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Despite the importance of antigen-specific T cells in infectious disease, characterizing and tracking clonally amplified T cells during the progression of a patient's symptoms remain unclear. Here, we performed a longitudinal, in-depth single-cell multiomics analysis of samples from asymptomatic, mild, usual severe, and delayed severe patients of SARS-CoV-2 infection. Our in-depth analysis revealed that hyperactive or improper T-cell responses were more aggressive in delayed severe patients. Interestingly, tracking of antigen-specific T-cell receptor (TCR) clonotypes along the developmental trajectory indicated an attenuation in functional T cells upon severity. In addition, increased glycolysis and interleukin-6 signaling in the cytotoxic T cells were markedly distinct in delayed severe patients compared to usual severe patients, particularly in the middle and late stages of infection. Tracking B-cell receptor clonotypes also revealed distinct transitions and somatic hypermutations within B cells across different levels of disease severity. Our results suggest that single-cell TCR clonotype tracking can distinguish the severity of patients through immunological hallmarks, leading to a better understanding of the severity differences in and improving the management of infectious diseases by analyzing the dynamics of immune responses over time., (© 2023 Wiley Periodicals LLC.)

Most patients with stroke suffer from complications and these include dysphagia. Dysphagia can cause malnutrition, and malnutrition affects prognosis and recovery. However, there is a lack of accurate studies on the nutritional status of stroke patients with dysphagia and its associated factors in different phases of stroke. This study retrospectively investigated 620 stroke patients who underwent a videofluoroscopic swallowing study (VFSS) due to dysphagia, from March 2018 to February 2021. The study aimed to evaluate the nutritional state and associated factors of malnutrition in acute and subacute stroke patients with dysphagia. Serum albumin and percentage of current weight to ideal weight were used to determine nutritional status. Malnutrition was observed in 58.9 and 78.9% of acute and subacute stroke patients. Exact logistic regression analysis revealed that old age and high penetration-aspiration scale score were significantly associated factors for malnutrition in patients with acute stroke. Old age, stroke history, bilateral hemiplegia, high modified Rankin score, low Korean Mini-Mental State Examination, pneumonia, and high functional dysphagia score were significantly associated factors for malnutrition in patients with subacute stroke. Patients with these associated factors in each phase of stroke require active nutritional assessment and care to decrease the risk of malnutrition.

Background: Little is known about the immune determinants for severe coronavirus disease 2019 (COVID-19) in individuals vaccinated against severe acute respiratory syndrome coronavirus 2. We therefore attempted to identify differences in humoral and cellular immune responses between patients with non-severe and severe breakthrough COVID-19., Methods: We prospectively enrolled hospitalized patients with breakthrough COVID-19 (severe and non-severe groups) and uninfected individuals who were vaccinated at a similar time (control group). Severe cases were defined as those who required oxygen therapy while hospitalized. Enzyme-linked immunosorbent assays and flow cytometry were used to evaluate humoral and cellular immune responses, respectively., Results: Anti-S1 IgG titers were significantly lower in the severe group than in the non-severe group within 1 week of symptom onset and higher in the non-severe group than in the control group. Compared with the control group, the cellular immune response tended to be diminished in breakthrough cases, particularly in the severe group. In multivariate analysis, advanced age and low anti-S1 IgG titer were associated with severe breakthrough COVID-19., Conclusions: Severe breakthrough COVID-19 might be attributed by low humoral and cellular immune responses early after infection. In the vaccinated population, delayed humoral and cellular immune responses may contribute to severe breakthrough COVID-19., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Lee, Choe, Kang, Lee, Song, Bang, Kim, Kim, Kim, Kim, Kim, Lee, Shin, Park, Park and Oh.)

The fourth vaccination dose confers additional protective immunity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in individuals with no prior coronavirus disease-19 (COVID-19). However, its immunological benefit against currently circulating BA.4/5 is unclear in individuals who have received a booster shot and been infected with Omicron variant BA.1/2. We analyzed immune responses in whom had been boosted once and did not have COVID-19 (n = 16), boosted once and had COVID-19 when BA.1/2 was dominant in Korea (Hybrid-6M group, n = 27), and boosted twice and did not have COVID-19 (Vx4 group, n = 15). Antibody binding activities against RBD o BA.1 and RBD o BA.4/5 , antigen-specific memory CD4 + and CD8 + T-cell responses against BA.4/5, and B-cell responses against SARS-CoV-2 wild-type did not differ statistically between the Hybrid-6M and Vx4 groups. The humoral and cellular immune responses of the Hybrid-6M group against BA.4/5 were comparable to those of the Vx4 group. Individuals who had been boosted and had an Omicron infection in early 2022 may not have high priority for an additional vaccination., (© 2023 Wiley Periodicals LLC.)

Background: Aducanumab (Adu), which is a human IgG1 monoclonal antibody that targets oligomer and fibril forms of beta-amyloid, has been reported to reduce amyloid pathology and improve impaired cognition after administration of a high dose (10 mg/kg) of the drug in Alzheimer's disease (AD) clinical trials. The purpose of this study was to investigate the effects of a lower dose of Adu (3 mg/kg) with enhanced delivery via focused ultrasound (FUS) in an AD mouse model., Methods: The FUS with microbubbles opened the blood-brain barrier (BBB) of the hippocampus for the delivery of Adu. The combined therapy of FUS and Adu was performed three times in total and each treatment was performed biweekly. Y-maze test, Brdu labeling, and immunohistochemical experimental methods were employed in this study. In addition, RNA sequencing and ingenuity pathway analysis were employed to investigate gene expression profiles in the hippocampi of experimental animals., Results: The FUS-mediated BBB opening markedly increased the delivery of Adu into the brain by approximately 8.1 times in the brains. The combined treatment induced significantly less cognitive decline and decreased the level of amyloid plaques in the hippocampi of the 5×FAD mice compared with Adu or FUS alone. Combined treatment with FUS and Adu activated phagocytic microglia and increased the number of astrocytes associated with amyloid plaques in the hippocampi of 5×FAD mice. Furthermore, RNA sequencing identified that 4 enriched canonical pathways including phagosome formation, neuroinflammation signaling, CREB signaling and reelin signaling were altered in the hippocami of 5×FAD mice receiving the combined treatment., Conclusion: In conclusion, the enhanced delivery of a low dose of Adu (3 mg/kg) via FUS decreases amyloid deposits and attenuates cognitive function deficits. FUS-mediated BBB opening increases adult hippocampal neurogenesis as well as drug delivery. We present an AD treatment strategy through the synergistic effect of the combined therapy of FUS and Adu., (© 2022. The Author(s).)

