Tadalafil is a potent selective phosphodiesterase 5 inhibitor used to treat pulmonary arterial hypertension. As tadalafil is a substrate of CYP3A, coadministration with a CYP inducer or inhibitor may affect the pharmacokinetics and pharmacological activity of tadalafil. Additionally, CYP expression is regulated by biological clocks and its activity fluctuates diurnally. Moreover, tadalafil's dosing time may also affect its pharmacokinetics. Therefore, this study aimed to investigate the effects of phenobarbital and tadalafil dosing time on the plasma concentration and pharmacological activity of tadalafil. Adult male ICR mice were treated with phenobarbital for 5 days, followed by oral administration of tadalafil perorally. Phenobarbital pretreatment decreased plasma tadalafil and pulmonary cGMP levels in a dose-dependent manner. In mice without phenobarbital pretreatment, the plasma tadalafil concentration tended to be higher when administered in the morning than in the evening. In contrast, plasma tadalafil concentration was higher when administered in the evening than in the morning in mice pretreated with phenobarbital. Similarly, pulmonary cGMP levels were higher when tadalafil was administered in the evening than in the morning in mice pretreated with phenobarbital. Notably, these effects on metabolism and pharmacological activity were observed at clinically relevant doses. Conclusively, these results indicate that the coadministration of phenobarbital and tadalafil may suppress the plasma levels and pharmacological activities of tadalafil. Additionally, dosing time should be carefully considered to maximize the efficacy of tadalafil.

Various molecules have been reported to be involved in the induction of HL-60 cell differentiation by all-trans retinoic acid (ATRA). Notably, the transfection of an antisense oligonucleotide targeting proteinase 3 (PRTN3, formerly known as myeloblastin) into HL-60 cells induces differentiation, though the underlying mechanism remains unclear. Here we found that the levels of secreted PRTN3 were significantly reduced when differentiation was initiated by ATRA treatment, suggesting that secreted PRTN3 inhibits HL-60 cell differentiation. The addition of anti-PRTN3 antibody to the culture medium of HL-60 cells promoted their differentiation. Conversely, when ATRA-treated HL-60 cells were cultured in conditioned media prepared from HEK293 cells transfected with the preproPRTN3 plasmid, differentiation was significantly suppressed. Moreover, conditioned media from HEK293 cells transfected with an inactive PRTN3 mutant (S203G) also suppressed HL-60 cell differentiation. These results suggest that PRTN3 secreted from HL-60 cells has an autocrine inhibitory effect on differentiation, independent of its enzymatic activity.

Introduction: Cephalosporins are widely used antimicrobials; however, their potential to induce coagulopathy-related events, particularly hypoprothrombinemia, raises safety concerns. Methods: This study aimed to comprehensively evaluate the risk of coagulopathy-related events associated with the use of intravenous cephalosporins using the Japanese Adverse Drug Event Report (JADER) database. We analyzed 783,788 cases from the JADER database to identify coagulopathy-related events associated with the use of eleven intravenous cephalosporins between 2004 and 2024. Reporting odds ratios (ROR) with 95% confidence intervals were calculated to detect potential signals. Subsequently, odds ratios were calculated to assess the influence of age and sex on the occurrence of coagulopathy-related events. Results: Cefmetazole (CMZ), cefoperazone/sulbactam (CPZ/SBT), flomoxef (FMOX), and ceftriaxone (CTRX) showed statistically significant signals for coagulopathy-related events (ROR: 10.87, 5.20, 2.67, and 1.38, respectively). Age emerged as a significant factor, with individuals aged >70 years exhibiting higher odds of experiencing coagulopathy-related events associated with CMZ, CPZ/SBT, FMOX, and CTRX use. However, no significant association was observed with sex. Our findings suggest that certain intravenous cephalosporins (CMZ, CPZ/SBT, FMOX, and CTRX) are associated with an increase of coagulopathy-related events, particularly in patients aged >70 years. Conclusions: These findings highlight the need for vigilance and careful consideration of patient-specific factors when prescribing these antibiotics.

