Efficacy and safety of TM5614 in combination with paclitaxel in the treatment of paclitaxel-resistant cutaneous angiosarcoma: Phase II study protocol.

Cutaneous angiosarcoma (CAS) is an endothelial cell-derived, highly aggressive type of vascular tumour. Although chemoradiotherapy with paclitaxel (PTX) is recognized as a first-line therapy for CAS, second-line therapy for CAS remains controversial, and there is no standard therapy for taxane-resistant CAS. Plasminogen activator inhibitor-1 (PAI-1) is associated with poor clinical outcomes, and elevated levels of PAI-1 in both tissue and serum are correlated with poor response to therapy in various cancers, including skin cancers. Since PAI-1 protects endothelial cells from Fas ligand-mediated apoptosis, PAI-1 inhibition might induce apoptosis of endothelial cell-derived tumours such as CAS. This is a single-arm, open-label, multi-institutional, Phase 2 clinical trial to assess the efficacy and safety of PTX in combination with TM5614 (PAI-1 inhibitor) in patients with PTX-resistant CAS. PTX will be administered for 28 weeks, with oral administration of TM5614. The primary endpoint of this study will be the overall response rate (ORR) at 28 weeks after starting treatment (central image evaluation). The secondary endpoint will include the ORR at 28 weeks after starting treatment (investigator evaluation), ORR at 8 weeks and 16 weeks after initiation of treatment (central and investigator evaluation), progression-free survival, overall survival, disease control rate and safety profiles. Assuming the null hypothesis of a response rate of 13.6% and an alternative hypothesis of 45%, a minimum of 15 patients are required to achieve a two-sided, Type I error of 5% and power of 70% based on the exact binomial distribution. Data quality control will be conducted by a combination of centralized (remote) and on-site monitoring. This study will contribute to the development of novel combination therapy for PTX-resistant CAS patients, which remains an unmet clinical need.

Phase II, multicenter study of plasminogen activator inhibitor-1 inhibitor (TM5614) plus nivolumab for treating anti-PD-1 antibody-refractory malignant melanoma: TM5614-MM trial.

BACKGROUND: Anti-programmed cell death 1 antibodies (anti-PD-1 Abs) are widely used for advanced melanoma, but the efficacy of an anti-PD-1 Abs is limited in the Asian population. There remains an unmet need to improve the therapeutic effects of anti-PD-1 Abs treatment, particularly in melanoma patients who are refractory to anti-PD-1 Abs. The aim was to evaluate anti-PD-1 Abs treatment in combination with TM5614 (plasminogen activator inhibitor-1: PAI-1 inhibitor) in patients with unresectable melanoma. METHODS: The TM5614-MM study was a multicentre, open-label, single-arm, phase 2 clinical trial to evaluate the efficacy and safety of nivolumab in combination with TM5614 in patients with advanced, unresectable malignant melanoma recruited at 7 Japanese institutes between 13 September 2021 and 31 March 2023. Patients with metastatic or unresectable melanoma previously treated with anti-PD-1 Abs were enrolled. Nivolumab 480 mg was administered intravenously every 4 weeks for 8 weeks, while TM5614 was administered orally at a dose of 120 mg (0-4 weeks) and 180 mg once daily (5-8 weeks). The primary endpoint was the overall response rate after 8 weeks of concomitant use of TM5614. RESULTS: Thirty nine patients were enrolled, and 34 patients in the anti-PD-1 Abs-refractory cohort. The overall response rate at 8 weeks was 25.9% (95% CI: 12.9-44.9%; P = .027) in 27 anti-PD-1-Abs refractory patients by investigator assessment in the protocol per set cohort. Seven patients discontinued treatment due to progressive disease or adverse events. Treatment-related grade 3 or higher adverse events occurred in 3 of 39 patients (7.7%) in the intention-to-treat cohort. CONCLUSIONS: TM5614 in combination with nivolumab is well-tolerated and effective in anti-PD-1 Abs-refractory, unresectable melanoma. TRIAL REGISTRATION: This trial was registered with Clinical Trial gov, jRCT2021210029.

Development and Initial Psychometric Validation of the COVID-19 Pandemic Burden Index for Healthcare Workers.