In this study, we investigated the effects of carbonated water concentration on swallowing function using surface electromyography (sEMG). Healthy subjects (n = 52, 26.77 ± 3.21 years old) were asked to perform two swallows each of noncarbonated water, low-concentration carbonated water, medium-concentration carbonated water, and high-concentration carbonated water. Onset time, the mean sEMG activity amplitude, and duration of muscle activity in each swallow were measured and analyzed for orbicularis oris, masseter, submental muscle complex and infrahyoid muscles. Onset time significantly decreased and mean sEMG activity amplitude significantly increased with carbonation concentration. Therefore, stimulation with carbonation can be effective for modulating a faster and stronger swallow in the oral and pharyngeal phases of swallowing, and its effect on amplitude was greater in the oral phase than in the pharyngeal phase.Clinical Trials Registration This study is registered with Clinical Research Information Service (KCT0005925)., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Introduction: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its variants brought waves of pandemics with breakthrough infections in vaccinated individuals. We analyzed the antibody responses after primary and booster vaccination in healthy controls (HC) and patients with early breast cancer (BC)., Methods: In this prospective longitudinal cohort study, the binding activity of serum antibody level against spike proteins and antigens of SARS-CoV-2 variants was measured within 21 days after each vaccination in the BC group and HC group., Results: All participants, 40 in the BC and 20 in the HC group, had increased antibody response after vaccination. BC group, however, had weaker humoral responses than the HC group (IgG: 1.5, 2.3, 2.5-folds in BC vs. 1.9, 3.6, 4.0-folds in HC after each dose; IgA: 2.1, 3.0, 3.6-folds in BC vs. 4.2, 10.4, 5.2-folds in HC after each dose, respectively). Those under concurrent cytotoxic chemotherapy had weaker antibody response than the non-cytotoxic treatment group and HC. Adjunct use of steroids and age were not significant risk factors. The levels of binding antibody against the Delta and the Omicron (BA1) variants were lower than the wild-type, especially in BC., Conclusion: In the waves of new sub-variants, our study suggests that an additional dose of vaccinations should be recommended according to the anti-cancer treatment modality in patients with BC who had received booster vaccination., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Kim, Jeong, Lee, Lee, Kang, Choe, Kim, Oh, Lee, Park, Lee and Im.)

Background: In recent years, many studies have found possible links between gene polymorphisms and venous thromboembolism (VTE). By identifying genetic risk factors before facing environmental risk factors such as surgical interventions and COVID-19 vaccination, we could rapidly respond to the risk of VTE. The aim of this study was to perform an umbrella review of genetic variants related to VTE. Integrative gene analysis of VTE was performed to identify critical genetic variations., Methods: This study conducted an umbrella review of systematic reviews and meta-analyses. All included studies were selected from the PubMed/MEDLINE database. To select eligible studies, the following variables were extracted: first author name; effect size of each study genetic variant; year of publication; the number of studies included in each article; ethnicity, sample size, P values, and heterogeneity estimates. To assess cumulative evidence in genetic epidemiology about effects of gene polymorphisms on VTE, Human Genome Epidemiology Network's Venice criteria were used. Methodological quality assessment was conducted with JBI Critical Appraisal Checklist for Systematic Reviews and Research Syntheses., Results: Genes provided in the present study with genetic variants associated with VTE were FVL (G1691A), Prothrombin (G20210A), MTHFR (C677T, A1298C), PAI-1 (4G/5G), factor VII activating protease (1601G > A), and endothelial protein C receptor (g.6936A_G, c.4600A_G). Among them, variants in FVL, Prothrombin, MTHFR, and PAI-1 showed high significance. Particularly, variants in Prothrombin (G20210A), MTHFR (C677T), and PAI-1 (4G/5G) had more than 2 types of model significance., Conclusion: The present study performed a systematic review of genetic variants associated with VTE. Our results could lead to a more comprehensive understanding of VTE etiology. These results could give a strategy of prediagnosis about evaluating individual risks of VTE who might be exposed to environmental risk factors., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2022 the Author(s). Published by Wolters Kluwer Health, Inc.)

Background: This study was performed to update the current evidence and evaluate the effects of robot-assisted rehabilitation (RAR) in comparison with conventional rehabilitation (CR) in patients following total knee (TKR) or hip replacements (THR)., Methods: PubMed Central, OVID Medline, Cochrane Collaboration Library, and EMBASE for a comprehensive search for all relevant studies, from database inception to July 2022. The following inclusion criteria were used to determine eligibility for studies: randomized and matched controlled trials recruiting men and women who underwent TKR and THR; and studies examining the effect of RAR on outcome measures of physical function and pain., Results: A total of 9 studies (230 patients) were included in this review and 4 were included in the meta-analysis. The meta-analysis of 2 studies showed that Hybrid Assistive Limb (HAL) training for 5 days, significantly improved pain measured on a visual analogue scale, compared to CR in patients following TKR (SMD = 1.05, 95% confidence interval [Cl] 0.39-1.71). Heterogeneity for I2 value was lower than moderate (tau^2 = 0.0121; I2 = 5%; P = .30). There were 2 studies that assessed self-selected walking speed. The meta-analysis of these studies showed that HAL training was significantly superior to CR in patients following TKR (SMD = 48.70, 95% Cl -50.53 to 147.94) at 2 months. A high heterogeneity was detected (P < .01; I2 = 97%)., Conclusion: The result of this systematic review and meta-analysis suggest that RAR may be an effective treatment in TKR and THR patients. However, high-quality studies are needed to verify the long-term effect on their recovery., Competing Interests: The authors have no funding and conflicts of interest to disclose., (Copyright © 2022 the Author(s). Published by Wolters Kluwer Health, Inc.)

