Targeting Intracellular Antigens with pMHC-Binding Antibodies: A Phage Display Approach.

Antibodies that bind peptide-MHC (pMHC) complex in a manner akin to T cell receptor (TCR) have not only helped in understanding the mechanism of TCR-pMHC interactions in the context of T cell biology but also spurred considerable interest in recent years as potential cancer therapeutics. Traditional methods to generate such antibodies using hybridoma and B cell sorting technologies are sometimes inadequate, possibly due to the small contribution of peptide to the overall B cell epitope space on the surface of the pMHC complex (typical peptide MW = 1 kDa versus MHC MW = 45 kDa) and to the multiple efficiency limiting steps inherent in these methods. In this chapter we describe phage display approaches, including a cell panning strategy, for the rapid generation of such antibodies with high specificity and affinity.

Uncovering the Gene Regulatory Network of Maize Hybrid ZD309 under Heat Stress by Transcriptomic and Metabolomic Analysis.

Maize is an important cereal crop but is sensitive to heat stress, which significantly restricts its grain yield. To explore the molecular mechanism of maize heat tolerance, a heat-tolerant hybrid ZD309 and its parental lines (H39_1 and M189) were subjected to heat stress, followed by transcriptomic and metabolomic analyses. After six-day-heat treatment, the growth of ZD309 and its parental lines were suppressed, showing dwarf stature and rolled leaf compared with the control plants. ZD309 exhibited vigorous growth; however, M189 displayed superior heat tolerance. By transcriptomic and metabolomic analysis, hundreds to thousands of differentially expressed genes (DEGs) and metabolites (DEMs) were identified. Notably, the female parent H39 shares more DEGs and DEMs with the hybrid ZD309, indicating more genetic gain derived from the female instead of the male. A total of 299 heat shock genes detected among three genotypes were greatly aggregated in sugar transmembrane transporter activity, plasma membrane, photosynthesis, protein processing in the endoplasmic reticulum, cysteine, and methionine metabolism. A total of 150 heat-responsive metabolites detected among three genotypes were highly accumulated, including jasmonic acid, amino acids, sugar, flavonoids, coumarin, and organic acids. Integrating transcriptomic and metabolomic assays revealed that plant hormone signal transduction, cysteine, and methionine metabolism, and α-linolenic acid metabolism play crucial roles in heat tolerance in maize. Our research will be facilitated to identify essential heat tolerance genes in maize, thereby contributing to breeding heat resistance maize varieties.

Microstructure Evolution and Mechanical Properties of Medium Carbon Martensitic Steel during Warm Rolling and Annealing Process.

The microstructure evolution and mechanical properties of medium carbon martensitic steel during the warm rolling and annealing process were studied by scanning electron microscope (SEM), electron back scattering diffraction (EBSD), and electronic universal testing machine. The results showed that the microstructure of ferrite matrix with mass dispersive cementite particles was obtained by decomposition of martensitic in medium-carbon martensitic steel after warm rolling. The grain size of ferrite was ~0.53 µm, the yield strength and tensile strength were 951 MPa and 968 MPa, respectively, and the total elongation rate was 11.5% after warm rolling at 600 °C. Additionally, after the next 4 h of annealing, the grain size of ferrite and particle size of cementite increased to ~1.35 µm and ~360 nm and the yield strength and tensile strength decreased to 600 MPa and 645 MPa, respectively, with a total elongation increases of 20.9%. The strength of the material increased with increasing strain rate in tension, and the yield-to-tensile strength ratio increased from 0.92 to 0.94 and maintained good plasticity.

Non-lab and semi-lab algorithms for screening undiagnosed diabetes: A cross-sectional study.

BACKGROUND: The terrifying undiagnosed rate and high prevalence of diabetes have become a public emergency. A high efficiency and cost-effective early recognition method is urgently needed. We aimed to generate innovative, user-friendly nomograms that can be applied for diabetes screening in different ethnic groups in China using the non-lab or noninvasive semi-lab data. METHODS: This multicenter, multi-ethnic, population-based, cross-sectional study was conducted in eight sites in China by enrolling subjects aged 20-70. Sociodemographic and anthropometric characteristics were collected. Blood and urine samples were obtained 2 h following a standard 75 g glucose solution. In the final analysis, 10,794 participants were included and randomized into model development (n = 8096) and model validation (n = 2698) group with a ratio of 3:1. Nomograms were developed by the stepwise binary logistic regression. The nomograms were validated internally by a bootstrap sampling method in the model development set and externally in the model validation set. The area under the receiver operating characteristic curve (AUC) was used to assess the screening performance of the nomograms. Decision curve analysis was applied to calculate the net benefit of the screening model. RESULTS: The overall prevalence of undiagnosed diabetes was 9.8% (1059/10794) according to ADA criteria. The non-lab model revealed that gender, age, body mass index, waist circumference, hypertension, ethnicities, vegetable daily consumption and family history of diabetes were independent risk factors for diabetes. By adding 2 h post meal glycosuria qualitative to the non-lab model, the semi-lab model showed an improved Akaike information criterion (AIC: 4506 to 3580). The AUC of the semi-lab model was statistically larger than the non-lab model (0.868 vs 0.763, P < 0.001). The optimal cutoff probability in semi-lab and non-lab nomograms were 0.088 and 0.098, respectively. The sensitivity and specificity were 76.3% and 81.6%, respectively in semi-lab nomogram, and 72.1% and 67.3% in non-lab nomogram at the optimal cut off point. The decision curve analysis also revealed a bigger decrease of avoidable OGTT test (52 per 100 subjects) in the semi-lab model compared to the non-lab model (36 per 100 subjects) and the existed New Chinese Diabetes Risk Score (NCDRS, 35 per 100 subjects). CONCLUSION: The non-lab and semi-lab nomograms appear to be reliable tools for diabetes screening, especially in developing countries. However, the semi-lab model outperformed the non-lab model and NCDRS prediction systems and might be worth being adopted as decision support in diabetes screening in China.