BACKGROUND: The burden of COVID-19 on healthcare workers (HCWs) is reported to be increasing, yet the psychometric scales now in use evaluate only single aspects; few measure the pandemic-specific burden on HCWs comprehensively. OBJECTIVE: To develop a scale to quantify the physical, mental, and socioeconomic burden of the COVID-19 pandemic on HCWs. DESIGN: Scale development and cross-sectional survey. PARTICIPANTS: Consenting HCWs aged ≥20. MAIN MEASURES: Development of an item-list based on literature reviews and HCW panel input, evaluation of content validity and item selection using the Delphi method, psychometric testing conducted on HCWs, validity assessment by factor analyses and hypothesis verification, internal consistency evaluation by Cronbach's alpha, test-retest analysis, and interpretability assessment. KEY RESULTS: Through the Delphi process, a 29-item pilot scale was generated. In psychometric testing, data from 863 HCWs contributed to the development of the final version of this scale, called Pandemic Burden Index twenty for HCWs (PBI-20), a 20-item scale to measure six domains: fatigue, fear of infection, inadequacy as a medical professional, mental health concerns, prejudice or discrimination, and anxiety about one's livelihood and daily life. Factor analysis showed each factor corresponded to the six domains of this scale. Hypothesis verification showed the PBI-20 total score to be moderately to highly correlated with the Short Form 36 vitality score and mental health score and with intention of turnover. The PBI-20 had good internal consistency (Cronbach's alpha 0.92). Test-retest analysis showed the intraclass correlation coefficient to be 0.70 and the minimal important change to be -7.0. CONCLUSIONS: The psychometrically sound questionnaire we developed to measure pandemic-specific burdens for HCWs provides an understanding of comprehensive burdens on HCWs and may serve to evaluate interventions to reduce the burdens.

Pyridoxamine reduces methylglyoxal and markers of glycation and endothelial dysfunction, but does not improve insulin sensitivity or vascular function in abdominally obese individuals: A randomized double-blind placebo-controlled trial.

AIM: To investigate the effects of pyridoxamine (PM), a B6 vitamer and dicarbonyl scavenger, on glycation and a large panel of metabolic and vascular measurements in a randomized double-blind placebo-controlled trial in abdominally obese individuals. MATERIALS AND METHODS: Individuals (54% female; mean age 50 years; mean body mass index 32 kg/m2 ) were randomized to an 8-week intervention with either placebo (n = 36), 25 mg PM (n = 36) or 200 mg PM (n = 36). We assessed insulin sensitivity, ß-cell function, insulin-mediated microvascular recruitment, skin microvascular function, flow-mediated dilation, and plasma inflammation and endothelial function markers. PM metabolites, dicarbonyls and advanced glycation endproducts (AGEs) were measured using ultra-performance liquid chromatography tandem mass spectrometry. Treatment effects were evaluated by one-way ANCOVA. RESULTS: In the high PM dose group, we found a reduction of plasma methylglyoxal (MGO) and protein-bound Nδ-(5-hydro-5-methyl-4-imidazolon-2-yl)-ornithine (MG-H1), as compared to placebo. We found a reduction of the endothelial dysfunction marker soluble vascular cell adhesion molecule-1 (sVCAM-1) in the low and high PM dose group and of soluble intercellular adhesion molecule-1 (sICAM-1) in the high PM dose, as compared to placebo. We found no treatment effects on insulin sensitivity, vascular function or other functional outcome measurements. CONCLUSIONS: This study shows that PM is metabolically active and reduces MGO, AGEs, sVCAM-1 and sICAM-1, but does not affect insulin sensitivity and vascular function in abdominally obese individuals. The reduction in adhesion markers is promising because these are important in the pathogenesis of endothelial damage and atherosclerosis.

TM5614, an Inhibitor of Plasminogen Activator Inhibitor-1, Exerts an Antitumor Effect on Chronic Myeloid Leukemia.

