a pegylated bilirubin nanomedicine for anticancer and anti … · 2019-10-06 · bilirubin...
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Sangyong Jon, Ph.D.
Bio-Nanomedicine Lab
Department of Biological Sciences
Korea Advanced Institute of Science and Technology (KAIST)
A PEGylated Bilirubin Nanomedicine
for Anticancer and Anti-inflammation
Therapy
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http://www.benbest.com/nutrceut/AntiOxidants.html
Bilirubin? A Final Metabolite of Heme
yellow, bile pigment
( ~ 1 mg/dL blood)
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Bilirubin as a Bad Guy
Eyes with Jaundice
(liver diseases?)
Neonatal jaundice
Jaundice itself is not a disease in adults,
but rather a sign of certain pathological conditions!
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Bilirubin: Water-insoluble Jaundice
Hydrophobic, Water insoluble!
(yellow colored pigment)
Deposition in various tissues
(skin, whites of the eyes (sclera), etc)
‘Unconjugated’ Bilirubin (BR)
More water soluble
‘Conjugated’ Bilirubin
glucuronyltransferase
Xin the liver
Display ‘Jaundice’ signature
Eyes with Jaundice
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• In 1929, Philip Hench, a rheumatologist, made a dramatic observation, correlating relief of incurable
symptoms of rheumatoid arthritis with the onset of jaundice.
Gilbert syndrome and Ischemic heart disease: a protective effect of elevated bilirubin
Inverse Relationship between serum bilirubin and atherosclerosis in men
Serum bilirubin and inverse correlation with colorectal cancer
Bilirubin as a Protective Factor for Rheumatoid arthritis
Relatively higher levels of bilirubin were associated with a lower risk of respiratory disease and all-
cause mortality
Exp Biol Med. (2003) Vol. 228(5):568-571.
Atherosclerosis (2002) Vol. 160:449–456.
JAMA. (2011) Vol. 305(7):691-697.
J Clin Med Res (2010) Vol.2(6):256-260.
Hepatology (2004) Vol.40: 827- 835.
Epidemiological Study (역학조사) on Bilirubin
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Bilirubin: A Potent Anti-oxidant in Vivo
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‘Jekyll & Hyde’
Character
‘Double-edged Sword’
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Bilirubin
Hydrophobic, Water insoluble!
Bilirubin
PEGylated Bilirubin
Amphiphillic, Water miscible!
Little chance to be remained
or accumulated in the body!
PEG2000-Bilirubin
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Bilirubin
EDC, mPEG2,000-NH2
DMSO
PEGylated bilirubin
(PEG-BR)
Self-assembly
Bilirubin nanoparticles (BRNPs)200 nm
SEM
0.5 mm100 nm
TEM
Bilirubin Nanoparticles (BRNPs)
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ROS & Inflammatory Diseases
ROS
Myocardiacinfarction model
Rat brain ischemic stroke model
BR NPs
Antioxidant effect Drug release
Drugreleased from
BR NPs by ROS stimulus
Another mechanism
Acute or Chronic Asthma model
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Bilirubin Nanoparticles: Universal anti-
inflammatory therapeutics without
immune suppression?
1) Inflammatory Bowel Disease: Acute colitis model
(Angew Chem Int Ed., 2016)
2) Hepatic ischemia-reperfusion injury:
Liver transplantation model (Biomaterials, 2017)
3) Islet xeno-transplantation (Biomaterials, 2017)
4) Acute asthma (Biomaterials, 2017)
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BRNPs: ROS/Light-responsive Drug
Delivery Carriers as well as a Medicine!
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Phototherapy for Neonatal Jaundice
blue light ~ 450 nm
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Bilirubin Metabolism During Phototherapy
Water insoluble!
Increased solubility!Increased solubility!
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Light-triggered Disruption of BRNPs
Light
Bilirubin
(water insoluble)
Photoisomers
(water soluble!)
