425 A Small Molecule Glycomimetic Antagonist of E-selectin and CXCR4 (GMI-1359)
Prevents Pancreatic Tumor Metastasis and Offers Improved Chemotherapy
Maria M. Steele1, BS, William E. Fogler2, PhD, John L. Magnani2, PhD, and Michael A. Hollingsworth1, PhD
1Eppley
Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha, NE
2GlycoMimetics, Inc., Gaithersburg, MD
Pancreas Orthotopic Metastasis Study
GMI-1359 Inhibits PDAC Binding to Lymphatic Endothelium
ABSTRACT
The tumor microenvironment's cellular and molecular composition includes a complex matrix that encases tumor cells and
presents a conundrum in respect to pancreatic tumor therapy. On one hand, the dense desmoplastic reaction that
accompanies growth of well differentiated pancreatic tumors reduces blood flow to tumors and creates an environment in
which it is difficult to deliver therapies. On the other hand, elimination of certain stromal elements enhances the
aggressiveness and metastatic potential of pancreatic cancer cells. Several current studies are attempting to target stroma in
an effort to enhance delivery and efficacy of therapeutic agents and to block metastasis. We have investigated the activity of
GMI-1359, a potent dual antagonist that targets both E-selectin and CXCR4. Adhesion protein E-selectin plays an important
role in the tumor microenvironment by regulating cell contacts, including tumor cell binding to vascular and lymphatic
endothelial cells during extravasation. Chemokine receptor CXCR4's role in the chemoattraction of tumor cells toward
endothelial cells (ECs) contributes to tumor microenvironment remodeling by influencing lymphangiogenesis/angiogenesis,
tumor cell survival/proliferation, and tumor stem cell mobilization. Our in vitro studies show that pancreatic ductal
adenocarcinoma (PDAC) cells do not attract growth of lymphatic nor vascular ECs toward themselves. However, tumor
associated fibroblasts, a major component of the PDAC tumor microenvironment, significantly increase EC directional
migration. GMI-1359 completely blocked lymphatic and vascular EC migration toward fibroblast cells and disrupted these
cell-cell interactions. In addition, GMI-1359 inhibited the capacity of invasive PDAC cell lines S2.013 and Colo357 to bind
and migrate across EC barriers. The dual antagonist was more effective than an independent small molecule E-selectin
inhibitor. We evaluated the capacity of GMI-1359 to inhibit growth and metastasis of orthotopically implanted S2.013 cells
with and without administration of Gemcitabine. Two weeks post implantation, mice were treated by intraperitoneal injection
for 4 weeks with either PBS once daily; 40 mg/kg GMI-1359 once daily; 100 mg/kg Gemcitabine every 4 days; or a
combination of GMI-1359 and Gemcitabine. GMI-1359 treatment slightly, but not significantly, decreased primary tumor size
as compared to the vehicle control. However, GMI-1359 in combination with Gemcitabine significantly reduced metastasis of
this tumor to liver and diaphragm as compared to mice that received only Gemcitabine. Further studies of GMI-1359 are
warranted to understand its potential for disrupting cellular contacts and blocking pancreatic tumor dissemination through the
vascular and lymphatic systems, and for enhancing the efficacy of chemotherapeutic approaches to pancreatic cancer.
All Mice:
1. Pretreat a confluent
endothelial monolayer with
drug for 1 hr then overlay
labeled tumor cells
2. Incubate for 1 hr to
allow tumor cells to
adhere to endothelial
layer
3. Wash away nonadherent tumor cells
and image adherent
cells
GMI-1359
GMI-1271
*
*
*
Tumor Volume
Also performed with a HUVEC monolayer with similar results
GMI-1359 Inhibits PDAC Transendothelial Migration
1. Plate a
monolayer of
endothelial cells in
the insert of a
Boyden chamber
2. Pretreat
monolayer for 1 hr
then add labeled
tumor cells
3. Allow tumor
cells to TEM
for 8 hours
4. Remove nonmigrated tumor
cells from inside of
the insert and
mount membrane
GMI-1271
GMI-1359 Inhibits PDAC and Fibroblast Proliferation
Only Mice Surviving at End of Study:
GMI-1359
n=8
GMI-1271
GMI-1359
Control
n=7
GMI-1359
*
*
*
n=8
*
*
S2.013
Control
10 ug/ml
100 ug/ml
1000 ug/ml
Also performed with a HUVEC monolayer with similar results
GMI-1359
13.34
Fibroblasts
GMI-1359 Inhibits Endothelial Migration Toward Fibroblast
Conditioned Media
Create Conditioned Media (CM):
Control
10 ug/ml
100 ug/ml
1000 ug/ml
Control
GMI-1359
1. Add conditioning cells
to a tissue culture plate
2. After adhere, add fresh
serum free media
Migration Toward CM:
4. Collect CM and add to
lower wells of Boyden
chamber. Plate migratory
cells in upper insert
3. Allow cells to condition
media for 24 hr
5. Following
migration, remove
non-migrated cells,
stain, and mount.
Conclusions
*
hLEC
*
Control
100 ug/ml
500 ug/ml
1000 ug/ml
Also performed with Colo357 and HUVECs with similar results
GMI-1359
Lymph
Node
Control
GMI-1271
Primary
Control
*
*
Also performed with a HUVEC monolayer with similar results
*
•
High dose GMI-1359, but not GMI-1271, inhibits PDAC cell and fibroblast proliferation in a dose
dependent manner. High dose GMI-1359 has no effect on endothelial cell proliferation.
•
GMI-1359 decreases the binding of PDAC cells to a lymphatic endothelium and also reduces these
cells’ ability to undergo transendothelial migration in a dose dependent manner.
•
GMI-1359 (and high dose GMI-1271) inhibits hLEC migration toward fibroblast conditioned media.
GMI-1359 also inhibits PDAC cell migration toward fibroblast conditioned media.
•
When used alone, GMI-1359 inhibits metastasis in a pancreas orthotopic tumor model. High dose
Gemcitabine also decreases metastasis. When used in combination with Gemcitabine, GMI-1271
further inhibits metastasis while GMI-1359 has no increased benefit.
*