolgus monkeys (1 animal/sex/group, 4 males and 4 females) for 2 weeks to determine the dose range. One male was moribund, and one female was found dead in the 100 mg/kg/day group. One male was also found dead in the 50 mg/kg/day group. The death was considered an adverse effect in both sexes since distal tubules/collecting duct dilation and hypertrophy in the epithelium of the papillary duct were observed in their kidneys. Based on dose range finding results, KDS2010 (10, 20, or 40 mg/kg/day) was administered orally for 4 weeks, and animals were given 2 weeks for recovery. No significant changes were observed during daily clinical observations and macro-and microscopic examinations, including body weight, food consumption, hematology, clinical chemistry, and organ weight. And, the kidney was seen as the primary target organ of KDS2010 in the 2 weeks study, but no adverse effect was observed in the 4 weeks study. Therefore, 40 mg/kg/day is considered the no-observed-adverse-effect level in both sexes of cynomolgus monkeys. Supplementary Information: The online version contains supplementary material available at 10.1007/s43188-023-00182-4.

Transcriptional control of motor pool formation and motor circuit connectivity by the LIM-HD protein Isl2.

The fidelity of motor control requires the precise positional arrangement of motor pools and the establishment of synaptic connections between them. During neural development in the spinal cord, motor nerves project to specific target muscles and receive proprioceptive input from these muscles via the sensorimotor circuit. LIM-homeodomain transcription factors are known to play a crucial role in successively restricting specific motor neuronal fates. However, their exact contribution to limb-based motor pools and locomotor circuits has not been fully understood. To address this, we conducted an investigation into the role of Isl2, a LIM-homeodomain transcription factor, in motor pool organization. We found that deletion of Isl2 led to the dispersion of motor pools, primarily affecting the median motor column (MMC) and lateral motor column (LMC) populations. Additionally, hindlimb motor pools lacked Etv4 expression, and we observed reduced terminal axon branching and disorganized neuromuscular junctions in Isl2-deficient mice. Furthermore, we performed transcriptomic analysis on the spinal cords of Isl2-deficient mice and identified a variety of downregulated genes associated with motor neuron (MN) differentiation, axon development, and synapse organization in hindlimb motor pools. As a consequence of these disruptions, sensorimotor connectivity and hindlimb locomotion were impaired in Isl2-deficient mice. Taken together, our findings highlight the critical role of Isl2 in organizing motor pool position and sensorimotor circuits in hindlimb motor pools. This research provides valuable insights into the molecular mechanisms governing motor control and its potential implications for understanding motor-related disorders in humans.

Temporal quantitative profiling of sialyllactoses and sialic acids after oral administration of sialyllactose to mini-pigs with osteoarthritis.

Sialyllactose (SL) is the most abundant acidic oligosaccharide in human breast milk and plays a primary role in various biological processes. Recently, SL has attracted attention as an excellent dietary supplement for arthritis because it is effective in cartilage protection and treatment. Despite the superior function of SL, there are few pharmacological studies of SL according to blood concentrations in arthritis models. In this study, we investigated quantitative changes in SL and sialic acids in the plasma obtained from mini-pigs with osteoarthritis throughout exogenous administration of SL using liquid chromatography-multiple reaction monitoring mass spectrometry. Plasma concentrations of SL and sialic acids in the SL-fed group showed a significant difference compared to the control group. Mini pigs were fed only Neu5Ac bound to SL, but the concentration patterns of the two types of sialic acid, Neu5Ac and Neu5Gc, were similar. In addition, the relative mRNA expression level of matrix metalloproteinases (MMPs), which is known as a critical factor in cartilage matrix degradation, was remarkably decreased in the synovial membrane of the SL-fed group. Consequently, the temporal quantitative profiling suggests that dietary SL can be metabolized and utilized in the body and may protect against cartilage degradation by suppressing MMP expression during osteoarthritis progression.

Multi-omic analysis of selectively vulnerable motor neuron subtypes implicates altered lipid metabolism in ALS.