Chronic myeloid leukemia (CML) is triggered by t(9;22)(q34;q11.2) translocation, leading to the formation of the BCR-ABL1 fusion gene. Although the development of BCR-ABL1 tyrosine kinase inhibitors (TKIs) has dramatically improved the prognosis of CML, the disease could often relapse, presumably because leukemic stem cell fraction of CML (CML-LSC) may reside in specific niches, and also acquire an ability to resist the cytotoxic agents. Recently a study indicated that pharmacological inhibition of plasminogen activator inhibitor-1 (PAI-1, also known as SERPINE1) would cause detachment of CML-LSCs from their niche by inducing maturation of membrane-type matrix metalloprotease-1 (MT1-MMP), leading to increased susceptibility of CML-LSCs against TKIs. However, the direct antitumor effect of PAI-1 inhibition in CML remains unclear. Because PAI-1 mRNA expression was lower in CML cell line (K562) than bone marrow mononuclear cells derived from CML patients, we established K562 cell clones stably expressing exogenous PAI-1 (K562/PAI-1). We found that TM5614 treatment significantly suppressed cell proliferation and induced apoptosis in K562/PAI-1 cells, accompanied by increased activity of Furin protease, which is a known target of PAI-1. Besides processing mature MT1-MMP, Furin is in charge of cleaving the NOTCH receptor to form a heterodimer before exporting it to the cell surface membrane. In K562/PAI-1 cells, TM5614 treatment increased NOTCH1 intracellular domain (NICD) protein expression as well as NOTCH1 target of HEY1 mRNA levels. Finally, forced expression of either Furin or NICD in K562/PAI-1 cells significantly inhibited cell proliferation and induced apoptosis. Collectively, PAI-1 inhibition may have an antitumor effect by modulating the Furin/NICD pathway.

Plasminogen activator inhibitor type-1 is a negative regulator of hematopoietic regeneration in the adipocyte-rich bone marrow microenvironment.

Bone marrow adipocytes (BMAs) have recently been recognized as a niche component with a suppressive function. Obese individuals with abundant BMAs exhibit impaired hematopoietic regeneration after hematopoietic stem cell transplantation (HSCT). We hypothesized that plasminogen activator inhibitor type-1 (PAI-1), an adipokine that regulates the fibrinolytic system, contributes to impaired hematopoiesis in bone marrow (BM) microenvironment with abundant BMAs. We demonstrated that BMAs differentiated in vitro could secrete PAI-1 and were positive for PAI-1 in vivo. In addition, the abundance of BMAs was associated with high levels of PAI-1 expression. The BMA-rich microenvironment exhibited impaired hematopoietic regeneration after HSCT when compared with a BMA-less microenvironment. The impaired hematopoietic regeneration in BMA-rich microenvironment was significantly alleviated by PAI-1 knockout or PAI-1 inhibitor treatment. Obese mice with abundant BMAs, compared with normal-weight mice, exhibited higher bone marrow PAI-1 concentrations, increased fibrinolytic system suppression, and lower stem cell factor (SCF) concentrations after HSCT. PAI-1 inhibitor administration significantly activated the fibrinolytic system in obese mice, contributing to the higher SCF concentration. Moreover, PAI-1 inhibitor treatment significantly alleviated the impaired hematopoietic regeneration in obese mice both after 5-fluorouracil injection and HSCT. These results indicate that PAI-1 hinders hematopoietic regeneration in BMA-rich microenvironments. The blockade of PAI-1 activity could be a novel therapeutic means of facilitating hematopoietic reconstitution in BMA-rich patients.

A novel PAI-1 inhibitor prevents ageing-related muscle fiber atrophy.

Sarcopenia is among the most common medical problems of the aging population worldwide and a major social concern. Here, we explored the therapeutic potential of TM5484, a novel orally available PAI-1 inhibitor, to prevent sarcopenia. The sarcopenic phenotypes of the calf muscle of 12- and 6-month-old middle-aged mice were compared. Although significant decline of isometric gastrocnemius muscle force was detected in the older untreated mice, those administered TM5484 had significantly greater calf muscle force, as determined using isometric measurements by electrical stimulation. Histological analysis indicated that cross-sectional gastrocnemius muscle fibers in untreated older mice were thinner than those in younger mice; however, TM5484-treated group showed thicker fibers than younger mice. Treatment with TM5484 for 6 months enhanced Igf1, Atrogin-1, Mt-Co1, and Chrna1 mRNA expression in the mice gastrocnemius muscle, with increased serum IGF-1 concentration. TM5484 induced dose-dependent Igf1, Atrogin-1, and Chrna1 expression in C2C12 myoblastic cells, confirming cell autonomous effect. Further, the presence of plasmin for 72 h caused significantly increased Igf1 expression in C2C12 cells. These findings suggest that oral PAI-1 inhibitors represent a promising therapeutic candidate for preventing sarcopenia progression in humans.

