Nonclinical pharmacodynamics of boron neutron capture therapy using direct intratumoral administration of a folate receptor targeting novel boron carrier.

Background: Boron neutron capture therapy (BNCT) is a precise particle radiation therapy known for its unique cellular targeting ability. The development of innovative boron carriers is crucial for the advancement of BNCT technologies. Our previous study demonstrated the potential of PBC-IP administered via convection-enhanced delivery (CED) in an F98 rat glioma model. This approach significantly extended rat survival in neutron irradiation experiments, with half achieving long-term survival, akin to a cure, in a rat brain tumor model. Our commitment to clinical applicability has spurred additional nonclinical pharmacodynamic research, including an investigation into the effects of cannula position and the time elapsed post-CED administration. Methods: In comprehensive in vivo experiments conducted on an F98 rat brain tumor model, we meticulously examined the boron distribution and neutron irradiation experiments at various sites and multiple time intervals following CED administration. Results: The PBC-IP showed substantial efficacy for BNCT, revealing minimal differences in tumor boron concentration between central and peripheral CED administration, although a gradual decline in intratumoral boron concentration post-administration was observed. Therapeutic efficacy remained robust, particularly when employing cannula insertion at the tumor margin, compared to central injections. Even delayed neutron irradiation showed notable effectiveness, albeit with a slightly reduced survival period. These findings underscore the robust clinical potential of CED-administered PBC-IP in the treatment of malignant gliomas, offering adaptability across an array of treatment protocols. Conclusions: This study represents a significant leap forward in the quest to enhance BNCT for the management of malignant gliomas, opening promising avenues for clinical translation.

Rapid growth of primary pulmonary meningioma with hemoptysis.

While lung cancer is the predominant neoplasm causing hemoptysis, rare benign neoplasms can also be associated with hemoptysis. A 60-year-old woman presented with cough and hemoptysis. Chest computed tomography revealed an oval-shaped, well-circumscribed solitary mass (10 cm in size) in the right lower lobe, which had grown rapidly over the past year. The presence of intramass air bubbles and a surrounding halo of ground-glass opacities suggested the hemorrhagic rupture of a circumscribed hematoma into the surrounding lung tissue. Subsequent right lower lobectomy revealed a well-demarcated hematoma; its wall consisted of nonatypical spindle tumor cells, which were histologically diagnosed as meningioma. No meningioma was observed in the central nervous system, leading to the diagnosis of primary pulmonary meningioma. This case highlights PPM as a rare benign tumor (World Health Organization grade 1) capable of rapid development due to intratumoral hemorrhage, presenting with hemoptysis.

High body mass index and triglyceride levels at health checkups increase the risk of new-onset chronic kidney disease and worsening renal function: the TAMA MED Project-CKD.

BACKGROUND: Health checkups are important in patients with chronic kidney disease (CKD), which is not easily accompanied by subjective symptoms. CKD can be caused or aggravated by factors that have not yet been identified. METHODS: This retrospective cohort study included 7 483 patients who underwent specific annual health checkups at a medical institution in Tama City, did not have CKD in 2012, and continued to undergo checkups (aged 40-74 years). We examined the risk factors for new-onset CKD and 1.5-fold increase in serum creatinine levels among laboratory values from 2012 to 2020. RESULTS: Age, body mass index (BMI), triglyceride levels, atrial fibrillation, and medication for hypertension (HT) and diabetes mellitus were independent risk factors for proteinuria, whereas current smoking, BMI, systolic blood pressure (SBP), and medication for HT were independent risk factors for estimated glomerular filtration rate < 60 mL/min/1.73 m2. SBP, triglyceride levels and medication for HT were risk factors for a 1.5-fold increase in serum creatinine levels during course of the study. The cut-off values of BMI for eGFR < 60 mL/min/1.73 m2 were 22.2 (men 24.7, women 22.1) kg/m2 and fasting triglyceride levels for a 1.5-fold increase in serum creatinine level were 171 (men 247, women 170) mg/dL, respectively. CONCLUSIONS: Health checkups provide information to prevent new-onset CKD and worsening of renal function. It is necessary to increase the rate of health checkups and visits to medical institutions after health checkups as well as to use these results for health guidance.

