Preclinical and clinical development of siRNA-based therapeutics.

The discovery of RNA interference, first in plants and Caenorhabditis elegans and later in mammalian cells, led to the emergence of a transformative view in biomedical research. Knowledge of the multiple actions of non-coding RNAs has truly allowed viewing DNA, RNA and proteins in novel ways. Small interfering RNAs (siRNAs) can be used as tools to study single gene function both in vitro and in vivo and are an attractive new class of therapeutics, especially against undruggable targets for the treatment of cancer and other diseases. Despite the potential of siRNAs in cancer therapy, many challenges remain, including rapid degradation, poor cellular uptake and off-target effects. Rational design strategies, selection algorithms, chemical modifications and nanocarriers offer significant opportunities to overcome these challenges. Here, we review the development of siRNAs as therapeutic agents from early design to clinical trial, with special emphasis on the development of EphA2-targeting siRNAs for ovarian cancer treatment.

[1]  Jim Euchner.  Design , 2014, Catalysis from A to Z.

[2]  A. Flenniken,et al.  Distinct and overlapping expression patterns of ligands for Eph-related receptor tyrosine kinases during mouse embryogenesis. , 1996, Developmental biology.

[3]  J. Burnett,et al.  Nanoparticle-Based Delivery of RNAi Therapeutics: Progress and Challenges , 2013, Pharmaceuticals.

[4]  J. Au,et al.  Delivery of siRNA Therapeutics: Barriers and Carriers , 2010, The AAPS Journal.

[5]  D. Bostwick,et al.  Overexpression of the EphA2 tyrosine kinase in prostate cancer , 1999, The Prostate.

[6]  M. Kinch,et al.  Activation of the EphA2 tyrosine kinase stimulates the MAP/ERK kinase signaling cascade , 2002, Oncogene.

[7]  A. Sood,et al.  EphA2 Expression Is Associated with Aggressive Features in Ovarian Carcinoma , 2004, Clinical Cancer Research.

[8]  M. Sioud.  Does the understanding of immune activation by RNA predict the design of safe siRNAs? , 2008, Frontiers in bioscience : a journal and virtual library.

[9]  A. Sood,et al.  Targeting melanoma growth and metastasis with systemic delivery of liposome-incorporated protease-activated receptor-1 small interfering RNA. , 2008, Cancer research.

[10]  L. Cekaite,et al.  Suppression of immunostimulatory siRNA-driven innate immune activation by 2'-modified RNAs. , 2007, Biochemical and biophysical research communications.

[11]  A. Sood,et al.  EphA2 as a target for ovarian cancer therapy , 2005, Expert opinion on therapeutic targets.

[12]  Liz Y. Han,et al.  EphA2 overexpression is associated with angiogenesis in ovarian cancer , 2007, Cancer.

[13]  Andrew D. Miller,et al.  Lipidic carriers of siRNA: differences in the formulation, cellular uptake, and delivery with plasmid DNA. , 2004, Biochemistry.

[14]  Yang Wang,et al.  Antitumor activity of poly(ethylene glycol)-camptothecin conjugate: the inhibition of tumor growth in vivo. , 2005, Journal of controlled release : official journal of the Controlled Release Society.

[15]  N. Dubrawsky.  Cancer statistics , 1989, CA: a cancer journal for clinicians.

[16]  M. Kinch,et al.  EphA2 overexpression causes tumorigenesis of mammary epithelial cells. , 2001, Cancer research.

[17]  J. Nesland,et al.  The clinical significance of EphA2 and Ephrin A-1 in epithelial ovarian carcinomas. , 2005, Gynecologic oncology.

[18]  Qi Zhou,et al.  Chemically Modified Short Interfering Hybrids (siHYBRIDS): Nanoimmunoliposome Delivery In Vitro and In Vivo for RNAi of HER-2 , 2006, Nucleosides, nucleotides & nucleic acids.

[19]  J. Kjems,et al.  Improved silencing properties using small internally segmented interfering RNAs , 2007, Nucleic acids research.

[20]  A. Sood,et al.  Therapeutic Silencing of Bcl-2 by Systemically Administered siRNA Nanotherapeutics Inhibits Tumor Growth by Autophagy and Apoptosis and Enhances the Efficacy of Chemotherapy in Orthotopic Xenograft Models of ER (−) and ER (+) Breast Cancer , 2013, Molecular therapy. Nucleic acids.

