Geldanamycin (GA) is an antibiotic targeting the ADP/ATP binding site of heat shock protein 90 (Hsp90).In screening for anti-herpes simplex virus type 1 (HSV-1) candidates, we found GA active against HSV-1. HSV-1 replication in vitro was significantly inhibited by GA with an 50% inhibitory concentration of 0.093 M and a concentration that inhibited cellular growth 50% in comparison with the results seen with untreated controls of 350 M. The therapeutic index of GA was over 3,700 (comparable to the results seen with acyclovir). GA did not inhibit HSV-1 thymidine kinase. Cells infected with HSV-1 demonstrated cell cycle arrest at the G 1 /S transition; however, treatment with GA resulted in a cell cycle distribution pattern identical to that of untreated cells, indicating a restoration of cell growth in HSV-1-infected cells by GA treatment. Accordingly, HSV-1 DNA synthesis was suppressed in HSV-1 ؉ cells treated with GA. The antiviral mechanism of GA appears to be associated with Hsp90 inactivation and cell cycle restoration, which indicates that GA exhibits broad-spectrum antiviral activity. Indeed, GA exhibited activities in vitro against other viruses, including severe acute respiratory syndrome coronavirus. Since GA inhibits HSV-1 through a cellular mechanism unique among HSV-1 agents, we consider it a new candidate agent for HSV-1.Geldanamycin (GA) is a benzoquinone ansamycin and was first isolated as a new entity from the fermentation of Streptomyces hygroscopicus (3). GA binds with a high level of specificity within the ADP/ATP binding pocket of heat shock protein 90 (Hsp90) and inhibits the function of this chaperone (27,36), resulting in inappropriately functioning and rapid degradation of Hsp90-associated client proteins (2, 28). The client proteins are mainly short-lived proteins, including several protein kinases (Raf-1, ErbB-2, and Bcr-Abl), p53, and pRb as well as cyclins and cyclin-dependent kinases (2, 25), which are degraded through the ubiquitin-proteasome pathway but protected by Hsp90 (4,19). As a specific inhibitor of Hsp90 function, GA demonstrated antitumor activity in a multitude of animal models (23) and is now in clinical trial (phase I) in the United States (26). Interference with the function of Hsp90 seems to be the major mechanism of action of GA (29).In a large-scale screening for novel candidates exhibiting activity against herpes simplex virus type 1 (HSV-1), we found (from the fermentation of S. hygroscopicus) a component active against HSV-1 replication. Chemical analysis of the purified compound showed a structure identical to that of GA. A study was then initiated to evaluate the potential of this compound in the treatment of HSV-1 infection. In the present study, the anti-HSV-1 effect of GA was examined in cell cultures. The possible molecular mechanism responsible for its activity against viral infection was also explored. Our investigation showed that the mode of action of GA was closely related to the inactivation of cellular Hsp90 and different from that seen with the vira...

The Bcl-2 protein has an anti-apoptotic effect in neuronal and other cell types. We show for the first time that the Bcl-2 promoter is activated by the neuronal survival factor nerve growth factor (NGF) and that this effect is dependent on a region of the promoter from -1472 to -1414. This activation requires the Rap-1 G protein and the MEK-1 and p42/p44 MAPK enzymes but is independent of other NGF-activated signalling pathways involving protein kinase A or protein kinase C.

MicroRNAs (miRNAs) play important roles in the cancer development and progression; overexpression of miR-103 has been identified in various tumors. However, its biological function and regulatory mechanism involved in modulation of human gastric cancer (GC) remain largely unknown. This study aimed to confirm clinical significance of miR-103 and investigate its biological role and underlying mechanism in GC. Real-time quantitative PCR (qRT-PCR) revealed miR-103 was highly expressed in GC tissues and cell lines. miR-103 expression was correlated closely with tumor size, Lauren’s classification, and lymph node metastasis. Importantly, Kaplan-Meier analysis revealed that high expression of miR-103 was significantly associated with poor overall survival and disease-free survival of GC patients. Downregulation of miR-103 by transfecting with miR-103 inhibitor significantly suppressed cell proliferation, induced apoptosis, inhibited migration and invasion in vitro and in vivo. Furthermore, miRNA target databases and luciferase reporter assay confirmed that Krüppel-like Factor-4 (KLF4) was a direct target of miR-103 in GC, and there was a significant inverse correlation between miR-103 and KLF4 expression in GC tissues. Moreover, KLF4 downregulation could rescue miR-103’s oncogenic effect on GC cell proliferation, apoptosis, migration, and invasion. Therefore, these results suggested that miR-103 overexpression could contribute to tumor progression by suppressing KLF4, and it might serve as a promising candidate for the prognosis of GC patients.

1Background: The colloidal gold immunochromatography assay (GICA) is a rapid 2 diagnostic tool for novel coronavirus disease 2019 (COVID-19) infections. However, 3 with significant numbers of false negatives, improvements to GICA are needed. 4Methods: Six recombinant HCoV-19 nucleocapsid and spike proteins were prepared 5 and evaluated. The optimal proteins were employed to develop a sandwich-format 6 GICA strip to detect total antibodies (IgM and IgG) against HCoV-19. GICA's 7 performance was assessed with comparison of viral RNA detection. 8Results: Recombinant HCoV-19 proteins were obtained, including three 9 prokaryotically expressed rN, rN1, rN2 nucleocapsid proteins, and three 10 eukaryotically expressed rS1, rS-RBD, rS-RBD-mFc spike proteins. The recombinant 11 proteins with the highest ELISA titers (rS1 and rS-RBD-mFc) against 12 coronavirus-specific IgM and IgG were chosen for GICA development. The GICA 13 has a sensitivity and specificity of 86.89% (106/122) and 99.39% (656/660), 14 respectively. Furthermore, 65.63% (21/32) of the clinically confirmed but RT-PCR 15 negative samples were GICA positive. 16 Conclusions:The eukaryotically-expressed spike proteins (rS1and rS-RBD-mFc) are 17 more suitable than the prokaryotically expressed nucleocapsid proteins for HCoV-19 18 serological diagnosis. The GICA sandwich used to detect total antibodies is a 19 powerful complement to the current standard RNA-based tests.

No abstract