Development of carbonic anhydrase IX-targeting molecular-targeted photodynamic therapy.

The efficacy of molecular-targeted photodynamic therapy (MT-PDT) targeting carbonic anhydrase (CA) IX, a cancer-specific molecule, was demonstrated. CA ligand-directed photosensitizers 1-3 were evaluated for their ability to deactivate CAIX protein in cells. Compounds 2 and 3 selectively deactivated CAIX protein under 540 nm light without affecting internal standard proteins. Mechanistic studies revealed that compound 3 not only induced CAIX-selective light inactivation via singlet oxygen but also induced cell membrane damage, resulting in an anti-tumor effect. In vivo studies of CAIX-targeting MT-PDT revealed that treatment with compound 3 followed by light irradiation exhibited remarkable anti-tumor activity, leading to tumor degeneration and necrosis.

Metastatic gallbladder cancer presenting as numb chin syndrome: A case report and literature review.

Gallbladder cancer (GBC) is an uncommon malignancy that is highly aggressive in the advanced stages. However, it rarely metastasizes to the mandible. Numb chin syndrome (NCS) is a rare neurological manifestation associated with various underlying causes, including occult primary cancers and distant metastases. It is often considered to be a significant indicator of malignancy, and thorough investigation is essential in the presence of unclear etiology. The current study reported on the case of a 69-year-old Japanese woman who presented with numbness and mild pain in the lower lip and chin area for three months. No other systemic symptoms were observed. Immunocytochemical examination revealed the presence of an adenocarcinoma and TNM staging as per the Union for International Cancer Control and the American Joint Committee on Cancer guidelines confirmed stage IVb GBC. Comprehensive full-body positron emission tomography-computed tomography examination using 18F-fluoro-2-deoxy-D-glucose revealed additional bone and soft-tissue metastases. Palliative chemotherapy and radiation treatment were initiated based on the advanced stage of disease at the time of diagnosis. However, the patient succumbed to multiple organ failure six months later. The simultaneous occurrence of GBC, mandibular metastasis and NCS is rare and associated with poor prognosis. Despite the widespread nature of the disease, it can often manifest as non-specific oral symptoms without any systemic indications. The current study emphasizes the critical importance of timely confirmatory testing for accurate diagnosis and initiation of appropriate management for such complex conditions.

Association between animal protein intake, oral frailty and calf circumference in middle-aged and older adults: a cross-sectional analysis from the Shika study.

OBJECTIVE: To investigate the relationship between oral frailty (OF), nutrient intake and calf circumference (CC) in middle-aged and older adults. DESIGN: Cross-sectional study. SETTING: Residents of four model districts of Shika town, Ishikawa Prefecture, Japan, using data from November 2017 to February 2018. PARTICIPANTS: One hundred and ninety-four residents aged ≥50 years in four model districts of Shika town. The OF total score ≥3 was defined as OF. Participants were divided into OF and non-OF groups and divided into the low-CC/kg and the high-CC/kg groups. OUTCOME MEASURES: The primary outcome is to use a two-way analysis of covariance to analyse the interaction between the two CC/kg groups and the two OF groups on nutrition intake. The secondary outcome is to use multiple regression analysis to investigate the nutrients significantly related to CC/kg when stratified by OF, with age, sex, body mass index, drinking status, smoking status and regular exercise as input covariates. RESULTS: A two-way analysis of covariance revealed a significant interaction between the two CC/kg groups and the two OF groups on animal protein intake (p=0.039). Multiple comparisons using the Bonferroni analysis revealed a significantly lower animal protein intake in the OF group than in the non-OF group with a low CC/kg (p=0.033) but not in the group with a high CC/kg. The multiple regression analysis stratified by OF revealed a positive correlation between animal protein intake and CC/kg (p=0.002). CONCLUSIONS: The present results revealed a significantly lower animal protein intake in the OF group than in the non-OF group in the low-CC/kg group, but no such difference was observed in the high-CC/kg group. Further longitudinal studies are needed to elucidate this relationship.

