Research ArticleHEALTH AND MEDICINE

Transient stealth coating of liver sinusoidal wall by anchoring two-armed PEG for retargeting nanomedicines

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Science Advances  26 Jun 2020:
Vol. 6, no. 26, eabb8133
DOI: 10.1126/sciadv.abb8133
  • Fig. 1 In situ stealth coating of liver sinusoidal wall using PEG-OligoLys.

    (A) OligoLys with 20 Lys units conjugated with two linear chains of 40-kDa PEG at its carboxyl end (two-arm-PEG-OligoLys). (B) Schematic illustration of in situ stealth coating of liver sinusoidal wall. Two-arm-PEG-OligoLys selectively attaches to the sinusoidal wall to prevent the attachment of nanomedicines, such as polyplex micelle (PM) and adeno-associated virus (AAV), to the wall via stealth property of PEG. Two-arm-PEG-OligoLys is gradually cleared to the bile to avoid prolonged disturbance of liver sinusoid functions.

  • Fig. 2 OligoLys distribution in connective tissue.

    (A) Chemical structures of one-arm-PEG-OligoLys (top, left), two-arm-PEG-OligoLys (top, right) with or without Alexa594 labeling (bottom). (B to D) Alexa594-labeled OligoLys with or without PEGylation was intravenously injected. Five minutes or 1 hour after the injection, earlobe dermis was observed using IVCLSM. (B) One-arm-PEG-OligoLys. (C) Two-arm-PEG-OligoLys. (D) Non-PEGylated OligoLys. Arrowheads, capillary walls. Two-way arrows, capillary lumen.

  • Fig. 3 Attachment of PEG-OligoLys to liver sinusoidal wall and its clearance.

    (A to D) IVCLSM images after injection of Alexa594-labeled one-arm-PEG-OligoLys (A) and two-arm-PEG-OligoLys (B). Green, autofluorescence of liver parenchyma. Red, one- and two-arm-PEG-OligoLys. Presumable regions of bile canaliculi are encircled with white dotted lines. Intensity profiles of Alexa594 in the white arrows in (A) and (B) are shown in (C) and (D), respectively. (C) One-arm-PEG-OligoLys. (D) Two-arm-PEG-OligoLys. (E) Bile ducts were visualized using 5-carboxyfluorescein (CF, green). Then, Alexa594-labeled two-arm-PEG-OligoLys (magenta) was injected for observation 7 hours later. Colocalization of these two colors is observed as white or cyan (encircled by yellow dotted lines). (F) Blood circulation profiles of PEG without OligoLys, and one- and two-arm-PEG-OligoLys. n = 4. Data are shown as means ± SEM.

  • Fig. 4 Coinjection of one- and two-arm-PEG-OligoLys.

    Alexa647-labeled one-arm-PEG-OligoLys (red) and Alexa594-labeled two-arm-PEG-OligoLys (green) were coinjected from the tail vein. (A) IVCLSM imaging of the liver. Presumable regions of bile canaliculi are encircled with white dotted lines. (B to D) Intensity profiles of Alexa594 and Alexa647 in the white arrows shown in (A). (B) 0.5 min, (C) 5 min, and (D) 6 hours after injection.

  • Fig. 5 Delivery of pDNA PM after preinjection of two-arm-PEG-OligoLys.

    Two-arm-PEG-OligoLys was intravenously injected to coat liver sinusoidal wall with PEG, followed by the intravenous injection of PM loading pDNA 5 min later. (A and B) IVCLSM imaging of PM loading Cy5-labeled pDNA (red) in the liver without PEG coating of sinusoid (A) or with the coating (B). Intensity profiles of Cy5 in the white arrows in (A) and (B) are shown in (C) and (D), respectively [(C) without coating and (D) with coating]. (E) Blood circulation profiles of PM with or without PEG coating of sinusoidal wall. n = 4. (F) PM loading Luc-expressing pDNA was injected to tumor-bearing mice with or without preinjection of two-arm-PEG-OligoLys. Luc expression in the tumor was measured 2 days after injection. n = 4. Data are shown as means ± SEM. Statistical analysis was performed using unpaired two-tailed Student’s t test.

  • Fig. 6 AAV transfection after preinjection of two-arm-PEG-OligoLys.

    Five minutes after intravenous injection of two-arm-PEG-OligoLys for PEG coating of liver sinusoidal wall, AAV8 expressing Luc was intravenously injected. Three weeks later, Luc expression in the liver (A), heart (B), and skeletal muscle (C) was measured. n = 6. Data are shown as means ± SEM. Statistical analysis was performed using unpaired two-tailed Student’s t test.

Supplementary Materials

  • Supplementary Materials

    Transient stealth coating of liver sinusoidal wall by anchoring two-armed PEG for retargeting nanomedicines

    Anjaneyulu Dirisala, Satoshi Uchida, Kazuko Toh, Junjie Li, Shigehito Osawa, Theofilus A. Tockary, Xueying Liu, Saed Abbasi, Kotaro Hayashi, Yuki Mochida, Shigeto Fukushima, Hiroaki Kinoh, Kensuke Osada, Kazunori Kataoka

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