Discussion
In traditional Chinese medicine, Ginkgo leaf extract is widely
used as medical herb to treat cardiovascular disorders
(Xu, Hu, Shen & McQuillan, 2015). It has
been reported that GGC isolated from Ginkgo leaf , can exhibit
diverse pharmacological properties (Huang
et al., 2014; Huang, Chen, Liu, Wu &
Liou, 2018; Liou, Lai, Chen, Wang, Wei &
Huang, 2015; Zhang et al., 2018). This
is the first investigation that has analyzed the potential impact of GGC
and its mechanism in NSCLC model. We found that GGC diminished
activation of STAT3 as well as that of JAK1, JAK2, and c-Src
phosphorylation. This inhibition led to an induction in the expression
of phosphatase PTPε. GGC also mitigated proliferation and promoted
apoptosis through down-regulating STAT3 signaling cascade. Moreover, it
attenuated tumor growth and survival in a xenograft mouse model without
displaying any toxicity.
Aberrant STAT3 activation has been associated with various tumors,
including head and neck, lung, breast, prostate, kidney, pancreas, liver
cancer, lymphomas, and multiple myeloma
(Banerjee & Resat, 2016;
Bharti, Donato & Aggarwal, 2003;
Chai et al., 2016;
Hwang et al., 2019;
Jung et al., 2018). We first investigated
if GGC could alter constitutive and inducible STAT3 phosphorylation in
NSCLC cells. We noted that GGC abrogated STAT3 phosphorylation and
localization of this protein into the nucleus. Phosphorylation of JAKs
has been linked to STAT3 activation (Ahn,
Sethi, Sung, Goel, Ralhan & Aggarwal, 2008;
Baek et al., 2017;
Bowman, Garcia, Turkson & Jove, 2000).
Interestingly, GGC mitigated JAK1, JAK2, as well as c-Src
phosphorylation substantially and overexpression of STAT3 not only
alleviated the inhibitory actions of GGC on p-STAT3 levels but also
relieved apoptosis caused by this agent.
Recently, many studies have reported that protein tyrosine phosphatases
(PTPs) may negatively control STAT3 signaling pathway
(Ahn, Sethi, Sung, Goel, Ralhan &
Aggarwal, 2008; Baek et al., 2016a;
Baek et al., 2016b;
Kim, Morales, Jang, Cho & Kim, 2018). We
noted that GGC may have altered STAT3 activation by causing the
modulation of PTPs. Interestingly, GGC promoted PTPε expression at
protein and mRNA levels. Moreover, GGC could increase PTPε M expression
also at mRNA levels. Moreover, the silencing of PTPε expression
repudiated GGC-driven attenuation of STAT3 activation and induction of
apoptosis. However, how GCC can stimulate PTPε levels and if GCC can
also alter, its activity needs additional investigations.
It has been reported that reduction of STAT3 activity may decrease the
survival ability of tumor cells (Aoki,
Feldman & Tosato, 2003). Hence, we observed if GGC could alter
proliferation and promote apoptosis in NSCLC cells. GGC induced
apoptotic death was detected by cell cycle, annexin V and TUNEL assays.
We demonstrated that GGC caused sub-G1 arrest, early apoptosis, and
mediated the activation of caspase-3 and PARP. In addition, we noted
that GGC dramatically reduced the expression of oncogenic molecules such
as anti-apoptotic and metastasis promoting proteins that may contribute
to its varying anti-neoplastic activities.
In a NSCLC preclinical model, GGC significantly suppressed lung tumor
growth, altered the levels of p-STAT3, PTPε, and caspase-3 in treated
groups. These modifications are consonant with its observed in
vitro actions and were noted in the absence of any adverse effects.
Thus, our preclinical studies imply that GGC may have a potential as a
novel therapeutic agent for the management of NSCLC.
In summary, it was noticed that GGC can exert pleiotropic
anti-neoplastic effects through modulating STAT3 signaling pathway by
affecting the PTPε expression. Taken together, GGC may act as an
effective inhibitor of STAT3 phosphorylation and its function can be
investigated further in alleviation of different malignancies.