Background: Humoral immunity to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) may wane rapidly in persons recovered from mild coronavirus disease 2019 (COVID-19), but little is known about the longevity., Methods: Serum samples were obtained 8, 12, and 18 months after infection from 20 patients with mild COVID-19. The binding activities of serum antibodies (immunoglobulin [Ig]A, IgG, and IgM) against SARS-CoV-2 antigens of the Wuhan-1 reference strain (wild-type) and the B.1.1.7, P.1, B.1.167.2, and B.1.1.529 variants were measured by enzyme-linked immunosorbent assays. Neutralizing antibody titers were measured using a cytopathic effect-based live virus neutralization assay., Results: Serum IgA and IgG antibodies against spike or receptor-binding domain (RBD) protein of wild-type SARS-CoV-2 were detected for up to 18 months, and neutralizing antibodies persisted for 8 to 18 months after infection. However, any significant antibody responses against RBD proteins of SARS-CoV-2 variants were not observed, and median neutralizing antibody titers against the Delta variant at 8, 12, and 18 months were 8- to 11-fold lower than against wild-type viruses (P<.001)., Conclusions: Humoral immunity persisted for up to 18 months after SARS-CoV-2 infection in patients with mild COVID-19. However, humoral immune activity against more recently circulating variants was reduced in this population., Competing Interests: Potential conflicts of interest. All authors: No reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest., (© The Author(s) 2022. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Background: To compare the incidence and severity of ipsilateral shoulder dysfunction and lymphedema of 2 groups of patients needing to undergo unilateral breast cancer surgery, one of which had only received printed education materials and the other group which had received educational materials plus preoperative education., Methods: We selected 61 patients who had been diagnosed with unilateral breast cancer and planned to undergo surgery. Before surgery, patients were randomly assigned, either to a control group that only received printed education materials about exercise for shoulder pain relief and lymphatic edema prevention following breast cancer surgery, or to an experimental group that received the printed education material with personal education. Participants were evaluated at 1, 3, 6, and 12 months after the surgery. To evaluate the impairment of shoulder function, we measured the passive shoulder range of motion (ROM), the degree of pain as visual analog scale (VAS), the short version of the disability of arm, shoulder, and hand (short DASH) scores, and the shoulder pain and disability index (SPADI). We checked arm circumferences to evaluate lymphedema., Results: There was no significant difference in demographic or clinical variables between the control and experimental groups. The experimental group showed significantly less limitation in abduction (P = .042) and forward flexion (P = .039) in the 6 months following surgery. Change in the VAS, short DASH, and SPADI scores were 1.633 (P < .001), 2.167 (P < .001), and 4.1 (P = .003) at 1 month following surgery, respectively. These then decreased with time. These changes started before shoulder ROM and arm circumference changes had occurred, which had started 3 months following surgery., Conclusions: Preoperative education might be helpful for the prevention of a shoulder ROM limitation, and we need to focus on pain and disability in patients immediately following breast cancer surgery, and then on ROM and lymphedema., Competing Interests: The authors have no funding and conflicts of interest to disclose., (Copyright © 2022 the Author(s). Published by Wolters Kluwer Health, Inc.)

Tumor acidosis, a common phenomenon in solid cancers such as breast cancer, is caused by the abnormal metabolism of cancer cells. The low pH affects cells surrounding the cancer, and tumor acidosis has been shown to inhibit the activity of immune cells. Despite many previous studies, the immune surveillance mechanisms are not fully understood. We found that the expression of PD-L1 was significantly increased under conditions of extracellular acidosis in MDA-MB-231 cells. We also confirmed that the increased expression of PD-L1 mediated by extracellular acidosis was decreased when the pH was raised to the normal range. Gene set enrichment analysis (GSEA) of public breast cancer patient databases showed that PD-L1 expression was also highly correlated with IL-6/JAK/STAT3 signaling. Surprisingly, the expression of both phospho-tyrosine STAT3 and PD-L1 was significantly increased under conditions of extracellular acidosis, and inhibition of STAT3 did not increase the expression of PD-L1 even under acidic conditions in MDA-MB-231 cells. Based on these results, we suggest that the expression of PD-L1 is increased by tumor acidosis via activation of STAT3 in MDA-MB-231 cells., (© 2022. The Author(s).)

Background: Practical guidance is needed regarding the vaccination of coronavirus disease 2019 (COVID-19) convalescent individuals in resource-limited countries. It includes the number of vaccine doses that should be given to unvaccinated patients who experienced COVID-19 early in the pandemic., Methods: We recruited COVID-19 convalescent individuals who received one or two doses of an mRNA vaccine within 6 or around 18 months after a diagnosis of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infection. Their samples were assessed for IgG-binding or neutralizing activity and cell-mediated immune responses against SARS-CoV-2 wild-type and variants of concern., Results: A total of 43 COVID-19 convalescent individuals were analyzed in the present study. The results showed that humoral and cellular immune responses against SARS-CoV-2 wild-type and variants of concern, including the Omicron variant, were comparable among patients vaccinated within 6 versus around 18 months. A second dose of vaccine did not significantly increase immune responses., Conclusion: One dose of mRNA vaccine should be considered sufficient to elicit a broad immune response even around 18 months after a COVID-19 diagnosis., (© 2022. The Author(s).)

Although endothelin receptor type A (ET A ) is emerging as a novel anticancer target, previously only small molecule drugs have been available as ET A antagonists. Ju and colleagues have successfully developed a functional anti-ET A antibody for colorectal cancer., Competing Interests: Declaration of interests The authors declare no conflicts of interest., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Background: Despite the fact of ongoing worldwide vaccination programs for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), understanding longevity, breadth, and type of immune response to coronavirus disease-19 (COVID-19) is still important to optimize the vaccination strategy and estimate the risk of reinfection. Therefore, we performed thorough immunological assessments 1 year post-COVID-19 with different severity., Methods: We analyzed peripheral blood mononuclear cells and plasma samples at 1 year post-COVID-19 in patients who experienced asymptomatic, mild, and severe illness to assess titers of various isotypes of antibodies (Abs) against SARS-CoV-2 antigens, phagocytic capability, and memory B- and T-cell responses., Findings: A total of 24 patients (7, 9, and 8 asymptomatic, mild, and severe patients, respectively) and eight healthy volunteers were included in this study. We firstly showed that disease severity is correlated with parameters of immune responses at 1 year post-COVID-19 that play an important role in protecting against reinfection with SARS-CoV-2, namely, the phagocytic capacity of Abs and memory B-cell responses., Interpretation: Various immune responses at 1 year post-COVID-19, particularly the phagocytic capacity and memory B-cell responses, were dependent on the severity of the prior COVID-19. Our data could provide a clue for a tailored vaccination strategy after natural infection according to the severity of COVID-19., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Kang, Kim, Hong, Kim, Lee, Chang, Choe, Kim, Kim, Seo, Song, Lee, Shin, Kim, Lee, Park, Kim and Oh.)

