Construction of bispecific antibodies by specific pairing between the heavy chain and the light chain using removable SpyCatcher/SnoopCatcher units.

During the production of bispecific antibodies (bsAbs), nonspecific pairing results in low yields of target bsAb molecules, an issue known as the "mispairing problem." Several antibody engineering techniques have been developed to overcome mispairing issues. Here, we introduce "bsAb by external pairing and excision" (BAPE), a novel chain pairing method that induces specific chain pairing by fusing external SpyCatcher/Tag and SnoopCatcher/Tag units. These tags are then removed via protease cleavage. In this study, we applied this method to force the correct pairings of heavy and light chains while the heavy-chain pairing was achieved by the Knobs-into-Holes mutation. We then confirmed the formation of interchain bridges with covalent isopeptide bonds. Both anti-CD3/anti-Her2 and anti-CD3/anti-EGFR bsAbs displayed satisfactory target binding activities and in vitro cell-killing activity with activated T-cells.

Efficacy of immune checkpoint inhibitors plus platinum-based chemotherapy as 1st line treatment for patients with non-small cell lung cancer harboring HER2 mutations: Results from LC-SCRUM-Asia.

INTRODUCTION: HER2 mutations are reported to occur in 2%-5% of all cases of non-small cell lung cancer (NSCLC). The clinical outcomes in patients with HER2-mutant NSCLC treated with immune checkpoint inhibitors (ICIs) plus platinum-based chemotherapy as 1st line treatment still remain unclear. METHODS: Using the large-scale clinico-genomic database of LC-SCRUM-Asia, the clinico-genomic characteristics and therapeutic outcomes of patients with HER2-mutant NSCLC were investigated. RESULTS: Of the 15,251 patients with NSCLC enrolled in the LC-SCRUM-Asia database, tumor HER2 mutations were detected in 402 patients (2.6 %). The most common subtype of HER2 mutations was exon 20 in-frame insertions (79 %), followed in frequency by mutations in the tyrosine kinase domain other than Exon20ins (10 %) and mutations in extracellular domains (7 %). NSCLCs harboring HER2 mutations showed a higher tumor mutation burden (TMB) as compared with NSCLCs harboring EGFR mutations or ALK fusions (median: 4.22 vs. 2.54 and 2.52 mutation per megabase, respectively). Of the 402 patients, 268 patients had received platinum-based chemotherapy with ICIs (Chemo-ICI, n = 95) or without ICI (Chemo-alone, n = 173) as 1st line treatment. The progression-free survival (PFS) was significantly longer in the Chemo-ICI group as compared with the Chemo-alone group (median 8.5 vs. 6.3 months; HR [95 %CI]: 0.66 [0.50-0.88]; P < 0.005). Multivariate analysis identified use of ICIs in addition to platinum-based chemotherapy as an independent favorable prognostic factor for PFS. There was no significant difference in the overall survival between the patients of the Chemo-ICI and Chemo-alone groups (median 31.1 vs. 23.3 months; HR [95 %CI]: 0.80 [0.57-1.12], P = 0.20). CONCLUSIONS: Addition of ICIs to platinum-based chemotherapy in 1st line treatment may improve the PFS in patients with HER2-mutant NSCLC. The relatively high TMB might be involved in the prolongation of the PFS in patients with HER2-mutant NSCLC receiving platinum-based chemotherapy with ICIs.

ULK1-regulated AMP sensing by AMPK and its application for the treatment of chronic kidney disease.

Adenosine monophosphate (AMP)-activated protein kinase (AMPK) is a central kinase involved in energy homeostasis. Increased intracellular AMP levels result in AMPK activation through the binding of AMP to the γ-subunit of AMPK. Recently, we reported that AMP-induced AMPK activation is impaired in the kidneys in chronic kidney disease (CKD) despite an increase in the AMP/ATP ratio. However, the mechanisms by which AMP sensing is disrupted in CKD are unclear. Here, we identified mechanisms of energy homeostasis in which Unc-51-like kinase 1 (ULK1)-dependent phosphorylation of AMPKγ1 at Ser260/Thr262 promoting AMP sensitivity of AMPK. AMPK activation by AMP was impaired in Ulk1 knockout mice despite an increased AMP/ATP ratio. ULK1 expression is markedly downregulated in CKD kidneys, leading to AMP sensing failure. Additionally, MK8722, an allosteric AMPK activator, stimulated AMPK in the kidneys of a CKD mouse model (5/6th nephrectomy) via a pathway that is independent of AMP sensing. Thus, our study shows that MK8722 treatment significantly attenuates the deterioration of kidney function in CKD and may be a potential therapeutic option in CKD therapeutics.