Targeting of plasminogen activator inhibitor-1 activity promotes elimination of chronic myeloid leukemia stem cells.

Therapeutic strategies that target leukemic stem cells (LSCs) provide potential advantages in the treatment of chronic myeloid leukemia (CML). Here, we show that selective blockade of plasminogen activator inhibitor-1 (PAI-1) enhances the susceptibility of CML-LSCs to tyrosine kinase inhibitor (TKI), which facilitates the eradication of CML-LSCs and leads to sustained remission of the disease. We demonstrated for the first time that TGF-ß-PAI-1 axis was selectively augmented in CML-LSCs in the bone marrow (BM), whereby protecting CML-LSCs from TKI treatment. Furthermore, the combined administration of TKI plus a PAI-1 inhibitor, in a mouse model of CML, significantly enhanced the eradication of CML cells in the BM and prolonged the survival of CML mice. The combined therapy of imatinib and a PAI-1 inhibitor prevented the recurrence of CML-like disease in serially transplanted recipients, indicating the elimination of CML-LSCs. Interestingly, PAI-1 inhibitor treatment augmented membrane-type matrix metalloprotease-1 (MT1-MMP)-dependent motility of CML-LSCs, and the anti-CML effect of PAI-1 inhibitor was extinguished by the neutralizing antibody for MT1-MMP, underlining the mechanistic importance of MT1-MMP. Our findings provide evidence of, and a rationale for, a novel therapeutic tactic, based on the blockade of PAI-1 activity, for CML patients.

Structural Insight into the Two-Step Mechanism of PAI-1 Inhibition by Small Molecule TM5484.

Plasminogen activator inhibitor-1 (PAI-1), a key regulator of the fibrinolytic system, is the main physiological inhibitor of plasminogen activators. By interacting with matrix components, including vitronectin (Vn), PAI-1 plays a regulatory role in tissue remodeling, cell migration, and intracellular signaling. Emerging evidence points to a role for PAI-1 in various pathological conditions, including cardiovascular diseases, cancer, and fibrosis. Targeting PAI-1 is therefore a promising therapeutic strategy in PAI-1-related pathologies. A class of small molecule inhibitors including TM5441 and TM5484, designed to bind the cleft in the central ß-sheet A of PAI-1, showed to be potent PAI-1 inhibitors in vivo. However, their binding site has not yet been confirmed. Here, we report two X-ray crystallographic structures of PAI-1 in complex with TM5484. The structures revealed a binding site at the flexible joint region, which is distinct from the presumed binding site. Based on the structural analysis and biochemical data we propose a mechanism for the observed dose-dependent two-step mechanism of PAI-1 inhibition. By binding to the flexible joint region in PAI-1, TM5484 might restrict the structural flexibility of this region, thereby inducing a substrate form of PAI-1 followed by a conversion to an inert form.

PAI-1 Regulation of TGF-ß1-induced Alveolar Type II Cell Senescence, SASP Secretion, and SASP-mediated Activation of Alveolar Macrophages.

