Proprotein convertase subtilisin/kexin type 9 (PCSK9) regulates low density lipoprotein receptor (LDLR) protein levels and function. Loss of PCSK9 increases LDLR levels in liver and reduces plasma LDL cholesterol (LDLc), whereas excess PCSK9 activity decreases liver LDLR levels and increases plasma LDLc. Here, we have developed active, cross-species, small interfering RNAs (siRNAs) capable of targeting murine, rat, nonhuman primate (NHP), and human PCSK9. For in vivo studies, PCSK9 and control siRNAs were formulated in a lipidoid nanoparticle (LNP). Liver-specific siRNA silencing of PCSK9 in mice and rats reduced PCSK9 mRNA levels by 50 -70%. The reduction in PCSK9 transcript was associated with up to a 60% reduction in plasma cholesterol concentrations. These effects were shown to be mediated by an RNAi mechanism, using 5-RACE. In transgenic mice expressing human PCSK9, siRNAs silenced the human PCSK9 transcript by >70% and significantly reduced PCSK9 plasma protein levels. In NHP, a single dose of siRNA targeting PCSK9 resulted in a rapid, durable, and reversible lowering of plasma PCSK9, apolipoprotein B, and LDLc, without measurable effects on either HDL cholesterol (HDLc) or triglycerides (TGs). The effects of PCSK9 silencing lasted for 3 weeks after a single bolus i.v. administration. These results validate PCSK9 targeting with RNAi therapeutics as an approach to specifically lower LDLc, paving the way for the development of PCSK9-lowering agents as a future strategy for treatment of hypercholesterolemia. plasma PCSK9 ͉ tissue LDLR levels P roprotein convertase subtilisin/kexin type 9 (PCSK9) is a member of the mammalian serine proprotein convertase family that typically functions in the proteolytic processing and maturation of secretory proteins (1, 2). PCSK9 was the first family member to be implicated in a dominantly inherited form of hypercholesterolemia (3). Mechanistic studies addressing the function of PCSK9 in mice and humans have demonstrated that overexpression or gain-of-function mutations in PCSK9 reduced low density lipoprotein receptor (LDLR) protein levels in liver, which significantly increased circulating plasma cholesterol both in mice and humans (4). Additional studies showed that the deletion of Pcsk9 in mice resulted in increased LDLR levels, accelerated the clearance of low density lipoprotein cholesterol (LDLc), and reduced circulating cholesterol levels (5). Recently, studies in mice have also shown that lowering PCSK9 transcript levels by antisense oligonucleotides resulted in reduced total cholesterol, LDLc, and HDL cholesterol (HDLc) in blood and increased LDLR levels in liver after 6 weeks of treatment (6). This effect was very similar to that observed in the Pcsk9 Ϫ/Ϫ mice (5). Collectively, these studies have clearly established a role for PCSK9 in cholesterol homeostasis.Validation of PCSK9 as an attractive therapeutic target for the treatment of hypercholesterolemia has come from genetic studies in humans. Cohen et al. (7) first identified loss-of-function mutations in PCSK9...

The growing water stress both in terms of water scarcity and quality deterioration promotes the development of reclaimed water as a new water resource use. This paper reviewed wastewater reuse practices in China, and the opportunities and challenges of expanding reclaimed water use were analyzed. Rapid urbanization with the increasing of water demand and wastewater discharge provides an opportunity for wastewater reuse. The vast amount of wastewater discharge and low reclaimed water production mean that wastewater reuse still has a great potential in China. Many environmental and economic benefits and successful reclamation technologies also provide opportunities for wastewater reuse. In addition, the overall strategy in China is also encouraging for wastewater reuse. In the beginning stage of wastewater reclamation and reuse, there are many significant challenges to expand wastewater reuse in China including slow pace in adopting urban wastewater reuse programs, the establishment of integrated water resources management framework and guidelines for wastewater reuse programs, incoherent water quality requirements, the limited commercial development of reclaimed water and the strengthening of public awareness and cooperation among stakeholders.

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