Testosterone Replacement Therapy Can Improve a Biomarker of Liver Fibrosis in Hypogonadal Men: A Subanalysis of a Prospective Randomized Controlled Study in Japan (EARTH Study).

PURPOSE: We investigated the effects of testosterone replacement therapy (TRT) on the Fibrosis-4 (FIB-4) index among hypogonadal patients who were extracted from a randomized controlled study in Japan (the EARTH study). MATERIALS AND METHODS: Data of 186 patients (88 in the TRT group; 98 in the control group) were collected. The patients in the TRT group received intramuscular administration of testosterone enanthate (250 mg) every 4 weeks for 12 months. The patients' background information such as current medical history and lifestyle habits were collected. Waist circumference, body mass index, and body fat volume were measured at baseline and 12-month visit. Fasting blood sugar (FBS), hemoglobin A1c, total cholesterol, triglyceride (TG), and high-density lipoprotein cholesterol levels were collected at baseline and 12-month visit. The FIB-4 index was calculated according to age, aspartate aminotransferase, alanine transaminase, and platelet count. RESULTS: Except for serum FBS values, most of baseline characteristics were comparable between the TRT and control groups. When comparing the changes of each variable from baseline at 12-month visit in both groups, significant differences were found in waist circumference (p=0.00248), fat volume (p=0.00812), and platelet counts (p=0.0478), whereas a FIB-4 index did not change. On the contrary, in a subanalysis including only patients with a FIB-4 index ≥1.30 at baseline, a significant difference in a FIB-4 index (-0.10±0.39 vs. 0.04±0.44; p=0.0311) was observed with significant decreases in waist circumference, body fat volume, and TG levels, and an increase in platelet counts. The FIB-4 index was significantly decreased by TRT from 1.98±0.52 to 1.87±0.60 (p=0.0277). CONCLUSIONS: TRT for 12 months improved the FIB-4 index among hypogonadal men with a higher baseline FIB-4 index.

Reaction Mechanism of the Terminal Plastoquinone QB in Photosystem II as Revealed by Time-Resolved Infrared Spectroscopy.

The secondary plastoquinone (PQ) electron acceptor QB in photosystem II (PSII) undergoes a two-step photoreaction through electron transfer from the primary PQ electron acceptor QA, converting into plastoquinol (PQH2). However, the detailed mechanism of the QB reactions remains elusive. Here, we investigated the reaction mechanism of QB in cyanobacterial PSII core complexes using two time-revolved infrared (TRIR) methods: dispersive-type TRIR spectroscopy and rapid-scan Fourier transform infrared spectroscopy. Upon the first flash, the ∼140 µs phase is attributed to electron transfer from QA•- to QB, while the ∼2.2 and ∼440 ms phases are assigned to the binding of an internal PQ in a nearby cavity to the vacant QB site and an external PQ traveling to the QB site through channels, respectively, followed by immediate electron transfer. The resultant QB•- is suggested to be in equilibrium with QBH•, which is protonated at the distal oxygen. Upon the second flash, the ∼130 µs and ∼3.3 ms phases are attributed to electron transfer to QBH• and the protonation of QB•- followed by electron transfer, respectively, forming QBH-, which then immediately accepts a proton from D1-H215 at the proximal oxygen to become QBH2. The resultant D1-H215 anion is reprotonated in ∼22 ms via a pathway involving the bicarbonate ligand. The final ∼490 ms phase may reflect the release of PQH2 and its replacement with PQ. The present results highlight the importance of time-resolved infrared spectroscopy in elucidating the mechanism of QB reactions in PSII.