Senescence of alveolar type II (ATII) cells, progenitors of the alveolar epithelium, is a pathological feature and contributes importantly to the pathogenesis of idiopathic pulmonary fibrosis. Despite recognition of the importance of ATII cell senescence in idiopathic pulmonary fibrosis pathogenesis, how ATII cell senescence is regulated and how senescent ATII cells contribute to lung fibrogenesis remain unclear. In this study, we show that TGF-ß1 (transforming growth factor-ß1), a most ubiquitous and potent profibrotic cytokine, induces plasminogen activator inhibitor-1 (PAI-1), a cell senescence and fibrosis mediator, and p16 as well as senescence, but not apoptosis, in primary mouse ATII cells. We also found that senescent ATII cells secrete various cytokines and chemokines, including IL-4 and IL-13, which stimulate the expression of genes associated with a profibrotic phenotype in alveolar macrophages. Similar responses were also observed in TGF-ß1-treated rat ATII (L2) and rat macrophage NR8383 cells. Deletion of PAI-1 or inhibition of PAI-1 activity with a small molecule PAI-1 inhibitor, however, blocks TGF-ß1-induced senescence as well as a senescence-associated secretory phenotype in ATII and L2 cells and, consequently, the stimulatory effects of the conditioned medium from senescent ATII/L2 cells on macrophages. Moreover, we show that silencing p16 ameliorates PAI-1 protein-induced ATII cell senescence and secretion of profibrotic mediators. Our data suggest that PAI-1 mediates TGF-ß1-induced ATII cell senescence and secretion of profibrotic mediators through inducing p16, and they also suggest that senescent ATII cells contribute to lung fibrogenesis in part by activating alveolar macrophages through secreting profibrotic and proinflammatory mediators.

Blockade of plasminogen activator inhibitor-1 empties bone marrow niche sufficient for donor hematopoietic stem cell engraftment without myeloablative conditioning.

Upon hematopoietic stem cell transplantation (HSCT), the availability of recipients' niches in the bone marrow (BM) is one of the factors that influence donor HSC engraftment and hematopoietic reconstitution. Therefore, myeloablative conditioning, such as irradiation and/or chemotherapy, which creates empty niches in the recipients' BM, is required for the success of HSCT. However, the conventional myeloablation causes extensive damages to the patients' BM, which results in the treatment-induced severe complications and even mortality. Thus, alternative and mild conditioning could fulfill the need for safer HSCT-based therapies for hematological and nonhematological disorders. Recently, we have demonstrated that pharmacological inhibition of plasminogen activator inhibitor-1 (PAI-1) activity increases cellular motility and cause detachment of HSCs from the niches. In this study, we performed HSCT using a PAI-1 inhibitor without any myeloablative conditioning. Donor HSCs were transplanted to recipient mice that were pretreated with saline or a PAI-1 inhibitor. Saline pretreated nonmyeloablative recipients showed no engraftment. In contrast, donor cell engraftment was detected in the PAI-1 inhibitor pretreated recipients. Multilineage differentiation, including lymphoid and myeloid cells, was observed in the PAI-1 inhibitor pretreated recipients. Donor-derived cells that exhibited multilineage reconstitution as well as the existence of stem/progenitor cells were detected in the secondary recipients, confirming the maintenance of donor HSCs in the BM of PAI-1 inhibitor pretreated primary recipients. The results indicate that the PAI-1 blockade vacates functional niches in the recipients' BM, which allows the engraftment of long-term multilineage HSCs without myeloablative conditioning.

TGF-ß-induced intracellular PAI-1 is responsible for retaining hematopoietic stem cells in the niche.

Hematopoietic stem and progenitor cells (HSPCs) reside in the supportive stromal niche in bone marrow (BM); when needed, however, they are rapidly mobilized into the circulation, suggesting that HSPCs are intrinsically highly motile but usually stay in the niche. We questioned what determines the motility of HSPCs. Here, we show that transforming growth factor (TGF)-ß-induced intracellular plasminogen activator inhibitor (PAI)-1 activation is responsible for keeping HSPCs in the BM niche. We found that the expression of PAI-1, a downstream target of TGF-ß signaling, was selectively augmented in niche-residing HSPCs. Functional inhibition of the TGF-ß-PAI-1 signal increased MT1-MMP-dependent cellular motility, causing a detachment of HSPCs from the TGF-ß-expressing niche cells, such as megakaryocytes. Furthermore, consistently high motility in PAI-1-deficient HSPCs was demonstrated by both a transwell migration assay and reciprocal transplantation experiments, indicating that intracellular, not extracellular, PAI-1 suppresses the motility of HSPCs, thereby causing them to stay in the niche. Mechanistically, intracellular PAI-1 inhibited the proteolytic activity of proprotein convertase Furin, diminishing MT1-MMP activity. This reduced expression of MT1-MMP in turn affected the expression levels of several adhesion/deadhesion molecules for determination of HSPC localization, such as CD44, VLA-4, and CXCR4, which then promoted the retention of HSPCs in the niche. Our findings open up a new field for the study of intracellular proteolysis as a regulatory mechanism of stem cell fate, which has the potential to improve clinical HSPC mobilization and transplantation protocols.