Diabetes mellitus (DM) characterized by hyperglycemia is a chronic metabolic disorder that leads to many symptoms and vascular complications. Despite the close association between DM and cancer progression, the response and role of endothelial cells (ECs) under diabetic conditions in tumor metastasis remain to be elucidated. In this study, we sought to determine whether and how ECs under diabetic conditions contribute to tumor metastasis. We have taken advantage of syngeneic mouse tumor models of Lewis lung carcinoma (LLC) and melanoma (B16F10) cells and a streptozotocin (STZ)-induced hyperglycemia model. We demonstrated that hyperglycemia increased the metastasis of LLC and B16F10 cells in an experimental metastasis model with an intravenous injection of the tumor cells. We also found that hyperglycemia promoted lung metastasis of tumor cells by increasing the adhesiveness of ECs to facilitate the adhesion of tumor cells to ECs rather than affecting the metastatic behavior of tumor cells themselves. From the analysis of gene expression in primary lung ECs from STZ-treated mice, we identified that vWF promoted the adhesion of tumor cells to ECs and the transendothelial migration of tumor cells. Mechanistically, hyperglycemia-induced oxidative stress in ECs, and increased oxidative stress enhanced vWF expression in ECs through an increase in the transcription factor GATA1. These results provide evidence for the role of vWF in ECs in promoting hyperglycemia-induced tumor metastasis and potential therapeutic targets for the regulation of vWF expression in ECs and hyperglycemia-induced tumor metastasis., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

Fc-mediated effector functions are important for the clearance of pathologic cells by therapeutic IgG antibodies through two mechanisms: via the activation of the classical complement pathway and through the binding to Fcγ receptors (FcγRs) which mediate clearance of targeted cells by antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP) by effector cells such as macrophages, NK cells, and other leukocytes subsets. Complement activation results in direct cell killing through the formation of the membrane attack complex (MAC, complement-dependent cytotoxicity or CDC) and in the deposition of complement opsonins on pathogen surfaces. The latter are recognized by complement receptors on effector cells in turn triggering complement-dependent cell cytotoxicity and phagocytosis (CDCC and CDCP, respectively). Little is known about the role of CDCC and CDCP on therapeutic antibody function because on the one hand, IgG isotype antibodies bind to both FcγR and C1q to activate the complement pathway, and on the other, immune cells express complement receptor as well as FcγRs. We engineered IgG1 Fc domains that bind with high affinity to C1q but have very little or no binding to FcγR. To this end, we employed display of IgG in E. coli (which lack protein glycosylation machinery) for the screening of very large libraries (>2 × 10 9 ) of randomly mutated human Fc domains to isolate Fc variants that bind to C1q. Herein we introduce and describe the method., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Non-invasive painless signaling therapy (NPST) is an electro-cutaneous treatment that converts endogenous pain information into synthetic non-pain information. This study explored whether pain improvement by NPST in failed back surgery syndrome (FBSS) patients is related to cerebral modulation., Methods: Electroencephalography (EEG) analysis was performed in 11 patients with FBSS. Subjects received daily NPST for 5 days. Before the first treatment, patients completed the Brief Pain Inventory (BPI) and Beck Depression Inventory and underwent baseline EEG. After the final treatment, they responded again to the BPI, reported the percent pain improvement (PPI), and then underwent post-treatment EEG. If the PPI grade was zero, they were assigned to the ineffective group, while all others were assigned to the effective group. We used standardized low-resolution brain electromagnetic tomography (sLORETA) to explore the EEG current-source distribution (CSD) associated with pain improvement by NPST., Results: The 11 participants had a median age of 67.0 years, and 63.6% were female. The sLORETA images revealed a beta-2 CSD increment in 12 voxels of the right anterior cingulate gyrus (ACG) and the right medial frontal area. The point of maximal CSD changes was in the right ACG. The alpha band CSD increased in 2 voxels of the left transverse gyrus., Conclusions: Pain improvement by NPST in FBSS patients was associated with increased cerebral activity, mainly in the right ACG. The change in afferent information induced by NPST seems to be associated with cerebral pain perception.

Background: Understanding the memory T-cell response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is crucial for assessing the longevity of protective immunity after SARS-CoV-2 infection or coronavirus disease 2019 (COVID-19) vaccination. However, the longitudinal memory T-cell response up to 8 months post-symptom onset (PSO) according to the severity of illness is unknown., Methods: We analyzed peripheral blood mononuclear cells (PBMCs) from healthy volunteers or patients with COVID-19 who experienced asymptomatic, mild, or severe illness at 2, 5, and 8 months PSO. SARS-CoV-2 spike, nucleocapsid, and membrane protein-stimulated PBMCs were subjected to flow cytometry analysis., Results: A total of 24 patients (7 asymptomatic, 9 with mild disease, and 8 with severe disease) and 6 healthy volunteers were analyzed. SARS-CoV-2-specific OX40+CD137+CD4+ T cells and CD69+CD137+CD8+ T cells persisted at 8 months PSO. Also, antigen-specific cytokine-producing or polyfunctional CD4+ T cells were maintained for up to 8 months PSO. Memory CD4+ T-cell responses tended to be greater in patients who had severe illness than in those with mild or asymptomatic disease., Conclusions: Memory response to SARS-CoV-2, based on the frequency and functionality, persists for 8 months PSO. Further investigations involving its longevity and protective effect from reinfection are warranted., (© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Objective: To determine whether distinct aquaporin-4 (AQP4)-IgG lineages play a role in neuromyelitis optica spectrum disorder (NMOSD) pathogenesis, we profiled the AQP4-IgG polyclonal serum repertoire and identified, quantified, and functionally characterized distinct AQP4-IgG lineages circulating in 2 patients with NMOSD., Methods: We combined high-throughput sequencing and quantitative immunoproteomics to simultaneously determine the constituents of both the B-cell receptor (BCR) and the serologic (IgG) anti-AQP4 antibody repertoires in the peripheral blood of patients with NMOSD. The monoclonal antibodies identified by this platform were recombinantly expressed and functionally characterized in vitro., Results: Multiple antibody lineages comprise serum AQP4-IgG repertoires. Their distribution, however, can be strikingly different in polarization (polyclonal vs pauciclonal). Among the 4 serum AQP4-IgG monoclonal antibodies we identified in 2 patients, 3 induced complement-dependent cytotoxicity in a model mammalian cell line ( p < 0.01)., Conclusions: The composition and polarization of AQP4-IgG antibody repertoires may play an important role in NMOSD pathogenesis and clinical presentation. Here, we present a means of coupling both cellular (BCR) and serologic (IgG) antibody repertoire analysis, which has not previously been performed in NMOSD. Our analysis could be applied in the future to clinical management of patients with NMOSD to monitor disease activity over time as well as applied to other autoimmune diseases to facilitate a deeper understanding of disease pathogenesis relative to autoantibody clones., (Copyright © 2021 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.)

