title: The efficacy of nuclease-resistant Chol-siRNA in primary breast tumors following complexation with PLL-PEG(5K).
abstract: Modifying the sense strand of nuclease-resistant siRNA with 3'-cholesterol (Chol-*siRNA) increases mRNA suppression after i.v. administration but with relatively low efficacy. We previously found evidence in vitro that suggests complexation of Chol-siRNA with PLL-PEG(5K), a block copolymer of poly-l-lysine and 5 kDa polyethylene glycol, may increase the efficacy of Chol-siRNA in vivo in a PLL block length-dependent manner. In this study, the extent that polyplexes of PLL10-PEG(5K), PLL30-PEG(5K), and PLL50-PEG(5K) protect complexed Chol-siRNA in high concentrations of murine serum and affect the activity of Chol-*siRNA in murine 4T1 breast tumor epithelial cells in vitro and in primary orthotopic tumors of 4T1 was compared. PLL-PEG(5K) required 3'-Chol to protect full-length siRNA from nuclease degradation in 90% (v/v) murine serum and protection was increased by increasing PLL block length and nuclease resistance of Chol-siRNA. Polyplexes of Chol-*siLuc suppressed stably expressed luciferase in 4T1-Luc cells to different levels in vitro where PLL30 > PLL50 > PLL10. In contrast, only polyplexes of Chol-*siLuc and PLL30-PEG(5K) or PLL50-PEG(5K) suppressed high levels of luciferase in primary orthotopic tumors of 4T1-Luc after i.v. administration, whereas polyplexes of Chol-*siLuc and PLL10-PEG(5K), inactive Chol-*siCtrl polyplexes of PLL-PEG(5K), or Chol-*siLuc alone had no detectable activity. As a whole, these results indicate that polyplexes of PLL-PEG(5K) increase the efficacy of nuclease-resistant Chol-siRNA in primary breast tumors after i.v. administration in a PLL block length-dependent manner. Thus, complexation of Chol-siRNA with PLL-PEG(5K) may be a promising approach to increase the efficacy of Chol-siRNA in a wide range of primary tumors, metastases, and other tissues but likely requires a PLL block length that balances polymer-related adverse effects, Chol-siRNA bioavailability, and subsequent activity in the target cell.
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Nebraska Medical Center, 986025 Nebraska Medical Center, Omaha, NE 68198-6025, USA.
authors.forename: Vishakha V
authors.initials: VV
authors.lastname: Ambardekar
authors.forename: Rajesh R
authors.initials: RR
authors.lastname: Wakaskar
authors.forename: Bhawna
authors.initials: B
authors.lastname: Sharma
authors.forename: Joy
authors.initials: J
authors.lastname: Bowman
authors.forename: Willy
authors.initials: W
authors.lastname: Vayaboury
authors.forename: Rakesh K
authors.initials: RK
authors.lastname: Singh
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Vetro
mesh_terms: D000818:Animals; D001835:Body Weight / Q000187:drug effects; D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D002784:Cholesterol / Q000737:chemistry*; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D005260:Female; D006801:Humans; D057446:Hydrodynamics; D008156:Luciferases / Q000378:metabolism; D015674:Mammary Neoplasms, Animal / Q000378:metabolism* / Q000473:pathology*; D051379:Mice; D011092:Polyethylene Glycols / Q000737:chemistry*; D011107:Polylysine / Q000737:chemistry*; D034741:RNA, Small Interfering / Q000097:blood / Q000378:metabolism*; D012260:Ribonucleases / Q000378:metabolism*; D047368:Tumor Burden / Q000187:drug effects
chemical_list: D034741:RNA, Small Interfering; D011107:Polylysine; D011092:Polyethylene Glycols; D002784:Cholesterol; D008156:Luciferases; D012260:Ribonucleases
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