Metformin, the most prescribed antidiabetic medicine, has shown other benefits such as anti-ageing and anticancer effects
. For clinical doses of metformin, AMP-activated protein kinase (AMPK) has a ...major role in its mechanism of action
; however, the direct molecular target of metformin remains unknown. Here we show that clinically relevant concentrations of metformin inhibit the lysosomal proton pump v-ATPase, which is a central node for AMPK activation following glucose starvation
. We synthesize a photoactive metformin probe and identify PEN2, a subunit of γ-secretase
, as a binding partner of metformin with a dissociation constant at micromolar levels. Metformin-bound PEN2 forms a complex with ATP6AP1, a subunit of the v-ATPase
, which leads to the inhibition of v-ATPase and the activation of AMPK without effects on cellular AMP levels. Knockout of PEN2 or re-introduction of a PEN2 mutant that does not bind ATP6AP1 blunts AMPK activation. In vivo, liver-specific knockout of Pen2 abolishes metformin-mediated reduction of hepatic fat content, whereas intestine-specific knockout of Pen2 impairs its glucose-lowering effects. Furthermore, knockdown of pen-2 in Caenorhabditis elegans abrogates metformin-induced extension of lifespan. Together, these findings reveal that metformin binds PEN2 and initiates a signalling route that intersects, through ATP6AP1, the lysosomal glucose-sensing pathway for AMPK activation. This ensures that metformin exerts its therapeutic benefits in patients without substantial adverse effects.
More than 60% of nonsmall cell lung cancer (NSCLC) patients show a positive response to the first ALK inhibitor, crizotinib, which has been used as the standard treatment for newly diagnosed patients ...with ALK rearrangement. However, most patients inevitably develop crizotinib resistance due to acquired secondary mutations in the ALK kinase domain, such as the gatekeeper mutation L1196M and the most refractory mutation, G1202R. Here, we develop XMU‐MP‐5 as a new‐generation ALK inhibitor to overcome crizotinib resistance mutations, including L1196M and G1202R. XMU‐MP‐5 blocks ALK signaling pathways and inhibits the proliferation of cells harboring either wild‐type or mutant EML4‐ALK in vitro and suppresses tumor growth in xenograft mouse models in vivo. Structural analysis provides insights into the mode of action of XMU‐MP‐5. In addition, XMU‐MP‐5 induces significant regression of lung tumors in two genetically engineered mouse (GEM) models, further demonstrating its pharmacological efficacy and potential for clinical application. These preclinical data support XMU‐MP‐5 as a novel selective ALK inhibitor with high potency and selectivity. XMU‐MP‐5 holds great promise as a new therapeutic against clinically relevant secondary ALK mutations.
Synopsis
Despite the clinical success of ALK inhibitors in NSCLC, multiple drug‐resistant mutations in ALK are inevitably reported. XMU‐MP‐5 overcomes resistance to first and second generation ALK inhibitors in vitro and in vivo, thus holds great promise for the therapeutic use against ALK‐positive NSCLC.
XMU‐MP‐5 is a new ALK inhibitor with high potency and selectivity.
XMU‐MP‐5 overcomes acquired resistance to first and second generation ALK inhibitors, including ALKL1196M and ALKG1202R.
XMU‐MP‐5 induces significant regression of lung tumors in ALK wild‐type and L1196M GEM models.
Despite the clinical success of ALK inhibitors in NSCLC, multiple drug‐resistant mutations in ALK are inevitably reported. XMU‐MP‐5 overcomes resistance to first and second generation ALK inhibitors in vitro and in vivo, thus holds great promise for the therapeutic use against ALK‐positive NSCLC.
Rational design and fabrication of small interfering RNA (siRNA) delivery system with simple production scheme, specific targeting capability, responsiveness to endogenous stimuli and potential ...multi-functionalities remains technically challenging. Herein, we screen and design a virus-mimicking polysaccharide nanocomplex that shows specific gene delivery capability in a selective subset of leukocytes. A virus-inspired poly (alkyl methacrylate-co-methacrylic acid) fragment was conjugated on barley β-glucans (EEPG) to endow the nanocomplex with pH-dependent endosomal membrane destabilization capabilities, as confirmed both biologically and computationally. siRNA loaded EEPG nanocomplex is feasibly fabricated in a single-step manner, which exhibit efficient gene silencing efficacy towards Dectin-1+ monocytes/macrophages. The inherent targeting affinity and feasible gene silencing potency of EEPG nanocomplex are investigated in three independent murine inflammation models, including myocardial infarction, lung fibrosis and acute liver damage. Significant enhanced accumulation level of EEPG nanocomplex is observed in cardiac lesion site, indicating its exclusive targeting capability for ischemic heart diseases. As a proof of concept, siTGF-β based gene therapy is confirmed in murine model with heart fibrosis. Overall, our findings suggest the designed EEPG nanocomplex is favorable for siRNA delivery, which might have translational potential as a versatile platform in inflammation-related diseases.