Mesenchymal stem cells (MSCs) are multipotent cells with critical roles in homeostasis and regeneration. MSCs undergo aging in response to various stresses, and this causes many diseases including degenerative disorders. Thus, regulation of aging factors is crucial for healthy aging. Mitochondrial open reading frame of the 12S rRNA-c (MOTS-c) was recently reported to regulate metabolic homeostasis. Here, we investigated the restorative effects of MOTS-c on aged human placenta-derived MSCs (hPD-MSCs). MOTS-c promoted the morphology of old hPD-MSCs. MOTS-c significantly activated AMP-activated protein kinase, which is the main target pathway of MOTS-c, and inhibited its antagonistic effector mTORC1. MOTS-c considerably enhanced mitochondrial homeostasis by decreasing oxygen consumption and reactive oxygen species production. The mitochondrial state of MOTS-c-treated old hPD-MSCs was more similar to that of young hPD-MSCs than the mitochondrial state of non-treated old hPD-MSCs. MOTS-c also decreased lipid synthesis. In conclusion, we demonstrated that MOTS-c promotes homeostasis in aged hPD-MSCs., (Copyright © 2021 Elsevier B.V. and Mitochondria Research Society. All rights reserved. All rights reserved.)

Monoclonal antibodies (mAb) have been used as therapeutic agents for various diseases, and immunoglobulin G (IgG) is mainly used among antibody isotypes due to its structural and functional properties. So far, regardless of the purpose of the therapeutic antibody, wildtype IgG has been mainly used, but recently, the engineered antibodies with various strategies according to the role of the therapeutic antibody have been used to maximize the therapeutic efficacy. In this review paper, first, the overall structural features and functional characteristics of antibody IgG, second, the old and new techniques for antibody discovery, and finally, several antibody engineering strategies for maximizing therapeutic efficacy according to the role of a therapeutic antibody will be introduced., Competing Interests: The authors declare no conflict of interest., (© The Korean Society for Biotechnology and Bioengineering and Springer 2021.)

Despite harboring mutations in oncogenes and tumor suppressors that promote cancer growth, T-cell acute lymphoblastic leukemia (T-ALL) cells require exogenous cells or signals to survive in culture. We previously reported that myeloid cells, particularly dendritic cells, from the thymic tumor microenvironment support the survival and proliferation of primary mouse T-ALL cells in vitro. Thus, we hypothesized that tumor-associated myeloid cells would support T-ALL in vivo. Consistent with this possibility, in vivo depletion of myeloid cells results in a significant reduction in leukemia burden in multiple organs in 2 distinct mouse models of T-ALL and prolongs survival. The impact of the myeloid compartment on T-ALL growth is not dependent on suppression of antitumor T-cell responses. Instead, myeloid cells provide signals that directly support T-ALL cells. Transcriptional profiling, functional assays, and acute in vivo myeloid-depletion experiments identify activation of IGF1R as a critical component of myeloid-mediated T-ALL growth and survival. We identify several myeloid subsets that have the capacity to directly support survival of T-ALL cells. Consistent with mouse models, myeloid cells derived from human peripheral blood monocytes activate IGF1R and directly support survival of primary patient T-ALL cells in vitro. Furthermore, enriched macrophage gene signatures in published clinical samples correlate with inferior outcomes for pediatric T-ALL patients. Collectively, these data reveal that tumor-associated myeloid cells provide signals critical for T-ALL growth in multiple organs in vivo and implicate tumor-associated myeloid cells and associated signals as potential therapeutic targets., (© 2020 by The American Society of Hematology.)

Background: To compare long-term effects of antigravity treadmill (AGT) combined with conventional rehabilitation (CR) and CR after hip fracture in patients with sarcopenia., Methods: Forty-five patients were randomly allocated to AGT combined with CR (experimental group) or CR (control group) for 10 consecutive working days. Participants were evaluated prior to treatment, 3 weeks, 3 months, and 6 months after treatment. Outcome measurement included Koval walking ability scores functional ambulatory category (FAC), Berg Balance Scale (BBS), Korean version of Mini-Mental State Examination, Euro Quality of Life Questionnaire Five-Dimensional Classification, Korean version of modified Barthel index, and grip strength., Results: At 3 weeks and 3 months, the comparison of change scores in KOVAL between two groups revealed difference of 0.84 (95% CI: -1.19, -0.49; p for trend = .000) and 1.21 (95% CI: -2.05, -0.36; p for trend = .006), respectively. At 3 weeks, comparison of change score in FAC between two groups revealed a difference of 0.73 (95% CI: 0.28, 1.19; p for trend = .003). The comparison of change scores between two groups also showed a difference in the 6 months in KOVAL and in the 3 and 6 months in FAC. The comparison of changes in scores in BBS between two groups revealed difference of 11.63 (95% CI: 5.85, 17.40; p for trend = .001), 9.00 (95% CI: 2.28, 15.71; p for trend = .006), and 11.05 (95% CI: 3.62, 18.48; p for trend = .006), respectively, at each follow-up., Conclusions: Both groups were improved after intervention. As additional benefits were evident among those who carried out AGT, it may be appropriate for patients with sarcopenia after hip fracture surgery., (© The Author(s) 2020. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objectives: To report the effects of scrambler therapy in patients diagnosed with failed back surgery syndromes and to analyze the factors affecting pain before and after the therapy., Methods: This study included 26 patients (12 males and 14 females). The Oswestry Disability Index (ODI) and Brief Pain Inventory (BPI) before and after scrambler therapy, Beck Depression Inventory (BDI) score before therapy, and residual pain after therapy were assessed. The changes in the ODI, BPI, and residual pain before and after the therapy were analyzed using the Wilcoxon signed rank test. Spearman correlation analysis and Fisher's exact test were used to confirm the correlation between BDI and other factors. Multiple regression analysis was used to identify independent factors predicting residual pain, posttherapy ODI, and posttherapy BPI., Results: The ODI changed from 25.69 ± 7.98 to 21.80 ± 9.41 ( p < 0.05), and the BPI changed from 68.96 ± 18.00 to 61.62 ± 20.27 after scrambler therapy ( p < 0.05). In addition, residual pain changed from 100 to 76.15 ( p < 0.05). The BDI was negatively correlated with the duration of scrambler therapy and positively correlated with the initial OPD and BPI. In multiple regression analysis, residual pain was significantly correlated with the BDI ( p < 0.05)., Conclusion: Scrambler therapy can be used to change the total scores of the ODI and BPI after 5 sessions of treatment. Also, residual pain was significantly related to the BDI. Clinical significance of depression severity on pain should be further investigated via prospective studies., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2020 Hayoung Byun et al.)