Amyotrophic lateral sclerosis (ALS) is a devastating disorder in which motor neurons degenerate, the causes of which remain unclear. In particular, the basis for selective vulnerability of spinal motor neurons (sMNs) and resistance of ocular motor neurons to degeneration in ALS has yet to be elucidated. Here, we applied comparative multi-omics analysis of human induced pluripotent stem cell-derived sMNs and ocular motor neurons to identify shared metabolic perturbations in inherited and sporadic ALS sMNs, revealing dysregulation in lipid metabolism and its related genes. Targeted metabolomics studies confirmed such findings in sMNs of 17 ALS (SOD1, C9ORF72, TDP43 (TARDBP) and sporadic) human induced pluripotent stem cell lines, identifying elevated levels of arachidonic acid. Pharmacological reduction of arachidonic acid levels was sufficient to reverse ALS-related phenotypes in both human sMNs and in vivo in Drosophila and SOD1G93A mouse models. Collectively, these findings pinpoint a catalytic step of lipid metabolism as a potential therapeutic target for ALS.

Impairment of motor coordination and interneuron migration in perinatal exposure to glufosinate-ammonium.

Glufosinate-ammonium (GLA) is a broad-spectrum herbicide for agricultural weed control and crop desiccation. Due to many GLA-resistant crops being developed to effectively control weeds and increase harvest yields, herbicide usage and the residual GLA in food has increased significantly. Though perinatal exposure by the residual GLA in food might affect brain development, the developmental neurotoxicity of GLA is still unclear. Therefore, this study aimed to investigate the effects of perinatal exposure to GLA on cortical development. The analysis revealed that perinatal GLA exposure altered behavioral changes in offspring, especially motor functional behavior. Moreover, perinatal GLA exposure affected cortical development, particularly by disrupting interneuron migration. These results provide new evidence that early life exposure to GLA alters cortical development.

Thirteen-week oral toxicity study of fermented ginseng, GBCK25, in Sprague-Dawley rats.

Ginseng (Panax ginseng) is commonly used in Asia as a medicinal herb. In particular, fermented ginseng, GBCK25, has been recently developed to increase ginsenoside absorption. It also has other beneficial biological effects such as hemodynamic and anti-inflammation functions. Here, we investigated the potential toxicity of GBCK25 in Sprague-Dawley rats following 13 weeks of GBCK25 treatment by oral gavage at doses of 250, 500, or 1000 mg/kg/day and reversible toxic effects over a 4-week recovery phase. Ten male and female rats per group were randomly allocated to the main toxicology groups and five male and female rats per group were allocated to the 0 and 1000 mg/kg/day recovery groups, respectively. There was no mortality; significant clinical toxicity or microscopic findings; and changes in body weight, food consumption, hematological parameters, serum biochemistry, or absolute and relative organ weights in any of the groups. In conclusion, there were no toxicological changes upon repeated oral gavage of GBCK25 at doses of 250, 500, or 1000 mg/kg/day in Sprague-Dawley rats over 13 weeks. The no-observed-adverse-effect level of GBCK25 was 1000 mg/kg/day in both sexes of Sprague-Dawley rat.

Four-week repeated dose oral toxicity study of KDS2010, a novel selective monoamine oxidase B inhibitor, in Sprague Dawley rats.

Repeated dose oral toxicity and toxicokinetic of KDS2010, a new drug for Parkinson's disease, was investigated after 4-week repeated oral administration at 30, 50, 75, or 100 mg/kg/day in rats. Body weight and body weight gain decreased in rats of both sexes in the 75 and 100 mg/kg groups, and food consumption was reduced in male rats of the 75 and 100 mg/kg male groups. Histological alterations were observed in the kidney (urothelial hyperplasia, inflammatory cell infiltration in the renal pelvis, tubular vacuolation/degeneration, basophilic tubules, and hyaline droplets in the proximal tubules) of the 75 and 100 mg/kg male groups and the 50 and 100 mg/kg female groups. The 75 and 100 mg/kg male groups showed adverse effect in the testes (degeneration/exfoliation of germ cells, seminiferous tubules atrophy) and epididymis (cellular debris, oligospermia). These changes were partially recovered after a 2-week recovery period. However, basophilic tubules and hyaline droplets in the proximal tubules in the kidney and germ cell degeneration/exfoliation in the testis were not recovered. In toxicokinetics study, systemic exposure to KDS2010 increased proportionally in both sexes by in a dose -dependent manner. In addition, repeated administration for 4 weeks led to increased tendency of systemic exposure in both sexes compared with that in Day 1. In conclusion, KDS2010 was shown to target the kidney and testis with a no-observed-adverse-effect level of 50 and 30 mg/kg/day for males and females, respectively.

