Development of an enzyme-linked immunosorbent assay for Keap1-Nrf2 interaction inhibitors identification.

Development of Keap1-Nrf2 interaction inhibitors is a promising strategy for the discovery of therapeutic agents against oxidative stress-mediated diseases. Two motifs of Nrf2, ETGE and DLG motif, are responsible for Keap1-Nrf2 binding. Previously, ETGE peptide or ETGE-derived peptide-based approaches were used to detect Keap1-Nrf2 interaction; however, these approaches are not able to monitor Keap1-DLG motif binding. We first report here a novel Enzyme-linked Immunosorbent Assay (ELISA) approach to detect the protein-protein interaction of full length Keap1 and Nrf2. In our assay, the test compounds can target either ETGE or DLG binding site, therefore facilitating the exploration of diverse Keap1-Nrf2 inhibitors. Three FDA-approved drugs, zafirlukast, dutasteride and ketoconazole, were found to inhibit the Keap1-Nrf2 interaction with IC50 of 5.87, 2.81 and 1.67 µM, respectively. Additionally, these three drugs also activated Nrf2 pathway in neuroblasts and lipopolysaccharide (LPS)-challenged mice. The results presented here indicate that the ELISA approach has the capacity to identify Keap1-Nrf2 inhibitors.

Rapid screening and identification of α-amylase inhibitors from Garcinia xanthochymus using enzyme-immobilized magnetic nanoparticles coupled with HPLC and MS.

α-Amylase inhibitors play an important role in management of diabetes and obesity. In order to rapidly discover potent α-amylase inhibitors from medicinal plants, a ligands-screening method based on enzyme-immobilized magnetic nanoparticles integrated with HPLC was developed. Amine-terminated magnetic nanoparticles were prepared for the immobilization of α-amylase. Based on the affinity theory, the α-amylase-coated magnetic nanoparticles were employed to fish out the ligands from the extracts of Garcinia xanthochymus, and the elutes were examined by HPLC. As a result, three ligands were screened out. Isolation and identification were carried out subsequently. By analyzing the UV, MS and NMR spectra, they were identified as three biflavonoids including GB2a glucoside (2), GB2a (3) and fukugetin (4). The IC50 values of the three compounds were also determined. The results suggest the proposed approach is efficient and accurate, and has great potential in rapid discovery of drug candidates from medical plants.

Successful treatment with bortezomib and thalidomide for POEMS syndrome associated with multicentric mixed-type Castleman's disease.

Polyneuropathy, organomegaly, endocrinopathy, M-protein and skin changes syndrome is a rare multi-systematic disorder of uncertain etiology, if associated with multicentric Castleman's disease, it can lead to a more serious condition. We here presented a case of polyneuropathy, organomegaly, endocrinopathy, M-protein and skin changes syndrome in a 37-year-old male patient who initially presented with progressive lower limb weakness accompanied by pain, low skin temperature, and hyperpigmentation. He was admitted with increasingly serious dyspnea and lower leg edema. Fluid of serous cavities in the patient were also indicated in ultrasonic inspection and X-ray. Furthermore, biopsy of a left axillary lymph node showed mixed hyaline-vascular and plasma cell type of multicentric Castleman's disease. Administration of bortezomib (Velcade) (1.3 mg/m(2) on days 1, 4, 8 and 11 of a 21-day cycle) combined with thalidomide (100 mg/day and 21-day cycle) dramatically improved the condition of this disease. Of note, in our study, combination therapy of bortezomib and thalidomide successfully improved the condition of the patient with polyneuropathy, organomegaly, endocrinopathy, M-protein and skin changes syndrome associated with multicentric Castleman's disease, suggesting that the combination therapy may be an effective therapeutic strategy for the intractable polyneuropathy, organomegaly, endocrinopathy, M-protein and skin changes syndrome associated with multicentric Castleman's disease.