Plasminogen activator inhibitor-1 regulates macrophage-dependent postoperative adhesion by enhancing EGF-HER1 signaling in mice.

Adhesive small bowel obstruction remains a common problem for surgeons. After surgery, platelet aggregation contributes to coagulation cascade and fibrin clot formation. With clotting, fibrin degradation is simultaneously enhanced, driven by tissue plasminogen activator-mediated cleavage of plasminogen to form plasmin. The aim of this study was to investigate the cellular events and proteolytic responses that surround plasminogen activator inhibitor (PAI-1; Serpine1) inhibition of postoperative adhesion. Peritoneal adhesion was induced by gauze deposition in the abdominal cavity in C57BL/6 mice and those that were deficient in fibrinolytic factors, such as Plat-/- and Serpine1-/- In addition, C57BL/6 mice were treated with the novel PAI-1 inhibitor, TM5275. Some animals were treated with clodronate to deplete macrophages. Epidermal growth factor (EGF) experiments were performed to understand the role of macrophages and how EGF contributes to adhesion. In the early phase of adhesive small bowel obstruction, increased PAI-1 activity was observed in the peritoneal cavity. Genetic and pharmacologic PAI-1 inhibition prevented progression of adhesion and increased circulating plasmin. Whereas Serpine1-/- mice showed intra-abdominal bleeding, mice that were treated with TM5275 did not. Mechanistically, PAI-1, in combination with tissue plasminogen activator, served as a chemoattractant for macrophages that, in turn, secreted EGF and up-regulated the receptor, HER1, on peritoneal mesothelial cells, which led to PAI-1 secretion, further fueling the vicious cycle of impaired fibrinolysis at the adhesive site. Controlled inhibition of PAI-1 not only enhanced activation of the fibrinolytic system, but also prevented recruitment of EGF-secreting macrophages. Pharmacologic PAI-1 inhibition ameliorated adhesion formation in a macrophage-dependent manner.-Honjo, K., Munakata, S., Tashiro, Y., Salama, Y., Shimazu, H., Eiamboonsert, S., Dhahri, D., Ichimura, A., Dan, T., Miyata, T., Takeda, K., Sakamoto, K., Hattori, K., Heissig, B. Plasminogen activator inhibitor-1 regulates macrophage-dependent postoperative adhesion by enhancing EGF-HER1 signaling in mice.

Identification of novel plasminogen activator inhibitor-1 inhibitors with improved oral bioavailability: Structure optimization of N-acylanthranilic acid derivatives.

Novel plasminogen activator inhibitor-1 (PAI-1) inhibitors with highly improved oral bioavailability were discovered by structure-activity relationship studies on N-acyl-5-chloroanthranilic acid derivatives. Because lipophilic N-acyl groups seemed to be important for the anthranilic acid derivatives to strongly inhibit PAI-1, synthesis of compounds in which 5-chloroanthranilic acid was bound to a variety of highly lipophilic moieties with appropriate linkers was investigated. As the result it appeared that some of the derivatives possessing aryl- or heteroaryl-substituted phenyl groups in the acyl chain had potent in vitro PAI-1 inhibitory activity. Oral absorbability of typical compounds was also evaluated in rats, and compounds 40, 55, 60 and 76 which have diverse chemical structure with each other were selected for further pharmacological evaluation.

Pyridoxamine: A novel treatment for schizophrenia with enhanced carbonyl stress.