Eliglustat exerts anti-fibrotic effects by activating SREBP2 in TGF-ß1-treated myofibroblasts derived from patients with idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a progressive chronic lung disease. Myofibroblasts play a critical role in fibrosis. These cells produce the extracellular matrix (ECM), which contributes to tissue regeneration; however, excess ECM production can cause fibrosis. Transforming growth factor-ß (TGF-ß)/Smad signaling induces ECM production by myofibroblasts; therefore, the inhibition of TGF-ß/Smad signaling may be an effective strategy for IPF treatment. We recently reported that miglustat, an inhibitor of glucosylceramide synthase (GCS), ameliorates pulmonary fibrosis by inhibiting the nuclear translocation of Smad2/3. In the present study, we examined the anti-fibrotic effects of another GCS inhibitor, eliglustat, a clinically approved drug for treating Gaucher disease type 1, in myofibroblasts derived from patient with IPF (IPF-MyoFs). We found that eliglustat exerted anti-fibrotic effects independent of GCS inhibition, and inhibited TGF-ß1-induced expression of α-smooth muscle actin, a marker of fibrosis, without suppressing the phosphorylation and nuclear translocation of Smad2/3. RNA sequencing analysis of eliglustat-treated human lung fibroblasts identified sterol regulatory element-binding protein 2 (SREBP2) activation. Transient overexpression of SREBP2 attenuated the TGF-ß1-induced increase in the expression of Smad target genes in IPF-MyoFs, and SREBP2 knockdown nullified the inhibitory effect of eliglustat on TGF-ß1-induced expression of α-SMA. These results suggested that eliglustat exerts its anti-fibrotic effects through SREBP2 activation. The findings of this study may contribute to the development of novel therapeutic strategies for IPF treatment.

Development of a Gadolinium-Boron-Conjugated Albumin for MRI-Guided Neutron Capture Therapy.

Noninvasive monitoring of boron agent biodistribution is required in advance of neutron capture therapy. In this study, we developed a gadolinium-boron-conjugated albumin (Gd-MID-BSA) for MRI-guided neutron capture therapy. Gd-MID-BSA was prepared by labeling bovine serum albumin with a maleimide-functionalized gadolinium complex and a maleimide-functionalized closo-dodecaborate orthogonally. The accumulation of Gd-MID-BSA in tumors in CT26 tumor-bearing mice reached a maximum at 24 h after the injection, as confirmed by T1-based MRI and biodistribution analysis using inductively coupled plasma optical emission spectrometry. The concentrations of boron and gadolinium in the tumors exceeded the thresholds required for boron neutron capture therapy (BNCT) and gadolinium neutron capture therapy (GdNCT), respectively. The boron concentration ratios of tumor to blood and tumor to normal tissues satisfied the clinical criteria, indicating the reduction of undesired nuclear reactions of endogenous nuclei. The molar ratio of boron to gadolinium in the tumor was close to that of Gd-MID-BSA, demonstrating that the accumulation of Gd-MID-BSA in the tumor can be evaluated by MRI. Thermal neutron irradiation with Gd-MID-BSA resulted in significant suppression of tumor growth compared to the group injected with a boron-conjugated albumin without gadolinium (MID-BSA). The neutron irradiation with Gd-MID-BSA did not cause apparent side effects. These results demonstrate that the conjugation of gadolinium and boron within the albumin molecule offers a novel strategy for enhancing the therapeutic effect of BNCT and the potential of MRI-guided neutron capture therapy as a promising treatment for malignant tumors.

Evaluation of the Effectiveness of Boron Neutron Capture Therapy with Iodophenyl-Conjugated closo-Dodecaborate on a Rat Brain Tumor Model.

