Inhibition of tissue factor signaling suppresses tumor growth

Reported by: Versteeg HH et al. Blood. 2008;111:190-199.

ABSTRACT
Background: The tissue factor (TF)-initiated coagulation pathway is
implicated in cancer progression, cancer-associated thrombosis, and
metastasis. However, the mechanisms by which TF exerts its effects
in cancer remain incompletely understood. This study examined how
TF contributes to primary tumor growth using antibodies that inhibit
RESULTS ! In nonmalignant human keratinocytes, VIIa induced association of Figure 5. Inhibition of TF-VIIa signaling with Mab-10H10 reduced breast
! Mab-10H10 and Mab-5G9 had different effects in an experimental TF with α1 integrin cancer tumor volume and weight in the mouse mammary fat pad. *P<.01
model of hematogenous metastasis ! In contrast, the association between TF- α1 integrin was found to be
! Blockade of TF-dependent coagulation (Mab-5G9) profoundly constitutive and not regulated by VIIa in cancer cells 700 0.8
reduced the load of surviving tumor cells after 24 hours
! Blockade of TF-VIIa signaling did not influence metastasis ! Inhibition of TF-VIIa signaling by Mab-10H10 attenuated tumor 600 MDA-MB-231mfp in mfp 0.7
growth in vivo in a highly aggressive breast cell cancer line

Tumor volume (mm3)

0.6
! These data suggest that TF-VIIa signaling does not contribute

Tumor weight (g)

500
either coagulation or direct TF-VIIa signaling. (Figure 3)
to initial survival of metastases 0.5
400
SFM 0.4
Methods: Two prototypic antibodies were identified with relative 300
0.3
specificity to block either direct TF-VIIa signaling or coagulation Figure 1. Inhibition of breast cancer metastasis by coagulation-blocking Figure 3. Inhibition of TF-VIIa signaling by Mab-10H10 inhibited 200
*
0.2
without displacing VIIa. These antibodies were used to determine the Mab-5G9. “Photon flux” is a measure of the tumor cell load subcutaneous tumor growth in a highly aggressive breast cancer cell line
100 10H10
contributions of TF signaling versus coagulation to tumor growth (MDA-MB-231mfp). **P<.001 0.1

in vivo. 0 0
0 5 10 15 20 25 30 SFM 10H10
3 Days after injection
600
Results: Blockade of direct TF-VIIa signaling inhibits TF-VIIa-mediated 2.5 MDA-MB-231mfp 0.4
signaling of PAR2 and disrupts interaction of TF with integrins. In 500

Tumor weight (g)

Tumor volume (mm3)
epithelial and TF-expressing endothelial cells, association of TF with 2 0.3

photons flux (x106)

400 IgG1
α1 integrins is regulated by TF extracellular ligand binding and is 1.5
300
independent of PAR2 signaling or proteolytic activity of VIIa. α1β3 ** 0.2
! Breast tumor growth was also attenuated by blockade of PAR2
1 5G9
integrin association of TF is constitutive in highly aggressive breast 200
signaling
0.1 **
cancer cells, whereas TF-integrin interaction is regulated in 0.5
100
noncancerous cells by extracellular ligand binding. The interaction ** ** **
0 10H10
0
between α1β3 and TF in breast cancer cells is blocked by antibodies 0

10H10
5G9

10H10
5G9

10H10
5G9

10H10
5G9
Buffer

Buffer

Buffer

Buffer
0 10 20 30 IgG1 10H10 5G9
that inhibit direct signaling, but not by antibodies that block Days after injection CONCLUSIONS
coagulation. Both antibodies had roughly equivalent antitumor ! The results of this study show that tumor cell TF-PAR2 signaling is
4h day 1 day 2 day 7
activity in vivo. critical for tumor growth
! Anti-TF strategies may be applied in cancer therapy with minor
Conclusions: Tumor growth is dependent, at least in part, on tumor
impairment of TF-dependent hemostatic pathways
cell-PAR2 signaling. Anti-TF strategies can be applied in cancer
Mab-10H10 and Mab-5G9 selectively blocked PAR2 signaling in ! Inhibition of TF-VIIa signaling also attenuated tumor growth in a
therapy with minor impairment of TF-dependent hemostatic pathways. !
breast cancer cells (Figure 2) melanoma cell model; Mab-5G9 inhibited melanoma primary tumor
growth; Mab-10H10 substantially more potent (Figure 4)

INTRODUCTION Figure 2. Both Mab10H10 and Mab5G9 inhibited TF-VIIa signaling in

! It is known that TF-initiated coagulation plays a key role in normal human breast cancer cells. Induction of the nuclear orphan receptor TR3 Figure 4. Inhibition of TF-VIIa signaling by either Mab-5G9 or
homeostasis, cardiovascular disease, and thrombosis. Activation of provides a sensitive readout of PAR signaling; induction of IL-8 Mab-10H10 retarded final tumor melanoma volumes and weights.
coagulation in the vicinity of TF-expressing tumor cells and shedding measures TF signaling **P<.001; *P<.01
of procoagulant activity into the circulation promote cancer-
associated thrombosis and its downstream complications 1400 1.2
! TF expression has also been found to correlate with tumor control M24met
5G9
1200 1.0
VIIa

progression and metastasis

Tumor volume (mm3)

10H10

Tumor weight (g)

1000 PBS 0.8
! Previous studies have found that overexpression of TF in anti-PAR2
800 *
several cancer types enhances tumor cell growth; knock-down control
0.6
5G9
VIIa/X

of TF attenuates tumor expansion 600

5G9 **
10H10 0.4
! Tumor growth has been shown to be attenuated by treatment with anti-PAR2 400
control 0.2
the direct thrombin inhibitor hirudin 200 10H10
5G9
IIa

! These data suggest that TF has an indirect, positive effect 10H10 0 0

on tumor growth by enhancing local angiogenesis, fibrin anti-PAR2 0 5 10 15 20 PBS 10H10 5G9
Days after injection
deposition, or platelet activation 0 10 20 30 40 50 60
! These activities may be mediated through PAR1; however, TR3 induction (fold)
PAR1-deficient animals show normal growth of transplanted control
tumors and metastases, suggesting thrombin-dependent PAR1 5G9
VIIa

signaling is dispensable for tumor growth 10H10

anti-PAR2 ! Mab10H10 was effective in reducing tumor volume and weight in the
! TF is known to be involved in 2 cellular signaling pathways control
mammary fat pad
! TF regulates integrins and, when bound by VIIa, reverses
VIIa/X

5G9
10H10
suppression cell migration mediated by integrin α1β3 anti-PAR2
! TF-VIIa also mediates cleavage of PAR2. PAR2 signaling control
stimulates migration by a feedback pathway that releases 5G9
IIa

10H10
integrin suppression
anti-PAR2

0 10 20 30 40 50 60 70 80 90
METHODS IL8 induction (fold)
! Two antibodies were identified:
! Mab-10H10: Blocks direct TF-VIIa signaling
! Mab-5G9: Blocks coagulation without displacing VIIa
! Tumor growth and metastasis were assessed in 6-week old, female
SCID mice

For educational purposes only. These were not prepared or reviewed by the primary author.