Sapanisertib attenuates pulmonary fibrosis by modulating Wnt5a/mTOR signalling.

Sapanisertib is an orally bioavailable ATP-dependent high-potential raptor-mTOR (TORC1) inhibitor with antineoplastic activity. Here, the impact of sapanisertib was assessed on transforming growth factor-ß1 (TGF-ß1)-treated L929 and A549 cells and on a rat model of bleomycin pulmonary fibrosis. First, in A549 cells treated with TGF-ß1, sapanisertib significantly suppressed the TGF-ß1-induced epithelial-mesenchymal transition, with elevated and reduced E-cadherin and vimentin expression, respectively. In L929 cells treated with TGF-ß1, sapanisertib significantly blocked the TGF-ß1-induced cell proliferation, with decreases in the extracellular matrix-related proteins collagens I and III and smooth muscle actin and in the mechanism-related proteins hypoxia-inducing factor, mTOR, p70S6K, and Wnt5a. Compared with bleomycin alone, continuous gavage administration of sapanisertib for 14 days reduced pathological scores in bleomycin-induced pulmonary fibrosis rats, with decreases in collagen deposition and in the same proteins as in L929 and A549 cells. Accordingly, our findings show that sapanisertib can ameliorate experimental pulmonary fibrosis by inhibiting Wnt5a/mTOR/HIF-1α/p70S6K.

Development and validation of a novel reporter gene assay for determination of recombinant human thrombopoietin.

Recombinant human thrombopoietin (rhTPO) was approved by the National Medical Products Administration in 2010 for the treatment of thrombocytopenia in patients with immune thrombocytopenic purpura and chemotherapy-induced thrombocytopenia. Nevertheless, no method for determining rhTPO bioactivity has been recorded in different national/regional pharmacopoeia. Novel methods for lot release and stability testing are needed that are simpler, quicker, and more accurate. Here, we developed a novel reporter gene assay (RGA) for rhTPO bioassay with Ba/F3 cell lines that stably expressed human TPO receptor and luciferase reporter driven by sis-inducible element, gamma response region, and gamma-interferon activated sequence. During careful optimization, the RGA method demonstrated high performance characteristics. According to the International Council for Harmonization Q2 (R1) guidelines and the Chinese Pharmacopoeia 2020 edition, the validation results demonstrated that this method is highly time-saving, sensitive, and robust for research, development, manufacture, and quality control of rhTPO.

Design, synthesis and biological evaluation of 1-hydroxy-2-phenyl-4-pyridyl-1H-imidazole derivatives as xanthine oxidase inhibitors.

In our previous study, we reported a series of 1-hydroxy-2-phenyl-1H-imidazole-5-carboxylic acid derivatives that presented excellent in vitro xanthine oxidase inhibitory potency. As a continuation study, a series of 1-hydroxy-2-phenyl-1H-imidazole derivatives containing a pyridine moiety (4a-g and 5a-g) at the 4-position was designed and synthesized. Evaluation of in vitro xanthine oxidase inhibition demonstrated that the 4a-g series was more potent than the 5a-g series. Compound 4f was the most promising derivative in the series with an IC50 value of 0.64 µM. A Lineweaver-Burk plot revealed that compound 4f acted as a mixed-type xanthine oxidase inhibitor. An iso-pentyloxy group at the 4'-position improved the inhibitory potency. More interestingly, structure-activity relationship analysis indicated that the pyridine para-N atom played a crucial role in the inhibition. Molecular modeling provided a reasonable explanation for the structure-activity relationships observed in this study. In addition, a three dimensional quantitative structure-activity relationships model which possessed reasonable statistics (q2 = 0.885 and r2 = 0.993) was conducted to further understand the structural basis of these compounds as xanthine oxidase inhibitors. These compounds, especially compound 4f, have good potential for further investigations.