[21]  A. Deisseroth,et al.  Liposomal delivery of oligodeoxynucleotides. , 1996, Leukemia & lymphoma.

[22]  Shubiao Zhang,et al.  Toxicity of cationic lipids and cationic polymers in gene delivery. , 2006, Journal of controlled release : official journal of the Controlled Release Society.

[23]  C. Echeverri,et al.  LNA incorporated siRNAs exhibit lower off-target effects compared to 2'-OMethoxy in cell phenotypic assays and microarray analysis. , 2008, Nucleic acids symposium series.

[24]  Howard Y. Chang,et al.  Long Noncoding RNAs: Cellular Address Codes in Development and Disease , 2013, Cell.

[25]  A. Sood,et al.  Nanomedicine based approaches for the delivery of siRNA in cancer , 2010, Journal of internal medicine.

[26]  Bernd Jagla,et al.  Sequence characteristics of functional siRNAs. , 2005, RNA.

[27]  D. Bennett,et al.  Protein tyrosine kinases in malignant melanoma. , 2000, Melanoma research.

[28]  Liz Y. Han,et al.  Effect of interleukin-8 gene silencing with liposome-encapsulated small interfering RNA on ovarian cancer cell growth. , 2008, Journal of the National Cancer Institute.

[29]  Bulent Ozpolat,et al.  Liposomal siRNA nanocarriers for cancer therapy. , 2014, Advanced drug delivery reviews.

[30]  A. Jemal,et al.  Cancer statistics, 2014 , 2014, CA: a cancer journal for clinicians.

[31]  M. Hendrix,et al.  Molecular regulation of tumor cell vasculogenic mimicry by tyrosine phosphorylation: role of epithelial cell kinase (Eck/EphA2). , 2001, Cancer research.

[32]  Judy Lieberman,et al.  Interfering with disease: a progress report on siRNA-based therapeutics , 2007, Nature Reviews Drug Discovery.

[33]  Phillip A Sharp,et al.  siRNAs can function as miRNAs , 2003 .

[34]  M. Andreeff,et al.  Cellular uptake and localization of liposomal-methylphosphonate oligodeoxynucleotides , 1996, Journal of Molecular Medicine.

[35]  Ming Wu,et al.  Designing highly active siRNAs for therapeutic applications , 2010, The FEBS journal.

[36]  A. Sood,et al.  Highly Specific Targeting of the TMPRSS2/ERG Fusion Gene Using Liposomal Nanovectors , 2012, Clinical Cancer Research.

[37]  Matthias John,et al.  Therapeutic silencing of an endogenous gene by systemic administration of modified siRNAs , 2004, Nature.

[38]  A. Klippel,et al.  Structural variations and stabilising modifications of synthetic siRNAs in mammalian cells. , 2003, Nucleic acids research.

[39]  L. Ellis,et al.  Therapeutic targeting of neuropilin-2 on colorectal carcinoma cells implanted in the murine liver. , 2008, Journal of the National Cancer Institute.

[40]  M. Sioud,et al.  Potential design rules and enzymatic synthesis of siRNAs. , 2004, Methods in molecular biology.

[41]  J. Swenberg,et al.  Quantification of 2'-fluoro-2'-deoxyuridine and 2'-fluoro-2'-deoxycytidine in DNA and RNA isolated from rats and woodchucks using LC/MS/MS. , 2002, Chemical research in toxicology.

[42]  Ola Snøve,et al.  Designing effective siRNAs with off-target control. , 2004, Biochemical and biophysical research communications.

[43]  Han-Oh Park,et al.  Chemical modification of siRNAs to improve serum stability without loss of efficacy. , 2006, Biochemical and biophysical research communications.

[44]  Matthias John,et al.  RNAi-mediated gene silencing in non-human primates , 2006, Nature.

[45]  R. Schachar,et al.  Phase 1 dose-escalation study of a siRNA targeting the RTP801 gene in age-related macular degeneration patients , 2012, Eye.

[46]  B. Bettencourt,et al.  Effect of an RNA interference drug on the synthesis of proprotein convertase subtilisin/kexin type 9 (PCSK9) and the concentration of serum LDL cholesterol in healthy volunteers: a randomised, single-blind, placebo-controlled, phase 1 trial , 2014, The Lancet.

[47]  Mark E. Davis,et al.  Effect of siRNA nuclease stability on the in vitro and in vivo kinetics of siRNA‐mediated gene silencing , 2007, Biotechnology and bioengineering.

