Kei Tashiro

Prostate cancer (PCa) is a leading malignancy in men, and understanding its molecular mechanisms is crucial for advancing therapeutic strategies. Ubiquitination, a key post-translational modification, regulates protein degradation and signaling, playing a vital role in cancer progression. This study focuses on HECTD4, a HECT-type E3 ubiquitin ligase, to identify its ubiquitination targets and understand its role in PCa. HECTD4 knockdown was performed in LNCaP, PC-3, and DU145 PCa cell lines. A combination of semi-quantitative PCR and liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to identify proteins with altered expression and ubiquitination profiles. Gene ontology analysis, pathway analysis, and a proliferation assay were conducted to explore the biological significance of HECTD4. We identified 1,605 downregulated and 1,736 upregulated proteins upon HECTD4 knockdown. Key proteins involved in tumor suppression and cell cycle regulation, such as NUSAP1, CDK6, and MED13L, were ubiquitinated by HECTD4. Functional annotations revealed that these targets are associated with critical pathways, including phosphoinositide 3-kinase (PI3K)-AKT, Ras-mitogen-activated protein kinase (MAPK), and mammalian target of rapamycin (mTOR), as well as immune infiltration, drug response, and survival analysis. HECTD4 regulates protein stability and activation through ubiquitination, impacting cell cycle progression, tumor suppression, and immune response in PCa. These findings suggest that HECTD4 is a promising therapeutic target, with potential applications in drug development aimed at disrupting oncogenic signaling and enhancing treatment efficacy.

Background/Aim: Identification of cancer biomarkers for early detection is required. However, little is known about which candidate cell signaling pathway markers can be identified and which pathways may serve as therapeutic targets. We focused on the disulfidptosis among numerous signaling pathways, because it is a mechanism that causes cell death and is associated with iron-dependent cell death or ferroptosis, the tricarboxylic acid cycle, energy metabolism, and glucose uptake. The aim of the study was to detect the disulfidptosis-linked gene signatures associated with stage-specific makers and prognosis. Materials and Methods: We examined the expression of 106 related genes in 324 patients with prostate cancer for disulfidptosis, a type of cell death triggered by disulfide stress resulting in disulfide bond-induced collapse of the cytoskeleton. Results: The expression levels of UBASH3B, ANP32E, PRC1, ACTB, SPG20, and DBN1 increased with cancer progression. Of these, UBASH3B, PRC1, and ANP32E were strongly expressed in cases with Gleason score ≥8. Conversely, the expression levels of MYH13, FLNC, GLUD1, SAMM50, CHCHD3, and CAPZB decreased. Of these, GLUD1, CAPZB, and SAMM50 were decreased in cases with Gleason score ≥8. In addition, UBASH3B, ANP32E, PRC1, DBN1, FLNC, and GLUD1 enabled the estimation of biochemical recurrence (BCR)-free survival. In particular, the prognostic formula comprising ZHX2, SMPD4, and CHD4 using the Lasso-Cox regression model properly distinguished the BCR-free survival curves, indicating that these genes could be signatures for disulfidptosis. Conclusion: Decoding disulfidptosis-related data in the transcriptome would provide crucial clues for finding novel approaches to personalized cancer medicine in prostate cancer.

Identification of cancer biomarkers for early detection is required. However, little is known about which candidate cell signaling pathway markers can be identified and which pathways may serve as therapeutic targets. We focused on the disulfidptosis among numerous signaling pathways, because it is a mechanism that causes cell death and is associated with iron-dependent cell death or ferroptosis, the tricarboxylic acid cycle, energy metabolism, and glucose uptake. The aim of the study was to detect the disulfidptosis-linked gene signatures associated with stage-specific makers and prognosis. We examined the expression of 106 related genes in 324 patients with prostate cancer for disulfidptosis, a type of cell death triggered by disulfide stress resulting in disulfide bond-induced collapse of the cytoskeleton. The expression levels of UBASH3B, ANP32E, PRC1, ACTB, SPG20, and DBN1 increased with cancer progression. Of these, UBASH3B, PRC1, and ANP32E were strongly expressed in cases with Gleason score ≥8. Conversely, the expression levels of MYH13, FLNC, GLUD1, SAMM50, CHCHD3, and CAPZB decreased. Of these, GLUD1, CAPZB, and SAMM50 were decreased in cases with Gleason score ≥8. In addition, UBASH3B, ANP32E, PRC1, DBN1, FLNC, and GLUD1 enabled the estimation of biochemical recurrence (BCR)-free survival. In particular, the prognostic formula comprising ZHX2, SMPD4, and CHD4 using the Lasso-Cox regression model properly distinguished the BCR-free survival curves, indicating that these genes could be signatures for disulfidptosis. Decoding disulfidptosis-related data in the transcriptome would provide crucial clues for finding novel approaches to personalized cancer medicine in prostate cancer.

