Page 33 - D. Cancer biology

Replenishment Of SOCS3 Suppresses Angiogenesis
By Negative Regulation Of JAK/STAT Signaling
Jaehyeon Kim, Sukyeong Jeong, Shinyoung Park, Chohyun Kim, Mikyung Kim, Youngsil Choi and Daewoong Jo
Anti-Cancer Drug Development Team, Cellivery R&D Institute, Cellivery Therapeutics, Inc., Seoul 03929, Korea.
BACKGROUND AIM

Massive upregulation of cytokines and growth factors drive pathologic
new vessel formation whereas physiologic angiogenesis is more The aim of this study is to develop an improved cell-permeable (iCP)-
controlled. Suppressor of cytokine signaling 3 (SOCS3) is an inducible SOCS3 and determine its applicability as an anti-angiogenic agent to
negative feedback regulator acting downstream of janus kinase/signal treat cancer by negatively regulating JAK/STAT signaling.
transducer and activator of transduction (JAK/STAT) signaling.

METHODS
iCP-SOCS3 has been developed by fusing sequence-optimized advanced macromolecule transduction domain (aMTD) and SOCS3 to
determine its applicability as an anti-angiogenic agent. Uptake of a fluorescently tagged iCP-SOCS3 by cells and tissues was monitored by flow
cytometry and confocal microscopy. Biological activities of iCP-SOCS3 were determined by angiogenesis assays and changes in biomarker
expression and anti-tumor activities were analyzed in mice bearing human tumor xenografts.

RESULTS

Figure 1. iCP-SOCS3 Shows High Solubility/Yield Figure 4. iCP-SOCS3 Significantly Inhibits Figure 6. iCP-SOCS3 Inhibits Cancer Cell (U-87 MG) -
& High Cell-/Tissue-Permeability Angiogenesis Ex Vivo & In Vivo Induced Angiogenesis In Vivo
A A A
Chick Embryonic Aortic Ring Assay New Vessel Formation
VEGF (20 ng/ml)
Diluent Non-CP-SOCS3 iCP-SOCS3 Avastin®
iCP-SOCS3
-
Non-CP-SOCS3
B Non-CP- Vehicle
SOCS3 iCP-SOCS3
M - + P - + P FITC only
FITC-Non-CP-SOCS3
FITC-iCP-SOCS3
300
Counts 200 100 120
Control
Solubility ++++ +++++ 0 100 Vehicle 20 Diluent (n=10)
10 2
Yield (mg/L) 27 50 10 0 10 1 Fluorescence 10 3 10 4 80 55% Non-CP-SOCS3 15 Non-CP-SOCS3 (60 mg/kg, n=10)
iCP-SOCS3
iCP-SOCS3 (60 mg/kg, n=10)
C FITC only FITC-Non- FITC-iCP-SOCS3 Rel. Sprouting Length (%) 60 * Avastin® Hemoglobin (mg/ml) 10 5 84% Avastin® (2 mg/kg, n=8) *: p<0.05
*: p<0.05
CP-SOCS3
40
*
20
FITC 0 0 Treatment
B
Chick Chorioallantoic Membrane (CAM) Assay
B Diluent Non-CP-SOCS3 iCP-SOCS3
Diluent Non-CP-SOCS3 iCP-SOCS3
Figure 2. iCP-SOCS3 Suppresses
Angiogenesis Activity of Endothelial Cells Trichrome
A B
C
120 120
(Protein: 10 µM) 90 (Protein: 10 µM) 120 Diluent CD31 VEGF Merge
Rel. Cell Viability (%) 60 30 60% Rel. Migrated Cells (%) 60 30 * NeovasculizedEggs (%) 100 Retinoic Acid Diluent
90
Non-CP-SOCS3
54%
iCP-SOCS3
80
*
60
0
Vehicle Non-CP- iCP-SOCS3 0 Vehicle Non-CP- iCP-SOCS3 40
SOCS3 SOCS3 20 83%
C 0 iCP-SOCS3
1
150 (Protein: 10 µM) (Protein: 10 µM) 2 0.5 Treatment (μg) 2 1
Rel. Tube Length (%) 100 60% No. of Branch Points 150 50 79% Figure 5. iCP-SOCS3 Suppresses Tumor - Induced Figure7. iCP-SOCS3 Suppresses Expression of
100
Angiogenesis in Matrigel Plug Assay
*
50
*
0 0 Pro-Angiogenic Factors in CDX Models
Vehicle Non-CP- iCP-SOCS3 Vehicle Non-CP- iCP-SOCS3
SOCS3 SOCS3 A
Diluent Cancer Cell: Glioblastoma (U87-MG)
Figure 3. iCP-SOCS3 Inhibits Cancer Cells-Mediated Protein: 6 ㎍ Pancreatic Cancer Glioblastoma Hepatoma
(HepG2)
(U87-MG)
(PANC-1)
Invasion in Endothelial Cells
Non-CP-SOCS3
A B
PANC-1 Conditioned Media 120 U87MG Conditioned Media iCP-SOCS3 VEGF
120
Relative Invaded Cells (%) 90 38% Relative Invaded Cells (%) 90 60 30 95% B Angiopoietin 2 Diluent Non-CP-SOCS3 iCP-SOCS3 p-ERK
(Protein: 10 µM)
(Protein: 10 µM)
*
ERK
60
Angt I
30
*
0
Vehicle Non-CP- iCP-SOCS3 0 Vehicle Non-CP- iCP-SOCS3 β-actin β-actin
SOCS3 SOCS3
CONCLUSION REFERENCES Contact information
iCP-SOCS3 significantly suppresses tumor Chung et al. (2020) Science Advances, 6: eaba 1193 Minyong Jung
progression associated with angiogenesis. Lim et al. (2013) Clinical Cancer Research, 19: 680-690 New Drug & Business Development
Overall, these results suggest that iCP- Lim et al. (2013) Biomaterials, 34: 6261-6271 Cellivery Therapeutics, Inc.
SOCS3, inhibitor of JAK/STAT signaling, is Lim et al. (2012) Molecular Therapy, 20: 1540-1549
an effective anti-angiogenic agent to reduce Jo et al. (2005) Nature Medicine, 11: 892-898 jungmy@cellivery.com
angiogenesis in various types of tumors. +82-2-3151-8900
Jo et al. (2001) Nature Biotechnology, 19: 929-933