[48]  A. Fire,et al.  Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans , 1998, Nature.

[49]  M. Zamora,et al.  RNA interference therapy in lung transplant patients infected with respiratory syncytial virus. , 2011, American journal of respiratory and critical care medicine.

[50]  D. Harpole,et al.  Predictive value of the EphA2 receptor tyrosine kinase in lung cancer recurrence and survival. , 2003, Clinical cancer research : an official journal of the American Association for Cancer Research.

[51]  Mark E. Davis,et al.  Correlating animal and human phase Ia/Ib clinical data with CALAA-01, a targeted, polymer-based nanoparticle containing siRNA , 2014, Proceedings of the National Academy of Sciences.

[52]  M. Sioud.  Deciphering the code of innate immunity recognition of siRNAs. , 2009, Methods in molecular biology.

[53]  R. Juliano,et al.  Mechanisms and strategies for effective delivery of antisense and siRNA oligonucleotides , 2008, Nucleic acids research.

[54]  Anil K Sood,et al.  Therapeutic EphA2 gene targeting in vivo using neutral liposomal small interfering RNA delivery. , 2005, Cancer research.

[55]  Qi Zhou,et al.  Materializing the potential of small interfering RNA via a tumor-targeting nanodelivery system. , 2007, Cancer research.

[56]  S. Akira,et al.  Sequence-specific potent induction of IFN-α by short interfering RNA in plasmacytoid dendritic cells through TLR7 , 2005, Nature Medicine.

[57]  Brendan J. Frey,et al.  Multi-way clustering of microarray data using probabilistic sparse matrix factorization , 2005, ISMB.

[58]  M. V. González,et al.  In vitro and in vivo efficacy of SYL040012, a novel siRNA compound for treatment of glaucoma. , 2014, Molecular therapy : the journal of the American Society of Gene Therapy.

[59]  T. Hunter,et al.  cDNA cloning and characterization of eck, an epithelial cell receptor protein-tyrosine kinase in the eph/elk family of protein kinases , 1990, Molecular and cellular biology.

[60]  Prudence W. H. Wong,et al.  Filtering of Ineffective siRNAs and Improved siRNA Design Tool , 2004, APBC.

[61]  E. Chargaff,et al.  Nucleic Acids , 2020, Definitions.

[62]  Gorjan Alagic,et al.  #p , 2019, Quantum information & computation.

[63]  Fran Lewitter,et al.  siRNA Selection Server: an automated siRNA oligonucleotide prediction server , 2004, Nucleic Acids Res..

[64]  Erik L L Sonnhammer,et al.  Improved and automated prediction of effective siRNA. , 2004, Biochemical and biophysical research communications.

[65]  T. Tuschl,et al.  Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells , 2001, Nature.

[66]  M. Zavolan,et al.  Strand-specific 5'-O-methylation of siRNA duplexes controls guide strand selection and targeting specificity. , 2007, RNA.

[67]  George A. Calin,et al.  RNA interference in the clinic: challenges and future directions , 2011, Nature Reviews Cancer.

[68]  John J. Rossi,et al.  Strategies for silencing human disease using RNA interference , 2007, Nature Reviews Genetics.

[69]  M. Hendrix,et al.  Differential regulation of EphA2 in normal and malignant cells. , 2003, The American journal of pathology.

[70]  C. Glass,et al.  Non-coding RNAs as regulators of gene expression and epigenetics. , 2011, Cardiovascular research.

[71]  J. Burnett,et al.  Current progress of siRNA/shRNA therapeutics in clinical trials , 2011, Biotechnology journal.

[72]  Mikiko C. Siomi,et al.  The Discovery of Rna Interference (rnai) Biogenesis of Small Rnas on the Road to Reading the Rna-interference Code Insight Review , 2022 .

[73]  K. Hirokawa,et al.  Overexpression of protein tyrosine kinases in human esophageal cancer. , 1997, Pathobiology : journal of immunopathology, molecular and cellular biology.

[74]  E. Partridge,et al.  Cyclophosphamide and cisplatin compared with paclitaxel and cisplatin in patients with stage III and stage IV ovarian cancer , 1996, New England Journal of Medicine.

[75]  T. Tuschl,et al.  Mechanisms of gene silencing by double-stranded RNA , 2004, Nature.

[76]  M. Stoffel,et al.  Mechanisms and optimization of in vivo delivery of lipophilic siRNAs , 2007, Nature Biotechnology.