Reactive microglia and astrocytes in neonatal intraventricular hemorrhage model are blocked by mesenchymal stem cells.

Severe intraventricular hemorrhage (IVH) in premature infants triggers reactive gliosis, causing acute neuronal death and glial scar formation. Transplantation of mesenchymal stem cells (MSCs) has often showed improved CNS recovery in an IVH model, but whether this response is related to reactive glial cells is still unclear. Herein, we suggest that MSCs impede the response of reactive microglia rather than astrocytes, thereby blocking neuronal damage. Astrocytes alone showed mild reactiveness under hemorrhagic conditions mimicked by thrombin treatment, and this was not blocked by MSC-conditioned medium (MSC-CM) in vitro. In contrast, thrombin-induced microglial activation and release of proinflammatory cytokines were inhibited by MSC-CM. Interestingly, astrocytes showed greater reactive response when co-cultured with microglia, and this was abolished in the presence of MSC-CM. Gene expression profiles in microglia revealed that transcript levels of genes for immune response and proinflammatory cytokines were altered by thrombin treatment. This result coincided with the robust phosphorylation of STAT1 and p38 MAPK, which might be responsible for the production and release of proinflammatory cytokines. Furthermore, application of MSC-CM diminished thrombin-mediated phosphorylation of STAT1 and p38 MAPK, supporting the acute anti-inflammatory role of MSCs under hemorrhagic conditions. In line with this, activation of microglia and consequent cytokine release were impaired in Stat1-null mice. However, reactive response in Stat1-deficient astrocytes was maintained. Taken together, our results demonstrate that MSCs mainly block the activation of microglia involving STAT1-mediated cytokine release and subsequent reduction of reactive astrocytes.

Cdk5 regulates N-cadherin-dependent neuronal migration during cortical development.

Cyclin-dependent kinase 5 (Cdk5) controls neuronal migration in the developing cortex when multipolar newborn neurons transform to become bipolar. However, by which mechanisms Cdk5 controls cell adhesion in migrating neurons are not fully understood. In this study, we examined the functional interaction between Cdk5 and N-cadherin (Ncad) in newborn neurons when they undergo the multipolar to bipolar transition in the intermediate zone (IZ). Detailed expression analysis revealed that both Cdk5 and Ncad were present in GFP-electroporated migrating neurons in the IZ. Misexpression of dominant negative Cdk5 into the embryonic brains stalled neuronal locomotion in the lower IZ in which arrested cells were round or multipolar. When Ncad was co-introduced with Cdk5DN, however, cells continue to migrate into the cortical plate (CP) and migrating neurons acquired typical bipolar morphology with a pia-directed leading process. Similarly, downregulation of CDK5 resulted in lesser aggregation ability, reversed by the expression of Ncad in vitro. Down-regulation of activity or protein level of CDK5 did not alter the total amount of NCAD proteins but lowered its surface expression in cells. Lastly, expression of CDK5 and NCAD overlapped in the IZ of the human fetal cortex, indicating that the role of Cdk5 and Ncad in neuronal migration is evolutionarily conserved.

ISL1-based LIM complexes control Slit2 transcription in developing cranial motor neurons.