To clarify the factors contributing to the interictal burden of migraine, we conducted a questionnaire survey among women with migraine. A questionnaire survey was conducted on the Internet of 400 women between the ages of 20 and 50 who tested positive on the migraine screener. The subjects were divided into a high burden group (n = 260) and a low burden group (n = 140) according to their Migraine Interictal Burden Scale-4 (MIBS-4) score. Using multivariate stepwise logistic regression analysis, we demonstrated that Headache Impact Test-6 (HIT-6) score, type of migraine, the causes of headaches (weather changes and anxiety) and Epworth sleepiness scale (ESS) score influenced to the interictal burden of migraine. Odds ratios of HIT-6 score, type of migraine, weather changes and anxiety in causes of headaches and ESS score were 3.669 (≤55 vs. ≥56; 95% CI = 1.900 – 7.090), 2.327 (migraine without aura vs. migraine with aura; 95% CI = 1.300 – 4.168), 1.810 (yes vs. no; 95% CI = 1.086 – 3.015), 1.703 (no vs. yes; 95% CI = 1.009 – 2.874) and 1.625 (≤10 vs. ≥11; 95% CI = 1.028 – 2.568), respectively. Our findings suggest that multiple factors independently contribute to the interictal burden of migraine.

Three flame retardants, tris(1-aziridinyl) phosphine oxide (APO), tris(2,3-dibromopropyl) phosphate (TDBPP), and bis(2,3-dibromopropyl) phosphate (BDBPP), are prohibited for use in textile products under the “Act on the Control of Household Products Containing Harmful Substances” in Japan. The currently used method for analyzing these substances by the Ministry of Health, Labor and Welfare in Japan (MHLWJ), is GC, which uses carcinogenic solvents and hazardous reagents during sample processing. Furthermore, due to the global shortage of helium gas used as carrier gas for GC, the development of an alternative method is necessary. In this study, we developed a method to simultaneously analyze the three flame retardants using LC–MS/MS. In addition, this method is an improvisation of ISO 17881-2, which involves extraction with acetone under acidic conditions and replacing it with acetonitrile. This prevents the acid decomposition of APO and the thermal decomposition of BDBPP. For accurate and sensitive quantification, deuterated compounds, APO-d₁₂, TDBPP-d₁₅, and BDBPP-d₁₀, were used as surrogate standards. The calibration curve displayed linearity within the 0.005–1.0 µg/mL range for APO and the 0.01–2.0 µg/mL range for TDBPP and BDBPP. The limits of detection for APO, TDBPP, and BDBPP were 0.19, 0.87, and 1.0 µg/g, respectively. These values were 2.2, 9.2, and 10 times more sensitive than the current detection limits of 0.4, 8, and 10 µg/g, respectively. The recovery of the three flame retardants in various textile products using the developed analytical method was 83.7–120.8%, which confirms their satisfactory analyses.

USP and MHLW-PMDA held a joint workshop on September 10-11, 2024 in Tokyo, Japan¹⁾. This workshop was organized by the United States Pharmacopeia (USP) in collaboration with Japan’s Ministry of Health, Labour, and Welfare (MHLW) and the Pharmaceuticals and Medical Devices Agency (PMDA). This workshop aimed to address critical issues in pharmaceutical quality and foster international collaboration between our two organizations and critical stakeholders. The workshop covered a range of topics, including the harmonization of standards with a specific focus on the inaugural pilot between USP and JP on the harmonization of a drug substance monograph and a drug product monograph, as well as on ICH Q2(R2)/Q14. Participants discussed advancements in quality testing, particularly for high-risk excipients like ethylene glycol and diethylene glycol, and shared insights on managing contamination issues. The event also explored the integration of emerging technologies such as quantitative nuclear magnetic resonance (qNMR) and the challenges associated with complex generics. Overall, the USP-MHLW/PMDA joint workshop underscored the commitment of both organizations to advancing public health through the development and harmonization of high-quality standards. By fostering collaboration and innovation, the workshop aimed to enhance the safety, efficacy, and accessibility of medicines globally.