Background/Aim: Prostate cancer (PCa) is the most frequently diagnosed cancer in men and a leading cause of cancer-related death. While prostate-specific antigen is a widely used biomarker, its specificity is limited. This study investigated the prognostic significance of gene subsets associated with the ubiquitin-proteasome pathway in PCa. Materials and Methods: We analyzed transcriptomic and clinical data of 94 early-onset (age IKBKB, UBQLN3, TMUB2, UBE2S, and BRCA1 was observed at relative-early stages of pT3a and Gleason 3+4. Similarly, HERPUD1, CDC20, UHRF1, PSMD7, PIAS3, MALT1, TNF, UBD, CD3E, CD247, SOCS1, UBE2C, CARD16, ZAP70, UBA7, and UBE3C expression levels also changed at pT3b and Gleason 4+3. At metastatic stages (pT4 and Gleason ≥8) OASL expression was up-regulated, whereas that of DDB1, RPN1, UBE3B, UBE2H, PPIL2, WWP2, and CDH1 was down-regulated. In addition, higher expression of PSMD2, CDC20, NFKB1, and STIP1 or lower expression of HERPUD2, NEDD4, ANAPC16, LNX1, and HERPUD1 was associated with poor prognoses according to the Kaplan–Meier or receiver operating characteristic analyses for biochemical recurrence-free survival. A LASSO-Cox model identified six gene candidates including LNX1, PSMD2, SUMO4, UBE2C, UBR5, and UHRF1. Conclusion: The identified gene subset provides novel prognostic insights into PCa progression and survival. These findings highlight potential biomarkers and therapeutic targets within the ubiquitin-proteasome pathway, offering new avenues for personalized treatment strategies.

In this study, we analyzed a relatively large subset of proteins, including 109 kinds of blood-circulating cytokines, and precisely described a cytokine storm in the expression level and the range of fluctuations during hospitalization for COVID-19. Of the proteins analyzed in COVID-19, approximately 70% were detected with Bonferroni-corrected significant differences in comparison with disease severity, clinical outcome, long-term hospitalization, and disease progression and recovery. Specifically, IP-10, sTNF-R1, sTNF-R2, sCD30, sCD163, HGF, SCYB16, IL-16, MIG, SDF-1, and fractalkine were found to be major components of the COVID-19 cytokine storm. Moreover, the 11 cytokines (i.e., SDF-1, SCYB16, sCD30, IL-11, IL-18, IL-8, IFN-γ, TNF-α, sTNF-R2, M-CSF, and I-309) were associated with the infection, mortality, disease progression and recovery, and long-term hospitalization. Increased expression of these cytokines could be explained in sequential pathways from hematopoietic progenitor cell differentiation to Th1-derived hyperinflammation in COVID-19, which might also develop a novel strategy for COVID-19 therapy with recombinant interleukins and anti-chemokine drugs.

In addition to its role in the treatment of pancreatitis, the serine protease inhibitor nafamostat exhibits a retinal protective effect. However, the exact mechanisms underlying this effect are unknown. In this study, the neuroprotective effects of nafamostat and its orally active derivative sepimostat against excitotoxicity were further characterised in vitro and in vivo. In primary rat cortical neurons, nafamostat completely suppressed N-methyl-D-aspartate (NMDA)-induced cell death. Intravitreal injection of nafamostat and sepimostat protected the rat retina against NMDA-induced degeneration, whereas the structurally related compounds, gabexate and camostat, did not. The neuroprotective effects of nafamostat and the NR2B antagonist ifenprodil were remarkably suppressed by spermidine, a naturally occurring polyamine that modulates the NR2B subunit. Both nafamostat and sepimostat inhibited [³H]ifenprodil binding to fractionated rat brain membranes. Thus, nafamostat and sepimostat may exert neuroprotective effects against excitotoxic retinal degeneration through NMDA receptor antagonism at the ifenprodil-binding site of the NR2B subunit.

