title: Synthesis, gene silencing, and molecular modeling studies of 4'-C-aminomethyl-2'-O-methyl modified small interfering RNAs.
abstract: The linear syntheses of 4'-C-aminomethyl-2'-O-methyl uridine and cytidine nucleoside phosphoramidites were achieved using glucose as the starting material. The modified RNA building blocks were incorporated into small interfering RNAs (siRNAs) by employing solid phase RNA synthesis. Thermal melting studies showed that the modified siRNA duplexes exhibited slightly lower T(m) (∼1 °C/modification) compared to the unmodified duplex. Molecular dynamics simulations revealed that the 4'-C-aminomethyl-2'-O-methyl modified nucleotides adopt South-type conformation in a siRNA duplex, thereby altering the stacking and hydrogen-bonding interactions. These modified siRNAs were also evaluated for their gene silencing efficiency in HeLa cells using a luciferase-based reporter assay. The results indicate that the modifications are well tolerated in various positions of the passenger strand and at the 3' end of the guide strand but are less tolerated in the seed region of the guide strand. The modified siRNAs exhibited prolonged stability in human serum compared to unmodified siRNA. This work has implications for the use of 4'-C-aminomethyl-2'-O-methyl modified nucleotides to overcome some of the challenges associated with the therapeutic utilities of siRNAs.
authors.affiliation: Department of Chemistry, Indian Institute of Technology Bombay, Mumbai 400076, India.
authors.forename: Kiran R
authors.initials: KR
authors.lastname: Gore
authors.forename: Ganesh N
authors.initials: GN
authors.lastname: Nawale
authors.forename: S
authors.initials: S
authors.lastname: Harikrishna
authors.forename: Vinita G
authors.initials: VG
authors.lastname: Chittoor
authors.forename: Sushil Kumar
authors.initials: SK
authors.lastname: Pandey
authors.forename: Claudia
authors.initials: C
authors.lastname: Höbartner
authors.forename: Swati
authors.initials: S
authors.lastname: Patankar
authors.forename: P I
authors.initials: PI
authors.lastname: Pradeepkumar
mesh_terms: D001483:Base Sequence; D003562:Cytidine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry; D020868:Gene Silencing; D006801:Humans; D006860:Hydrogen Bonding; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D009690:Nucleic Acid Conformation; D009711:Nucleotides / Q000138:chemical synthesis* / Q000235:genetics*; D034741:RNA, Small Interfering / Q000737:chemistry* / Q000235:genetics*; D014529:Uridine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry
chemical_list: C574408:4'-C-aminomethyl-2'-O-methyl cytidine; C574407:4'-C-aminomethyl-2'-O-methyl uridine; D009711:Nucleotides; D034741:RNA, Small Interfering; D003562:Cytidine; D014529:Uridine