Dry eye disease is an ocular disease in which the stability of tear fluid decreases, causing ocular discomfort and abnormal visual function as well as damage to the ocular surface. It has been reported that specific types of food ingredients can promote lachrymal secretion, which is expected to prevent and improve dry eye. Here, we evaluated the effects of a heat-killed form of Fructobacillus fructosus FMO-85, a species of fructophilic lactic acid bacteria (FLAB) that is derived from the digestive tract of honeybees, on lacrimal fluid secretion using a stress-induced dry eye mouse model. Male C57BL/6J mice were fed a 3% FLAB-mixed diet 3 weeks before stress loading. We observed that the tear fluid volume was decreased after stress loading, which was significantly improved by FLAB treatment after 7 and 11 days. Mechanistically, the mRNA levels of brain-derived neurotrophic factor (Bdnf), one of the important growth factors involved in lacrimal fluid secretion, isoform-2 and -6 were increased in the hippocampus of FLAB-treated mice. Furthermore, the plasma levels of an anti-inflammatory cytokine, interleukin-10 (IL-10), increased in the FLAB-treated mice. These results suggested that the ingredients contained in dried FLAB increase tear fluid volume by affecting the lacrimal secretion mechanism and the production of inhibitory cytokines. In conclusion, the decrease in tear fluid volume after stress loading was suppressed by FLAB intake. These results indicated that FLAB supplementation may be a useful strategy for the prevention and treatment of dry eye.

The Committee on Sick House Syndrome: Indoor Air Pollution, established by the Ministry of Health, Labour and Welfare of Japan, is reviewing indoor air quality guidelines. A comprehensive exposure assessment is essential for pollutants with revised guideline values or newly developed candidate pollutants, necessitating the development of standardized test methods for an accurate evaluation. However, the available test methods that have been provided as a standard test method (measurement manual) were introduced over 20 years ago. Its applicability to pollutants for which guideline values have been established since then had not been examined. Therefore, we established a test method for six compounds based on the current guideline values and three candidate compounds that underwent initial risk assessment. This method considered the new guideline values established after 2001 using solid-phase adsorption-thermal desorption-gas chromatography/mass spectrometry, as indicated in the measurement manual for volatile organic compounds. This method was validated at four institutions using samples at approximately 1/10^th the concentration of the current, revised, and newly proposed guideline values, as of 2017. Results revealed that the average recovery of the four laboratories ranged from 84.2 to 95.6%, the repeatability ranged from 0.43 to 16%, which was <20%, thereby effectively achieving the target evaluation criteria. Therefore, this method could be presented as a standard test method for nine volatile organic compounds.

To avoid harm caused by falsified medicines, we aimed to devise a method to identify falsified medicines in Japan using visual observation among medicines obtained from personal import agency websites, which are the main conduits through which falsified medicines are obtained. We recorded details regarding the information provided on personal import agency websites used to purchase medicines, the outer package received, the customs declaration description, and the product appearance for 212 samples of medicines obtained through personal import via the Internet. We investigated the relationship between each observed item and the rate of falsified medicines. We developed a classification and prediction model to identify falsified medicines using items that could be visually observed. The results showed that the rate of falsified medicines was significantly higher for websites that did not contain identifying information such as the name and address of the contact or import agency, as well as for products that did not contain the name and address of the manufacturer, indicating that these items may be useful in the identification of falsified medicines. In the prediction model constructed, we extracted features such as the country of dispatch and address of the import agency, and a prediction model was created to identify falsified medicines and websites selling these medicines. Careful observation of the identified features and use of our prediction model will help to prevent harm owing to the use of falsified medicines.

Bisulfite modification of cytosine residues is a widely used method for analyzing genomic DNA methylation. Despite its robustness, degradation of DNA during modification has hampered the application to the analysis of small amounts of DNA. We show that the addition of large volume bisulfite-treated heterogeneous DNA promotes the amplification of the targeted region in bisulfite-treated DNA by PCR. The addition of untreated DNA did not promote the amplification. The addition of large volume bisulfite-treated heterogeneous DNA neither promoted the amplification when untreated DNA was used as template. PCR products were detected when a ten thousandth of aliquot of bisulfite-treated 1 μg of human genomic DNA (0.1 ng) was used as template. This figure is equivalent to that of human genomic DNA derived from as little as 15 cells. This procedure permits the analysis of genomic DNA methylation when only limited numbers of cells are available.