LIM-homeodomain (HD) transcription factors form a multimeric complex and assign neuronal subtype identities, as demonstrated by the hexameric ISL1-LHX3 complex which gives rise to somatic motor (SM) neurons. However, the roles of combinatorial LIM code in motor neuron diversification and their subsequent differentiation is much less well understood. In the present study, we demonstrate that the ISL1 controls postmitotic cranial branchiomotor (BM) neurons including the positioning of the cell bodies and peripheral axon pathfinding. Unlike SM neurons, which transform into interneurons, BM neurons are normal in number and in marker expression in Isl1 mutant mice. Nevertheless, the movement of trigeminal and facial BM somata is stalled, and their peripheral axons are fewer or misrouted, with ectopic branches. Among genes whose expression level changes in previous ChIP-seq and microarray analyses in Isl1-deficient cell lines, we found that Slit2 transcript was almost absent from BM neurons of Isl1 mutants. Both ISL1-LHX3 and ISL1-LHX4 bound to the Slit2 enhancer and drove endogenous Slit2 expression in SM and BM neurons. Our findings suggest that combinations of ISL1 and LHX factors establish cell-type specificity and functional diversity in terms of motor neuron identities and/or axon development.

Contralateral migration of oculomotor neurons is regulated by Slit/Robo signaling.

BACKGROUND: Oculomotor neurons develop initially like typical motor neurons, projecting axons out of the ventral midbrain to their ipsilateral targets, the extraocular muscles. However, in all vertebrates, after the oculomotor nerve (nIII) has reached the extraocular muscle primordia, the cell bodies that innervate the superior rectus migrate to join the contralateral nucleus. This motor neuron migration represents a unique strategy to form a contralateral motor projection. Whether migration is guided by diffusible cues remains unknown. METHODS: We examined the role of Slit chemorepellent signals in contralateral oculomotor migration by analyzing mutant mouse embryos. RESULTS: We found that the ventral midbrain expresses high levels of both Slit1 and 2, and that oculomotor neurons express the repellent Slit receptors Robo1 and Robo2. Therefore, Slit signals are in a position to influence the migration of oculomotor neurons. In Slit 1/2 or Robo1/2 double mutant embryos, motor neuron cell bodies migrated into the ventral midbrain on E10.5, three days prior to normal migration. These early migrating neurons had leading projections into and across the floor plate. In contrast to the double mutants, embryos which were mutant for single Slit or Robo genes did not have premature migration or outgrowth on E10.5, demonstrating a cooperative requirement of Slit1 and 2, as well as Robo1 and 2. To test how Slit/Robo midline repulsion is modulated, we found that the normal migration did not require the receptors Robo3 and CXCR4, or the chemoattractant, Netrin 1. The signal to initiate contralateral migration is likely autonomous to the midbrain because oculomotor neurons migrate in embryos that lack either nerve outgrowth or extraocular muscles, or in cultured midbrains that lacked peripheral tissue. CONCLUSION: Overall, our results demonstrate that a migratory subset of motor neurons respond to floor plate-derived Slit repulsion to properly control the timing of contralateral migration.

Light-induced Notch activity controls neurogenic and gliogenic potential of neural progenitors.

Oscillations in Notch signaling are essential for reserving neural progenitors for cellular diversity in developing brains. Thus, steady and prolonged overactivation of Notch signaling is not suitable for generating neurons. To acquire greater temporal control of Notch activity and mimic endogenous oscillating signals, here we adopted a light-inducible transgene system to induce active form of Notch NICD in neural progenitors. Alternating Notch activity saved more progenitors that are prone to produce neurons creating larger number of mixed clones with neurons and progenitors in vitro, compared to groups with no light or continuous light stimulus. Furthermore, more upper layer neurons and astrocytes arose upon intermittent Notch activity, indicating that dynamic Notch activity maintains neural progeny and fine-tune neuron-glia diversity.

Chemoradiation with paclitaxel and carboplatin in high-risk cervical cancer patients after radical hysterectomy: a Korean Gynecologic Oncology Group study.

