Science

Platforms that
Accelerate Drug Discovery

RNAi triggers

Overview

Arrowhead has established a robust and versatile drug discovery and development platform based on proprietary technologies, broad licenses to fundamental intellectual property and extensive scientific expertise after more than a decade working on RNAi-based therapies. This platform enables the company to develop effective new therapies rapidly, cost effectively and potentially with lower risk relative to traditional approaches.

RNA Interference

One of the most important recent advances in biology, the discovery of RNA interference, or RNAi, led to a Nobel Prize in 2006. RNAi refers to a natural cellular mechanism whereby short oligonucleotide molecules called RNAi triggers silence gene expression and regulate the production of proteins. Arrowhead’s RNAi-based therapeutics leverage this natural pathway of gene silencing to target and shut down specific genes that cause disease. This mechanism offers many potential advantages in the development of disease therapies, including the ability to target a broad range of genes and proteins with high specificity, and also target disease pathways that have proven difficult to address with traditional small molecule and biologic therapeutics.

This figure depicts the mechanism by which gene silencing occurs. Double stranded RNAi triggers are introduced into a cell and get loaded into the RNA-induced silencing complex, or RISC. The strands are separated leaving an active RISC/RNAi trigger complex. This complex can then pair with and degrade the complementary messenger RNAs, or mRNA, and stop the production of the target proteins. RNAi is a catalytic process, so each RNAi trigger can degrade mRNA hundreds of times, which results in a relatively long duration of effect for RNAi therapeutics.

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Targeted RNAi Molecule (TRiMTM) Platform

Arrowhead’s Targeted RNAi Molecule, or TRiMTM, platform utilizes ligand-mediated delivery and is designed to enable tissue-specific targeting while being structurally simple. Targeting has been core to Arrowhead’s development philosophy and the TRiMTM platform builds on more than a decade of work on actively targeted drug delivery vehicles. Arrowhead scientists have discovered ways to progressively “TRiM” away extraneous features and chemistries and retain optimal pharmacologic activity.

The TRiMTM platform comprises a highly potent RNA trigger with the following components optimized, as needed, for each drug candidate: a high affinity targeting ligand; various linker and chemistries; structures that enhance pharmacokinetics; and highly potent RNAi triggers with sequence specific stabilization chemistries.

Therapeutics developed with the TRiMTM platform offer several advantages: simplified manufacturing and reduced costs; multiple routes of administration; potential for improved safety because there are less metabolites from smaller molecules, thereby reducing the risk of intracellular buildup.

At Arrowhead, we also believe that for RNAi to reach its true potential, it must target organs outside the liver. Arrowhead is leading this expansion with the TRiMTM platform that holds the promise of reaching multiple tissues, including liver, lung, and tumor.

RNA Chemistries

The structure and chemistries of the oligonucleotide molecules used to trigger the RNAi mechanism can be tailored for optimal activity. Arrowhead’s broad portfolio of RNA trigger structures and chemistries, including some proprietary structures, enable the company to optimize each drug candidate on a target-by-target basis and utilize the combination of structure and chemical modifications that yield the most potent RNAi trigger.

As a component of the TRiM platform, Arrowhead’s design philosophy on RNA chemical modifications is to start with a structurally simple molecule and only add selective modification and stabilization chemistries as necessary to achieve the desired level of target knockdown and duration of effect. The conceptual framework for the stabilization strategy starts with a more sophisticated RNAi trigger screening and selection process that identifies potent sequences rapidly in locations that others may miss. We pursue chemical stabilization strategies with a target duration of effect of 30-90 days, and typically limit the use of strategies that produce longer activity because we anticipate that such strategies will increase long-term safety risks.

Intellectual Property

Our intellectual property portfolio provides broad freedom to apply multiple RNAi delivery technologies, chemistries, structures and manufacturing techniques in the development of novel therapeutics. Access to a broad range of technologies allows us to choose the best approach for a wide range of gene targets and disease indications.

Publications

2015

Sebestyén MG, Wong SC, Trubetskoy V, Lewis DL, Wooddell CI (2015) “Targeted in vivo delivery of siRNA and an endosome-releasing agent to hepatocytes.” Chapter in RNA Interference: Challenges and Therapeutic Opportunities, Mouldy Sioud (ed.), Methods in Molecular Biology vol. 1218.

Gish RG, Yuen MF, Chan HLY, Given BD, Lai CL, Locarnini SA, Lau JYN, Wooddell CI, Schluep T, Lewis DL (2015) “Synthetic RNAi triggers and their use in chronic hepatitis B therapies with curative intent.” Antiviral Research 2015 Sep;121:97-108.

Gish RG, Given BD, Lai CL, Locarnini SA, Lau JY, Lewis DL, Schluep T. (2015) “Chronic hepatitis B: Virology, natural history, current management and a glimpse at future opportunities.” Antiviral Research 2015 Sep;121:47-58.

