{"id":{"repo_id":"utswmed","oai_identifier":"oai:utswmed-ir.tdl.org:2152.5/3332"},"canonical_url":"https://search.dev.ndltd.org/etd/utswmed/oai:utswmed-ir.tdl.org:2152.5/3332","repository":{"repo_id":"utswmed","name":"University of Texas Southwestern Medical Center","base_url":"https://utswmed-ir.tdl.org/server/oai/request"},"display":{"title":"Using Chemically Modified Oligonucleotides to Modulate Gene Expression, Treat Genetic Diseases, and Probe Novel Mechanisms of RNA Interference","abstract":"A number of inherited neurological disorders remain incurable despite having well-defined monogenic etiologies. One example is Huntington's disease (HD), which is caused by CAG trinucleotide expansion in the gene HUNTINGTIN (HTT) and production of toxic glutamine-expanded protein. Targeting HTT with siRNAs could be a powerful approach, but allele-selectivity is a major challenge: nearly all HD patients are heterozygous at the HTT locus, and expression of wild-type HTT may need to be preserved. One way to achieve allele-selectivity is by exploiting the fact that the mutant HTT allele contains more CAG repeats. Previous work with double-stranded siRNAs (dsRNA) and chemically modified antisense oligonucleotides (ASO) that target the poly-CAG sequence both showed promise but each had significant limitations. To combine the simplicity of ASO and high selectivity of dsRNA, we tested chemically modified, single-stranded small-interfering RNA (ss-siRNA) of sequences targeting CAG repeats in collaboration with ISIS Pharmaceutical, and showed them to have high potency (IC50 ~2 nM) and allele-selectivity (>30-fold) against mutant HTT in HD-patient-derived cell-lines. Mechanistically, CAG-targeting ss-siRNA functions through endogenous RNAi by recruiting Ago2 and GW182 to HTT mRNA in the absence of a passenger strand and reducing mutant HTT protein level without affecting its mRNA level. Selectivity is achieved through preferential cooperative binding of multiple RISC units to the longer poly-CAG tract on the mutant HTT mRNA versus that of the wild-type. Structural-activity relationship (SAR) studies showed that several ss-siRNAs tolerated significant structural modifications and still retained high potency and selectivity. Furthermore, intraventricular infusion of a candidate ss-siRNA in a HD knock-in mouse model yielded selective inhibition of mutant HTT in a wide range of brain regions. Finally, we showed that a subset of ss-siRNAs were also potent, allele-selective inhibitors of ATAXIN-3, the mutated gene in spinocerebellar ataxia type 3 (SCA3). Taken together, we have identified and characterized a novel class of mechanistically interesting and therapeutically promising nucleic-acid-based compounds that could open new doors to finding a cure for genetic diseases such as HD.","abstract_html":"A number of inherited neurological disorders remain incurable despite having well-defined monogenic etiologies. One example is Huntington's disease (HD), which is caused by CAG trinucleotide expansion in the gene HUNTINGTIN (HTT) and production of toxic glutamine-expanded protein. Targeting HTT with siRNAs could be a powerful approach, but allele-selectivity is a major challenge: nearly all HD patients are heterozygous at the HTT locus, and expression of wild-type HTT may need to be preserved. One way to achieve allele-selectivity is by exploiting the fact that the mutant HTT allele contains more CAG repeats. Previous work with double-stranded siRNAs (dsRNA) and chemically modified antisense oligonucleotides (ASO) that target the poly-CAG sequence both showed promise but each had significant limitations. To combine the simplicity of ASO and high selectivity of dsRNA, we tested chemically modified, single-stranded small-interfering RNA (ss-siRNA) of sequences targeting CAG repeats in collaboration with ISIS Pharmaceutical, and showed them to have high potency (IC50 ~2 nM) and allele-selectivity (>30-fold) against mutant HTT in HD-patient-derived cell-lines. Mechanistically, CAG-targeting ss-siRNA functions through endogenous RNAi by recruiting Ago2 and GW182 to HTT mRNA in the