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Drug Loading Capability of BRNPs
Fraction (1 per 2ml)
0 5 10 15 20 25
Re
lati
ve
Flu
ore
sc
en
ce
0
20
40
60
80
100PBN (PBN loaded with DOX)
DOX (PBN loaded with DOX)
Doxorubicin (Free DOX)
Bilirubin PEG-2000
Hydrophilic Drug
(Doxorubicin)
Hydrophilic drug
(DOX) in Water
(e.g Distilled
water or PBS)
Hydrophilic Drug
Encapsulated
Nanoparticles
Hydrophilic drug
elimination
By column separation
1) Dissolved
in CHCl3
2) CHCl3
EvaporationPEG-BR
film layer
100 % Encapsulation
efficiency
9.09 % Drug loading
percentage
(Maximun 23%)
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Light-Induced Disruption of BRNPs
(450nm)
122.4 nm1.1 nm
Photo irradiation for 1 min at 450 nm
(10 mW/cm2) Light-induced drug release profile
Time (min)
0 2 4 6 8 10
% S
pec
ies
0
20
40
60
80
100
DTX
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Light-Induced Disruption of BRNPs
(650nm)
Photo irradiation for 1 min at 650 nm
(90 mW/cm2)
98.9 nm
Time (min)
0 2 4 6 8 10
% S
pecie
s
0
20
40
60
80
100
DTX
Light-induced drug release profile
0.96 nm
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Xenograft model : Human lung adenocarcinoma epithelial cell line (A549)
in Balb/c nude mice
Dose: BRNPs (600 μg) in PBS, I.V injection
Control group: PBS
Cancer Targeting Ability of BRNPs
Control BNVs group
TumorHeart
HeartTumor
Liver
Spleen
Spleen
Liver
KidneyKidney
LungLung
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Day
0 5 10 15 20 25 30
% B
od
y W
eig
ht
60
70
80
90
100
110
Control
DOX
BRNV
DOX loaded BRNV
DOX loaded BRNV with light
Mouse : Balb/c nude mouse 7 weeks
Tumor : A549 Human lung carcinoma cell line (1*10^6)
Group : Control group (PBS), DOX (2mg/kg), BRNPs (20mg/kg),
DOX(2mg/kg)/BRNPs (20mg/kg), DOX(2mg/kg)/BRNPs (20mg/kg) with
laser [Laser : 650nm laser (5min, 200 mW/cm2) 30 min after injection of Dox
loaded BRNPs]
Dosing schedule : 0, 3, 6, 9,12 days
Day
0 5 10 15 20 25 30
Tu
mo
r vo
lum
e (
mm
3)
100
200
300
400
500Control
DOX (2mg/kg)
BRNV itself (20mg/kg)
DOX loaded BRNV
DOX loaded BRNV with light
*: p<0.01
#: p<0.05
##: p<0.05
Antitumor Efficacy of BRNPs in Vivo
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Adapted from Overchuk M. and Zheng G., Biomaterials, 2018.
Tumor microenvironment & nanomedicine
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Adapted from Zhang Y. et al., Oxidative Medicine and Cellular Logevity, 2016.
Tumor microenvironment & ROS
ROS are overproduced in the TME!
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Bilirubin: A Redox Active Compound
Water insoluble!More water soluble!
greenish pigment yellowish pigment
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ROS-triggered Disruption of BRNPs
ROS
Bilirubin
(water insoluble)
Biliverdin
(increased water solubility)
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37.8 nm1.5 nm0 min 10 min 60 min
60 min
(N.D.)0 min
ROS-triggered Disruption of BRNPs
The size of BRNPs drastically decreased!
Solution color of BRNPs became changed
Upon oxidation by peroxy radicals
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bt-BRNPs as a TME ROS-targeting nanomedicine
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Preparation of ROS-responsive bt-BRNPs
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Cell targeting and drug release of Dox@bt-BRNPs
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Tumor targeting of Cyp@bt-BRNPs
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Antitumor Efficacy of BRNPs in Vivo
Adv. Science, 2018.