JaponiconeA induces apoptosis of bortezomib-sensitive and -resistant myeloma cells in vitro and in vivo by targeting IKKβ

Objective:Multiple myeloma(MM)remains incurable with high rates of relapse.New therapeutic drugs are therefore urgently needed to improve the prognosis.JaponiconeA(JA),a natural product isolated from Inula japonica Thunb,has shown good anti-MM potential.A comprehensive study should therefore be conducted to identify both the in vitro and in vivo mechanisms of the anti-MM effects of JA.Methods:CCK8 assays and flow cytometry were used to detect the proliferation,apoptosis,and cell cycle of MM cell lines when treated with JA.In vivo experiments were conducted using subcutaneous xenograft mouse models.We also identified possible targets and the mechanism of JA using RNA-seq and c-Map databases,and identified the specific targets of JA in bortezomib-sensitive and-resistant MM cell lines using CETSA,DARTS,and rescue experiments.Furthermore,JA and bortezomib were used separately or together to characterize their possible synergistic effects.Results:In vitro,JA inhibited proliferation,and induced apoptosis and G2/M phase arrest in MM cell lines,and selectively killed primary CD138+MM cells.In vivo,JA also demonstrated a strong anti-tumor effect with no observable toxicity.In addition,JA showed synergetic effects in combination with bortezomib,and enhanced the anti-tumor effect of bortezomib in bortezomibresistant cells.CETSA and DARTS confirmed direct binding of JA to NF-κB inhibitor kinase beta(IKKβ),and overexpression of IKKβor knockdown of IκBαpartially rescued the apoptosis induced by JA.Conclusions:JA exhibited strong anti-tumor effects in MM.It sensitized myeloma cells to bortezomib and overcame NF-κB-induced drug resistance by inhibiting IKKβ,providing a new treatment strategy for MM patients.

Research review of the cement sand and gravel(CSG)dam

The cement sand and gravel(CSG)dam is a new style of dam that owes the advantages both of the concrete faced rock-fill dam(CRFD)and roller compacted concrete(RCC)gravity dam,because of which it has attracted much attention of experts home and abroad.At present,some researches on physic-mechanical property of CSG material and work behavior of CSG dam have been done.This papcr introduces the development and characteristics of CSG dam systematically,and summarizes the progress of the study on basic tests,constitutive relation of CSG material and numerical analysis of CSG dam,in addition,indicates research and application aspect of the dam.

Ubiquitin-specific protease 47 regulates intestinal inflammation through deubiquitination of TRAF6 in epithelial cells

Deubiquitinates(DUBs) alter the stabilities, localizations or activities of substrates by removing their ubiquitin conjugates,which are closely related to the development of inflammatory response. Here, we show that ubiquitin-specific protease 47(USP47) prevents inflammation development in inflammatory bowel disease(IBD). Compared with wild-type mice, Usp47 knockout mice are more susceptible to dextran sodium sulfate(DSS)-induced acute and chronic colitis with higher inflammatory cytokines expression and severe intestinal tissue damage. Chimeric mouse experiments suggest that non-hematopoietic cells mainly contribute to the phenotype. And, DSS-induced colitis of the Usp47 knockout mice depends on commensal bacteria.Mechanistically, down-regulation of USP47 aggravates the activation of NF-κB signaling pathway by increasing the K63-linked poly-ubiquitination of tumor necrosis factor receptor-associated factor 6(TRAF6) in intestinal epithelial cells. Furthermore, the expression of USP47, negatively correlated with the degree of inflammation, is lower at colonic inflammatory lesions than that non-inflammatory sites from the intestine from ulcerative colitis(UC) and Crohn's disease(CD) patients. These data, taken together, indicate that USP47 regulates intestinal inflammation through de-ubiquitination of K63-linked poly-ubiquitination TRAF6 in intestinal epithelial cells.

USP7 deubiquitinates and stabilizes NOTCH1 in T-cell acute lymphoblastic leukemia

