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Background Impaired multi-ciliated cell (MCC) differentiation, observed in various airway diseases, causes prolonged inflammation and further airway complications due to malfunction of the airway defense mechanism. YAP/TAZ-TEAD signaling mediates multiple mechanisms associated with MCC differentiation. However, YAP reportedly has both negative and positive roles in MCC differentiation, putatively due to its dual functions in the cytoplasm and nucleus. We hypothesize inhibition of nuclear YAP, while preserving cytoplasmic YAP functions, restores MCCs in various airway disorders. Here, we investigated the relationship between YAP nuclear localization and the presence of MCCs across patients. In addition, we identified compounds that promote MCC differentiation based on the inhibitory function of YAP nuclear localization. Method Tracheal specimens were collected from patients with different conditions (smoking, irradiation, tracheotomy, dysphagia), and YAP nuclear localization in MCCs was assessed by immunohistochemistry at areas with varying MCC compositions. To identify compounds that efficiently inhibit nuclear localization of YAP, airway epithelial cells were treated with multiple YAP inhibitors and YAP nuclear localization was assessed by immunofluorescence-based image analysis. The effects of the compounds on MCC differentiation were evaluated using air-liquid interface (ALI) culture and rat airway squamous metaplasia models. Results In the human trachea, YAP nuclear localization in MCCs was negatively correlated with the proportion of MCCs in the epithelium, regardless of patient condition. Among known YAP inhibitors, verteporfin and atorvastatin efficiently inhibited YAP nuclear localization in cultured airway epithelial cells. Additionally, verteporfin promoted MCC differentiation in ALI culture and in vivo squamous metaplasia models. Conclusion Our findings suggest YAP nuclear localization in human airway MCCs is broadly associated with decreased MCCs regardless of patient condition, and verteporfin improves airway MCC differentiation.

Staphylococcus aureus lipase (SAL), a triacylglycerol esterase, is an important virulence factor and may be a therapeutic target for infectious diseases. Herein, we determined the 3D structure of native SAL, the mutated S116A inactive form, and the inhibitor complex using the anti-obesity drug orlistat to aid in drug development. The determined crystal structures showed a typical α/β hydrolase motif with a dimeric form. Fatty acids bound near the active site in native SAL and inactive S116A mutant structures. We found that orlistat potently inhibits SAL activity, and it covalently bound to the catalytic Ser116 residue. This is the first report detailing orlistat–lipase binding. It provides structure-based information on the production of potent anti-SAL drugs and lipase inhibitors. These results also indicated that orlistat can be repositioned to treat bacterial diseases.

Androgen‐deprivation therapy is a standard treatment for advanced prostate cancer. However, most patients eventually acquire resistance and progress to castration‐resistant prostate cancer (CRPC). In this study, we established new CRPC cell lines, AILNCaP14 and AILNCaP15, from LNCaP cells under androgen‐deprived conditions. Unlike most pre‐existing CRPC cell lines, both cell lines expressed higher levels of androgen receptor (AR) and prostate‐specific antigen (PSA) than parental LNCaP cells. Moreover, these cells exhibited primary resistance to enzalutamide. Since AR signaling plays a significant role in the development of CRPC, PSA promoter sequences fused with GFP were introduced into AILNCaP14 cells to conduct GFP fluorescence‐based chemical screening. We identified flavopiridol, a broad‐spectrum CDK inhibitor, as a candidate drug that could repress AR transactivation of CRPC cells, presumably through the inhibition of phosphorylation of AR on the serine 81 residue (pARSer81). Importantly, this broad‐spectrum CDK inhibitor inhibited the proliferation of AILNCaP14 cells both in vitro and in vivo. Moreover, a newly developed liver metastatic model using AILNCaP15 cells revealed that the compound attenuated tumor growth of CRPC harboring highly metastatic properties. Finally, we developed a patient‐derived xenograft (PDX) model of CRPC and DCaP CR from a patient presenting therapeutic resistance to enzalutamide, abiraterone, and docetaxel. Flavopiridol successfully suppressed the tumor growth of CRPC in this PDX model. Since ARSer81 was found to be phosphorylated in clinical CRPC samples, our data suggested that broad‐spectrum CDK inhibitors might be a potent candidate drug for the treatment of CRPC, including those exhibiting primary resistance to enzalutamide. We established new castration‐resistant prostate cancer (CRPC) cell lines, AILNCaP14 and AILNCaP15. Using high‐throughput chemical screening, we identified a broad‐spectrum CDK inhibitor, flavopiridol, as a candidate drug for enzalutamide‐resistant CRPC. We developed preclinical models of CRPC and assessed the effect of flavopiridol on CRPC both in vitro and in vivo. Flavopiridol successfully attenuated tumor growth in liver metastasis as well as in a patient‐derived xenograft (PDX) model of CRPC.

