Discovery and preclinical evaluation of KYS202004A, a novel bispecific fusion protein targeting TNF-α and IL-17A, in autoimmune disease models.

The treatment of autoimmune and inflammatory diseases often requires targeting multiple pathogenic pathways. KYS202004A is a novel bispecific fusion protein designed to antagonize TNF-α and IL-17A, pivotal in the pathophysiology of autoimmune and inflammatory diseases. Our initial efforts focused on screening for optimal structure by analyzing expression levels, purity, and binding capabilities. The binding affinity of KYS202004A to TNF-α and IL-17A was evaluated using SPR. In vitro, we assessed the inhibitory capacity of KYS202004A on cytokine-induced CXCL1 expression in HT29 cells. In vivo, its efficacy was tested using a Collagen-Induced Arthritis (CIA) model in transgenic human-IL-17A mice and an imiquimod-induced psoriasis model in cynomolgus monkeys. KYS202004A demonstrated significant inhibition of IL-17A and TNF-α signaling pathways, outperforming the efficacy of monotherapeutic agents ixekizumab and etanercept in reducing CXCL1 expression in vitro and ameliorating disease markers in vivo. In the CIA model, KYS202004A significantly reduced clinical symptoms, joint destruction, and serum IL-6 concentrations. The psoriasis model revealed that KYS202004A, particularly at a 2  mg/kg dose, was as effective as the combination of ixekizumab and etanercept. This discovery represents a significant advancement in treating autoimmune and inflammatory diseases, offering a dual-targeted therapeutic approach with enhanced efficacy over current monotherapies.

Formic Acid Dehydrogenation through Ligand Design Strategy of Amidation in Half-Sandwich Ir Complexes.

A series of Cp*Ir (Cp* = pentamethylcyclopentadienyl) complexes with amidated 8-aminoquinoline ligands were synthesized and tested for formic acid (FA) dehydrogenation. These complexes showed improved activities compared to pristine 8-anminquinoline (L1). Specially, amidation changed the outer coordination sphere of the complex (3) bearing N-8-quinolinylformamide (L3), and 3 was proved to be a proton-responsive catalyst. Our experimental results and DFT calculations demonstrated that the deprotonated carbanion in L3 could interact with a water molecule to stabilize the transition states and lower the reaction energy barrier, which improved the reaction activity. A turnover frequency of 206250 h-1 was achieved by 3 under optimized conditions. This study presents a method to develop new ligands and modify the existing ligands for efficient FA dehydrogenation.

A ligand design strategy to enhance catalyst stability for efficient formic acid dehydrogenation.

New Ir complexes bearing N-(methylsulfonyl)-2-pyridinecarboxamide (C1) and N-(phenylsulfonyl)-2-pyridinecarboxamide (C2) were employed as catalysts for aqueous formic acid dehydrogenation (FADH). The ligands were designed to maintain the picolinamide skeleton and introduce strong sigma sulfonamide moieties. C1 and C2 exhibited good stability towards air and concentrated formic acid (FA). During 20 continuous cycles, C1 and C2 could achieve the complete conversion of FA with TONs of 172 916 and 172 187, respectively. C1 achieved a high TOF of 19 500 h-1 at 90 °C and an air-stable Ir-H species was observed by 1H NMR spectroscopy.

Formic Acid as a Potential On-Board Hydrogen Storage Method: Development of Homogeneous Noble Metal Catalysts for Dehydrogenation Reactions.

Hydrogen can be used as an energy carrier for renewable energy to overcome the deficiency of its intrinsically intermittent supply. One of the most promising application of hydrogen energy is on-board hydrogen fuel cells. However, the lack of a safe, efficient, convenient, and low-cost storage and transportation method for hydrogen limits their application. The feasibility of mainstream hydrogen storage techniques for application in vehicles is briefly discussed in this Review. Formic acid (FA), which can reversibly be converted into hydrogen and carbon dioxide through catalysis, has significant potential for practical application. Historic developments and recent examples of homogeneous noble metal catalysts for FA dehydrogenation are covered, and the catalysts are classified based on their ligand types. The Review primarily focuses on the structure-function relationship between the ligands and their reactivity and aims to provide suggestions for designing new and efficient catalysts for H2 generation from FA.

Chiral N,N'-dioxide/Co(ii)-promoted asymmetric 1,3-dipolar cycloaddition of nitrones with methyleneindolinones.

A chiral N,N'-dioxide/Co(BF4)2·6H2O complex catalytic system has been developed to efficiently catalyze the asymmetric 1,3-dipolar cycloaddition of nitrones with methyleneindolinones. The corresponding chiral multisubstituted spiroisoxazolidines with three contiguous quaternary-tertiary stereocenters were obtained in moderate to high yields with excellent dr and ee values (up to 97% yield, >19 : 1 dr and 98% ee).

Kinetic Resolution of Oxaziridines via Chiral Bifunctional Guanidine-Catalyzed Enantioselective α-Hydroxylation of ß-Keto Esters.

An efficient kinetic resolution of racemic oxaziridines has been realized via catalytic asymmetric α-hydroxylation of available ß-keto esters. In the presence of a chiral bifunctional guanidine catalyst, a variety of optically active oxaziridines and chiral α-hydroxy ß-keto esters were generated with excellent results (ee's of up to 99% and 97% and yields of up to 44% and 54%, respectively).