High-grade gliomas present a significant challenge in neuro-oncology because of their aggressive nature and resistance to current therapies. Boron neutron capture therapy (BNCT) is a potential treatment method; however, the boron used by the carrier compounds-such as 4-borono-L-phenylalanine (L-BPA)-have limitations. This study evaluated the use of boron-conjugated 4-iodophenylbutanamide (BC-IP), a novel boron compound in BNCT, for the treatment of glioma. Using in vitro drug exposure experiments and in vivo studies, we compared BC-IP and BPA, with a focus on boron uptake and retention characteristics. The results showed that although BC-IP had a lower boron uptake than BPA, it exhibited superior retention. Furthermore, despite lower boron accumulation in tumors, BNCT mediated by BC-IP showed significant survival improvement in glioma-bearing rats compared to controls (not treated animals and neutrons only). These results suggest that BC-IP, with its unique properties, may be an alternative boron carrier for BNCT. Further research is required to optimize this potential treatment modality, which could significantly contribute to advancing the treatment of high-grade gliomas.

Proposal of recommended experimental protocols for in vitro and in vivo evaluation methods of boron agents for neutron capture therapy.

Recently, boron neutron capture therapy (BNCT) has been attracting attention as a minimally invasive cancer treatment. In 2020, the accelerator-based BNCT with L-BPA (Borofalan) as its D-sorbitol complex (Steboronine®) for head and neck cancers was approved by Pharmaceutical and Medical Devices Agency for the first time in the world. As accelerator-based neutron generation techniques are being developed in various countries, the development of novel tumor-selective boron agents is becoming increasingly important and desired. The Japanese Society of Neutron Capture Therapy believes it is necessary to propose standard evaluation protocols at each stage in the development of boron agents for BNCT. This review summarizes recommended experimental protocols for in vitro and in vivo evaluation methods of boron agents for BNCT based on our experience with L-BPA approval.

Discovery of three-dimensional bicyclo[3.3.1]nonanols as novel heat shock protein 90 inhibitors.

We developed an efficient synthetic method for constructing bicyclo[3.3.1]nonane, an sp3-carbon-rich three-dimensional scaffold consisting of two fused six-membered rings. Among the bicyclo[3.3.1]nonanols synthesized, several bicyclo[3.3.1]nonane derivatives were found to inhibit gene transcription by hypoxia-inducible factor-1 (HIF-1). The structure-activity relationship study revealed that the number of hydrophobic functional groups and a carboxylic acid moiety in the bicyclo[3.3.1]nonanols are important for inhibitory activities of both gene transcription by HIF-1 and cell growth. Bicyclo[3.3.1]nonanols fluctuated the amounts of client proteins of heat shock protein (HSP) 90 without inducing a heat shock response in cells and specifically inhibited the ATPase activity of HSP90. These results indicate that bicyclo[3.3.1]nonanols are novel HSP90 inhibitors with a different mechanism of action from conventional HSP90 inhibitors.

Mangostin enhances efficacy of aminolevulinic acid-photodynamic therapy against cancer through inhibition of ABCG2 activity.

BACKGROUND: Aminolevulinic acid-photodynamic therapy (ALA-PDT) is gaining attention as a potential method for treating select cancers due to its high specificity and low side effect feature. ALA enters cancer cells and accumulate as protoporphyrin IX (PpIX), which will then trigger phototoxicity following light irradiation. However, it is reported that some cancer cells have reduced efficacy of ALA-PDT due to high expression of ABCG2, a transporter involved in the PpIX efflux. In this study, we evaluated the effect of mangostin, a natural compound containing anti-tumor property, on the efficacy of ALA-PDT against cancer and the mechanism involved. METHODS: We utilized TMK1 gastric cancer cell line, which has high ABCG2 expression, to evaluate the PpIX accumulation and phototoxicity exerted by ALA and mangostin co-addition. RESULTS: We found that co-addition of ALA and mangostin significantly increase the phototoxicity and PpIX accumulation in TMK1 cells. We also investigated the effect of mangostin on porphyrin-heme pathway enzymes and ABCG2 and found that the addition of mangostin reduce the activity of ABCG2, reducing PpIX efflux. CONCLUSION: These findings suggest that mangostin enhances the efficacy of ALA-PDT in cancer through inhibition of ABCG2 activity.

