Engineering T cell receptor fusion proteins using nonviral CRISPR/Cas9 genome editing for cancer immunotherapy.

Manufacture of chimeric antigen receptor (CAR)-T cells usually involves the use of viral delivery systems to achieve high transgene expression. However, it can be costly and may result in random integration of the CAR into the genome, creating several disadvantages including variation in transgene expression, functional gene silencing and potential oncogenic transformation. Here, we optimized the method of nonviral, CRISPR/Cas9 genome editing using large donor DNA delivery, knocked-in an anti-tumor single chain variable fragment (scFv) into the N-terminus of CD3ε and efficiently generated fusion protein (FP) T cells. These cells displayed FP integration within the TCR/CD3 complex, lower variability in gene expression compared to CAR-T cells and good cell expansion after transfection. CD3ε FP T cells were predominantly CD8+ effector memory T cells, and exhibited anti-tumor activity in vitro and in vivo. Dual targeting FP T cells were also generated through the incorporation of scFvs into other CD3 subunits and CD28. Compared to viral-based methods, this method serves as an alternative and versatile way of generating T cells with tumor-targeting receptors for cancer immunotherapy.

Anion Storage of MXenes.

Recently, anion storage materials have gained significant attention owing to the widened cell voltage and additional anion storing capacity for a large energy density. MXenes are considered as the emerging anion storing materials owing to their sufficient interlayer spacing, rich surface chemistries, tunable structures, remarkable electrochemical properties, and mechanical integrity. Herein, a comprehensive review on the anion storage of MXenes covering their anion storage mechanism and state-of-the-art chemical strategies for the improved anion storage performances is reported. The recent progress of MXenes on aluminum ion batteries, metal halogen batteries, halogen ion batteries, and electrochemical electrode deionization is addressed. The scientific and technical challenges and the research direction into the anion storage of MXenes are also addressed and finally the authors' perspective on anion storage of MXenes is provided. Therefore, this review offers an insight into the rational design of MXenes for anion storage materials and the correlation of surface chemistries and structural modifications with anion storage properties for the applications into electrochemical energy storage and water purification.

Machine learning-driven dynamic risk prediction for highly pathogenic avian influenza at poultry farms in Republic of Korea: Daily risk estimation for individual premises.

Highly pathogenic avian influenza (HPAI) is a fatal zoonotic disease that damages the poultry industry and endangers human lives via exposure to the pathogen. A risk assessment model that precisely predicts high-risk groups and occurrence of HPAI infection is essential for effective biosecurity measures that minimize the socio-economic losses of massive outbreaks. However, the conventional risk prediction approaches have difficulty incorporating the broad range of factors associated with HPAI infections at poultry holdings. Therefore, it is difficult to accommodate the complexity of the dynamic transmission mechanisms and generate risk estimation on a real-time basis. We proposed a continuous risk prediction framework for HPAI occurrences that used machine learning algorithms (MLAs). This integrated environmental, on-farm biosecurity, meteorological, vehicle movement tracking, and HPAI wild bird surveillance data to improve accuracy and timeliness. This framework consisted of (i) the generation of 1788 predictors from six types of data and reconstructed them with an outcome variable into a data mart based on a temporal assumption (i.e. infected period and day-ahead forecasting); (ii) training of the predictors with the temporally rearranged outcome variable that corresponded to HPAI H5N6 infected state at each individual farm on daily basis during the 2016-2017 HPAI epidemic using three different MLAs [Random Forest, Gradient Boosting Machine (GBM), and eXtreme Gradient Boosting]; (iii) predicting the daily risk of HPAI infection during the 2017-2018 HPAI epidemic using the pre-trained MLA models for each farm across the country. The models predicted the high risk to 8-10 out of 19 infected premises during the infected period in advance. The GBM MLAs outperformed the 7-day forecasting of HPAI prediction at individual poultry holdings, with an area under the curve (AUC) of receiver operating characteristic of 0.88. Therefore, this approach enhances the flexibility and timing of interventions against HPAI outbreaks at poultry farms.

Microspinning: Local Surface Mixing via Rotation of Magnetic Microparticles for Efficient Small-Volume Bioassays.

