title: Annexin A2 facilitates endocytic trafficking of antisense oligonucleotides.
abstract: Chemically modified antisense oligonucleotides (ASOs) designed to mediate site-specific cleavage of RNA by RNase H1 are used as research tools and as therapeutics. ASOs modified with phosphorothioate (PS) linkages enter cells via endocytotic pathways. The mechanisms by which PS-ASOs are released from membrane-enclosed endocytotic organelles to reach target RNAs remain largely unknown. We recently found that annexin A2 (ANXA2) co-localizes with PS-ASOs in late endosomes (LEs) and enhances ASO activity. Here, we show that co-localization of ANXA2 with PS-ASO is not dependent on their direct interactions or mediated by ANXA2 partner protein S100A10. Instead, ANXA2 accompanies the transport of PS-ASOs to LEs, as ANXA2/PS-ASO co-localization was observed inside LEs. Although ANXA2 appears not to affect levels of PS-ASO internalization, ANXA2 reduction caused significant accumulation of ASOs in early endosomes (EEs) and reduced localization in LEs and decreased PS-ASO activity. Importantly, the kinetics of PS-ASO activity upon free uptake show that target mRNA reduction occurs at least 4 hrs after PS-ASOs exit from EEs and is coincident with release from LEs. Taken together, our results indicate that ANXA2 facilitates PS-ASO trafficking from early to late endosomes where it may also contribute to PS-ASO release.
authors.affiliation: Department of Core Antisense Research, Ionis Pharmaceuticals, Inc. 2855 Gazelle Court, Carlsbad, CA 92010, USA.
authors.forename: Shiyu
authors.initials: S
authors.lastname: Wang
authors.affiliation: Department of Core Antisense Research, Ionis Pharmaceuticals, Inc. 2855 Gazelle Court, Carlsbad, CA 92010, USA.
authors.forename: Hong
authors.initials: H
authors.lastname: Sun
authors.affiliation: Department of Medicinal Chemistry, Ionis Pharmaceuticals, Inc. 2855 Gazelle Court, Carlsbad, CA 92010, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Tanowitz
authors.affiliation: Department of Core Antisense Research, Ionis Pharmaceuticals, Inc. 2855 Gazelle Court, Carlsbad, CA 92010, USA Lliang@ionisph.com.
authors.forename: Xue-Hai
authors.initials: XH
authors.lastname: Liang
authors.affiliation: Department of Core Antisense Research, Ionis Pharmaceuticals, Inc. 2855 Gazelle Court, Carlsbad, CA 92010, USA.
authors.forename: Stanley T
authors.initials: ST
authors.lastname: Crooke
mesh_terms: D017306:Annexin A2 / Q000172:deficiency / Q000378:metabolism*; D001692:Biological Transport; D002460:Cell Line; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism*; D006801:Humans; D016376:Oligonucleotides, Antisense / Q000378:metabolism*; D009418:S100 Proteins / Q000378:metabolism; D012492:Salts; D000099248:S100 Calcium Binding Protein A10
chemical_list: D017306:Annexin A2; D016376:Oligonucleotides, Antisense; D009418:S100 Proteins; D012492:Salts; D000099248:S100 Calcium Binding Protein A10