Use of Phenoxyaniline Analogues To Generate Biochemical Insights into the Interactio n of Polybrominated Diphenyl Ether with CYP2B Enzymes.

Human hepatic cytochromes P450 (CYP) are integral to xenobiotic metabolism. CYP2B6 is a major catalyst of biotransformation of environmental toxicants, including polybrominated diphenyl ethers (PBDEs). CYP2B substrates tend to contain halogen atoms, but the biochemical basis for this selectivity and for species specific determinants of metabolism has not been identified. Spectral binding titrations and inhibition studies were performed to investigate interactions of rat CYP2B1, rabbit CYP2B4, and CYP2B6 with a series of phenoxyaniline (POA) congeners that are analogues of PBDEs. For most congeners, there was a <3-fold difference between the spectral binding constants (KS) and IC50 values. In contrast, large discrepancies between these values were observed for POA and 3-chloro-4-phenoxyaniline. CYP2B1 was the enzyme most sensitive to POA congeners, so the Val-363 residue from that enzyme was introduced into CYP2B4 or CYP2B6. This substitution partially altered the protein-ligand interaction profiles to make them more similar to that of CYP2B1. Addition of cytochrome P450 oxidoreductase (POR) to titrations of CYP2B6 with POA or 2'4'5'TCPOA decreased the affinity of both ligands for the enzyme. Addition of cytochrome b5 to a recombinant enzyme system containing POR and CYP2B6 increased the POA IC50 value and decreased the 2'4'5'TCPOA IC50 value. Overall, the inconsistency between KS and IC50 values for POA versus 2'4'5'TCPOA is largely due to the effects of redox partner binding. These results provide insight into the biochemical basis of binding of diphenyl ethers to human CYP2B6 and changes in CYP2B6-mediated metabolism that are dependent on POA congener and redox partner identity.

Structural and biophysical characterization of human cytochromes P450 2B6 and 2A6 bound to volatile hydrocarbons: analysis and comparison.

X-ray crystal structures of complexes of cytochromes CYP2B6 and CYP2A6 with the monoterpene sabinene revealed two distinct binding modes in the active sites. In CYP2B6, sabinene positioned itself with the putative oxidation site located closer to the heme iron. In contrast, sabinene was found in an alternate conformation in the more compact CYP2A6, where the larger hydrophobic side chains resulted in a significantly reduced active-site cavity. Furthermore, results from isothermal titration calorimetry indicated a much more substantial contribution of favorable enthalpy to sabinene binding to CYP2B6 as opposed to CYP2A6, consistent with the previous observations with (+)-α-pinene. Structural analysis of CYP2B6 complexes with sabinene and the structurally similar (3)-carene and comparison with previously solved structures revealed how the movement of the F206 side chain influences the volume of the binding pocket. In addition, retrospective analysis of prior structures revealed that ligands containing -Cl and -NH functional groups adopted a distinct orientation in the CYP2B active site compared with other ligands. This binding mode may reflect the formation of Cl-π or NH-π bonds with aromatic rings in the active site, which serve as important contributors to protein-ligand binding affinity and specificity. Overall, the findings from multiple techniques illustrate how drugs metabolizing CYP2B6 and CYP2A6 handle a common hydrocarbon found in the environment. The study also provides insight into the role of specific functional groups of the ligand that may influence the binding to CYP2B6.

Design, synthesis, and evaluation of 2-substituted ethenesulfonic acid ester derivatives as protein tyrosine phosphatase 1B inhibitors.

Thirty-two 2-substituted ethenesulfonic acid ester derivatives were designed, synthesized, and evaluated for their inhibitory activities against protein tyrosine phosphatase 1B (PTP1B) and selectivity over T-Cell protein tyrosine phosphatase (TCPTP). Preliminary structure-activity relationship studies demonstrated that the substitution at the aromatic center and the length of linker between the hydrophobic tail and aromatic center markedly affected the inhibitory activity against PTP1B and the selectivity over TCPTP. Specifically, compounds 43 and 36 revealed excellent inhibitory activity to PTP1B with IC(50) = 1.3 µM and 1.5 µM, respectively, and marked 10- and 20-fold selectivity over TCPTP. Cytotoxicity data showed low cytotoxicity for COS-7 cell with IC(50) values >100 µM for most synthesized chemicals.

[Recent advances in the study of bioreductive drugs targeted tumor hypoxia].

Tumor hypoxia is the necessary process in the development of solid tumors, which is the key factor for drug resistance, recurrence, attack and shift of tumor. Hypoxic tumor cells have a certain extent of tolerance to radiation and chemotherapy. Tumor hypoxia is an important target for medication therapy. In the recent years, the bioreductive drugs targeted tumor hypoxia has made great process in the treatment of tumors. The latest advances of bioreductive drugs targeted hypoxia were reviewed in this paper.