The need for high-throughput screening has led to the miniaturization of the reaction volume of the chamber in bioassays. As the reactor gets smaller, surface tension dominates the gravitational or inertial force, and mixing efficiency decreases in small-scale reactions. Because passive mixing by simple diffusion in tens of microliter-scale volumes takes a long time, active mixing is needed. Here, we report an efficient micromixing method using magnetically rotating microparticles with patterned magnetization induced by magnetic nanoparticle chains. Because the microparticles have magnetization patterning due to fabrication with magnetic nanoparticle chains, the microparticles can rotate along the external rotating magnetic field, causing micromixing. We validated the reaction efficiency by comparing this micromixing method with other mixing methods such as simple diffusion and the use of a rocking shaker at various working volumes. This method has the potential to be widely utilized in suspension assay technology as an efficient mixing strategy.

A novel missense variant in the DIAPH1 gene in a Korean family with autosomal dominant nonsyndromic hearing loss.

Hair cells in the cochlea display highly regulated actin polymerization, which is mediated by the human diaphanous-related formin 1 gene (DIAPH1; also called DFNA1, DIA1). DFNA1, the first type of autosomal dominant nonsyndromic hearing loss (ADNSHL), is known to be associated with mutations in DIAPH1. However, no genetic study of DFNA1 in Koreans with hearing loss has yet been reported. A 51-year-old patient in a Korean family with ADNSHL was examined by pure-tone audiometry, and genetic analysis of DIAPH1 was performed. A novel variant, p.I530S (c.1589T > G), was identified in the DIAPH1 gene, and the mutation was located in the highly conserved coiled-coil domain of the DIA1 protein, where an amino acid substitution was predicted to change the domain structure. Further functional investigations will provide more information to help us understand the role of DIAPH1 in maintenance of hair cell function in the auditory pathway.

Genetic association of MYH genes with hereditary hearing loss in Korea.

BACKGROUND: Myosin is a key protein involved in regulating the shape and motility of cells. The MYH9 and MYH14 genes, which encode non-muscle myosin heavy chain IIA (NMMHC II-A) and IIC (NMMHC II-C), respectively, are expressed in the inner ear. These myosin genes are known to be associated with autosomal dominant non-syndromic hearing loss (ADNSHL); however, genetic studies in patients with ADNSHL in Korea have rarely been reported. METHODS: We analyzed the MYH9 and MYH14 genes in 75 Korean patients with ADNSHL. RESULTS: We identified 4 possible pathogenic variants: a novel variant p.F1303L and 2 previously reported variants (p.R1730C and p.R1785C) in the MYH9 gene, and a novel variant p.A1868T in the MYH14 gene. All the variants were located in the myosin tail domain, which is essential for the interaction of myosin with actin. These variants were predicted to be possibly pathogenic by functional prediction tools and were absent in 100 unrelated normal controls. CONCLUSION: These results suggest that all the variants identified in this study have a strong potential to affect the structural stability and/or function of non-muscle myosin in the inner ear, which might lead to ADNSHL. This study establishes the link between the genotype and development of ADNSHL and contributes to the establishment of Korean database for hereditary hearing loss.

An all-joint-control master device for single-port laparoscopic surgery robots.

PURPOSE: Robots for single-port laparoscopic surgery (SPLS) typically have all of their joints located inside abdomen during surgery, whereas with the da Vinci system, only the tip part of the robot arm is inserted and manipulated. A typical master device that controls only the tip with six degrees of freedom (DOFs) is not suitable for use with SPLS robots because of safety concerns. METHODS: We designed an ergonomic six-DOF master device that can control all of the joints of an SPLS robot. We matched each joint of the master, the slave, and the human arm to decouple all-joint motions of the slave robot. Counterbalance masses were used to reduce operator fatigue. Mapping factors were determined based on kinematic analysis and were used to achieve all-joint control with minimal error at the tip of the slave robot. RESULTS: The proposed master device has two noteworthy features: efficient joint matching to the human arm to decouple each joint motion of the slave robot and accurate mapping factors, which can minimize the trajectory error of the tips between the master and the slave. CONCLUSIONS: We confirmed that the operator can manipulate the slave robot intuitively with the master device and that both tips have similar trajectories with minimal error.