Exploring stage­specific embryonic antigen 3 involvement in oral cancer progression and as a potential target for taxane­based chemotherapy.

Despite significant advancements in therapeutic approaches, oral neoplasms remain formidable and life­threatening conditions that affect a substantial number of individuals worldwide. Within oral malignancies, a subset of cancer stem cells (CSCs) represent a crucial population responsible for tumor initiation and progression. The identification of reliable markers for the detection and characterization of CSCs in solid tumors, particularly in the context of oral cancers, remains an ongoing challenge. Stage­specific embryonic antigen 3 (SSEA3), previously associated with mesenchymal stem cells and linked to the progression of breast neoplasms and poor prognosis, has yet to be comprehensively elucidated in the context of oral malignancies. The present study aimed to investigate the expression and properties of SSEA3 in 16 distinct subsets of human oral neoplastic cell lines, classified as either CD44 positive (+) or CD44 negative (­). For the first time, SSEA3 was examined as an indicator of tumorigenicity and resistance to taxane­derived chemotherapeutic agents. In the majority of oral neoplastic cell lines analyzed, SSEA3 was expressed in a small population of CD44(+) cells. Significantly, SSEA3(+) cells exhibited heightened proliferative activity and upregulated expression of genes associated with stem cells compared with SSEA3(­) cells. The aforementioned findings suggested that SSEA3 may contribute to the evolution and progression of oral malignancies by fostering tumor growth. Furthermore, SSEA3(+) cells displayed increased sensitivity to taxane­based pharmaceuticals, indicating the potential for SSEA3 to be a viable target in the treatment schema for oral cavity neoplasms. In conclusion, the present study provides novel insight into the role of SSEA3 in the progression and management of oral neoplasms, potentially paving the way for more effective therapeutic approaches.

Bronchoscopic visualization of the inner cavity caused by Pasteurella multocida infection.

We report a 69-year-old man who presented to our hospital with cough and sputum production. He had been in close contact with six domestic cats. He had a smoking history of 40 pack-years and had been in close contact with six domestic cats. A chest computed tomography scan revealed multiple consolidations with cavities in both lung fields. Pasteurella multocida was cultured from his sputum. On bronchoscopic evaluation, the flexible bronchoscope was navigated through the right middle lobe bronchus, which opened inside the cavity, allowing visualization of a spider-web-appearing architecture consisting of many cord-like lung tissues loosely adherent to the cavity lumen. Using these findings, a diagnosis of cavity formation was made secondary to Pasteurella multocida infection. Pasteurella infection should be considered as a cause of a lung cavity in patients with chronic lung disease. History taking regarding animal exposure is important for its diagnosis.

Fenofibrate attenuates the cytotoxic effect of cisplatin on lung cancer cells by enhancing the antioxidant defense system in vitro.

Fenofibrate (FF) is a peroxisome proliferator- activated receptor (PPAR)-α agonist that is widely used for the treatment of hyperlipidemia. It has been shown to have pleiotropic actions beyond its hypolipidemic effect. FF has been shown to exert a cytotoxic effect on some cancer cells when used at higher than clinically relevant concentrations; on the other hand, its cytoprotective effect on normal cells has also been reported. The present study assessed the effect of FF on cisplatin (CDDP) cytotoxicity to lung cancer cells in vitro. The results demonstrated that the effect of FF on lung cancer cells depends on its concentration. FF at ≤50 µM, which is a clinically achievable blood concentration, attenuated CDDP cytotoxicity to lung cancer cells, whereas FF at ≥100 µM, albeit clinically unachievable, had an anticancer effect. The mechanism of FF attenuation of CDDP cytotoxicity involved PPAR-α-dependent aryl hydrocarbon receptor (AhR) expression, which in turn stimulated nuclear factor erythroid 2-related factor 2 (Nrf2) expression and antioxidant production, resulting in lung cancer cell protection from CDDP-evoked oxidative damage. In conclusion, the present study revealed that FF, at clinically relevant concentrations, attenuated CDDP cytotoxicity to lung cancer cells by enhancing the antioxidant defense system through activation of a pathway that involves the PPAR-α-PPAR response element-AhR xenobiotic response element-Nrf2-antioxidant response element. These findings suggested that concomitant use of FF with CDDP may compromise the efficacy of chemotherapy. Although the anticancer property of FF has recently attracted much attention, concentrations that exceed clinically relevant concentrations are required.