A virus-mimicking polysaccharide-based multifunctional nanocomplex shows considerable endosomal escape activity for siRNA delivery, which is manifested via computational and biological tools. In vivo, the inherent targeting capability and feasible gene silencing potency of produced nanocomplex are confirmed in three independent inflammation murine models, suggesting the potential value of this versatile platform for inflammation-related diseases. Created with BioRender.com. Display omitted
•On-demand designed, screen and synthesis of β-glucan-based nanoparticles for gene delivery.•Computational simulation to understand the mechanism of endosome escape capability from the developed nanosystem.•One-step production scheme without further surface chemical modification to achieve potent in vivo targeting.•Targeting efficiency and gene silencing capability were tested in three independent inflammatory murine models.•As a proof of concept, siTGF-β based nanoformulation showed potential therapeutic effect in murine cardiac fibrosis model.
Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related death worldwide and targeted therapeutics exhibit limited success. Polo-like kinase 1 (PLK1), a Ser/Thr kinase, plays a ...pivotal role in cell-cycle regulation and is considered a promising target in HCC. Here, via structural optimization using both biochemical kinase assays and cellular antiproliferation assays, we discovered a potent and selective PLK1 kinase inhibitor, compound 31. Compound 31 exhibited biochemical activity with IC50 of < 0.508 nM against PLK1 and a KINOMEscan selectivity score (S(1)) of 0.02 at a concentration of 1 μM. Furthermore, 31 showed broad antiproliferative activity against a variety of cancer cell lines, with the lowest antiproliferative IC50 (11.1 nM) in the HCC cell line HepG2. A detailed mechanistic study of 31 revealed that inhibition of PLK1 by 31 induces mitotic arrest at the G2/M phase checkpoint, thus leading to cancer cell apoptosis. Moreover, 31 exhibited profound antitumor efficacy in a xenograft mouse model. Collectively, these results establish compound 31 as a good starting point for the development of PLK1 targeted therapeutics for HCC.
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•31 is a potent and selective PLK1 inhibitor with biochemical IC50 of < 0.508 nM.•31 shows a differential inhibition profile against a broad panel of cancer cells.•31 exhibits profound anti-tumor efficacy in HCC xenograft mouse model.
Cardiac Repair
In article number 2204666, Qiang Zhao, Xiaofeng Ye, Xianming Deng, Hélder A. Santos, and co‐workers introduce a microfluidics‐assisted single step, green synthesis method for producing ...targeting ligands free heart homing nanoparticles in a tailored manner. The developed microfluidic platform facilitates an economically feasible and simple yet robust synthesis procedure for nanoparticle production, which shows great clinical translational potential as an alternative post‐conditioning routine after revascularization surgery.
Nanoparticle (NP)‐based intravenous administration represents the most convenient cardiac targeting delivery routine, yet, there are still therapeutic issues due to the lack of targeting efficiency ...and specificity. Active targeting methods using functionalization of ligands onto the NPs’ surface may be limited by trivial modification procedures and reduced targeting yield in vivo. Here, a microfluidics assisted single step, green synthesis method is introduced for producing targeting ligands free heart homing NPs in a tailored manner. The generated β‐glucan‐based NPs exhibit precise and efficient targeting capability toward Dectin‐1+ monocytes/macrophages, which are confirmed as main pathogenesis mediators for cardiac ischemic/reperfusion (I/R) injury, with a sequentially enhanced cardiac NP accumulation, and this targeting strategy is exclusively suitable for cardiac I/R but not for other cardiovascular diseases, as confirmed both in murine and human model. Comparing to FDA‐approved nano‐micelles formulation, β‐glucan NPs loaded with NACHT, LRR, and PYD domains‐containing protein 3 (NLRP3) inflammasome inhibitor (CY‐09) exhibit better efficiency in ameliorating myocardial injury and heart failure induced by surgically induced I/R. These findings indicate a simple production of targeting‐ligand free NPs, and demonstrate their potential therapeutic applications for preclinical I/R‐induced cardiac injury amelioration.