T-cell acute lymphoblastic leukemia(T-ALL)is a highly aggressive leukemia that is primarily caused by aberrant activation of the NOTCH1 signaling pathway.Recent studies have revealed that posttranslational modifications,such as ubiquitination,regulate NOTCH1 stability,activity,and localization.However,the specific deubiquitinase that affects NOTCH1 protein stability remains unestablished.Here,we report that ubiquitin-specific protease 7(USP7)can stabilize NOTCH1.USP7 deubiquitinated NOTCH1 in vivo and in vitro,whereas knockdown of USP7 increased the ubiquitination of NOTCH1.USP7 interacted with NOTCH1 protein in T-ALL cells,and the MATH and UBL domains of USP7 were responsible for this interaction.Depletion of USP7 significantly suppressed the proliferation of T-ALL cells in vitro and in vivo,accompanied by downregulation of the NOTCH1 protein level.Similarly,pharmacologic inhibition of USP7 led to apoptosis of T-ALL cells.More importantly,we found that USP7 was significantly upregulated in human T-ALL cell lines and patient samples,and a USP7 inhibitor exhibited cell cytotoxicity toward primary T-ALL cells,indicating the clinical relevance of these findings.Overall,our results demonstrate that USP7 is a novel deubiquitinase that stabilizes NOTCH1.Therefore,USP7 may be a promising therapeutic target in the currently incurable T-ALL.

NES1/KLK10 and hNIS gene therapy enhanced iodine-131 internal radiation in PC3 proliferation inhibition

NES1 gene is thought to be a tumor-suppressor gene.Our previous study found that overexpression of NES1 gene in PC3 cell line could slow down the tumor proliferation rate,associated with a mild decrease in BCL-2 expression.The BCL-2 decrease could increase the sensitivity of radiotherapy to tumors.Thus,we supposed to have an“enhanced firepower”effect by combining overexpressed NES1 gene therapy and 131I radiation therapy uptake by overexpressed hNIS protein.We found a weak endogenous expression of hNIS protein in PC3 cells and demonstrated that the low expression of hNIS protein in PC3 cells might be the reason for the low iodine uptake.By overexpressing hNIS in PC3,the radioactive iodine uptake ability was significantly increased.Results of in vitro and in vivo tumor proliferation experiments and 18F-fluorothymidine(18F-FLT)micro-positron emission tomography/computed tomography(micro-PET/CT)imaging showed that the combined NES1 gene therapy and 131I radiation therapy mediated by overexpressed hNIS protein had the best tumor proliferative inhibition effect.Immunohistochemistry showed an obvious decrease of Ki-67 expression and the lowest BCL-2 expression.These data suggest that via inhibition of BCL-2 expression,overexpressed NES1 might enhance the effect of radiation therapy of 131I uptake in hNIS overexpressed PC3 cells.

YL064 activates proteasomal-dependent degradation of c-Myc and synergistically enhances the anti-tumor activity of ABT-199 in diffuse large B cell lymphoma

Dear Editor,c-Myc is highly associated with poor prognosis and aggressive progression of diffuse large B cell lymphoma(DLBCL)and is thus a desirable drug target.Moreover,studies indicate that 5–15%of DLBCL patients harbor MYC and BCL-2 translocations,while 20–35%DLBCL patients simultaneously overexpress of c-Myc and BCL-2 proteins without gene rearrangements.1 These two types of DLBCL are referred as“double-hit”lymphoma(DHL)and“double-expressor”lymphoma(DEL),respectively.Both DHL and DEL lymphomas have inferior clinical outcomes and are refractory to R-CHOP or even hematopoietic stem cell transplant.2 Thus,targeting both c-Myc and BCL-2 is a promising strategy to treat high-risk DLBCLs.3 Although BCL-2 inhibitors are clinically available,c-Myc remains to be“undruggable”owing to its lack of kinase activity and intrinsically disordered structure.4 Thus,developing clinically applicable c-Myc inhibitor remains challenging.

Hsp90/C terminal Hsc70-interacting protein regulates the stability of Ikaros in acute myeloid leukemia cells

The stability of Ikaros family zinc finger protein 1(Ikaros),a critical hematopoietic transcription factor,can be regulated by cereblon(CRBN)ubiquitin ligase stimulated by immunomodulatory drugs in multiple myeloma.However,other stabilization mechanisms of Ikaros have yet to be elucidated.In this study,we show that the pharmacologic inhibition or knockdown of Hsp90 downregulates Ikaros in acute myeloid leukemia(AML)cells.Proteasome inhibitor MG132 but not autophagy inhibitor chloroquine could suppress the Hsp90 inhibitor STA-9090-induced reduction of Ikaros,which is accompanied with the increased ubiquitination of Ikaros.Moreover,Ikaros interacts with E3 ubiquitin-ligase C terminal Hsc70 binding protein(CHIP),which mediates the STA-9090-induced ubiquitination of Ikaros.In addition,the knockdown of Ikaros effectively inhibits the proliferation of leukemia cells,but this phenomenon could be rescued by Ikaros overexpression.Collectively,our findings indicate that the interplay between HSP90 and CHIP regulates the stability of Ikaros in AML cells,which provides a novel strategy for AML treatment through targeting the HSP90/Ikaros/CHIP axis.