Background: To compare the muscle strength of patients with a hip fracture according to the presence of sarcopenia after surgery and the correlation of measured values between a Biodex and surface electromyography (sEMG) in postoperative measurement of muscle strength., Methods: Seventy-one patients who underwent hip fracture surgery were included in this study. Muscle mass was measured using dual energy X-ray absorptiometry and the grip strength was evaluated using a dynamometer. The diagnosis of sarcopenia followed the Asian Working Group for Sarcopenia criteria. We evaluated the Biodex to assess muscle strength according to the presence of sarcopenia and at the same time measured the sEMG to evaluate the correlation of muscle strength between Biodex and sEMG., Results: We assigned 34 patients with sarcopenia and 37 without sarcopenia to 2 groups. In the comparison of muscle strength using Biodex and sEMG between the 2 groups, it was confirmed that muscle strength of sarcopenia group was decreased compared with that of the non-sarcopenia group, although there was no statistical significance between the groups. However, Biodex and sEMG showed very close correlation with muscle strength in all variables., Conclusions: We suggest that using sEMG for the evaluation of muscle strength after hip fracture surgery may be an excellent tool alternative to isokinetic testing machines such as the Biodex., Competing Interests: Conflict of interest: No potential conflict of interest relevant to this article was reported., (Copyright © 2020 The Korean Society for Bone and Mineral Research.)

Background: The complications after stroke inhibit functional recovery and worsen the prognosis of patients. The implementation of a critical pathway (CP) can facilitate functional recovery after stroke by enabling comprehensive and systematic structured rehabilitation., Objective: To evaluate the effects of the implementation of CP in stroke patients for 10 years., Methods: The data were collected from 960 patients who were diagnosed with a stroke at the university hospital emergency room, who were transferred to the rehabilitation center after the acute phase, and who were discharged after undergoing comprehensive rehabilitation. Based on data collected over a period of 10 years, changes in demographic and stroke characteristics, preexisting medical conditions, poststroke complications, and functional states, as well as length of stay (LOS), were evaluated before and after CP implementation. The modified Rankin Scale (mRS) and the Korean version of the Modified Barthel Index (K-MBI) were used to evaluate functional states., Results: There were no significant differences in demographic and stroke characteristics before and after CP implementation. For those with preexisting medical conditions, there was no significant difference between before and after CP implementation. The majority of the complications were significantly decreased after the implementation of CP. Except for hemorrhagic stroke patients, the Brunnstrom stage in the ischemic and total stroke patients after CP implementation was significantly increased in the upper and lower extremities. The total hospitalization LOS and rehabilitation center hospitalization times were significantly reduced in ischemic and total stroke patients. There was no statistically significant difference in the functional gain of K-MBI and the efficiency of rehabilitation between before and after CP implementation., Conclusion: The implementation of CP allows for better application of evidence- and guideline-based key interventions and helps to provide early, comprehensive, organized, and more specialized care to stroke patients. Despite limited evidence, CP is still recommended as a means of promoting best practices in hospital care for stroke patients., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2020 Yun Jeong Jang et al.)

Background: Myofascial pain syndrome (MPS) is commonly seen in clinical settings and negatively influences a patient's daily life. Recently, the application of extracorporeal shock wave therapy (ESWT) has been utilized as one of the treatment methods for MPS. The aim of this systematic review and meta-analysis was to summarize the current evidence for the short-term effect of ESWT on MPS of trapezius., Methods: PubMed, EMBASE, Web of Science, and Cochrane Central Register of Controlled Trials were searched from the database inception to March 2019. Two reviewers independently screened articles, evaluated methodological quality, and extracted data. The primary outcome was post-interventional pain intensity., Results: Randomized controlled trials (RCTs) were conducted to determine whether ESWT was used as the main treatment on MPS. The 5 studies reviewed in this meta-analysis were evaluated for changes in pain intensity. Compared with other treatments, focused ESWT in MPS was more effective in reducing the scores of visual analog scale (VAS) (standardized mean difference [SMD] = -0.48, 95% CI -0.74 to -0.22)., Conclusions: There is very low level evidence that focused ESWT is effective for short-term relief of neck pain in MPS. The limited sample size and poor quality of these studies highlight and support the need for large scale, good quality placebo controlled trials in this area.

The pharmacokinetic properties of antibodies are largely dictated by the pH-dependent binding of the IgG fragment crystallizable (Fc) domain to the human neonatal Fc receptor (hFcRn). Engineered Fc domains that confer a longer circulation half-life by virtue of more favorable pH-dependent binding to hFcRn are of great therapeutic interest. Here we developed a pH Toggle switch Fc variant containing the L309D/Q311H/N434S (DHS) substitutions, which exhibits markedly improved pharmacokinetics relative to both native IgG1 and widely used half-life extension variants, both in conventional hFcRn transgenic mice and in new knock-in mouse strains. engineered specifically to recapitulate all the key processes relevant to human antibody persistence in circulation, namely: (i) physiological expression of hFcRn, (ii) the impact of hFcγRs on antibody clearance and (iii) the role of competing endogenous IgG. DHS-IgG retains intact effector functions, which are important for the clearance of target pathogenic cells and also has favorable developability.