Significance Familial dysautonomia (FD) is caused by missplicing of the IκB kinase complex-associated protein (IKAP) gene, which results in the skipping of exon 20, especially in neurons. FD would be treatable if exon 20 inclusion were increased correctly to reestablish correct splicing. Here, we have established a dual-color splicing reporter that recapitulates FD-type splicing. By using this reporter, we have identified a small chemical compound, named rectifier of aberrant splicing (RECTAS), that rectifies the aberrant splicing of FD. RECTAS promotes both exon 20 inclusion and the product IKAP expression in cells of patients with FD. Furthermore, we have demonstrated that modification levels of wobble uridine residues of several tRNAs are reduced in FD cells and that RECTAS can recover not only tRNA modifications but also cell viability of FD cells. Familial dysautonomia (FD), a hereditary sensory and autonomic neuropathy, is caused by missplicing of exon 20, resulting from an intronic mutation in the inhibitor of kappa light polypeptide gene enhancer in B cells, kinase complex-associated protein ( IKBKAP ) gene encoding IKK complex-associated protein (IKAP)/elongator protein 1 (ELP1). A newly established splicing reporter assay allowed us to visualize pathogenic splicing in cells and to screen small chemicals for the ability to correct the aberrant splicing of IKBKAP . Using this splicing reporter, we screened our chemical libraries and identified a compound, rectifier of aberrant splicing (RECTAS), that rectifies the aberrant IKBKAP splicing in cells from patients with FD. Here, we found that the levels of modified uridine at the wobble position in cytoplasmic tRNAs are reduced in cells from patients with FD and that treatment with RECTAS increases the expression of IKAP and recovers the tRNA modifications. These findings suggest that the missplicing of IKBKAP results in reduced tRNA modifications in patients with FD and that RECTAS is a promising therapeutic drug candidate for FD.

Autophosphorylation of amino-acid residues is part of the folding process of various protein kinases. Conventional chemical screening of mature kinases has missed inhibitors that selectively interfere with the folding process. Here we report a cell-based assay that evaluates inhibition of a kinase at a transitional state during the folding process and identify a folding intermediate-selective inhibitor of dual-specificity tyrosine-phosphorylation-regulated kinase 1A (DYRK1A), which we refer to as FINDY. FINDY suppresses intramolecular autophosphorylation of Ser97 in DYRK1A in cultured cells, leading to its degradation, but does not inhibit substrate phosphorylation catalysed by the mature kinase. FINDY also suppresses Ser97 autophosphorylation of recombinant DYRK1A, suggesting direct inhibition, and shows high selectivity for DYRK1A over other DYRK family members. In addition, FINDY rescues DYRK1A-induced developmental malformations in Xenopus laevis embryos. Our study demonstrates that transitional folding intermediates of protein kinases can be targeted by small molecules, and paves the way for developing novel types of kinase inhibitors. Kinase inhibitors are important drugs and usually target the ATP binding pocket of kinases. Here, Kii et al. report a completely new type of kinase inhibitor that specifically targets the protein folding intermediate state, but not the mature form, of the protein kinase DYRK1A.