Semaphorin family belongs to secreted or membrane anchored proteins. Recent studies have demonstrated the involvement of semaphorins and their receptors in cancer biology, but it remains unclear how each semaphorin molecule regulates tumorigenesis. Previously we reported that Semaphorin 3A (Sema3A) and its receptor Plexin A1 (PlxnA1) regulate the malignant phenotypes of mouse-derived lewis lung cancer (LLC) cells constitutively expressing GFP (LLC-GFP). Here we show that Semaphorin 6B (Sema6b) serves as one of the oncogenic semaphorin molecules in lung cancer using LLC-GFP. Sema3a or Plxna1 knockdown downregulated Sema6b, and their suppressive effect on proliferation was significantly recovered by recombinant SEMA6B (rSEMA6B) treatment. Sema6b knockdown suppressed the proliferation and tumorigenicity of LLC-GFP. Interestingly, the self-renewal capacity of LLC-GFP derived cancer stem-like cells (LLC-GFPstem) was completely lost by Sema6b knockdown. These results demonstrate that Sema6B would be the novel therapeutic target of lung cancer.

Iron is an essential nutrient for bacterial survival. Vibrio alginolyticus is a pathogenic Vibrio species that produces vibrioferrin, a cognate siderophore for efficient iron acquisition in iron-limited environments. Many bacteria have developed mechanisms to utilize xenosiderophores produced by other microorganisms, in addition to using self-produced siderophores for iron acquisition. In this study, we found through a homology search using the whole genome sequence of V. alginolyticus NBRC 15630 that this bacterium has genes similar to those involved in the utilization of hydroxamate-based xenosiderophores, desferri-ferrichrome (DFC), desferrioxamine B (DFOB), and aerobactin (AERO), possessed by V. parahaemolyticus, V. cholerae, and V. vulnificus. In growth assays using an iron-limiting medium supplemented with each xenosiderophore, it was found that the N646_3157 (fhuA1), N646_0489 (fhuA2), N646_0777 (desA), and N646_4356 (iutA) are outer membrane receptor genes involved in the utilization of DFC, DFOB, and AERO and N646_3158-3160 (fhuC1D1B1), and N646_0486-0488 (fhuC2D2B2) are ATP-binding cassette transporter genes for both DFC and DFOB. Additionally, we demonstrated by reverse transcriptase-quantitative PCR and electrophoretic mobility shift assay that the fhuC2D2B2A2 genes, which were newly identified in pathogenic Vibrio species, are an operon whose expression is probably regulated by Fur in response to iron availability.

OSW-1, a promising compound that is toxic to diverse tumor cell lines, is a saponin from Ornithogalum saundersia. In this study, we analyzed the stress responses induced by OSW-1 using the human colon cancer cell line HT-29 cells and compared it with the commonly used ER and Golgi stress inducers brefeldin A (BFA), thapsigargin (Tg), and tunicamycin (Tm). OSW-1 induced few ER stress-related factors, but there was an increase in expression of TFE3 protein, one of the Golgi stress response factors. A shift in the molecular weight of TFE3 was also found, likely attributable to dephosphorylation. Conversely, the impact of OSW-1 on the expression of the TFEB protein, another member of the MiTF/TFE family, was minimal. Cleavage of CREB3, another Golgi stress sensor, was apparently induced only by BFA. LC3-II and p62, autophagy-related factors, were increased in all drug treatments. Unexpectedly, OSBP protein levels, one of the targets of OSW-1, were increased by not only three reagents but also OSW-1. Taken together, OSW-1 treatment of HT-29 cells induced atypical Golgi stress that strongly activated the TFE3 pathway and did not involve the CREB3 pathway or the ER stress response. Although OSW-1 was also found to affect the autophagy system, it was suggested that the effects of OSW-1 may not be mediated by OSBP depletion. These findings will contribute to the development of OSW-1-based cancer therapies and to our understanding of Golgi stress responses.