Rozema DB, Blokhin AV, Wakefield DH, Benson JD, Carlson JC, Klein JJ, Almeida LJ, Nicholas AL, Hamilton HL, Chu Q, Hegge JO, Wong SC, Trubetskoy VS, Hagen CM, Kitas E, Wolff JA, Lewis DL. “Protease-triggered siRNA delivery vehicles.” Journal of Controlled Release. 14 April 2015; doi:10.1016/j.jconrel.2015.04.012.

2013

Wooddell CI, Rozema DB, Hossbach M, John M, Hamilton HL, Chu Q, Hegge JO, Klein JJ, Wakefield DH, Oropeza CE, Deckert J, Roehl I, Jahn-Hofmann K, Hadwiger P, Vornlocher HP, McLachlan A, Lewis DL, “Hepatocyte-targeted RNAi Therapeutics for the Treatment of Chronic Hepatitis B Virus Infection.” Molecular Therapy. 26 February 2013; doi: 10.1038/mt.2013.31.

2012

Schneider B, Grote M, John M, Haas A, Bramlage B, Ickenstein LM, Jahn-Hofmann K, Bauss F, Cheng W, Croasdale R, Daub K, Dill S, Hoffmann E, Lau W, Burtscher H, Ludtke JL, Metz S, Mundigl O, Neal ZC, Scheuer W, Stracke J, Herweijer H, Brinkmann U. “Targeted siRNA Delivery and mRNA Knockdown Mediated by Bispecific Digoxigenin-binding Antibodies.” Molecular Therapy Nucleic Acids. 18 September 2012; 1, e45; doi:10.1038/mtna.2012.39

Wong SC, Klein JJ, Hamilton HL, Chu Q, Frey CL, Trubetskoy VS, Hegge J, Wakefield D, Rozema DB, Lewis DL. “Co-Injection of a Targeted, Reversibly Masked Endosomolytic Polymer Dramatically Improves the Efficacy of Cholesterol-Conjugated Small Interfering RNAs In Vivo.” Nucleic Acid Therapeutics. December 2012; 22(6): 380-390. doi:10.1089/nat.2012.0389.

2011

Wooddell CI, Hegge JO, Zhang G, Sebestyén MG, Noble M, Griffin JB, Pfannes LV, Herweijer H, Hagstrom JE, Braun S, Huss T, Wolff JA “Dose response in rodents and nonhuman primates after hydrodynamic limb vein delivery of naked plasmid DNA”. Human Gene Therapy 2011 July; 22(7):889-903. doi: 10.1089/hum.2010.160. Epub 2011 Apr 18.

Wooddell CI, Subbotin VM, Sebestyén MG, Griffin JB, Zhang G, Schleef M, Braun S, Huss T, Wolff JA “Muscle damage after delivery of naked plasmid DNA into skeletal muscles is batch dependent”. Human Gene Therapy February 2011; 22:225-235. DOI: 10.1089/hum.2010.113

Wooddell CI, Radley-Crabb HG, Griffin JB, Zhang G “Myofiber damage evaluation by evans blue dye injection”. Current Protocols in Mouse Biology 1: 1-26, December 2011. DOI: 10.1002/9780470942390.mo110141

Pichavant C, Aartsma-Rus A, Clemens PR, Davies KE, Dickson G, Takeda S, Wilton SD, Wolff JA, Wooddell CI, Xiao X, Tremblay JP “Current status of pharmaceutical and genetic therapeutic approaches to treat DMD”. Molecular Therapy 2011 May; 19(5): 830–840.  doi:10.1038/mt.2011.59

2010

Zhang G, Wooddell CI, Hegge JO, Griffin JB, Huss T, Braun S, Wolff JA “Functional efficacy of dystrophin expression from plasmids delivered to mdx mice by hydrodynamic limb vein injection”. Human Gene Therapy February 2010; 21:221-237. DOI: 10.1089/hum.2009.133

Wooddell CI, Zhang G, Griffin JB, Hegge JO, Huss T, Wolff JA “Use of evans blue dye to compare limb muscles in exercised young and old mdx mice”.  Muscle and Nerve, April 2010. DOI 10.1002/mus.21527

Hegge JO, Wooddell CI, Zhang G, Hagstrom JE, Braun S, Huss T, Sebestyén MG, Emborg ME, Wolff JA “Evaluation of hydrodynamic limb vein injections in nonhuman primates”. Human Gene Therapy July 2010; 21:829-842. DOI: 10.1089/hum.2009.172

Mudd SR, Trubetskoy VS, Blokhin AV, Weichert JP, Wolff JA “Hybrid PET/CT for noninvasive pharmacokinetic evaluation of dynamic PolyConjugates, a synthetic siRNA delivery system.” Bioconjug Chem. 2010 Jul 21;21(7):1183-9. doi: 10.1021/bc900558z.