Trajectories of Liver Fibrosis and Gene Expression Profiles in Nonalcoholic Fatty Liver Disease Associated With Diabetes.

Understanding the mechanisms linking steatosis to fibrosis is needed to establish a promising therapy against nonalcoholic fatty liver disease (NAFLD). The aim of this study was to clarify clinical features and hepatic gene expression signatures that predict and contribute to liver fibrosis development during the long-term real-world histological course of NAFLD in subjects with and without diabetes. A pathologist scored 342 serial liver biopsy samples from 118 subjects clinically diagnosed with NAFLD during a 3.8-year (SD 3.45 years, maximum 15 years) course of clinical treatment. At the initial biopsy, 26 subjects had simple fatty liver, and 92 had nonalcoholic steatohepatitis (NASH). In the trend analysis, the fibrosis-4 index (P < 0.001) and its components at baseline predicted the future fibrosis progression. In the generalized linear mixed model, an increase in HbA1c, but not BMI, was significantly associated with fibrosis progression (standardized coefficient 0.17 [95% CI 0.009-0.326]; P = 0.038) for subjects with NAFLD and diabetes. In gene set enrichment analyses, the pathways involved in zone 3 hepatocytes, central liver sinusoidal endothelial cells (LSECs), stellate cells, and plasma cells were coordinately altered in association with fibrosis progression and HbA1c elevation. Therefore, in subjects with NAFLD and diabetes, HbA1c elevation was significantly associated with liver fibrosis progression, independent of weight gain, which may be a valuable therapeutic target to prevent the pathological progression of NASH. Gene expression profiles suggest that diabetes-induced hypoxia and oxidative stress injure LSECs in zone 3 hepatocytes, which may mediate inflammation and stellate cell activation, leading to liver fibrosis. ARTICLE HIGHLIGHTS: It remains uncertain how diabetes and obesity contribute to histological courses of nonalcoholic fatty liver disease (NAFLD). Clinical features and gene expression signatures that predict or are associated with future liver fibrosis development were assessed in a serial liver biopsy study of subjects with NAFLD. An increase in HbA1c, but not BMI, was associated with liver fibrosis progression in the generalized linear mixed model. Considering hepatic gene set enrichment analyses, diabetes may enhance liver fibrosis via injuring central liver sinusoidal endothelial cells that mediate inflammation and stellate cell activation during NAFLD development.

Pulmonary miliary sarcoidosis masquerading the reactivation of tuberculosis 30 years later.

Key Clinical Message: Sarcoidosis may occur after treatment with pulmonary tuberculosis and requires differential diagnosis from tuberculosis reactivation. Miliary sarcoidosis should be promptly differentiated from miliary tuberculosis associated with high mortality. Abstract: Clinical, histological, and radiological similarities between sarcoidosis and tuberculosis render differential diagnosis challenging. The association between these two diseases has long been discussed, although the coexistence or subsequent occurrence of tuberculosis and sarcoidosis is rare. We report a case of miliary sarcoidosis that developed 30 years after tuberculous pleurisy treatment. Sarcoidosis may occur after treatment with pulmonary tuberculosis and requires differential diagnosis from tuberculosis reactivation. Although miliary sarcoidosis is uncommon, it should be promptly differentiated from miliary tuberculosis associated with high mortality. This study reignites the debate on the causal association between tuberculosis and sarcoidosis.

