TY - JOUR
T1 - Development of quantitative molecular clinical end points for siRNA clinical trials
AU - Hickerson, Robyn P.
AU - Leachman, Sancy A.
AU - Pho, Lana N.
AU - Gonzalez-Gonzalez, Emilio
AU - Smith, Frances J.D.
AU - McLean, W. H.Irwin
AU - Contag, Christopher H.
AU - Leake, Devin
AU - Milstone, Leonard M.
AU - Kaspar, Roger L.
N1 - Funding Information:
We thank Manuel Flores, Heini Ilves, Sally Tran, Travis Canova, Robert Kaspar, and Rebecca Kaspar for technical assistance. We thank Jing Zhou, Daniel DiMaio, and Anne Edwards for assistance and advice regarding immortalization of keratinocytes. We thank Alexander Mankin, Zigurts Majumdar, and Joseph Carroll for critical comments and suggestions. This work was supported by grants from the PC Project (to SAL, LMM, and FJDS), Huntsman Cancer Foundation (to SAL), Food and Drug Administration (FDA OOPD R01-FD-003553-01, to SAL), National Institutes of Health (1R43AR056559-01, to RLK), and Medical Research Council (to WHIM and FJDS). EGG is the recipient of a PC Project fellowship. We are grateful for the generous support, collegiality, and useful suggestions from members of the PC Project and the International PC Consortium (IPCC). We also thank Tracey Lewis from ARUP Laboratories for assistance with QRT-PCR assay validation. Most of all, we thank the PC patients who have offered their unwavering support, including providing skin biopsies and tissue samples that have made this study possible.
PY - 2011/5
Y1 - 2011/5
N2 - 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.
AB - 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.
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U2 - 10.1038/jid.2010.372
DO - 10.1038/jid.2010.372
M3 - Article
C2 - 21191405
AN - SCOPUS:79954616664
SN - 0022-202X
VL - 131
SP - 1029
EP - 1036
JO - Journal of Investigative Dermatology
JF - Journal of Investigative Dermatology
IS - 5
ER -