for Anticancer and Anti-inflammation Therapy Sangyong Jon, Ph.D. - - PowerPoint PPT Presentation

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

http://www.benbest.com/nutrceut/AntiOxidants.html

Bilirubin? A Final Metabolite of Heme

yellow, bile pigment ( ~ 1 mg/dL blood)

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!

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

X

in the liver

Display ‘Jaundice’ signature

Eyes with Jaundice

• 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

Bilirubin: A Potent Anti-oxidant in Vivo

‘Jekyll & Hyde’ Character ‘Double-edged Sword’

Bilirubin

Hydrophobic, Water insoluble! Bilirubin

 PEGylated Bilirubin

Amphiphillic, Water miscible!  Little chance to be remained

• r accumulated in the body!

PEG2000-Bilirubin

Bilirubin EDC, mPEG2,000-NH2 DMSO PEGylated bilirubin (PEG-BR) Self-assembly Bilirubin nanoparticles (BRNPs)

200 nm

SEM

0.5 mm 100 nm

TEM

Bilirubin Nanoparticles (BRNPs)

ROS & Inflammatory Diseases

ROS ROS

Myocardia iac infa nfarctio ion mod

• del

Rat Rat brain rain isch chemic ic st strok roke mod

• del

BR BR NPs NPs

Ant ntio ioxid idant ef effe fect Dru rug re rele lease

Dr Drug

released from BR NPs by ROS stimulus

Ano nother r mechanis ism Acu cute or

• r Chr

Chronic ic Asthma mod

• del

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)

BRNPs: ROS/Light-responsive Drug Delivery Carriers as well as a Medicine!

Phototherapy for Neonatal Jaundice

blue light ~ 450 nm

Bilirubin Metabolism During Phototherapy

Water insoluble! Increased solubility! Increased solubility!

Light-triggered Disruption of BRNPs

Light Bilirubin (water insoluble) Photoisomers (water soluble!)

Drug Loading Capability of BRNPs

Fraction (1 per 2ml)

5 10 15 20 25

Relative Fluorescence

20 40 60 80 100

PBN (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 Evaporation PEG-BR film layer

100 % Encapsulation efficiency 9.09 % Drug loading percentage (Maximun 23%)

Light-Induced Disruption of BRNPs (450nm)

122.4 nm 1.1 nm

 Photo irradiation for 1 min at 450 nm (10 mW/cm2)  Light-induced drug release profile

Time (min)

2 4 6 8 10

% Species

20 40 60 80 100

DTX

Light-Induced Disruption of BRNPs (650nm)

 Photo irradiation for 1 min at 650 nm (90 mW/cm2)

98.9 nm

Time (min)

2 4 6 8 10

% Species

20 40 60 80 100

DTX

 Light-induced drug release profile

0.96 nm

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

Tumor Heart Heart Tumor Liver Spleen Spleen Liver Kidney Kidney Lung Lung

Day

5 10 15 20 25 30

% Body Weight

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

5 10 15 20 25 30

Tumor volume (mm3)

100 200 300 400 500

Control 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

Adapted from Overchuk M. and Zheng G., Biomaterials, 2018.

Tumor microenvironment & nanomedicine

Adapted from Zhang Y. et al., Oxidative Medicine and Cellular Logevity, 2016.

Tumor microenvironment & ROS

ROS are overproduced in the TME!

Bilirubin: A Redox Active Compound

Water insoluble! More water soluble! greenish pigment yellowish pigment

ROS-triggered Disruption of BRNPs

ROS Bilirubin (water insoluble) Biliverdin (increased water solubility)

37.8 nm 1.5 nm 0 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

bt-BRNPs as a TME ROS-targeting nanomedicine

Preparation of ROS-responsive bt-BRNPs

Cell targeting and drug release of Dox@bt-BRNPs

Tumor targeting of Cyp@bt-BRNPs

Antitumor Efficacy of BRNPs in Vivo

• Adv. Science, 2018.