absence of a passenger strand and reducing mutant HTT protein level without affecting its mRNA level. Selectivity is achieved through preferential cooperative binding of multiple RISC units to the longer poly-CAG tract on the mutant HTT mRNA versus that of the wild-type. Structural-activity relationship (SAR) studies showed that several ss-siRNAs tolerated significant structural modifications and still retained high potency and selectivity. Furthermore, intraventricular infusion of a candidate ss-siRNA in a HD knock-in mouse model yielded selective inhibition of mutant HTT in a wide range of brain regions. Finally, we showed that a subset of ss-siRNAs were also potent, allele-selective inhibitors of ATAXIN-3, the mutated gene in spinocerebellar ataxia type 3 (SCA3). Taken together, we have identified and characterized a novel class of mechanistically interesting and therapeutically promising nucleic-acid-based compounds that could open new doors to finding a cure for genetic diseases such as HD.","abstract_has_math":false,"creators":["Yu, Dongbo 1984-"],"institution":null,"degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":null,"school":null,"contributors":["Yu, Hongtao","Liu, Qinghua","Conrad, Nicholas","Corey, David R."],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2016,"date_issued":"2016-06-27T20:19:40Z","date_published":"2016-06-27T20:19:40Z","updated_at":"2026-07-24T05:52:34Z","subjects":["Huntington Disease","Nerve Tissue Proteins","RNA Interference","RNA, Small Interfering"],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier","label":"Identifier","values":["https://hdl.handle.net/2152.5/3332","952355705"],"render_values":[{"text":"https://hdl.handle.net/2152.5/3332","href":"https://hdl.handle.net/2152.5/3332","code":true},{"text":"952355705","href":null,"code":true}]}]},"links":{"outbound_url":"https://hdl.handle.net/2152.5/ETD-UTSWMED-2014-05-59","outbound_label":"Handle","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Yu, Hongtao","Liu, Qinghua","Conrad, Nicholas","Corey, David R."]},{"key":"dc:creator","label":"Author","values":["Yu, Dongbo 1984-"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2016-06-27T20:19:40Z","2014-05","2013-02-20","May 2014","2016-06-27T19:46:10Z"]},{"key":"dc:type","label":"Dc Type","values":["Thesis","text"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Huntington Disease","Nerve Tissue Proteins","RNA Interference","RNA, Small Interfering"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["https://hdl.handle.net/2152.5/ETD-UTSWMED-2014-05-59","https://hdl.handle.net/2152.5/3332","952355705"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["A number of inherited neurological disorders remain incurable despite having well-defined monogenic etiologies. One example is Huntington's disease (HD), which is caused by CAG trinucleotide expansion in the gene HUNTINGTIN (HTT) and production of toxic glutamine-expanded protein. Targeting HTT with siRNAs could be a powerful approach, but allele-selectivity is a major challenge: nearly all HD patients are heterozygous at the HTT locus, and expression of wild-type HTT may need to be preserved. One way to achieve allele-selectivity is by exploiting the fact that the mutant HTT allele contains more CAG repeats. Previous work with double-stranded siRNAs (dsRNA) and chemically modified antisense oligonucleotides (ASO) that target the poly-CAG sequence both showed promise but each had significant limitations. To combine the simplicity of ASO and high selectivity of dsRNA, we tested chemically modified, single-stranded small-interfering RNA (ss-siRNA) of sequences targeting CAG repeats in collaboration with ISIS Pharmaceutical, and showed them to have high potency (IC50 ~2 nM) and allele-selectivity (>30-fold) against mutant HTT in HD-patient-derived cell-lines. Mechanistically, CAG-targeting ss-siRNA functions through endogenous RNAi by recruiting Ago2 and GW182 to HTT mRNA in the absence of a passenger strand and reducing mutant HTT protein level without affecting its mRNA level. Selectivity is achieved through preferential cooperative binding of