2007

Vigen KK, Hegge JO, Zhang G, Mukherjee R, Braun S, Grist TM, Wolff JA “Magnetic resonance imaging-monitored plasmid DNA delivery in primate limb muscle”.Human Gene Therapy March 2007; 18:257-268. DOI: 10.1089/hum.2006.115

Lewis, D. L., Wolff, J. A. (2007) “Systemic siRNA delivery via hydrodynamic intravascular injection.” Advanced Drug Delivery Reviews, 59, 115-123.

Sebestyen, M. G., Hegge, J. O., Noble, M. A., Lewis, D. L., Herweijer, H.,Wolff, J. A. (2007) “Progress toward a nonviral gene therapy protocol for the treatment of anemia.” Human Gene Therapy, 18, 269-85.

Dai, X., De Souza, A.T., Dai, H., Lewis, D. L., Lee, C., Spencer, A. G., Herweijer, H., Hagstrom, J. E., Linsley, P. S., Bassett, D. E., Ulrich, R. G.,  He, Y. D. (2007) “PPARα siRNA–Treated Expression Profiles Uncover the Causal Sufficiency Network for Compound-Induced Liver Hypertrophy.” PLoS Computational Biology, 3 (3).

Rozema DB, Lewis DL, Wakefield DH, Wong SC, Klein JJ, Roesch PL, Bertin SL, Reppen TW, Chu Q, Blokhin AV, Hagstrom JE, Wolff JA. “Dynamic PolyConjugates for targeted in vivo delivery of siRNA to hepatocytes.” Proceedings of the National Academy of Sciences. 104(32): 12982-87.

2006

De Souza, A. T., Dai, X., Spencer, A. G., Reppen, T., Menzie, A., Roesch, P. L., He, Y., Caguyong, M. J., Bloomer, S., Herweijer, H., Wolff, J. A., Hagstrom, J. E., Lewis, D. L., Linsley, P. S., Ulrich, R. G. (2006) “Transcriptional and phenotypic comparisons of Ppara knockout and siRNA knockdown mice.” Nucleic Acids Research, 34, 4486-4494.

Wong S.  C., Wakefield D., Klein J., Monahan S. D., Rozema D. B., Lewis D. L., Higgs L., Ludtke J., Sokoloff A. V., Wolff J. A. (2006) “Hepatocyte Targeting of Nucleic Acid Complexes and Liposomes by a T7 Phage p17 Peptide.” Molecular Pharmacology, 3, 386-397.

Sebestyen, M. G., Budker, V. G., Budker, T., Subbotin, V. M., Zhang, G., Monahan, S. D., Lewis, D. L., Wong, S. C., Hagstrom,  J. E., Wolff, J.A. (2006) “Mechanism of plasmid delivery by hydrodynamic tail vein injection. I. Hepatocyte uptake of various molecules.” Journal of Gene Medicine, 8, 852-873.

2005

Lewis, D. L. and Wolff, J. A. (2005) “Delivery of siRNA and siRNA Expression Constructs to Adult Mammals Using Hydrodynamic Intravascular Injection.” In Methods in Enzymology, 392, RNA Interference, Engelke, D. R. and Rossi, J. (Eds.). Elsevier/Academic Press, San Diego, CA.

Wolff, J., Lewis, D. L., Herweijer, H., Hegge, J., Hagstrom, J. (2005) “Non-viral approaches for gene transfer.” Acta Myol., 24, 202-8.

Wooddell, C. I., Van Hout, C. V., Reppen, T., Lewis, D. L., Herweijer, H. (2005) “Long-term RNA interference from optimized siRNA expression constructs in adult mice. Biochemical and Biophysical Research Communications,” 334, 117–127.

2004

Hagstrom, J. E., Hegge, J., Zhang, G., Noble, M., Budker, V., Lewis, D. L., Herweijer, H., and Wolff, J. A. (2004) “A facile method for delivering genes and siRNAs to skeletal muscle of mammalian limbs.” Molecular Therapy, 10, 386-398.

2003

Rozema, D. B., and Lewis, D. L., (2003) “siRNA delivery technologies for mammalian systems.” Targets, 2, 253-260.

Rozema, D. B., Ekena, K., Lewis, D. L., Loomis, A. G., and Wolff, J. A.  (2003) “Endosomolysis by masking of a membrane-active agent (EMMA) for cytoplasmic release of macromolecules.” Bioconjugate Chemistry, 14, 51-57.

2002

Lewis, D. L., Hagstrom, J. E., Loomis, A. G., Wolff, J. A. and Herweijer, H. (2002) “Efficient delivery of siRNA for inhibition of gene expression in postnatal mice.” Nature Genetics 32, 107-108.