Efficient neutron capture therapy of glioblastoma with pteroyl-closo-dodecaborate-conjugated 4-(p-iodophenyl)butyric acid (PBC-IP).

Boron neutron capture therapy (BNCT) has been applied for clinical trials on glioblastoma patients since 1950s, however, the low survival rate under the treatments has hampered the widespread use of BNCT. In this study, we developed a novel boron agent, PBC-IP, which consists of three functional groups: FRα-targeting, 10B resource (twelve 10B atoms in the molecule), and albumin-binding moieties. PBC-IP was selectively taken up by glioma cell lines such as C6, F98, and U87MG cells and accumulated 10- to 20-fold higher than L-4­boronophenylalanine (BPA). PBC-IP administrated intravenously to the human glioblastoma (U87MG) xenograft model showed higher boron accumulation in tumors (29.8 µg [10B]/g at 6 h) than BPA (9.6 µg [10B]/g at 3 h) at a 25 mg [10B]/kg dose, effectively suppressing tumor growth after thermal neutron irradiation. PBC-IP administrated via convection-enhanced delivery (CED) accumulated in the F98 glioma orthotopic rat model, achieving 26.5 µg [10B]/g in tumors with tumor/normal (T/N) brain and tumor/blood (T/B) boron ratios of 37.8 and 94.6, respectively, 3 h after CED. Survival at 180 days after BNCT was 50% in the PBC-IP group and 70% in the combined BPA and PBC-IP groups, with no residual brain tumors.

Peptide Cyclization by the Use of Acylammonium Species.

Although cyclic peptides have become increasingly important as drugs, the most conventional peptide cyclization method using moderately active coupling agents suffers from a lot of waste and high cost as well as long reaction times and burdensome purification. Herein, we report an unconventional approach to peptide cyclization that uses acylammonium species generated from inexpensive and less wasteful Me2 NBn and ClCO2 i-Pr. Using this approach, we observed the desired rapid activation of the C-terminus of cyclization precursors by an acylammonium ion for rapid and epimerization/dimerization-free cyclization of synthetically challenging peptides, including a difficult cyclization involving N-methyl amide bond formation. The ease of purification, productivities, and reaction mass efficiencies of our approach were significantly superior to those in previous reports. We synthesized a previously reported versicotide D analogue, and our data indicated that its assigned stereostructure should be revised.

Development of a Peptide Sensor Derived from Human ACE2 for Fluorescence Polarization Assays of the SARS-CoV-2 Receptor Binding Domain.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the continuing emergence of infectious variants have caused a serious pandemic and a global economic slump since 2019. To overcome the situation and prepare for future pandemic-prone diseases, there is a need to establish a convenient diagnostic test that is quickly adaptable to unexpected emergence of virus variants. Here we report a fluorescent peptide sensor 26-Dan and its application to the fluorescence polarization (FP) assay for the highly sensitive and convenient detection of SARS-CoV-2. The 26-Dan sensor was developed by fluorescent labeling of the 26th amino acid of a peptide derived from the N-terminal α-helix of human angiotensin-converting enzyme 2 (hACE2) receptor. The 26-Dan sensor maintained the α-helical structure and showed FP changes in a concentration-dependent manner of the receptor binding domain (RBD) of the virus. The half maximal effective concentrations (EC50's) for RBD of Wuhan-Hu-1 strain, Delta (B.1.617.2), and Omicron (BA.5) variants were 51, 5.2, and 2.2 nM, respectively, demonstrating that the 26-Dan-based FP assay can be adaptable to virus variants that evade standard diagnostic tests. The 26-Dan-based FP assay could also be applied to model screening of a small molecule that inhibits RBD binding to hACE2 and identified glycyrrhizin as a potential inhibitor. The combination of the sensor with a portable microfluidic fluorescence polarization analyzer allowed for the detection of RBD in a femtomolar range within 3 min, demonstrating the assay could be a promising step toward a rapid and convenient test for SARS-CoV-2 and other possible future pandemic-prone diseases.