title: Identification of the endosomal sorting complex required for transport-I (ESCRT-I) as an important modulator of anti-miR uptake by cancer cells.
abstract: Mechanisms of unassisted delivery of RNA therapeutics, including inhibitors of microRNAs, remain poorly understood. We observed that the hepatocellular carcinoma cell line SKHEP1 retains productive free uptake of a miR-21 inhibitor (anti-miR-21). Uptake of anti-miR-21, but not a mismatch (MM) control, induces expression of known miR-21 targets (DDAH1, ANKRD46) and leads to dose-dependent inhibition of cell growth. To elucidate mechanisms of SKHEP1 sensitivity to anti-miR-21, we conducted an unbiased shRNA screen that revealed tumor susceptibility gene 101 (TSG101), a component of the endosomal sorting complex required for transport (ESCRT-I), as an important determinant of anti-proliferative effects of anti-miR-21. RNA interference-mediated knockdown of TSG101 and another ESCRT-I protein, VPS28, improved uptake of anti-miR-21 in parental SKHEP1 cells and restored productive uptake to SKHEP1 clones with acquired resistance to anti-miR-21. Depletion of ESCRT-I in several additional cancer cell lines with inherently poor uptake resulted in improved activity of anti-miR-21. Finally, knockdown of TSG101 increased uptake of anti-miR-21 by cancer cells in vivo following systemic delivery. Collectively, these data support an important role for the ESCRT-I complex in the regulation of productive free uptake of anti-miRs and reveal potential avenues for improving oligonucleotide free uptake by cancer cells.
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Timothy R
authors.initials: TR
authors.lastname: Wagenaar
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Tatiana
authors.initials: T
authors.lastname: Tolstykh
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Chaomei
authors.initials: C
authors.lastname: Shi
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Lan
authors.initials: L
authors.lastname: Jiang
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: JingXin
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Zhifang
authors.initials: Z
authors.lastname: Li
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Qunyan
authors.initials: Q
authors.lastname: Yu
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Hui
authors.initials: H
authors.lastname: Qu
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Fangxian
authors.initials: F
authors.lastname: Sun
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Hui
authors.initials: H
authors.lastname: Cao
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Jack
authors.initials: J
authors.lastname: Pollard
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Shujia
authors.initials: S
authors.lastname: Dai
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Gao
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Bailin
authors.initials: B
authors.lastname: Zhang
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Heike
authors.initials: H
authors.lastname: Arlt
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: May
authors.initials: M
authors.lastname: Cindhuchao
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Dietmar
authors.initials: D
authors.lastname: Hoffmann
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Madelyn
authors.initials: M
authors.lastname: Light
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Karin
authors.initials: K
authors.lastname: Jensen
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Joern
authors.initials: J
authors.lastname: Hopke
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Richard
authors.initials: R
authors.lastname: Newcombe
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Carlos
authors.initials: C
authors.lastname: Garcia-Echeverria
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA.
authors.forename: Christopher
authors.initials: C
authors.lastname: Winter
authors.affiliation: Regulus Therapeutics, San Diego, CA 92121, USA.
authors.forename: Sonya
authors.initials: S
authors.lastname: Zabludoff
authors.affiliation: Sanofi Oncology, Cambridge, MA 02139, USA dmitri.wiederschain@sanofi.com.
authors.forename: Dmitri
authors.initials: D
authors.lastname: Wiederschain
mesh_terms: D000818:Animals; D001692:Biological Transport; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000502:physiology; D056827:Endosomal Sorting Complexes Required for Transport / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000502:physiology; D005260:Female; D006801:Humans; D016513:Mice, SCID; D035683:MicroRNAs / Q000037:antagonists & inhibitors* / Q000378:metabolism; D009369:Neoplasms / Q000235:genetics / Q000378:metabolism*; D009841:Oligonucleotides / Q000378:metabolism*; D014157:Transcription Factors / Q000502:physiology; D000099177:Tsg101 Protein
chemical_list: D004268:DNA-Binding Proteins; D056827:Endosomal Sorting Complexes Required for Transport; D035683:MicroRNAs; D009841:Oligonucleotides; D014157:Transcription Factors; D000099177:Tsg101 Protein; C524940:MIRN21 microRNA, human