Long-circulating siRNA nanoparticles for validating Prohibitin1-targeted non-small cell lung cancer treatment

Abstract

RNA interference (RNAi) represents a promising strategy for identification and validation of putative therapeutic targets and for treatment of a myriad of important human diseases including cancer. However, the effective systemic in vivo delivery of small interfering RNA (siRNA) to tumors remains a formidable challenge. Using a robust self-assembly strategy, we develop a unique nanoparticle (NP) platform composed of a solid polymer/cationic lipid hybrid core and a lipid-poly(ethylene glycol) (lipid-PEG) shell for systemic siRNA delivery. The new generation lipid-polymer hybrid NPs are small and uniform, and can efficiently encapsulate siRNA and control its sustained release. They exhibit long blood circulation (t 1/2 ∼8 h), high tumor accumulation, effective gene silencing, and negligible in vivo side effects. With this RNAi NP, we delineate and validate the therapeutic role of Prohibitin1 (PHB1), a target protein that has not been systemically evaluated in vivo due to the lack of specific and effective inhibitors, in treating non-small cell lung cancer (NSCLC) as evidenced by the drastic inhibition of tumor growth upon PHB1 silencing. Human tissue microarray analysis also reveals that high PHB1 tumor expression is associated with poorer overall survival in patients with NSCLC, further suggesting PHB1 as a therapeutic target. We expect this long-circulating RNAi NP platform to be of high interest for validating potential cancer targets in vivo and for the development of new cancer therapies.siRNA delivery | nanoparticle | Prohibitin1 | non-small cell lung cancer W ith the capability to silence any gene of interest, RNA interference (RNAi) technology has demonstrated enormous potential in medical research and applications (1, 2). RNAi-mediated gene silencing has revealed the functionality of specific genetic alterations in cancers (3-5). Many of these genes and pathways are considered "undruggable" targets or require complex and time-consuming development of effective inhibitors. The ubiquitous application of RNAi in cancer research and therapy is nevertheless hindered by the challenge of effective systemic in vivo delivery of siRNA to tumors, which requires overcoming of multiple physiological barriers, such as enzymatic degradation, rapid elimination by renal excretion or by the mononuclear phagocyte system (MPS), and poor cellular uptake and endosomal escape (2, 6, 7). To this end, a great number of cationic lipid/polymer-based nanoparticles (NPs) have been developed to protect siRNA from serum nucleases and facilitate its cytosolic delivery (8). Surface PEGylation has also been applied extensively to improve NP stability and reduce MPS recognition (9, 10). Several RNAi nanotherapeutics are now in clinical trial in cancer patients. However, the clinical stage anticancer RNAi NPs have shown relatively rapid clearance in blood (11,12), which may reduce their extravasation into tumor tissue through the enhanced permeability and retention (EPR) effect (13). This could result in decreased in vivo silencing efficacy...

Lipophilic siRNA targets albumin in situ and promotes bioavailability, tumor penetration, and carrier-free gene silencing

“…The lower tumor:liver ratio observed with in vivo jetPEI and nanoparticle systems in the literature is consistent with reported challenges in achieving efficient nanoparticle delivery to tumor sites; in a comprehensive analysis of nanoparticle delivery to solid tumors, the median injected dose delivered to the tumor site was 0.7% (47). It is notable that even in recent, advanced, and promising nanoparticle systems, including those that use modifications for "stealth" or targeting mechanisms, the ratio of tumor:liver accumulation is consistently close to or below 1:1 (3,5,(57)(58)(59)(60)(61)(62). The marked improvement of siRNA-L 2 in relative tumor accumulation supports its translational promise.…”

Theranostic near-infrared fluorescent nanoplatform for imaging and systemic siRNA delivery to metastatic anaplastic thyroid cancer

“…1A). Because the high hydrophobicity of polyPCPDTBT also enables effective hydrophobic interaction with the amphiphilic cationic lipids that are widely used for complexing with siRNA, the NIR RNAi NPs can be formulated through a modified selfassembly nanoprecipitation method that we recently developed (15), with an siRNA encapsulation efficiency of ∼50%. With a surface coating of 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)], the resulting NPs were highly dispersed in the aqueous solution and remained stable within 1 month.…”

Recent progress in development of siRNA delivery vehicles for cancer therapy

“…10 using polymeric micelles, inorganic nanoparticles, and lipid nanoparticles [31][32][33]. Of importance in this regard is that the tumor accumulation behavior of nanoparticles is significantly affected by the pathophysiology of tumor tissues [34].…”

“…ACCEPTED MANUSCRIPT 33 The length of spacer between ligand and a nanoparticle surface affects the binding chances due to flexibility of the spacer. Antibody-installed nanoparticles equipped with PEG 2000 or PEG 3000 exhibited greater binding to cultured dendritic cells, compared to those with PEG 6000 , PEG 10000 , or PEG 20000 [160].…”