4. Discussion
Currently, HCC is still one of the primary causes of cancer deaths,
although the survival rates and prognosis of HCC have been greatly
improved by liver transplantation and surgical resection [2]. TACE
procedures are currently widely used for various cases of HCC, mostly
patients at BCLC stage B. Gelatin sponge microparticle (GSM) is an
effective embolization agent, and a study reported that when compared
with the ethiodized oil which is a permanent embolization agent, GSMs
had many advantages, such as a higher tumor response rate at 1 month
after treatment, especially in large tumors [13]. Our previous
clinical practices have confirmed the safety and efficacy of GSMs-TACE
[5, 14, 15]. It was found that 350-560 μm GSMs combined with single
chemotherapy drug TACE procedures was effective and safe in treating
elderly HCC without surgical resection, and more elderly patients may
have better prognosis [5]. GSMs combined with trans-arterial
p53-gene-embolization also achieved satisfactory efficacy in treating
BCLC stage B HCC as evidenced by favorable survival rates (100% for
both 6 and 12 months) and no significant complications [14].
Therefore, GSMs were adopted in this study, and imaging scan confirmed
the efficacy of GSMs-TACE. DSA angiography during the procedure revealed
that after GSMs embolization, the blood flow velocity of tumor feeding
arteries, slowed down, and the tumor staining basically disappeared.
Moreover, 4 days after GSMs-TACE, CT plain scan showed some
honeycomb-like low-density changes in tumors compared with before the
procedure. In addition, in a previous study, after analyzing the data of
37 patients who received GSMs‑TACE for liver metastases after
gastrointestinal tumor surgery, it was observed that the treatment
produced various degrees of necrosis and shrinkage of lesions, and even
2 patients achieved a complete response [16]. Considering the close
relationship between tumor metastasis and the immunosuppressive TME,
these findings prompted us to wonder whether tumor necrosis after
GSMs-TACE improved the therapeutic efficacy via regulating the
immune-mediated elimination of tumor cells.
Regulatory T (Treg) cells are a subset of CD4+ T cells
in immune system known to function as “immune-suppressor”. It is well
recognized that Treg cells play an important role in protecting against
excessive inflammation and immune response in physiological or some
pathological conditions. Furthermore, it has been noticed that Treg
cells are involved in liver diseases including HCC [17]. The
infiltration of Treg cells into tumor sites can exert a suppressive
effect on host anticancer immunity and thereby become an obstacle to
curative anticancer therapy [10, 18, 19]. In the current study, we
first compared the Treg cell proportion in peripheral blood between HCC
patients and healthy volunteers, and it was found that there was an
increased proportion of Treg cells in peripheral blood of patients with
HCC (11.26 ± 1.31% and 7.32 ±1.63%, respectively), which was in
accordance with the previous study [19]. Besides, we analyzed the
association between the Treg cell proportion and various clinical
indicators of HCC, and it was found that the Treg cell proportion in
peripheral blood of HCC patients was closely related to tumor stage, AFP
content, and tumor diameter, which further confirmed the association
between the Treg cell proportion and HCC. However, the mechanism of the
association between peripheral Treg cells and HCC pathophysiological
characteristics has not been fully elucidated. Shi et al.proposed that Tregs could modify HCC in the way that potentiates the
metastasis since it was found that Treg cell proportions in HCC patients
were significantly correlated with biomarkers of tumor cell metastasis
such as E-cadherin, vimentin and SNAIL, although these findings were
based on the analysis of limited cases [20]. In addition, tumor
encapsulation was an important clinicopathological marker of HCC
invasiveness, and it was observed that the high Treg cell proportion in
peripheral blood of HCC patients was significantly correlated with the
presence of tumor encapsulation, which was inconsistent with the
published literature, where Gao et al. reported that a high Tregs
proportion was associated with the absence of tumor encapsulation
[21]. Nevertheless, these await further studies.
The 28 patients enrolled in this study received GSMs-TACE, and the
imaging techniques confirmed the success of the procedure. To testify
our hypothesis that TACE procedures may also have an impact on
anti-cancer immunity, the Treg cell proportion in peripheral blood of
these 28 HCC patients was determined before and at 10 and 30 days after
TACE. It was found that the peripheral Treg cell proportion at 10 days
after TACE was significantly lower than that of before the procedure,
and the percentage slightly increased 30 days after TACE but was still
significantly lower than that of before the procedure. These results
indicated that GSMs-TACE had a positive regulatory effect on immune
function. The possible reason is that after the effective embolization
of tumor feeding artery, the tumor cell necrosis is induced
significantly in a short time, and the tumor load is also significantly
reduced, which is related to the significant reduction of the
immunosuppressive effect on the body. It is well recognized that many
tumors bear tumor antigens and can induce T-cell cytotoxic responses
[22]. The release of tumor antigens then led to local
microenvironment immune response and a subsequent reduction of Treg
cells in peripheral blood. Our results showed that the Treg cells
proportion in peripheral blood at 10 days after operation was the
lowest, which indicated that it may be the optimal time for
immunoadjuvant therapy. The improvement of immune function can be
maintained within one month, but with the further proliferation of tumor
cells, the immune function gradually recovers to the state of
inhibition. Therefore, the follow-up treatment including TACE should be
carried out subsequently to further reduce the burden of tumors in order
to better restore and enhance the body’s anti-tumor immunity.
In recent years, tumor immunotherapies have gained promising results.
Programmed cell death protein 1 (PD‐1), expressed by various activated
immune cells, is an immune checkpoint and can protect against autoimmune
responses. It has been revealed that in cancer immunity, tumor cells
express PD-L1, the ligand of PD-1, and combine with PD-1 in T cells,
thereby suppressing T cell activities and promoting the differentiation
of Treg cells [23]. As a result, the anticancer immune responses are
inhibited in tumor microenvironment. The immunotherapies targeting
PD-1/PD-L1 have successfully improved the outcomes of various types of
cancer worldwide, including HCC [24]. A pioneering study analyzed
the PD-L1 expression in 240 HCC patients who received surgical
resection, and it was found that patients who were positive for PD-L1
showed significant shorter disease-free survival or overall survival
than those PD-L1 negative patients [25]. Antibodies of PD-1 or PD-L1
in HCC immunology have shown promising efficacies [26]. However,
there are still unmet clinical needs considering that many patients show
no response. Since one of the important mechanisms of PD-1/PD-L1 pathway
is the promotion of Treg cells differentiation, an interesting and
promising suggestion was drawn from this study that GSMs-TACE has the
potential to be used in combination with immune adjuvant therapies such
as PD-1/PD-L1 pathway targeting therapies to increase the efficacy of
HCC treatment.
In conclusion, the Treg cell proportion in peripheral blood of HCC
patients was higher than that of healthy controls and was closely
related to the clinical characteristics of HCC. The GSMs-TACE procedure
significantly reduced the peripheral Treg cell proportions at 10 and 30
days after operation. These results indicated that GSMs-TACE could
reduce the proportion of Treg cells in peripheral blood after operation
and exert a positive regulatory effect on the immune function of HCC
patients. This study provides a piece of preliminary evidence that
GSMs-TACE has the potential to be used in combination with immune
adjuvant therapies such as therapies targeting PD-1 or PDL-1 to increase
the efficacy of HCC treatment.
However, there are also some limitations of this study, and the
expression of other molecules or biomarkers involved in tumor immunity
should be assessed along with the Treg cell proportion to better explain
the mechanism of GSMs-TACE procedures affecting tumor immunity, so as to
provide references for the treatment design of HCC patients.