Asymmetric [3 + 2] Cycloaddition of Methyleneindolinones with N,N'-Cyclic Azomethine Imines Catalyzed by a N,N'-Dioxide-Mg(OTf)2 Complex.

A highly efficient chiral N,N'-dioxide-Mg(OTf)2 catalyst system has been developed for the asymmetric 1,3-dipolar cycloaddition between methyleneindolinones and N,N'-cyclic azomethine imines. The desired pyrazolidine products with contiguous quaternary-tertiary stereocenters were obtained in up to 99% yields with up to 99% ee and >19:1 dr under mild reaction conditions.

The N,N'-dioxide/Ni(II)-catalyzed asymmetric inverse-electron-demand hetero-Diels-Alder reaction of methyleneindolinones with hetero-substituted alkenes.

The highly efficient catalytic asymmetric inverse-electron-demand hetero-Diels-Alder reaction of methyleneindolinones with hetero-substituted alkenes has been accomplished under mild reaction conditions. In the presence of chiral N,N'-dioxide/Ni(II) complexes, a wide range of optically active dihydropyran-fused indoles were obtained in up to 99% yield, >95 : 5 dr and 99% ee.

Catalytic hetero-ene reactions of 5-methyleneoxazolines: highly enantioselective synthesis of 2,5-disubstituted oxazole derivatives.

An efficient catalytic asymmetric hetero-ene reaction of 5-methyleneoxazolines with 1,2-dicarbonyl compounds (including α-ketoesters and glyoxal derivatives) was realized using Ni(II)-N,N'-dioxide complexes as the catalysts. It provides a rapid, high yielding (up to 99%) route for the preparation of 2,5-disubstituted oxazole derivatives in a highly enantioenriched form (up to >99% ee) under mild conditions.

A neutralization scFv antibody against IL-1ß isolated from a NIPA-based bacterial display library.

OBJECTIVE: RA is one of autoimmune diseases, has drawn great attention of the world. Currently, the anti- IL-1ß monoclonal antibody Canakinumab (ACZ885) for treatment of RA has entered into clinical trials. However, Full length antibody has large molecular weight, and is difficult to penetrate the tissue or the nidus. In contrast, scFv has low molecular weight and strong penetration ability, and is favorable to increase the drug concentration in the indus, hence improving the efficacy of the drug. The aim of this study is to obtain a neutralizing scFv antibody from a combinatorial scFv library against hIL-1ß by the modified NLPA-based bacterial display system, for further development of the small molecule antibody drug for treatment of RA. METHODS: The modified NIPA-based bacterial display system was used to construct the combinatorial scFv library derived from the spleen cDNA of immunized mice with hIL-1ß. FACS was used to screen hIL- 1ß-binding clones with FITC-labeled hIL-1ß protein. Three clones were randomly selected from the third round of screening, and their nucleotide sequences were aligned with mouse immunoglobulin genes. The single chain antibody genes of the hIL-1ß-binding clones were subcloned into the prokaryotic expression vector pET-27b for expression. The molecular mass of the purified anti-hIL-1ß single chain antibody was about 28ku. The hIL-1ß-binding ability of antibody were examined by ELISA and Western blot assays. Ability of the scFv antibody to neutralize hIL-1ß was evaluated by the MTT test. CONCLUSIONS: In this study, it is the first time to use the NIPA-based bacteria display system to construct and screen the combinatorial scFv library. Three scFvs against hIL-1ß were obtained from the scFv library of the immunized mice. Prokaryotically expressed and purified scFvs demonstrate binding ability with hIL-1ß. Among the three clones. The MTT test suggests that scFv-20 is a neutralization antibody against hIL-1ß. The study provides a lead candidate for further development of small molecule therapeutic antibodies for treatment of RA.

[Establishment of bacteria display technology for Fab antibody library screening].

AIM: To establish bacterial display technology for the purpose of Fab antibody library screening, by using six amino acids (CDQSSS) of the amino termimus of NlpA protein to anchore antibodies to the periplasmic side of the bacterial inner membrane. METHODS: The NlpA Leader sequences (encoding CDQSSS) was amplified from pNAD plasmid. The PCR product was subcloned into pComb3 expression vector to generate Fab display vector pBFD. The heavy chains of the Fab gene fragments and the light chains of the anti-human IL-1ß (hIL-1ß) antibody were inserted downstream of the NlpA leader and pelB leader respectively to construct the pBFD-Fab for Fab antibody display. Then pBFD-Fab transformed E.coli DH5α was induced by IPTG to express the Fab antibodies, as detected by flow cytometry (FCM), and positive populations were sorted. Instead of PCR, plasmids were extracted for rescue purpose. The rescue plasmids were retransformed to E.coli DH5α and FCM was performed again. RESULTS: The pBFD-Fab-transformed bacteria were incubated with antigen and antigen specific FITC-antibody, and showed strong fluorescence as detected by FCM in a dose-dependent manner. The rescued pBFD-Fab displayed similar fluorescence intensity, indicating the reliability of this technology. CONCLUSION: The Fab expressed by the bacterial display system folds efficiently and binds to hIL-1ß specifically. The plasmid rescue works well and it can avoid mutation and mis-pairing chains. This bacterial display technology has the stability of antibody expression. This study has used the technology to screen anti-hIL-1ß Fab antibody Library successfully.