Background Exercise is known to improve physical fitness, functional performance, muscle strength, quality of life (QoL), and treatment outcomes in patients with cancer. However, the feasibility and impact of implementing an exercise program from the first visit for suspected advanced lung cancer to treatment initiation remain unclear. This study aimed to evaluate the feasibility of a structured exercise program during this period. Methods This prospective, single-center study included patients with suspected advanced lung cancer who were instructed to perform a pre-treatment 4-week exercise program consisting of slow movement training with tonic force generation and aerobic exercise. Attendance rates and adverse events were assessed, and changes in functional performance, exercise capacity, balance function, muscle strength and mass, physical activity levels, nutritional status, and QoL were evaluated. Patient feedback on the exercise program was collected. Results A total of 17 patients were analyzed, with a mean age of 70 years. The overall exercise attendance rate was 159.7 ± 54.4%. A high attendance rate (≥70.8%) was achieved by 15 patients without any adverse events. Significant improvements from baseline to after-exercise intervention were observed in the five-time sit-to-stand test and 30-second chair stand test. No significant changes in 6-minute walk distance, one-leg standing time, isometric handgrip and knee extensor strengths, skeletal muscle mass or self-reported measures were observed. Patient feedback indicated that exercise was beneficial in alleviating anxiety and maintaining physical condition during the waiting period before treatment. Conclusions A structured exercise program initiated from the first visit to treatment initiation was feasible and safe in patients with advanced lung cancer. Notably, improvements in the five-time sit-to-stand test and 30-second chair stand test were observed, suggesting potential benefits for enhancing lower-limb functional performance during this period. Further studies, including randomized controlled trials, are warranted to assess its impact on clinical outcomes and long-term patient wellbeing. Trial registration University Hospital Medical Information Network, UMIN000055155. Date of registration: August 5, 2024 Graphical Abstract

It is now well-established that Staphylococcus aureus can produce a range of toxin proteins, resulting in a spectrum of pathological conditions when it infects individuals with pre-existing medical conditions or immunocompromised. Among these, MRSA is one of the most prominent antimicrobial-resistant organisms and a significant cause of mortality in many patients. It has been demonstrated that Staphylococcus aureus lipase (SAL) is a vital factor in the proliferation of this bacterium. A combination of in silico screening and X-ray crystallography was employed to analyze inhibitors of SAL, and the results were highly significant. In silico screening identified a number of compounds, and the enzyme activity assay demonstrated that the antipsychotic drug penfluridol exhibited potent inhibitory activity against SAL. We have conducted co-crystallization of penfluridol and SAL on the ground and in space. The resulting co-crystals were subjected to data measurement using the synchrotron radiation facility at SPring-8, and the complex structure was determined. The crystal structure of the penfluridol-SAL complex was determined at 2.2 Å resolution, thereby providing the structural basis for developing new anti-infective agents that inhibit the growth of Staphylococcus aureus . These findings are anticipated to facilitate the development of compounds with potent inhibitory activity.

Staphylococcus aureus produces large amounts of toxins and virulence factors. In patients with underlying diseases or compromised immune systems, this bacterium can lead to severe infections and potentially death. In this study, the crystal structure of the complex of S. aureus lipase (SAL), which is involved in the growth of this bacterium, with petroselinic acid (PSA), an inhibitor of unsaturated fatty acids, was determined by X‐ray crystallography. Recently, PSA was shown to inhibit S. aureus biofilm formation and the enzymatic activity of SAL. To further characterize the inhibitory mechanism, we determined the half‐inhibitory concentration of SAL by PSA and the crystal structure of the complex. The IC50 of the inhibitory effect of PSA on SAL was 3.4 μm. SAL and PSA inhibitors were co‐crystallized, and diffraction data sets were collected to 2.19 Å resolution at SPring‐8 to determine the crystal structure and elucidate the detailed structural interactions. The results show that the fatty acid moiety of PSA is tightly bound to a hydrophobic pocket extending in two directions around the catalytic residue Ser116. Ser116 was also covalently bonded to the carbon of the unsaturated fatty acid moiety, and an oxyanion hole in SAL stabilized the electrons of the double bond. The difference in inhibitory activity between PSA and ester compounds revealed a structure–activity relationship between SAL and PSA. Additional research is required to further characterize the clinical potential of PSA. Staphylococcus aureus lipase (SAL) is a glycerol ester hydrolase involved in the growth of this bacterium and petroselinic acid (PSA) has been shown to inhibit SAL. In this study, we determined the IC50 of PSA for SAL and the crystal structure of the SAL‐PSA complex. Further research is required to uncover the clinical potential of PSA.