Alexander disease (AxD) is an extremely rare neurodegenerative disorder caused by glial fibrillary acidic protein (GFAP) gene mutations. Compared with the cerebral type, which is characterized by infantile onset, the bulbospinal type and intermediate form are associated with a late onset, spanning from juveniles to the elderly, and more diverse clinical spectrum, suggesting the existence of factors contributing to phenotypic diversity. To build a foundation for future genetic studies of this rare disease, we obtained genomic data by whole exome-sequencing (WES) and DNA microarray derived from thirty-one AxD patients with the bulbospinal type and intermediate form. Using this data, we aimed to identify genetic variations determining the age at onset (AAO) of AxD. As a result, WES- or microarray-based association studies between younger (GAN, SLC1A2, CASP3, HDACs, and PI3K. Although we need to replicate the results using an independent population, this is the first step towards constructing a database, which may serve as an important tool to advance our understanding of AxD.

We investigated clinical and genetic characteristics of high-risk follicular lymphoma (FL), that lacked evidence of large cell transformation at diagnosis, in the rituximab era. First, we retrospectively analysed the clinical features of 100 patients with non-transformed FL that were consecutively treated with rituximab-containing therapies in a discovery cohort. The presence of either peripheral blood and/or bone involvement was associated with short progression-free survival. This was confirmed in a validation cohort of 66 FL patients. Then, whole exome sequencing was performed on randomly selected 5 high- and 9 standard-risk FL tumours. The most common mutational signature was a CG > TG substitution-enriched signature associated with spontaneous deamination of 5-methylcytosine at CpG, but mutations in WA and WRC(Y) motifs (so-called activation-induced cytidine deaminase (AID) motifs) were also enriched throughout the whole exome. We found clustered mutations in target sequences of AID in the IG and BCL2 loci. Importantly, high-risk FLs harboured more somatic mutations (mean 190 vs. 138, P = 0.04), including mutations in WA (33 vs. 22, P = 0.038), WRC (34 vs. 22, P = 0.016) and WRCY motifs (17 vs. 11, P = 0.004). These results suggest that genomic instability that allows for emergence of distinct mutations through AID activity underlies development of the high-risk FL phenotype.

Lymphoblastoid cell lines (LCLs) transformed by Epstein-Barr virus (EBV) serve as an unlimited resource of human genomic DNA. The protocol that is widely used to establish LCLs involves peripheral blood mononuclear cell isolation by density gradient centrifugation, however, that method requires as much as 5 ml of peripheral blood. In this study, in order to provide a more simple and efficient method for the generation of LCLs, we developed a new protocol using hemolytic reaction to enrich white blood cells for EBV transformation and found that the hemolytic protocol successfully generated LCLs from a small volume (i.e., 0.1 ml) of peripheral blood. To assess the quality of genomic DNA extracted from LCLs established by the hemolytic protocol (LCL-hemolytic), we performed single nucleotide polymorphism (SNP) microarray genotyping using the GeneChip^® 100 K Array Set (Affymetrix, Inc.). The concordances of the SNP genotyping resulting from genomic DNA from LCL-hemolytic (99.92%) were found to be as good as the technical replicate (99.90%), and Kappa statistics results confirmed the reliability. The findings of this study reveal that the hemolytic protocol is a simple and reliable method for the generation of LCLs, even from a small volume of peripheral blood.

We read with great interest the article by Ng et al. 1 entitled ''Genetic Association at the 9p21 Glaucoma Locus Contributes to Sex Bias in Normal-Tension Glaucoma,'' that was recently published in Investigative Ophthalmology & Visual Science. As the 9p21 locus was identified by a genome-wide association study (GWAS) to be associated with primary open-angle glaucoma (POAG), and the association was found to be significantly stronger among the normal-tension glaucoma (NTG) subgroup (for review see Ref. 2), Ng et al. 1 extended the analysis to investigate the influence of sex difference on the genetic association at the 9p21 locus. They carefully analyzed the genotype data of 13 variants linked with clinical information and clearly presented the results that: (1) the sex bias was present within advanced NTG cases, (2) the strongest associated variant in all POAG cases showed stronger association in females than in males with a statistically significant difference in female-to-male odds ratio (OR) comparison, and (3) the significant association was observed only in females under the NTG to high-tension glaucoma (HTG) subanalysis along with a significant difference in OR comparison. Although they used a large reliable number of samples and obtained robust results, they honestly pointed out the limitation of the