AIM: The aim of this clinical trial was to obtain proof of concept for high-dose pyridoxamine as a novel treatment for schizophrenia with enhanced carbonyl stress. METHODS: Ten Japanese schizophrenia patients with high plasma pentosidine, which is a representative biomarker of enhanced carbonyl stress, were recruited in a 24-week, open trial in which high-dose pyridoxamine (ranging from 1200 to 2400 mg/day) was administered using a conventional antipsychotic regimen. Main outcomes were the total change in Positive and Negative Syndrome Scale score and the Brief Psychiatric Rating Scale score from baseline to end of treatment at week 24 (or at withdrawal). RESULTS: Decreased plasma pentosidine levels were observed in eight patients. Two patients showed marked improvement in their psychological symptoms. A patient who harbors a frameshift mutation in the Glyoxalase 1 gene also showed considerable reduction in psychosis accompanied with a moderate decrease in plasma pentosidine levels. A reduction of greater than 20% in the assessment scale of drug-induced Parkinsonism occurred in four patients. Although there was no severe suicide-related ideation or behavior, Wernicke's encephalopathy-like adverse drug reactions occurred in two patients and were completely suppressed by thiamine supplementation. CONCLUSION: High-dose pyridoxamine add-on treatment was, in part, effective for a subpopulation of schizophrenia patients with enhanced carbonyl stress. Further randomized, placebo-controlled trials with careful monitoring will be required to validate the efficacy of high-dose pyridoxamine for these patients.

Advanced Glycation Endproducts Are Increased in the Animal Model of Multiple Sclerosis but Cannot Be Reduced by Pyridoxamine Treatment or Glyoxalase 1 Overexpression.

Multiple sclerosis (MS) is a demyelinating autoimmune disease of the central nervous system (CNS). The immune response in MS patients leads to the infiltration of immune cells in the CNS and their subsequent activation. Immune cell activation induces a switch towards glycolysis. During glycolysis, the dicarbonyl product methylglyoxal (MGO) is produced. MGO is a glycating agent that can rapidly form advanced glycation endproducts (AGEs). In turn, AGEs are able to induce inflammatory responses. The glyoxalase system is the endogenous defense system of the body to reduce the burden of MGO thereby reducing AGE formation. This system consists of glyoxalase-1 and glyoxalase-2 which are able to detoxify MGO to D-lactate. We investigated whether AGE levels are induced in experimental autoimmune encephalitis (EAE), an inflammatory animal model of MS. Twenty seven days post EAE induction, MGO and AGE (Nε-(carboxymethyl)lysine (CML), Nε-(carboxyethyl)lysine (CEL), 5-hydro-5-methylimidazolone (MG-H1)) levels were significantly increased in the spinal cord of mice subjected to EAE. Yet, pyridoxamine treatment and glyoxalase-1 overexpression were unable to counteract AGE production during EAE and did not influence the clinical course of EAE. In conclusion, AGEs levels increase during EAE in the spinal cord, but AGE-modifying treatments do not inhibit EAE-induced AGE production and do not affect disease progression.

Furan- and Thiophene-2-Carbonyl Amino Acid Derivatives Activate Hypoxia-Inducible Factor via Inhibition of Factor Inhibiting Hypoxia-Inducible Factor-1.

Induction of a series of anti-hypoxic proteins protects cells during exposure to hypoxic conditions. Hypoxia-inducible factor-α (HIF-α) is a major transcription factor that orchestrates this protective effect. To activate HIF exogenously, without exposing cells to hypoxic conditions, many small-molecule inhibitors targeting prolyl hydroxylase domain-containing protein have been developed. In addition, suppression of factor inhibiting HIF-1 (FIH-1) has also been shown to have the potential to activate HIF-α. However, few small-molecule inhibitors of FIH-1 have been developed. In this study, we synthesized a series of furan- and thiophene-2-carbonyl amino acid derivatives having the potential to inhibit FIH-1. The inhibitory activities of these compounds were evaluated in SK-N-BE(2)c cells by measuring HIF response element (HRE) promoter activity. Several furan- and thiophene-2-carbonyl amino acid derivatives inhibited FIH-1 based on correlations among the docking score of the FIH-1 active site, the chemical structure of the compounds, and biological HIF-α/HRE transcriptional activity.

[Continuation of Axitinib for Advanced Renal Cell Carcinoma by The Application of Homecare Urology - A Case Report and Literature Review].