[77]  O. Slabý,et al.  Novel classes of non-coding RNAs and cancer , 2012, Journal of Translational Medicine.

[78]  B. Monia,et al.  Antisense oligonucleotides containing locked nucleic acid improve potency but cause significant hepatotoxicity in animals , 2006, Nucleic acids research.

[79]  A. Judge,et al.  Sequence-dependent stimulation of the mammalian innate immune response by synthetic siRNA , 2005, Nature Biotechnology.

[80]  L. Lim,et al.  Widespread siRNA "off-target" transcript silencing mediated by seed region sequence complementarity. , 2006, RNA.

[81]  M. Fromm,et al.  Functionalized silicon quantum dots tailored for targeted siRNA delivery. , 2009, Biochemical and biophysical research communications.

[82]  J. Neijt,et al.  First line chemotherapy with carboplatin plus paclitaxel in advanced ovarian cancer--a new standard of care? , 1999, Annals of oncology : official journal of the European Society for Medical Oncology.

[83]  Robert Langer,et al.  A combinatorial library of lipid-like materials for delivery of RNAi therapeutics , 2008, Nature Biotechnology.

[84]  Shigeru Takasaki,et al.  Methods for selecting effective siRNA sequences by using statistical and clustering techniques. , 2009, Methods in molecular biology.

[85]  Y. Rojanasakul,et al.  Oxygen Radical-Mediated Pulmonary Toxicity Induced by Some Cationic Liposomes , 2000, Pharmaceutical Research.

[86]  B. Bettencourt,et al.  Safety and efficacy of RNAi therapy for transthyretin amyloidosis. , 2013, The New England journal of medicine.

[87]  Mark E. Davis,et al.  Administration in non-human primates of escalating intravenous doses of targeted nanoparticles containing ribonucleotide reductase subunit M2 siRNA , 2007, Proceedings of the National Academy of Sciences.

[88]  V. Vlassov,et al.  Selective protection of nuclease-sensitive sites in siRNA prolongs silencing effect. , 2009, Oligonucleotides.

[89]  Lin He,et al.  MicroRNAs: small RNAs with a big role in gene regulation , 2004, Nature Reviews Genetics.

[90]  Frank Baas,et al.  UvA-DARE ( Digital Academic Repository ) In vivo tumor growth inhibition and biodistribution studies of locked nucleic acid ( LNA ) antisense oligonucleotides , 2017 .

[91]  J. Burnett,et al.  RNA-based therapeutics: current progress and future prospects. , 2012, Chemistry & biology.

[92]  Amy A. Caudy,et al.  Post-transcriptional gene silencing by double-stranded RNA , 2001, Nature Reviews Genetics.

[93]  Julia C. Engelmann,et al.  siPools: highly complex but accurately defined siRNA pools eliminate off-target effects , 2014, Nucleic acids research.

[94]  Ook.,et al.  CYCLOPHOSPHAMIDE AND CISPLATIN COMPARED WITH PACLITAXEL AND CISPLATIN IN PATIENTS WITH STAGE III AND STAGE IV OVARIAN CANCER , 2000 .

[95]  K. Ui-Tei,et al.  Functional dissection of siRNA sequence by systematic DNA substitution: modified siRNA with a DNA seed arm is a powerful tool for mammalian gene silencing with significantly reduced off-target effect , 2008, Nucleic acids research.

[96]  B. Usmani,et al.  Design, manufacture, and assay of the efficacy of siRNAs for gene silencing. , 2008, Methods in molecular biology.

[97]  Liz Y. Han,et al.  Focal Adhesion Kinase Targeting Using In vivo Short Interfering RNA Delivery in Neutral Liposomes for Ovarian Carcinoma Therapy , 2006, Clinical Cancer Research.

[98]  Shan Jiang,et al.  Quantum-dot based nanoparticles for targeted silencing of HER2/neu gene via RNA interference. , 2007, Biomaterials.

[99]  Z. Paroo,et al.  Biodistribution of phosphodiester and phosphorothioate siRNA. , 2004, Bioorganic & medicinal chemistry letters.

[100]  Prahlad T. Ram,et al.  2′-OMe-phosphorodithioate-modified siRNAs show increased loading into the RISC complex and enhanced anti-tumour activity , 2014, Nature Communications.