Introduction Extensive small cell lung cancer (ES‐SCLC) are currently managed using first‐line chemotherapy options, including atezolizumab (Atezo) plus etoposide and carboplatin (CE) or durvalumab (Durva) plus etoposide with either cisplatin (PE) or carboplatin (CE). However, a definitive distinction in therapeutic effects between Atezo and Durva in these regimens remains unestablished. Methods We analyzed data from 100 patients diagnosed with ES‐SCLC who received immune checkpoint inhibitors (ICIs) as first‐line chemotherapy. Among them, 70 were administered Atezo + CE, 12 received Durva + PE, and 18 received Durva + CE. We assessed the efficacy of the two ICIs across various factors. Results The progression‐free survival (PFS) and overall survival (OS) did not significantly differ between Atezo + CE and Durva + CE/PE as first‐line chemotherapy treatments for SCLC. We observed no significant differences in age, sex, performance status (PS), liver metastasis, bone metastasis, or platinum‐based agent usage between the treatment cohorts. However, a marked improvement in PFS and OS was observed in the solitary patient with brain metastasis treated with Atezo + CE. Conclusion The primary distinction between these treatments was observed in the management of patients with brain metastasis. The literature lacks comparative studies on the effects of first‐line ICI treatment on the central nervous system, rendering our findings significant in clinical practice. Despite the retrospective nature of this study and the potential for various biases, we recommend the preferential use of Atezo + CE in patients with brain metastasis to potentially enhance prognosis. This is a comparison of 1st line Chemotherapy for small cell lung cancer. There was a difference in efficacy for brain metastasis.

Background Lineage plasticity in prostate cancer primarily induces transdifferentiation of adenocarcinoma into neuroendocrine prostate cancer (NEPC). Lineage alteration is largely driven by epigenome and is potentially reversible. Nonetheless, evidence supporting NEPC reversibility is lacking in materials derived from clinical specimens. Hence, we established KUCaP13, a novel NEPC cell line derived from the tissue of a patient initially diagnosed with adenocarcinoma, which later recurred as NEPC. We aimed to prove the reversibility of cellular lineage by high‐throughput chemical screening using KUCaP13 cells. Methods Compounds responsible for androgen receptor (AR) re‐expression in KUCaP13 cells were screened. A reporter gene, androgen response element luciferase (AREluc), was transduced into KUCaP13 cells to detect AR activity using luciferase assay. Results Positive control cells (KUCaP13_AREluc_AR) overexpressing AR showed enhanced luminescence upon administration of synthetic androgen. An initial screen of 1552 compounds revealed 30 candidate molecules potentially enhancing luciferase luminescence. In the second screening, we eliminated false positives and validated the findings using luciferase assay and quantitative real‐time polymerase chain reaction. However, all hit compounds were confirmed as false positives, probably due to their inhibitory activity on luciferase. Conclusions We developed a highly sensitive screening system to evaluate the reversibility of plasticity in NEPC using a novel cell line. However, no compound demonstrated the ability to re‐express AR in this cell line. Nevertheless, in future studies, this screening system could prove valuable for elucidating the mechanism of lineage plasticity in NEPC and for developing novel therapies focused on reversing plasticity. We established a robust and sensitive androgen response element luciferase reporter assay to monitor androgen receptor (AR) activity using KUCaP13 cells, a novel human‐derived treatment‐related neuroendocrine prostate cancer (t‐NEPC) cell line. A high‐throughput screening using a chemical library to identify potential compounds that induce AR re‐expression in KUCaP13 cells revealed 30 candidate molecules potentially enhancing luciferase luminescence; however, subsequent validation steps demonstrated these signals to be false‐positives. Despite not achieving the goal of AR re‐expression, this study stands as a significant proof‐of‐concept for the application of high‐throughput screening approaches in t‐NEPC research.