This study evaluated the efficiency of the removal of heavy metals from contaminated water via adsorption isotherm and kinetic experiments on two composite mineral adsorbents, CMA1 and CMA2. The developed CMA1 (zeolite with clinoptilolite of over 20 weight percent and feldspar of ~10 percent, with Portland cement) and CMA2 (zeolite with feldspar of over 15 weight percent and ~9 percent clinoptilolite, with Portland cement) were applied to remove Cu, Cd, and Pb ions. Based on the adsorption isotherm and kinetic experiments, the adsorbents CMA1 and CMA2 exhibited high removal efficiency for Cu, Cd, and Pb ions in solution compared to other adsorbents. In the adsorption kinetic experiment, CMA2 demonstrated better adsorption than CMA1 with the same initial concentration and reaction time, and Cu, Cd, and Pb ions almost reached equilibrium within 180 min for both CMA1 and CMA2. The results of the adsorption kinetic experiments with pseudo-first-order (PFO) and pseudo-second-order (PSO) models indicated that the PSO model was more suitable than the PFO model. Comparing the Langmuir and Freundlich adsorption isotherm models, the former showed a very slightly higher correlation coefficient (r 2 ) than the latter, indicating that the two models can both be applied to heavy metal solutions on a spherical monolayer surface with a weak heterogeneity of the surface. Additionally, the adsorbents CMA1 and CMA2 demonstrated different removal abilities depending on which heavy metals were used.

Variations in biofeedback coding schemes for postural control, in recent research, have shown significant differences in performance outcomes due to variations in coding schemes. However, the application of vibrotactile biofeedback coding schemes to gait symmetry training is not well explored. In this paper, we devised various vibrotactile biofeedback modes and identified their efficacy during gait symmetry training of individuals suffering from hemiparesis due to stroke. These modes are composed of variations in vibration type (on-time or intensity), and relation type (proportional or inversely-proportional) with the error in symmetry ratio. Eight individuals with stroke participated in walking trials. From dependent t-tests on the collected data, we found improved achievement of temporal gait symmetry while utilizing all the provided biofeedback modes compared to no biofeedback (P < 0.001). Furthermore, two-way repeated measures ANOVA revealed statistically significant difference in symmetry ratio for main effect of vibration type (P-value = 0.016, partial eta squared = 0.585). The participants performed better with modes of biofeedback with varying vibration on-times. Furthermore, participants showed better performance when the biofeedback varied proportionally with the error. These findings suggest that biofeedback coding schemes may have a significant effect on the performance of gait training.

Objective: To investigate the risk factors for fall in the elderly population residing in rural areas of Korea and provide useful data for their prevention., Methods: As part of farmers' health promotion project, a retrospective study was conducted with a total of 350 elderly people recruited from March 2016 to December 2016. These subjects were divided into two groups: 254 non-fallers and 96 fallers. A person who fell to the floor at least once in the past year was defined as a faller. Participants were asked to visit the hospital once. The demographic characteristics, social environment, and educational levels were surveyed using a questionnaire. Physical examination was performed in the following order: cognitive function, lower leg strength and torque, body composition, and knee image test., Results: Statistically significant factors for falls in univariate analysis were female gender, age, living alone, educational level less than middle school, skeletal muscle mass, Mini-Mental State Exam, knee osteoarthritis, hip torque, hip power mean, knee torque, and knee power mean. Multivariate analysis was performed to identify variables most relevant to falls among statistically significant factors in univariate logistic analysis. It was confirmed that female gender and age of 70-79 years were statistically significant factors related to falls., Conclusion: Female gender and elderly status (70-79 years) are important risk factors for falls in rural areas underscoring the need for special attention when considering risk factors for falls among the elderly living in rural areas.

IgG antibodies mediate the clearance of target cells via the engagement of Fc gamma receptors (FcγRs) on effector cells by eliciting antibody-dependent cellular cytotoxicity and phagocytosis (ADCC and ADCP, respectively). Because (i) the IgG Fc domain binds to multiple FcγRs with varying affinities; (ii) even low Fc:FcγRs affinity interactions can play a significant role when antibodies are engaged in high avidity immune complexes and (iii) most effector cells express multiple FcγRs, the clearance mechanisms that can be mediated by individual FcγR are not well-understood. Human FcγRIIIa (hFcγRIIIa; CD16a), which exists as two polymorphic variants at position 158, hFcγRIIIa V158 and hFcγRIIIa F158 , is widely considered to only trigger ADCC, especially with natural killer (NK) cells as effectors. To evaluate the role of hFcγRIIIa ligation in myeloid-derived effector cells, and in particular on macrophages and monocytes which express multiple FcγRs, we engineered an aglycosylated engineered human Fc (hFc) variant, Fc3aV, which binds exclusively to hFcγRIIIa V158 . Antibodies formatted with the Fc3aV variant bind to the hFcγRIIIa V158 allotype with a somewhat lower K D than their wild type IgG1 counterparts, but not to any other hFcγR. The exceptional selectivity for hFcγRIIIa V158 was demonstrated by SPR using increased avidity, dimerized GST-fused versions of the ectodomains of hFcγRs and from the absence of binding of large immune complex (IC) to CHO cells expressing each of the hFcγRs, including notably, the FcγRIIIa F158 variant or the highly homologous FcγRIIIb. We show that even though monocyte-derived GM-CSF differentiated macrophages express hFcγRIIIa at substantially lower levels than the other two major activating receptors, namely hFcγRI or hFcγRIIa, Fc3aV-formatted Rituximab and Herceptin perform ADCP toward CD20- and Her2-expressing cancer cells, respectively, at a level comparable to that of the respective wild-type antibodies. We further show that hFcγRIIIa activation plays a significant role on ADCC by human peripheral monocytes. Our data highlight the utility of Fc3aV and other similarly engineered exquisitely selective, aglycosylated Fc variants toward other hFcγRs as tools for the detailed molecular understanding of hFcγR biology.

