Introduction
Lung cancer is the leading cause of cancer-related deaths among humans
worldwide (Kyu et al., 2018), and
non-small
cell lung cancer
(NSCLC)
accounts for approximately 85% of all lung cancer cases (Yuan, Huang,
Chen, Wu & Xu, 2019). Even when NSCLC is diagnosed at an operable stage
and treatment with chemotherapy is applied, the incidence of recurrence
and metastasis remains high with a median survival of less than 10-12
months. Most patients initially respond to platinum or gemcitabine-based
chemotherapy (Mlak et al., 2016; Pelayo Alvarez, Westeel, Cortes-Jofre
& Bonfill Cosp, 2013; Wang et al., 2018), but usually relapse and
acquire
chemo-resistant
disease, such that the 5-year survival is < 0.5% (Morabito et
al., 2014). Thus, targeting tumor metastasis and drug-resistant may
represent a promising strategy to treat NSCLC (Vyse & Huang, 2019).
It has long been presumed that tumors may take advantage of the
hemostatic system. Many significant hemostatic abnormalities have been
described in cancer patients, including disseminated intravascular
coagulation, hemorrhagic events, and migratory thrombophlebitis (Soff,
2019; Tan et al., 2019). Indeed, hemostatic complications are a common
cause of death in patients with cancer(Joseph S. Palumbo, 2000).
Previous studies in mice have unequivocally shown that tumor
cell-associated tissue factor (Yokota et al., 2014), circulating
prothrombin (Horowitz et al., 2011), and several downstream thrombin
procoagulant targets (i.e.,
platelets,
fibrinogen,
factor XII) strongly promote tumor cell metastatic potential (Joseph S.
Palumbo, 2000; Plantureux, Crescence, Dignat-George, Panicot-Dubois &
Dubois, 2018; Yokoyama, Mori & Matsuura, 2008). However, at the level
of primary tumor growth, the contribution of hemostatic factors has been
less clear. Thrombin is an allosteric enzyme with an elaborate structure
that exerts diverse biological effects by interacting with various
receptors on the surface of vascular and nonvascular cells (Lane,
Philippou & Huntington, 2005).
Thrombin
has also been shown to contribute to tumor progression in manners both
coagulation-dependent and coagulation-independent (Xue et al., 2010). A
substantial amount of data supports the idea that thrombin plays
important roles in tumorigenesis, contributing to inflammation,
angiogenesis, and metastatic dissemination of tumor cells through its
PAR-1 receptor (Battinelli, Markens, Kulenthirarajan, Machlus,
Flaumenhaft & Italiano, 2014; Reddel et al., 2017; Yokota et al.,
2014). Nevertheless, thrombin expression in NSCLC primary tumor tissues
and
the association with prognosis of NSCLC patients remain largely unknown.
PAR-1, the prototypic member of the PAR family, has been shown to
respond to a highly select group of serine proteases. Cleavage of PAR-1
by thrombin initiates potent inflammatory responses, including the
up-regulation of cell surface adhesion molecules, induction of
hyperpermeability (Feistritzer & Riewald, 2005), and the activation of
the nuclear factor κB
(NF-κB)
pathways (Jeffers et al., 2015).
At present, chemotherapy is an established multimodal therapy for NSCLC,
however, its benefits are limited due to a low response rate or acquired
tumor resistance. In the present study, Arnold et. al have shown that
PAR-1 in the tumors induces the chemo-resistance of cancer. As thrombin
is the prototypical PAR-1 agonist, which indicates that targeting
thrombin may be particular effective in combination with routine
chemotherapy.
Direct
Thrombin Inhibitor Peptide
(DTIP)
and recombinant hirudin
(r-hirudin)
which are
derivatives
of wild-type hirudin variant 2, were developed by our group (Mo, Zhang,
Chen, Wang & Song, 2009; Zhao et al., 2017). DTIP and r-hirudin bind to
exosite I and to the apolar region of thrombin, while the N-terminal
moiety of r-hirudin and DTIP blocks access to the thrombin active site
to inhibit the activity of thrombin.
r-hirudin
has entered phase I clinical trials, and DTIP is a novel antithrombotic
agent that could be used to prevent thrombosis without conferring an
increased bleeding risk for subcutaneous injection.
Here, we investigated the protein levels of thrombin in clinical NSCLC
samples, explored the relationship between thrombin expression level and
clinicopathological features, prognosis of NSCLC patients. We evaluated
the effects of
r-hirudin
and DTIP on tumor progression, dissemination and spontaneous metastasis
in vitro and in vivo. We also demonstrated the presence of thrombin and
PAR-1 accounts for the majority of the invasive signal. The novel
findings presented here indicate the roles
of
r-hirudin
and DTIP, anticoagulant drugs, could be expanded for anti-tumor therapy.
We speculate that the use of r-hirudin and DTIP in combination would be
a new breakthrough in cancer treatment.