Impact of Pre-operative Lymphocyte and Albumin Combination on Adjuvant Chemotherapy and Prognosis of Gastric Cancer.

BACKGROUND/AIM: Cancer-associated systemic inflammatory response is a pivotal indicator of tumor progression and prognosis in various cancers. "Lymphocyte × albumin (LA)" is a prognostic inflammatory marker in rectal cancer. This study examined the correlation between LA, complete adjuvant chemotherapy (ACT), and prognosis in patients with gastric cancer (GC) who underwent radical gastrectomy. PATIENTS AND METHODS: We retrospectively evaluated 108 patients with stage II/III GC who underwent radical gastrectomy at our institute between January 2015 and December 2021. Survival was assessed using Kaplan-Meier and Cox regression analyses. Factors associated with complete ACT were identified using logistic regression analysis. RESULTS: Of the 108 patients with GC, 60 (55.6%) and 41 (38.0%) initiated and completed ACT, respectively. In multivariate analysis, the pre-operative LA was an independent factor for complete ACT [hazard ratio (HR)=0.35, 95% confidence interval (CI)=0.121-0.995; p=0.049]. In addition, age, pre-operative creatinine clearance, neutrophil-to-lymphocyte ratio, modified Glasgow prognostic score, and poor overall survival were significantly associated with low LA (<7,474). LA was an independent prognostic factor for overall survival in univariate analysis (HR=2.29, 95%CI=1.020-5.145; p=0.045) but not in multivariate analysis (HR=2.00, 95%CI=0.882-4.552; p=0.097). CONCLUSION: Pre-operative LA is a useful marker for predicting complete ACT and prognosis of patients with GC following radical gastrectomy.

Impact of Frailty Risk on Functional Outcome after Aneurysmal Subarachnoid Hemorrhage: A Historical Cohort Study.

We evaluated the utility of the Hospital Frailty Risk Score (HFRS) as a predictor of adverse events post-hospitalization in a retrospective analysis of patients undergoing neurosurgical procedures due to aneurysmal subarachnoid hemorrhage (SAH). This historical cohort study analyzed the data of patients hospitalized with aneurysmal SAH (n = 1,343) between April 2014 and August 2020 who were registered in the JMDC database. We used HFRS to classify the patients into the low-frailty risk group (HFRS < 5) and high-frailty risk group (HFRS ≥ 5). The primary outcome was a modified Rankin Scale (mRS) score of 0-2 points at discharge. Of 1,343 patients, 1,001 (74.5%) and 342 (25.5%) were in the low- and high-frailty risk groups, respectively. A high-frailty risk was negatively associated with a mRS score of 0-2 at discharge (high-frailty risk group: odds ratio 0.4; 95% confidence interval [CI]: 0.3-0.6) and home discharge (high-frailty risk group: odds ratio 0.5; 95% CI: 0.4-0.7). A high-frailty risk was negatively associated with Barthel Index gain (high-frailty risk group: coefficient -10.4, 95% CI: -14.7 to -6.2) and had a longer length of stay (high-frailty risk group: coefficient 8.4, 95% CI: 5.1-11.7). HFRS could predict adverse outcomes during hospitalization of aneurysmal SAH patients.

A comparison of the efficacy of dutasteride on reducing lower urinary tract symptoms among patients with small versus large benign prostatic hyperplasia.