multiple RISC units to the longer poly-CAG tract on the mutant HTT mRNA versus that of the wild-type. Structural-activity relationship (SAR) studies showed that several ss-siRNAs tolerated significant structural modifications and still retained high potency and selectivity. Furthermore, intraventricular infusion of a candidate ss-siRNA in a HD knock-in mouse model yielded selective inhibition of mutant HTT in a wide range of brain regions. Finally, we showed that a subset of ss-siRNAs were also potent, allele-selective inhibitors of ATAXIN-3, the mutated gene in spinocerebellar ataxia type 3 (SCA3). Taken together, we have identified and characterized a novel class of mechanistically interesting and therapeutically promising nucleic-acid-based compounds that could open new doors to finding a cure for genetic diseases such as HD."]},{"key":"dc:format","label":"Dc Format","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["Using Chemically Modified Oligonucleotides to Modulate Gene Expression, Treat Genetic Diseases, and Probe Novel Mechanisms of RNA Interference"]}]}],"canonical_facts":{"dc:contributor":["Yu, Hongtao","Liu, Qinghua","Conrad, Nicholas","Corey, David R."],"dc:creator":["Yu, Dongbo 1984-"],"dc:date":["2016-06-27T20:19:40Z","2014-05","2013-02-20","May 2014","2016-06-27T19:46:10Z"],"dc:description":["A number of inherited neurological disorders remain incurable despite having well-defined monogenic etiologies. One example is Huntington's disease (HD), which is caused by CAG trinucleotide expansion in the gene HUNTINGTIN (HTT) and production of toxic glutamine-expanded protein. Targeting HTT with siRNAs could be a powerful approach, but allele-selectivity is a major challenge: nearly all HD patients are heterozygous at the HTT locus, and expression of wild-type HTT may need to be preserved. One way to achieve allele-selectivity is by exploiting the fact that the mutant HTT allele contains more CAG repeats. Previous work with double-stranded siRNAs (dsRNA) and chemically modified antisense oligonucleotides (ASO) that target the poly-CAG sequence both showed promise but each had significant limitations. To combine the simplicity of ASO and high selectivity of dsRNA, we tested chemically modified, single-stranded small-interfering RNA (ss-siRNA) of sequences targeting CAG repeats in collaboration with ISIS Pharmaceutical, and showed them to have high potency (IC50 ~2 nM) and allele-selectivity (>30-fold) against mutant HTT in HD-patient-derived cell-lines. Mechanistically, CAG-targeting ss-siRNA functions through endogenous RNAi by recruiting Ago2 and GW182 to HTT mRNA in the absence of a passenger strand and reducing mutant HTT protein level without affecting its mRNA level. Selectivity is achieved through preferential cooperative binding of multiple RISC units to the longer poly-CAG tract on the mutant HTT mRNA versus that of the wild-type. Structural-activity relationship (SAR) studies showed that several ss-siRNAs tolerated significant structural modifications and still retained high potency and selectivity. Furthermore, intraventricular infusion of a candidate ss-siRNA in a HD knock-in mouse model yielded selective inhibition of mutant HTT in a wide range of brain regions. Finally, we showed that a subset of ss-siRNAs were also potent, allele-selective inhibitors of ATAXIN-3, the mutated gene in spinocerebellar ataxia type 3 (SCA3). Taken together, we have identified and characterized a novel class of mechanistically interesting and therapeutically promising nucleic-acid-based compounds that could open new doors to finding a cure for genetic diseases such as HD."],"dc:format":["application/pdf"],"dc:identifier":["https://hdl.handle.net/2152.5/ETD-UTSWMED-2014-05-59","https://hdl.handle.net/2152.5/3332","952355705"],"dc:subject":["Huntington Disease","Nerve Tissue Proteins","RNA Interference","RNA, Small Interfering"],"dc:title":["Using Chemically Modified Oligonucleotides to Modulate Gene Expression, Treat Genetic Diseases, and Probe Novel Mechanisms of RNA Interference"],"dc:type":["Thesis","text"]},"updated_at":"2026-07-24T05:52:34Z"}