title: Development of quantitative molecular clinical end points for siRNA clinical trials.
abstract: RNA interference (RNAi) is an evolutionarily conserved mechanism that results in specific gene inhibition at the mRNA level. The discovery that short interfering RNAs (siRNAs) are selective, potent, and can largely avoid immune surveillance has resulted in keen interest to develop these inhibitors as therapeutics. A single nucleotide-specific siRNA (K6a_513a.12, also known as TD101) was recently evaluated in a phase 1b clinical trial for the rare skin disorder, pachyonychia congenita (PC). To develop a clinical trial molecular end point for this type of trial, methods were developed to: (1) isolate total RNA containing amplifiable mRNA from human skin and callus material; (2) quantitatively distinguish the single-nucleotide mutant mRNA from wild-type K6a mRNA in both patient-derived keratinocytes and patient callus; and (3) demonstrate that repeated siRNA treatment results in sustained inhibition of mutant K6a mRNA in patient-derived keratinocyte cultures. These methods allow noninvasive sampling and monitoring of gene expression from patient-collected shavings and may be useful in evaluating the effectiveness of RNAi-based therapeutics, including inhibitors that specifically target single-nucleotide mutations.
authors.affiliation: TransDerm, Santa Cruz, California 95060, USA.
authors.forename: Robyn P
authors.initials: RP
authors.lastname: Hickerson
authors.forename: Sancy A
authors.initials: SA
authors.lastname: Leachman
authors.forename: Lana N
authors.initials: LN
authors.lastname: Pho
authors.forename: Emilio
authors.initials: E
authors.lastname: Gonzalez-Gonzalez
authors.forename: Frances J D
authors.initials: FJ
authors.lastname: Smith
authors.forename: W H Irwin
authors.initials: WH
authors.lastname: McLean
authors.forename: Christopher H
authors.initials: CH
authors.lastname: Contag
authors.forename: Devin
authors.initials: D
authors.lastname: Leake
authors.forename: Leonard M
authors.initials: LM
authors.lastname: Milstone
authors.forename: Roger L
authors.initials: RL
authors.lastname: Kaspar
mesh_terms: D002146:Bony Callus / Q000737:chemistry; D002478:Cells, Cultured; D002986:Clinical Trials as Topic; D006801:Humans; D053553:Keratin-6 / Q000235:genetics*; D015603:Keratinocytes / Q000737:chemistry; D053549:Pachyonychia Congenita / Q000628:therapy*; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D034622:RNA Interference; D012333:RNA, Messenger / Q000302:isolation & purification; D034741:RNA, Small Interfering / Q000235:genetics / Q000627:therapeutic use*; D012867:Skin / Q000737:chemistry
chemical_list: D053553:Keratin-6; D012333:RNA, Messenger; D034741:RNA, Small Interfering