Background: Only a few reports have currently studied the efficacy of dutasteride in patients with small benign prostatic hyperplasia (BPH). We investigated the efficacy of dutasteride on reducing lower urinary tract symptoms among them. Materials and methods: A total of 81 patients with BPH who completed 52weeks of 0.5?mg dutasteride treatment were enrolled. Each patient filled out the International Prostatic Symptom Score (IPSS) and overactive bladder symptom score (OABSS) at baseline and at the 6- and 12-month follow-up visits. Total testosterone, prostate-specific antigen, adenoma/prostate volume (PV), uroflowmetry analysis, and postvoid residual volume were evaluated at baseline and at the 12-month follow-up visit. The enrolled patients were divided into 2 groups according to PV at baseline. The groups were as follows: Group A (PV ≥ 30 mL) and Group B (PV < 30 mL). Results: Groups A and B had mean PVs of 52.1 and 23.6 mL and mean IPSS scores of 16.7 and 14.4, respectively. Group A had significantly higher OABSS and prostate-specific antigen levels at baseline than Group B, while no significant differences in any other baseline characteristics was observed. After dutasteride treatment, adenoma volume and PV decreased significantly, while testosterone level showed a significant increase in both groups. Group A showed significant improvements in the total IPSS, voiding and storage subscore of the IPSS, OABSS, maximum flow rate, and postvoid residual volume. Group B, on the other hand, also showed significant improvements only in the total IPSS, voiding subscore of the IPSS, and maximum flow rate. Conclusions: The present study suggests a possible beneficial effect of dutasteride treatment on the reduction of lower urinary tract symptoms in patients with small and large BPH. However, the effectiveness of dutasteride was limited compared to patients with large BPH (PV ≥ 30 mL).

Ultrasound-Guided Posteromedial Semitendinosus Tendon Harvest.

Hamstring tendon autografts have emerged as the graft of choice by over 50% of sports surgeons worldwide performing anterior cruciate ligament reconstruction. A more recent technique in harvesting the semitendinosus tendon, the posteromedial approach, afforded multiple benefits compared with the anteromedial approach. However, for the inexperienced surgeon, the current techniques may not be as simple because of decreased tendon tension after general anesthesia induction and subcutaneous layer dissection, making it difficult to palpate the semitendinosus tendon. By utilizing ultrasound to assist us during our harvest, we can perform the procedure with increased accuracy and efficiency, which leads to a safer, more proficient, and less invasive procedure.

Dose modification in enzalutamide and abiraterone plus prednisolone for castration-resistant prostate cancer: A subanalysis from the ENABLE study for PCa.

BACKGROUND: A head-to-head comparison between enzalutamide (ENZ) and abiraterone plus prednisolone (ABI) revealed similar survival benefits for castration-resistant prostate cancer (CRPC) in the ENABLE study for PCa. Considering that a dose reduction of ENZ and ABI has demonstrated sufficient inhibitory ability of androgen receptor (AR) signaling, we analyzed the efficacy of modified doses of these agents in the ENABLE study for PCa. METHODS: This investigator-initiated, multicenter, randomized controlled trial that was conducted in Japan analyzed the prespecified survival endpoints, prostate-specific antigen (PSA) response rate ( ≥50% decline from baseline), and safety profile in patients treated with modified doses (ENZ ≤ 120 mg/day, ABI ≤ 750 mg/day) compared with those treated with a standard dose (ENZ 160 mg/day, ABI 1000 mg/day) as a starting dose. RESULTS: In total, 92 patients in each arm were treated and analyzed; 16 patients were treated with a modified dose in both the ENZ and ABI arms, respectively. Moreover, 32 patients treated with modified doses showed a significantly better time to PSA progression (TTPP) and overall survival (OS) compared with the 152 patients treated with a standard dose (HR 0.47, 95%CI 0.27-0.83, p = 0.0379, and HR 0.35, 95%CI 0.19-0.63, p = 0.0162). Despite a significantly longer TTPP in the modified ABI group than in the standard ABI group (HR 0.29, 95%CI 0.14-0.62, p = 0.0248), no significant difference was observed in the TTPP between the modified and standard ENZ groups (p = 0.5366). Furthermore, similar adverse event rates and grades were observed in each treatment dose group. CONCLUSIONS: The modified doses of ABI showed better TTPP than the standard dose of ABI and may be a potential treatment option for CRPC patients; however, its mechanism is still unclear, although its ability to suppress AR signaling is equivalent to that of a standard dose.

Lipid Peroxidation Regulators GPX4 and FSP1 as Prognostic Markers and Therapeutic Targets in Advanced Gastric Cancer.