[101]  P. Linsley,et al.  Recognizing and avoiding siRNA off-target effects for target identification and therapeutic application , 2010, Nature Reviews Drug Discovery.

[102]  R. Hickerson,et al.  First-in-human mutation-targeted siRNA phase Ib trial of an inherited skin disorder. , 2010, Molecular therapy : the journal of the American Society of Gene Therapy.

[103]  M. Behlke.  Chemical modification of siRNAs for in vivo use. , 2008, Oligonucleotides.

[104]  L. Lim,et al.  Position-specific chemical modification of siRNAs reduces "off-target" transcript silencing. , 2006, RNA.

[105]  R. Jorgensen,et al.  Altered gene expression in plants due to trans interactions between homologous genes. , 1990, Trends in biotechnology.

[106]  Ola Snøve,et al.  Many commonly used siRNAs risk off-target activity. , 2004, Biochemical and biophysical research communications.

[107]  P. Kaiser,et al.  RNAi-based treatment for neovascular age-related macular degeneration by Sirna-027. , 2010, American journal of ophthalmology.

[108]  J. Hochman,et al.  Metabolite identification of small interfering RNA duplex by high-resolution accurate mass spectrometry. , 2008, Rapid communications in mass spectrometry : RCM.

[109]  Anastasia Khvorova,et al.  Off-target effects by siRNA can induce toxic phenotype. , 2006, RNA.

[110]  Mark E. Davis,et al.  Evidence of RNAi in humans from systemically administered siRNA via targeted nanoparticles , 2010, Nature.

[111]  Y. Kopelman,et al.  Mutant KRAS is a druggable target for pancreatic cancer , 2013, Proceedings of the National Academy of Sciences.

[112]  K. Ui-Tei,et al.  Designing functional siRNA with reduced off-target effects. , 2013, Methods in molecular biology.

[113]  J. Aronson.  Safety , 2009, BMJ : British Medical Journal.

[114]  G. G. Stokes.  "J." , 1890, The New Yale Book of Quotations.

[115]  Yuhan Lee,et al.  Cationic solid lipid nanoparticles reconstituted from low density lipoprotein components for delivery of siRNA. , 2008, Molecular pharmaceutics.

[116]  Mauro Ferrari,et al.  Enhancing Chemotherapy Response with Sustained EphA2 Silencing Using Multistage Vector Delivery , 2013, Clinical Cancer Research.

[117]  Jeong Won Lee,et al.  Targeted Gene Silencing Using RGD-Labeled Chitosan Nanoparticles , 2010, Clinical Cancer Research.

[118]  Erkki Ruoslahti,et al.  Targeted quantum dot conjugates for siRNA delivery. , 2007, Bioconjugate chemistry.

[119]  J. Belmont,et al.  Liposome-incorporated Grb2 antisense oligodeoxynucleotide increases the survival of mice bearing bcr-abl-positive leukemia xenografts. , 2007, International journal of oncology.

[120]  Jinwoo Cheon,et al.  All-in-one target-cell-specific magnetic nanoparticles for simultaneous molecular imaging and siRNA delivery. , 2009, Angewandte Chemie.

[121]  P. Mann,et al.  An Evaluation of the Toxicities of 2' -Fluorouridine and 2'-Fluorocytidine-HCI in F344 Rats and Woodchucks (Marmota monax) , 1999, Toxicologic pathology.

[122]  K. Giese,et al.  Phase I clinical development of Atu027, a siRNA formulation targeting PKN3 in patients with advanced solid tumors. , 2012, International journal of clinical pharmacology and therapeutics.

[123]  N. Kosaka,et al.  Atelocollagen-mediated synthetic small interfering RNA delivery for effective gene silencing in vitro and in vivo. , 2004, Nucleic acids research.

[124]  Sunit K. Singh,et al.  siRNAs: their potential as therapeutic agents--Part I. Designing of siRNAs. , 2009, Drug discovery today.

[125]  A. Tari,et al.  Safety, pharmacokinetics, and tissue distribution of liposomal P-ethoxy antisense oligonucleotides targeted to Bcl-2. , 1999, The Journal of pharmacology and experimental therapeutics.

[126]  Liz Y. Han,et al.  Efficacy and antivascular effects of EphA2 reduction with an agonistic antibody in ovarian cancer. , 2006, Journal of the National Cancer Institute.

[127]  Xiaohui Wang,et al.  Selection of hyperfunctional siRNAs with improved potency and specificity , 2009, Nucleic acids research.