A 56-year-old man with advanced RCC and a past medical history of type 2 diabetes underwent a radical left nephrectomy following a histological diagnosis of papillary RCC, G2, INF b, pT3, V1 in 1999. In 2008, sorafenib was started to treat multiple pulmonary metastases of RCC. In 2011, sorafenib was switched to sunitinib when radiologic progression was observed. In 2014, sunitinib was switched to axitinib when further radiologic progression was observed. In 2015, the patient was referred to Yazawa clinic for homecare urology when hospital visits became difficult due to cancer pain and bilateral lower-extremity muscle weakness. Cancer pain was controlled using acetaminophen and a fentanyl patch. During the administration of axitinib, a CTCAE grade 1 vocal disorder was detected. We reduced the axitinib dose from 10 mg to 6 mg, and valsartan and an antiflatulent were administered due to CTCAE grade 2 hypertension and diarrhea, respectively. Axitinib administration continued until the patient died. He had survived more than 11 years following the detection of lung metastasis. In this patient, a good balance between cancer treatment and palliative care was achieved through the application of homecare urology. In a super-aged society such as Japan, urologists with an awareness of Zaitaku Medicine, a Japanese style of homecare that provides continuing appropriate medical treatment and welfare support to patients with access barriers to hospital treatment to enable them to live out the remainder of their lives with dignity, may play a key role in the development of Zaitaku Medicine.

PAI-1-regulated extracellular proteolysis governs senescence and survival in Klotho mice.

Cellular senescence restricts the proliferative capacity of cells and is accompanied by the production of several proteins, collectively termed the "senescence-messaging secretome" (SMS). As senescent cells accumulate in tissue, local effects of the SMS have been hypothesized to disrupt tissue regenerative capacity. Klotho functions as an aging-suppressor gene, and Klotho-deficient (kl/kl) mice exhibit an accelerated aging-like phenotype that includes a truncated lifespan, arteriosclerosis, and emphysema. Because plasminogen activator inhibitor-1 (PAI-1), a serine protease inhibitor (SERPIN), is elevated in kl/kl mice and is a critical determinant of replicative senescence in vitro, we hypothesized that a reduction in extracellular proteolytic activity contributes to the accelerated aging-like phenotype of kl/kl mice. Here we show that PAI-1 deficiency retards the development of senescence and protects organ structure and function while prolonging the lifespan of kl/kl mice. These findings indicate that a SERPIN-regulated cell-nonautonomous proteolytic cascade is a critical determinant of senescence in vivo.

Erythropoietin Synthesis in Renal Myofibroblasts Is Restored by Activation of Hypoxia Signaling.

Erythropoietin (Epo) is produced by renal Epo-producing cells (REPs) in a hypoxia-inducible manner. The conversion of REPs into myofibroblasts and coincident loss of Epo-producing ability are the major cause of renal fibrosis and anemia. However, the hypoxic response of these transformed myofibroblasts remains unclear. Here, we used complementary in vivo transgenic and live imaging approaches to better understand the importance of hypoxia signaling in Epo production. Live imaging of REPs in transgenic mice expressing green fluorescent protein from a modified Epo-gene locus revealed that healthy REPs tightly associated with endothelium by wrapping processes around capillaries. However, this association was hampered in states of renal injury-induced inflammation previously shown to correlate with the transition to myofibroblast-transformed renal Epo-producing cells (MF-REPs). Furthermore, activation of hypoxia-inducible factors (HIFs) by genetic inactivation of HIF-prolyl hydroxylases (PHD1, PHD2, and PHD3) selectively in Epo-producing cells reactivated Epo production in MF-REPs. Loss of PHD2 in REPs restored Epo-gene expression in injured kidneys but caused polycythemia. Notably, combined deletions of PHD1 and PHD3 prevented loss of Epo expression without provoking polycythemia. Mice with PHD-deficient REPs also showed resistance to LPS-induced Epo repression in kidneys, suggesting that augmented HIF signaling counterbalances inflammatory stimuli in regulation of Epo production. Thus, augmentation of HIF signaling may be an attractive therapeutic strategy for treating renal anemia by reactivating Epo synthesis in MF-REPs.