Conventional methods for synthesizing heart‐targeting nanoparticles usually require complex surface modification of specific targeting ligands. Herein, a modular microfluidics device is designed and introduced for single‐step production of heart‐targeting nanoparticles. The generated targeting‐ligand free nanoparticles may feasibly be used for myocardial reperfusion and other heart diseases.
The receptor tyrosine kinase rearranged during transfection (RET) plays pivotal roles in several cancers, including thyroid carcinoma and non-small cell lung cancer (NSCLC). Currently, there are ...several FDA-approved RET inhibitors, but their indication is limited to thyroid cancer, and none can overcome their gatekeeper mutants (V804L and V804M). Here, we report the discovery of 9x representing a new chemotype of potent and selective RET inhibitors, using a rational design strategy of type II kinase inhibitors. 9x exhibited both superior antiproliferative activities against NSCLC-related carcinogenic fusions KIF5B-RET and CCDC6-RET and gatekeeper mutant-transformed Ba/F3 cells, with the lowest GI50 of 9 nM, and substantial inhibitory activities against wild-type RET and RET mutant proteins, with the best IC50 of 4 nM. More importantly, 9x also showed nanomole potency against RET-positive NSCLC cells LC-2/ad, but not against a panel of RET-negative cancer cells, such as A549, H3122, A375 or parental Ba/F3 cells, demonstrating its selective ‘on-target’ effect. In mouse xenograft models, 9x repressed tumor growth driven by both wild type KIF5B-RET-Ba/F3 and gatekeeper mutant KIF5B-RET(V804M)-Ba/F3 cells in a dose-dependent manner. Together, these data establish that 9x provides a good starting point for the development of targeted therapeutics against RET-positive cancers, especially NSCLC.
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•9x exhibits superior activities in both RET biochemical and cellular assays.•9x selectively inhibits cell growth of RET-positive NSCLC cell LC-2/ad.•9x dose-dependently represses tumor growth in mouse xenograft models.
Based on the review of the development situation of the ETC home and abroad and the ETC key technology, this paper put forward to apply ZigBee network in the freeway ETC system. CC2431 wireless ...location function is used in the high speed highway ETC system to realize data transmission between network centre node and terminal node safely, which records passing information of the vehicle terminal by the wireless data transmission between the coordinator and vehicle terminal. To improve the position accuracy, a weighted centroid position algorithm is introduced combination the RSSI with centroid position algorithm. The test results of the wireless communication between vehicle terminal and coordinator show that CC2431 wireless position system can realize vehicle omnibearing position and real time monitor in the superspeed highway. Index Terms-ZigBee; coordinator; vehicle terminal; electronic toll collection
试验以大豆为材料,采用水培方法,以β-微管蛋白基因为内参基因,用半定量逆转录聚合酶链式反应（se-mi RT-PCR法）检测在低磷胁迫下大豆根系质膜H. _ATPase基因表达量的变化,以期建立适于检测该基因表达的semi RT-PCR试验体系。结果表明：在低磷胁迫2 h时,与对照相比基因表达量有所增加,在4 h时相对表达量达到最大,6 h略有下降。这表明大豆根系质膜H. ..._ATPase基因表达量的增加可能与适应低磷胁迫的逆境有关,半定量RT-PCR法可以用来检测特定基因在不同条件下的表达量。著者文摘
In this experiment, a semi RT- PCR system was established to detect the gene expression of PM H. _-ATPase in soybean root under phosphorus deficiency quickly and conveniently, in which β- tubulin gene was used as reference gene. The results showed that the gene of PM H. _- ATPase was up-regulated under low P environment for 2 h and increased gradually until 4 h, but the gene expression decreased a little as the time delayed to 6 h. It was shown that gene expression changes of PM H. _-ATPase may be related to the adaptation of the phosphorus deficiency stress, furthermore, the semi-quantitative RT- PCR system could be used to detect gene expression under different conditions successfully.
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