PURPOSE: To evaluate the efficacy and toxicity of concurrent chemoradiation with paclitaxel and carboplatin in patients with high-risk cervical cancer. METHODS AND MATERIALS: Patients after radical hysterectomy for cervical cancer, with at least 1 high-risk characteristic, were administered paclitaxel 135 mg/m(2), carboplatin area under the curve = 5 every 3 weeks for 3 cycles concomitant with radiation therapy as adjuvant treatment. RESULTS: This prospective study enrolled 71 consecutive patients. Sixty-six patients (93%) completed the planned treatment. The majority of grade 3/4 neutropenia or nonhematologic toxicities were usually self-limited. Diarrhea grades 3/4 were observed in 4 patients (5.6%). One patient developed anaphylactic shock after infusion of paclitaxel. With a median follow-up of 57 months, recurrences occurred in 16 patients. Multivariable analysis indicated that common iliac lymph node involvement is an independent risk factor for disease recurrence (odds ratio 13.48; 95% confidence interval 2.93-62.03). In the intent-to-treat population (n=71), the estimated 5-year disease-free survival and overall survival rates were 77.3% and 80.3% respectively. In the per-protocol population (n=62), disease-free survival was 78.9% and overall survival was 83.9%. CONCLUSIONS: Concurrent chemoradiation with paclitaxel/carboplatin is well tolerated and seems to be effective for patients who undergo radical hysterectomy. Therefore, a prospective, randomized controlled study should be designed to evaluate efficacy of this approach for patients with high-risk cervical cancer.

Axonal neuropathy-associated TRPV4 regulates neurotrophic factor-derived axonal growth.

Spinal muscular atrophy and hereditary motor and sensory neuropathies are characterized by muscle weakness and atrophy caused by the degenerations of peripheral motor and sensory nerves. Recent advances in genetics have resulted in the identification of missense mutations in TRPV4 in patients with these hereditary neuropathies. Neurodegeneration caused by Ca(2+) overload due to the gain-of-function mutation of TRPV4 was suggested as the molecular mechanism for the neuropathies. Despite the importance of TRPV4 mutations in causing neuropathies, the precise role of TRPV4 in the sensory/motor neurons is unknown. Here, we report that TRPV4 mediates neurotrophic factor-derived neuritogenesis in developing peripheral neurons. TRPV4 was found to be highly expressed in sensory and spinal motor neurons in early development as well as in the adult, and the overexpression or chemical activation of TRPV4 was found to promote neuritogenesis in sensory neurons as well as PC12 cells, whereas its knockdown and pharmacologic inhibition had the opposite effect. More importantly, nerve growth factor or cAMP treatment up-regulated the expression of phospholipase A(2) and TRPV4. Neurotrophic factor-derived neuritogenesis appears to be regulated by the phospholipase A(2)-mediated TRPV4 pathway. These findings show that TRPV4 mediates neurotrophic factor-induced neuritogenesis in developing peripheral nerves. Because neurotrophic factors are essential for the maintenance of peripheral nerves, these findings suggest that aberrant TRPV4 activity may lead to some types of pathology of sensory and motor nerves.

Clinicopathologic characteristics of granulosa cell tumors of the ovary: a multicenter retrospective study.

OBJECTIVE: To evaluate the clinicopathologic characteristics and prognostic factors of ovarian granulosa cell tumors. METHODS: Medical records of 113 patients presenting between January 1995 and December 2007 were retrospectively reviewed. RESULTS: One-hundred two patients had adult type disease, with a mean age of 46.2 years (range, 18 to 83 years) and a mean follow-up period of 54.7 months (range, 1 to 155 months). The distribution of FIGO stages was 86 patients at stage I, 11 at stage II, and 5 at stage III. During follow-up, ten patients recurred at a mean time of 48 months (range, 4 to 109 months). Among them, three patients died after a mean of 57 months (range, 25 to 103 months). In recurrence analysis, advanced stage (p=0.032) and presence of residual disease (p=0.012) were statistically significant, and age<40 years, premenopause and positive washing cytology were marginally significant (p<0.1). In multivariate analysis, stage was the only factor associated with recurrence; adjuvant chemotherapy and fertility-sparing surgery were not statistically significant. Among 36 patients with fertility-sparing operations, eight patients had nine pregnancies and delivered seven babies. Eleven patients had juvenile type tumors; the mean age was 20.0 years (range, 8 to 45 years) and the mean follow-up period was 69.8 months (range, 20 to 156 months). The distribution of FIGO stage was nine patients at stage I and two at stage III. There were no recurrences or deaths reported. Four patients had seven pregnancies and delivered six babies. CONCLUSION: Stage is the only factor associated with disease-free survival, and fertility-sparing surgery may be a treatment option for women with early-stage disease who want to retain fertility.

