Accelerated blood clearance phenomenon upon cross-administration of PEGylated nanocarriers in beagle dogs

Figures & data

Table 1 Characterization data of nanocarriers

Formulation	Mean particle size (nm)	PDI	Zeta potential (mV)	FALT (nm)
PE	126.8±3.2	0.178±0.008	−36.1±2.6	3.5±0.2
PL	132.8±6.2	0.142±0.015	−47.4±4.2	3.3±0.1
PSLN-GDS	137.3±4.9	0.196±0.010	−28.9±1.8	2.8±0.4
PSLN-GMS	123.8±2.8	0.207±0.006	−32.2±2.7	2.1±0.2
PM	14.3±1.4	0.137±0.002	−16.4±1.9	–

Notes: Data are mean ± standard deviation of three repeats. “−” indicates that the value was not determined.

Abbreviations: PDI, polydispersity index; FALT, fixed aqueous layer thickness; GDS, glycerol distearate; GMS, glycerin monostearate; PE, PEGylated emulsions; PEG, poly(ethylene glycol); PL, PEGylated liposomes; PM, PEG micelles; PSLN, PEGylated solid lipid nanoparticles.

Table 2 Injection protocols used to administer nanocarriers to normal beagle dogs

Group	First treatment	Second treatment
A	PE	PE
B	PL	PE
C	PSLN-GMS	PE
D	PSLN-GDS	PE
E	PM	PE
F	PE	PL
G	PE	PSLN-GMS
H	PE	PM
I	PSLN-GMS	PSLN-GMS
J	PM	PM
K	PL	PL

Abbreviations: GDS, glycerol distearate; GMS, glycerin monostearate; PE, PEGylated emulsions; PEG, poly(ethylene glycol); PL, PEGylated liposomes; PM, PEG micelles; PSLN, PEGylated solid lipid nanoparticles.

Figure 1 The blood clearance profile of a single intravenous injection of various PEGylated nanocarriers at a phospholipid dose of 2.5 μmol/kg in beagle dogs.

Notes: Data shown over 0–4 hours (A). A magnified view of the 0–0.5 hour period (B). Data show mean ± standard deviation of three repeats.

Abbreviations: GMS, glycerin monostearate; PE, PEGylated emulsions; PEG, poly(ethylene glycol); PL, PEGylated liposomes; PM, PEG micelles; PSLN, PEGylated solid lipid nanoparticles.

Figure 2 Effect of various PEGylated nanocarriers on the induction of the ABC phenomenon in beagle dogs.

Notes: The beagle dogs were first given PE, PL, PSLN-GMS, or PM, at a dose of 2.5 μmol phospholipid/kg. Then, 7 days after the first injection, the beagle dogs were given PE. Data show mean ± standard deviation of three repeats.

Abbreviations: GMS, glycerin monostearate; PE, PEGylated emulsions; PEG, poly(ethylene glycol); PL, PEGylated liposomes; PM, PEG micelles; PSLN, PEGylated solid lipid nanoparticles.

Figure 3 Effect of PE on the pharmacokinetics of various PEGylated nanocarriers in beagle dogs.

Notes: The beagle dogs were first given PE at a dose of 2.5 μmol phospholipid/kg. Then, 7 days after the first injection, the beagle dogs were given PE (A), PSLN-GMS (B), PM (C), or PL (D). Data show mean ± standard deviation of three repeats.

Abbreviations: GMS, glycerin monostearate; PE, PEGylated emulsions; PEG, poly(ethylene glycol); PL, PEGylated liposomes; PM, PEG micelles; PSLN, PEGylated solid lipid nanoparticles.

Table 3 The ABC index_{(0–30 min)} of repeated injections and cross-administration of various PEGylated nanocarriers

Group	ABC index(0–30 min)a
Repeated injections
PE-PE	0.05±0.00***
PSLN-GMS-PSLN-GMS	0.51±0.03**
PM-PM	0.99±0.08
PL-PL	0.13±0.02***
Cross-administration
PL-PE	0.09±0.00***
PSLN-GMS-PE	0.42±0.08***
PM-PE	0.68±0.05**
PE-PM	1.04±0.09
PE-PSLN-GMS	0.08±0.01***
PE-PL	0.18±0.04***

Notes:

a The ABC index_{(0–30 min)} was calculated as follows: ABC index_{(0–30 min)} = AUC_{(0–30 min)} of the second dose/AUC_{(0–30 min)} of the single dose. Data are mean ± standard deviation of three repeats. P-values represent a significant difference compared with 1.

** P<0.01,

*** P<0.001.

Abbreviations: ABC, accelerated blood clearance; AUC, area under the curve; GMS, glycerin monostearate; min, minutes; PE, PEGylated emulsions; PEG, poly(ethylene glycol); PL, PEGylated liposomes; PM, PEG micelles; PSLN, PEGylated solid lipid nanoparticles.

Figure 4 Comparison of the effects of repeated injections and cross-administration of various PEGylated nanocarriers on immune response.

Notes: PE (A); PSLN-GMS (B); PM (C); and PL (D). Data show mean ± standard deviation of three repeats.

Abbreviations: GMS, glycerin monostearate; PE, PEGylated emulsions; PEG, poly(ethylene glycol); PL, PEGylated liposomes; PM, PEG micelles; PSLN, PEGylated solid lipid nanoparticles.

Figure 5 The production of anti-PEG IgM induced by the initial priming dose of various PEGylated nanocarriers (PE, PL, PSLN-GMS, and PM) at doses of 2.5 μmol phospholipid/kg.

Notes: The anti-PEG IgM in the serum was determined using ELISA. Data show mean ± standard deviation of three repeats. P-values represent a significant difference compared with 1. *P<0.05; **P<0.01.

Abbreviations: GMS, glycerin monostearate; ELISA, enzyme-linked immunosorbent assay; PE, PEGylated emulsions; PEG, poly(ethylene glycol); PL, PEGylated liposomes; PM, PEG micelles; PSLN, PEGylated solid lipid nanoparticles.

Figure 6 Diagram depicting the different PEG conformations on the surface of various nanocarriers.

Notes: PEG chains adopt a brush conformation on the surface of PE and PL, but have a mushroom-like conformation on the surface of PSLN-GMS. PL (A); PE (B), and PSLN-GMS (C).

Abbreviations: GMS, glycerin monostearate; PE, PEGylated emulsions; PEG, poly(ethylene glycol); PL, PEGylated liposomes; PSLN, PEGylated solid lipid nanoparticles.

Figure 7 Effect of the types of lipid used to prepare PSLN on the induction of ABC phenomenon in beagle dogs. The beagle dogs were first given PSLN-GMS and PSLN-GDS at a dose of 2.5 μmol phospholipid/kg. Then, 7 days after the first injection, the beagle dogs were given PE. Data show mean ± standard deviation of three repeats.

Abbreviations: GDS, glycerol distearate; GMS, glycerin monostearate; PE, PEGy lated emulsions; PEG, poly(ethylene glycol); PSLN, PEGylated solid lipid nanoparticles.

Figure S1 The percentage change in leucocytes, neutrophilic granulocytes, monocytes, and lymphocytes in left and right limbs after the intravenous injection.

Notes: The beagle dogs were treated with PE at a dose of 2.5 μmol phospholipid/kg. Right limb (A); left limb (B). Data show mean ± standard deviation of three repeats. Q% = (Q_t/Q₀)×100, where Q_t is the value for the blood sample and Q₀ is the control value.

Abbreviation: PE, PEGylated emulsions.