Novel Hsp90 inhibitor platycodin D disrupts Hsp90/Cdc37 complex and enhances the anticancer effect of mTOR inhibitor.

Heat shock protein 90 (Hsp90) is a critically conserved molecular chaperone protein and promising therapeutic target for cancer treatment. In this study, platycodin D (PD), a saponin isolated from traditional Chinese herb Platycodonis Radix, was identified as a novel Hsp90 inhibitor. We verified that PD did not affect the ATPase activity of Hsp90. However, PD disrupted the co-chaperone interaction of Hsp90/cell division cycle protein 37 (Cdc37) and subsequently degraded multiple Hsp90 client proteins without the feedback increase of Hsp70. In different genotypes of non-small cell lung cancer cells, co-treatment with the mTOR inhibitor Everolimus and PD enhanced antiproliferation activity and apoptotic effect. The feedback survival signal upon mTOR inhibition was fully terminated by the co-administration with PD through reduced epidermal growth factor receptor (EGFR) and insulin growth factor 1 receptor (IGF1R) expression, suppressed AKT activity, and reinforced 4E-BP1 inhibition. Our results not only identified PD as a novel Hsp90 inhibitor by disrupting the protein-protein interaction of Hsp90/Cdc37 complex, but also provided mechanistic insights into the ineffectiveness of mTOR inhibitors and identified therapeutic strategy for cancer treatment.

Delivery of therapeutic small interfering RNA: The current patent-based landscape.

Implementing small interfering RNA (siRNA) is a promising therapy because it silences disease-related genes theoretically. However, the efficient delivery of siRNA is challenging, which limits its therapeutic applications. Various pharmaceutical delivery systems containing key technologies have been developed and patented, which are of great concern to developers in the field. Despite numerous studies devoted to siRNA-delivery technologies, few researchers have systematically examined relevant patents. Patents, as bridges connecting academic progress with applicable innovation, encapsulate cumulative technological innovations and provide valuable information for academic research and commercial development. This study aims to analyze advances in therapeutic siRNA delivery technology from a patent perspective. A total of 11,509 patent documents from 3,309 patent families were collected, classified into 10 technological categories, and comprehensively analyzed. An overall patent landscape of siRNA delivery was presented from the temporal, spatial, organizational, and technological dimensions. This work is expected to help researchers and developers in the field of siRNA delivery form a basis for decision-making by combining our findings with supplementary data.

PI3KC3 complex subunit NRBF2 is required for apoptotic cell clearance to restrict intestinal inflammation.

NRBF2, a regulatory subunit of the ATG14-BECN1/Beclin 1-PIK3C3/VPS34 complex, positively regulates macroautophagy/autophagy. In this study, we report that NRBF2 is required for the clearance of apoptotic cells and alleviation of inflammation during colitis in mice. NRBF2-deficient mice displayed much more severe colitis symptoms after the administration of ulcerative colitis inducer, dextran sulfate sodium salt (DSS), accompanied by prominent intestinal inflammation and apoptotic cell accumulation. Interestingly, we found that nrbf2-/- mice and macrophages displayed impaired apoptotic cell clearance capability, while adoptive transfer of nrbf2+/+ macrophages to nrbf2-/- mice alleviated DSS-induced colitis lesions. Mechanistically, NRBF2 is required for the generation of the active form of RAB7 to promote the fusion between phagosomes containing engulfed apoptotic cells and lysosomes via interacting with the MON1-CCZ1 complex and regulating the guanine nucleotide exchange factor (GEF) activity of the complex. Evidence from clinical samples further reveals the physiological role of NRBF2 in maintaining intestinal homeostasis. In biopsies of UC patient colon, we observed upregulated NRBF2 in the colon macrophages and the engulfment of apoptotic cells by NRBF2-positive cells, suggesting a potential protective role for NRBF2 in UC. To confirm the relationship between apoptotic cell clearance and IBD development, we compared TUNEL-stained cell counts in the UC with UC severity (Mayo Score) and observed a strong correlation between the two indexes, indicating that apoptotic cell population in colon tissue correlates with UC severity. The findings of our study reveal a novel role for NRBF2 in regulating apoptotic cell clearance to restrict intestinal inflammation.Abbreviation: ANOVA: analysis of variance; ATG14: autophagy related 14; ATG16L1: autophagy related 16-like 1 (S. cerevisiae); BMDM: bone marrow-derived macrophage; BSA: bovine serum albumin; CD: Crohn disease; CD68: CD68 molecule; CFP: cyan fluorescent protein; CMFDA: 5-chloromethylfluorescein diacetate; Co-IP, co-immunoprecipitation; CPR: C-reactive protein; Cy7: cyanine 7 maleimide; DAB: diaminobezidine 3; DAI: disease activity indexes; DAPI: 4'6-diamidino-2-phenylindole; DMEM: dulbecco's modified eagle's medium; DMSO: dimethyl sulfoxide; DOC: sodium deoxycholate; DSS: dextran sulfate sodium; EDTA: ethylenediaminetetraacetic acid; EGTA: ethylenebis (oxyethylenenitrilo) tetraacetic acid; FBS: fetal bovine serum; FITC: fluorescein isothiocyanate; FRET: Förster resonance energy transfer; GDP: guanine dinucleotide phosphate; GEF: guanine nucleotide exchange factor; GFP: green fluorescent protein; GTP: guanine trinucleotide phosphate; GWAS: genome-wide association studies; HEK293: human embryonic kidney 293 cells; HRP: horseradish peroxidase; IBD: inflammatory bowel disease; IgG: immunoglobin G; IL1B/IL-1ß: interleukin 1 beta; IL6: interleukin 6; IRGM: immunity related GTPase M; ITGAM/CD11b: integrin subunit alpha M; KO: knockout; LRRK2: leucine rich repeat kinase 2; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MOI: multiplicity of infection; MPO: myeloperoxidase; NaCl: sodium chloride; NEU: neutrophil; NOD2: nucleotide binding oligomerization domain containing 2; NP40: nonidet-P40; NRBF2: nuclear receptor binding factor 2; PBS: phosphate buffer saline; PCR: polymerase chain reaction; PE: P-phycoerythrin; PIK3C3/VPS34: phosphatidylinositol 3-kinase catalytic subunit type 3; PtdIns3P: phosphatidylinositol-3-phosphate; PTPRC/CD45: protein tyrosine phosphatase receptor type C; SDS-PAGE: sodium dodecylsulphate-polyacrylamide gel electrophoresis; TBST: tris-buffered saline Tween-20; Tris-HCl: trihydroxymethyl aminomethane hydrochloride; TUNEL: TdT-mediated dUTP nick-end labeling; UC: ulcerative colitis; ULK1: unc-51 like autophagy activating kinase 1; WB: western blotting; WT: wild type; YFP: yellow fluorescent protein.