Gastric cancer is one of the most common cancers worldwide, and new therapeutic strategies are urgently needed. Ferroptosis is an intracellular iron-dependent cell death induced by the accumulation of lipid peroxidation, a mechanism different from conventional apoptosis and necrosis. Therefore, induction of ferroptosis is expected to be a new therapeutic strategy. Glutathione peroxidase 4 (GPX4) and ferroptosis suppressor protein 1 (FSP1) have been identified as the major inhibitors of ferroptosis. Herein, we performed immunohistochemistry for GPX4, FSP1, and 4-HNE using tissues from patients with gastric cancer and investigated the relationship between these factors and prognosis. Patients with high GPX4 expression or high GPX4 expression and low 4-HNE accumulation tended to have a poor prognosis (p = 0.036, 0.023), whereas those with low FSP1 expression and high 4-HNE accumulation had a good prognosis (p = 0.033). The synergistic induction of cell death by inhibiting GPX4 and FSP1 in vitro was also observed, indicating that the cell death was non-apoptotic. Our results indicate that the expression and accumulation of lipid peroxidation-related factors play an important role in the clinicopathological significance of gastric cancer and that novel therapeutic strategies targeting GPX4 and FSP1 may be effective in treating patients with gastric cancer who have poor prognosis.

Synaptotagmin 4 Supports Spontaneous Axon Sprouting after Spinal Cord Injury.

Injuries to the central nervous system (CNS) can cause severe neurological deficits. Axonal regrowth is a fundamental process for the reconstruction of compensatory neuronal networks after injury; however, it is extremely limited in the adult mammalian CNS. In this study, we conducted a loss-of-function genetic screen in cortical neurons, combined with a Web resource-based phenotypic screen, and identified synaptotagmin 4 (Syt4) as a novel regulator of axon elongation. Silencing Syt4 in primary cultured cortical neurons inhibits neurite elongation, with changes in gene expression involved in signaling pathways related to neuronal development. In a spinal cord injury model, inhibition of Syt4 expression in cortical neurons prevented axonal sprouting of the corticospinal tract, as well as neurological recovery after injury. These results provide a novel therapeutic approach to CNS injury by modulating Syt4 function.

Causative agent for perioperative anaphylaxis in a child with autism successfully identified using the intradermal test under general anesthesia.

BACKGROUND: The skin-prick and intradermal tests are the main diagnostic methods used to identify the causative agent in patients with suspected perioperative anaphylaxis. Although the intradermal test is more sensitive than the skin-prick test, multiple intradermal injections can be painful for children. Here, we present the case of a child with autism and suspected perioperative anaphylaxis. The causative agent was successfully identified using the intradermal test under general anesthesia. CASE PRESENTATION: An 8-year-old boy with autism developed anaphylaxis during general anesthesia for the fourth cleft lip and palate surgery. An allergic workout was performed, but both the skin-prick and basophil activation tests for suspected causative agents yielded negative results. The patient was afraid of multiple injections, and an intradermal test was performed under general anesthesia by anesthesiologists and allergists. Piperacillin was confirmed as the causative agent, and subsequent surgery using the same anesthetic agents without piperacillin was uneventful. CONCLUSIONS: Concerted efforts should be made to identify the causative agent for diagnosing perioperative anaphylaxis.

Turn-Over Technique for Inserting and Stabilizing Guiding Sheath without Straightening It during Carotid Artery Stenting via Trans-Brachial Approach in Cases with Severe Vascular Access Conditions.

Objective: In cases of severe atherosclerosis or tortuous arteries, inserting the guiding sheath into the target vessel is challenging. Here, we present the turn-over technique for inserting and stabilizing the guiding sheath without straightening it during carotid artery stenting (CAS). Case Presentation: Two patients with severe left internal carotid artery stenosis underwent CAS via the trans-brachial approach. Although inserting the guiding sheath into the common carotid artery using conventional techniques was challenging, we succeeded in inserting it into the target vessel using the "turn-over technique." At first, the guidewire was U-turned just above the aortic valve and inserted into the left external carotid artery. The inner catheter and guiding sheath were then followed along the guidewire to the left common carotid artery, and the guidewire and inner catheter were removed. The guiding sheath was stable in a U-turned position, and straightening the guiding sheath was difficult. Devices such as stents and balloons could be delivered without problems, and CAS was completed with the guiding sheath in a U-turned position. Conclusion: The turn-over technique for inserting and stabilizing the guiding sheath for CAS via the trans-brachial approach is an option in cases of difficult catheter access due to atherosclerosis or tortuous arteries.