Vaccination with a human papillomavirus (HPV)-16/18 AS04-adjuvanted cervical cancer vaccine in Korean girls aged 10-14 years.

The human papillomavirus (HPV)-16/18 AS04-adjuvanted cervical cancer vaccine has been demonstrated to be highly efficacious and immunogenic with a favorable safety profile. This study assessed the immunogenicity and safety of the HPV-16/18 AS04-adjuvanted vaccine in healthy Korean girls aged 10-14 yr. This multi-center, observer-blind trial randomly assigned 321 healthy girls to receive three doses (0, 1, 6-month schedule) of HPV-16/18 AS04-adjuvanted vaccine or hepatitis A vaccine. Immunogenicity against vaccine antigens was assessed one month post-Dose 3. Solicited and unsolicited adverse events (AEs) and serious AEs (SAEs) were recorded. In the according-to-protocol analysis, all initially seronegative subjects vaccinated with the HPV-16/18 AS04-adjuvanted vaccine had seroconverted at Month 7, with a peak geometric mean titer (GMT) that was 600-fold higher than the natural infection titer of 29.8 EU/mL for HPV-16 and a peak GMT that was 400-fold higher than the natural infection titer of 22.6 EU/mL for HPV-18. The vaccine was well tolerated with no increase in reactogenicity with subsequent doses and no reports of vaccine-related SAEs. In conclusion, the HPV-16/18 AS04-adjuvanted vaccine is shown to be highly immunogenic and generally well-tolerated in Korean girls aged 10-14 yr.

Clinicopathologic characteristics of malignant germ cell tumors in the ovaries of Korean women: a Korean Gynecologic Oncology Group Study.

From January 1995 to December 2000, medical records of 196 patients were collected from 14 hospitals nationwide and were reviewed retrospectively. We evaluated the clinicopathologic characteristics of malignant germ cell tumors in the ovaries of South Korean women and determined the prognostic factors affecting recurrence. The mean patient age was 23.8 years (range, 4-63 years), and the median follow-up period was 67 months (range, 1-128 months). The distribution of the International Federation of Gynecology and Obstetrics stage was as follows: 128 cases (65.3%) in stage I, 27 cases (13.8%) in stage II, 39 cases (19.9%) in stage III, and 2 cases (1.0%) in stage IV. Histologically, immature teratoma was the most common tumor type (n = 68), followed by dysgerminoma (n = 54), endodermal sinus tumor (n = 38), mixed form (n = 24), and choriocarcinoma (n = 12). A fertility-sparing operation was performed in 134 patients, staging operation in 43 patients, and hysterectomy and bilateral salpingo-oophorectomy in 19 patients. Postoperative chemotherapy was administered in 166 patients, and the most common regimen was bleomycin, etoposide, and cisplatin (n = 120). Recurrence was observed in 13 patients (6.8%) and was influenced by the stage of the tumor and patient age (>40 years). The 5-year survival rate was 96.7%. During the follow-up period, 20 patients had 22 pregnancies that resulted in 17 normal deliveries at term and 5 abortions. The results of this study demonstrate that most malignant germ cell tumors of the ovary in Korean women are detected in the early stage and have excellent survival outcomes with conservative operation and platinum-based chemotherapy.

Cervical cancer associated with pregnancy: results of a multicenter retrospective Korean study (KGOG-1006).

OBJECTIVE: The objective of the study was to analyze the characteristics of cervical cancer associated with pregnancy. STUDY DESIGN: Forty patients with cervical cancer associated with pregnancy were retrospectively identified between 1995-2003. Three controls for each case were matched on the basis of age, stage, histology, and date of treatment. RESULTS: Sampling of cervical cytology after the second trimester was the most common cause of delayed diagnosis. Among 12 patients who delayed treatment for fetal maturity, 2 died of disease. There was no difference in overall survival between pregnant and nonpregnant patients with stage Ib tumors. In contrast to nonpregnant patients, the depth of stromal invasion was not correlated with the incidence of lymph vascular space involvement and lymph node metastasis in pregnant patients. CONCLUSION: Thorough evaluation is warranted before deciding whether to delay treatment until fetal maturity. Pregnancy does not adversely affect the prognosis of early-stage cervical cancer significantly.