A better understanding of antitumor immune responses is the key to advancing the field of cancer immunotherapy. Endogenous immunity in cancer patients, such as circulating anticancer antibodies or tumor-reactive B cells, has been historically yet incompletely described. Here, we demonstrate that tumor-draining (sentinel) lymph node (SN) is a rich source for tumor-reactive B cells that give rise to systemic IgG anticancer antibodies circulating in the bloodstream of breast cancer patients. Using a synergistic combination of high-throughput B-cell sequencing and quantitative immunoproteomics, we describe the prospective identification of tumor-reactive SN B cells (based on clonal frequency) and also demonstrate an unequivocal link between affinity-matured expanded B-cell clones in the SN and antitumor IgG in the blood. This technology could facilitate the discovery of antitumor antibody therapeutics and conceivably identify novel tumor antigens. Lastly, these findings highlight the unique and specialized niche the SN can fill in the advancement of cancer immunotherapy.

Molecular understanding of serological immunity to influenza has been confounded by the complexity of the polyclonal antibody response in humans. Here we used high-resolution proteomics analysis of immunoglobulin (referred to as Ig-seq) coupled with high-throughput sequencing of transcripts encoding B cell receptors (BCR-seq) to quantitatively determine the antibody repertoire at the individual clonotype level in the sera of young adults before and after vaccination with trivalent seasonal influenza vaccine. The serum repertoire comprised between 40 and 147 clonotypes that were specific to each of the three monovalent components of the trivalent influenza vaccine, with boosted pre-existing clonotypes accounting for ∼60% of the response. An unexpectedly high fraction of serum antibodies recognized both the H1 and H3 monovalent vaccines. Recombinant versions of these H1 + H3 cross-reactive antibodies showed broad binding to hemagglutinins (HAs) from previously circulating virus strains; several of these antibodies, which were prevalent in the serum of multiple donors, recognized the same conserved epitope in the HA head domain. Although the HA-head-specific H1 + H3 antibodies did not show neutralization activity in vitro, they protected mice against infection with the H1N1 and H3N2 virus strains when administered before or after challenge. Collectively, our data reveal unanticipated insights regarding the serological response to influenza vaccination and raise questions about the added benefits of using a quadrivalent vaccine instead of a trivalent vaccine., Competing Interests: The authors declare no competing financial interests.

Ricin is a toxin that could potentially be used as a bioweapon. We identified anti-ricin A chain antibodies by sequencing the antibody repertoire from immunized mice and by selecting high affinity antibodies using yeast surface display. These methods led to the isolation of multiple antibodies with high (sub-nanomolar) affinity. Interestingly, the antibodies identified by the 2 independent approaches are from the same clonal lineages, indicating for the first time that yeast surface display can identify native antibodies. The new antibodies represent well-characterized reagents for biodefense diagnostics and therapeutics development.

The silencing of specific oncogenes via RNA interference (RNAi) holds great promise for the future of cancer therapy. RNAi is commonly carried out using small interfering RNA (siRNA) composed of a 19 bp duplex region with a 2-nucleotide overhang at each 3' end. This classical siRNA structure, however, can trigger non-specific effects, which has hampered the development of specific and safe RNAi therapeutics. Previously, we developed a novel siRNA structure, called asymmetric shorter-duplex siRNA (asiRNA), which did not cause the non-specific effects triggered by conventional siRNA, such as off-target gene silencing mediated by the sense strand. In this study, we first screened potent asiRNA molecules targeting the human c-MET gene, a promising anticancer target. Next, the activity of a selected asiRNA was further optimized by introducing a locked nucleic acid (LNA) to maximize the gene silencing potency. The optimized asiRNA targeted to c-MET may have potential as a specific and safe anticancer RNAi therapeutic.

Human kringle domains (KDs) are ubiquitously expressed binding modulators that fold into seven flexible loops and it has been previously demonstrated that KDs can be engineered toward target-specific binding proteins as a non-antibody protein scaffold. Here, we report a method for efficient expression of a KD derivative (KD548)-a promising anti-cancer agent-by high-cell-density culture of Escherichia coli at a preparative scale production. The correct folding of KD548 requires three disulfide bonds. Nevertheless, cytoplasmic expression of KD548 in E. coli led to good yields of highly soluble proteins with high activity. For efficient expression, four sets of expression systems consisting of different promoters (lac or T7) and fusion tags (His or FLAG) were examined. Of these, the expression system using a combination of the T7 promoter with the FLAG tag resulted in the highest production in shake flask cultivation as well as in high-cell-density cultivation performed in a 6.6-L jar bioreactor. When protein expression was induced at high-cell density (optical density [OD] = 100) and when complex feeding solutions were supplemented, cell density (maximum OD = 184) and production yield (approximately 5.4 g/L) were significantly enhanced to values that were much higher than those found previously with Pichia cultivation (<8 mg/L).

Here, we report the development of target-specific binding proteins based on the kringle domain (KD) ( approximately 80 residues), a ubiquitous modular structural unit occurring across eukaryotic species. By exploiting the highly conserved backbone folding by core residues, but using extensive sequence variations in the seven loop regions of naturally occurring human KDs, we generated a synthetic KD library on the yeast cell surface by randomizing 45 residues in the loops of a human KD template. We isolated KD variants that specifically bind to anticancer target proteins, such as human death receptor 4 (DR4) and/or DR5, and that function as agonists to induce apoptotic cell death in several cancer cell lines in vitro and inhibit tumor progression in mouse models. Combined treatments with KD variants possessing different recognition sites on the same target protein exerted synergisitic tumoricidal activities, compared to treatment with individual variants. In addition to the agonists, we isolated an antagonistic KD variant that binds human tumor necrosis factor-alpha (TNFalpha) and efficiently neutralizes TNFalpha-induced cytotoxicity in vitro and in vivo. The KD scaffold with seven flexible loops protruding from the central core was strongly sequence-tolerant to mutations in the loop regions, offering a potential advantage of distinct binding sites for target recognition on the single domain. Our results suggest that the KD scaffold can be used to develop target-specific binding proteins that function as agonists or antagonists toward given target molecules, indicative of their potential use as biotherapeutics.