Polyorchidism in a young Sprague-Dawley rat.

Duplicate testes lined in series were observed in the right scrotum of a 6-week-old Sprague-Dawley rat in a single-dose toxicity study. Of the two right testicles, one was spherical and less than half the size of a normal testis. The other was oval-shaped, slightly smaller than a normal testis, and possessed clear, tortuous blood vessels similar to those of a normal testis. Each right testis was grossly separated but faced the intertesticular adipose tissue and was sparsely joined by thin cord-like structures. Only one epididymis covered or encompassed the two right testes. The caput epididymis was attached to the smaller spherical testis, whereas the cauda epididymis was attached to the oval testis. Histopathological examination revealed that the smaller spherical testis on the right side and the testis on the left side were normal. The oval-shaped testis on the right exhibited markedly dilated degenerative seminiferous tubules with one to two layers of Sertoli or germ cells, and almost no spermatogenesis was observed. Multinucleated germ cells were observed in the lumen of the degenerated seminiferous tubules. The right epididymis was morphologically normal and contained few sperm in the epididymal duct of the tail. The cord-like structures between duplicate testes comprised fibrous and adipose tissues. Single efferent ductules, ectopic cartilage, and skeletal muscle tissues were buried in the adipose tissue. To our knowledge, this is the first report of spontaneous polyorchidism in a rodent.

Molecular microbiological profiling of bottled unsweetened tea beverages: A screening experiment.

To explore the potential storage and safety of drinking leftover bottled tea beverages from various manufacturers after direct drinking from bottles, we conducted a screening experiment on the growth of salivary bacteria in plastic bottles of tea. The diluted saliva samples from 10 participants were inoculated into the test bottled beverages, which resulted in bacteria, particularly former members of the genus Lactobacillus, growing in some green tea beverages with a neutral pH. In contrast, tea beverages with less bacterial growth contained Streptococcus spp., and the leftovers may be safe to store and drink again.

Impact of Early Rehabilitation after Endovascular Treatment for Peripheral Arterial Disease.

Objectives: The prevalence of peripheral arterial disease (PAD) is on the rise, with endovascular treatment being a widely accepted surgical intervention. Patients with PAD often experience reduced activities of daily living (ADL). Therefore, we conducted a retrospective cohort study to investigate the impact of early rehabilitation after endovascular treatment in patients with PAD. Methods: Using data from the JMDC hospital database, the study included 529 patients who were hospitalized for PAD and underwent endovascular treatment. Patients were classified into two independent variables: early rehabilitation group (rehabilitation started within 2 days postoperatively) and control group (rehabilitation started within 3-7 days postoperatively). The outcome measures were the occurrence of hospital-associated disability (HAD) and duration of hospitalization. Results: Unadjusted data showed that the early rehabilitation group (n=469) had fewer HAD events (8.5% vs. 23.3%, P <0.001) and a shorter mean hospitalization duration (4.4 vs. 18.9 days, P <0.001) than the control group (n=60). The difference remained significant after adjustment by propensity score analysis. Conclusions: In patients with PAD, early rehabilitation after endovascular treatment may be beneficial in preventing the development of HAD and reducing the duration of hospitalization.

Nutritional Care for Adults in Nursing Homes: A Scoping Review.

The need for improved nutrition in older adults requiring care has been acknowledged, but, to the best of our knowledge, there is a lack of systematic review and integration of nutritional care studies with older adults in nursing homes. This scoping review aimed to examine the scope and nature of nutritional care research for older adults in nursing homes and to identify research gaps, following the guidelines of the Joanna Briggs Institute. We found varied nutritional care for older adults living in nursing homes, including individualized sessions, such as nutrition counseling, the addition of foods and preparations for increased nutritional intake, and the maintenance of an eating environment, such as feeding assistance and calling. The nutritional care identified in this scoping review also included studies that have improved the nutritional status of older adults in nursing homes by implementing educational programs for care staff. For future research on effective nutritional care for older adults in nursing homes, we suggest evaluating both short- and long-term intervention effects with an adequate sample size.