4 Discussion
The interactions of PD-L1 binding to its inhibitory receptor PD-1
inactivate the T cells recognizing the antigen of tumour cells, and
consequently, the generation of population of cytotoxic T lymphocytes is
reduced that provide an opportunity to cancer cells for escaping immune
surveillance. This finding has triggered research to shift treatment
from targeting molecules directly on the surface of cancer cells to
non-contact methods for blocking the PD-L1/PD-1 pathway for better
activation of the immune system4. An increasing number
of studies have confirmed that the application of PD-1/PD-L1 treatment
efficiently enhance antitumor immunity that reduces tumour growth and
improves survival, whereas the prognostic roles of PD-L1 positivity in
OSCC are controversial.
Our results revealed a significant association between PD-L1 expression
levels and clinicopathological factors. The levels of PD-L1 expression
in TCs were relatively high in female patients, non-smoker,
non-drinkers, which in line with previous findings of meta-analysis
reported by Lenouvel et al52. Consumption of alcohol
and tobacco have been proved to be positively associated with OSCC
recurrence and poor prognosis, but have no significant association with
PD-L1 expression on TCs4. Previous study using PD-L1
antibody to detect the PD-L1 expression found that non-smokers or
non-alcohol consumers express higher frequency of PD-L1
expression12. Interestingly, the protective effect of
PD-L1 expression was observed in females for OS and DSS, which did not
weaken with smoking, or drinking1. It has been
reported that PD-L1 is overexpressed in never-smokers and never-drinkers
(NSND) who undergo the immunosuppressive therapy with pembrozulimab. The
enrichment of T-cell activation, interferon-γ(IFN-γ) and PD-1 signalling
were observed in OSCC from NSND. Overexpression of PD-L1 induced by
IFN-c, the unique feature in NSND, was associated with rate of TILs. It
could be inferred that PD1/PD-L1 blockade enhanced the immunity response
that provided potential benefit of PD1/PD-L1 inhibition in
NSND39. In addition, our meta-analysis also revealed
that PD-L1 upregulation in TC was associated with clinicopathological
features involving advanced stage, N+ status, in favour of the findings
of previous reports19,24,28,30.
To date, the prognostic value of PD-L1 in OSCC cells has been identified
in several meta-analysis. Consensus of them indicated that expressions
of PD-L1 in TCs had protective effects on OS with no
significance52-56, which was different from our study
identifying prognostic role of PD-L1 expression in TCs was insignificant
and unfavorable. As for DFS, four meta-analysis suggested that
expression of PD-L1 was associated with better survival regardless of no
statistical significance53-56. However, a previous
study hold the opposite perspective that expression of PD-L1 probably
lead to reduced survival52. In terms of DSS, our
results revealed that PD-L1 expression exhibited a trend towards worse
survival on DSS, which in line with the results reported by Troiano et
al and Lenouvel et al.52,56 There was a significant
association for worse LRFS in tumors characterized by high PD-L1
expression on TCs in our study. Geum et al. suggested locoregional
recurrence had a significantly lower survival rate, and simultaneously,
was significantly correlated with PD-L1 expression3.
The remarkable advantages of this study was not only the abundant
studies that was already enrolled in previous meta-analysis, but also in
the first exclusively comprehensive evaluation of the oral compartments
and immunostaining patterns as confounding factors for prognostic role
of PD-L1 expression in OSCC.
The role of PD-L1 in the prognosis of OSCC have been undetermined due to
expectable disparities in tissue of origin, location of staining,
variations in immunostaining patterns (ie, assay cutoffs, antibody
clones)9. In our study, high PD-L1 expression in TCs
was associated with an disoperative effect on prognosis in DFS when only
Asian patients of OSCC were analyzed. In OS and DSS of Asian patients,
high PD-L1 expression on TC also had a tendency to increase the risk of
poor prognosis. This might be attributed to the discrepancies in diet
and lifestyle between the Asia and the non-Asia.
Of the selected studies, it was not difficult to find that tongue as the
subsite of OSCC was more common than other subsite, suggesting that
tongue might be an important site affecting the prognosis and
clinicopathological factor of OSCC. In this study, we firstly compared
the PD-L1 expression in TCs between tongue and other sites of OSCC
patients, and the results showed no significant difference of any
clinicopathological factors between the two groups. Next, we pooled the
studies restricted only to tongue of OSCC with respect to prognosis and,
found a significant relationship between the high expression of PD-L1 in
TCs and worse DSS and OS, and the heterogeneity simultaneously decreased
from 81% to 0%. Previous study have demonstrated that PD-L1
overexpression in TCs of the tongue and the floor of the oral cavity was
associated with with a worse OS1. It was reasonable to
speculate that discrepancies of the results in regard to the role of
PD-L1 expressions in prognosis was attributed to skewed distribution of
cancer location.
The use of different antibodies clones for immunohistochemistry resulted
in discrepant results of association between PD-L1 expression and
prognosis in OSCC. A trial in lung cancer comparing four anti-PD-L1
antibodies (22C3, 28-8, SP142, E1L3N) found that the patients detected
as PD-L1-positive cases was similar for all antibodies except for SP142
which was only 50% as much as the other three of
them57. Interestingly, SP142 showed more sensitive
detection efficiency than 22C3 in another trail lung
cancer58. This study extracted antibody of PD-L1
including 22C3, 28-8, 5H1, E1L3N, SP142 as different subgroups. The
results indicated that OS was worse in 5H1, but greater in 22C3 in OSCC
patients with high PD-L1 expression in TCs. A study using two different
anti-PD-L1 antibodies (clones E1L3N and 22C3) to evaluate the prognostic
significance of tumor PD-L1 expression in OSCC. Consequently,
significant results of DSS only appeared in 22C3 but not in
E1L3N12, in contrast to our findings demonstrating DSS
was more likely to get worse with high PD-L1 expression detecting by
E1L3N. It was not ignored that the positive expression ratio of tumor
PD-L1 expression in OSCC tested by E1L3N was lower than 22C3 whether the
cut off is ≥ 1% or ≥ 10%12. Subgroup analysis of our
study revealed that almost no intra-heterogeneity was found in group of
E1L3N and 22C3 in aspects of OS, DFS. Combined with our results, in
OSCC, it can be believed that 22C3 seemed to be more delicate than
E1L3N, and the PD-L1 expression in TCs appeared to have the protective
role in OS via 22C3 as well as a damaging effect on DSS via E1L3N.
Therefore, using different antibodies to evaluate the relationship
between PD-L1 expression and prognosis probably lead to various results.
Due to the lack of studies comparing the various antibodies specific to
OSCC, our results needed to be interpreted with caution and were
expected to be validated by more high-quality studies.
We evaluated the influence of PD-L1 expression on prognosis by using
separate scoring methods. In this study, both DSS and DFS showed a
significant result at a 5% cut-off of PPC. As for OS, 5% cut-off of 4
scores cut-off of H score presented a reduced survival. The cut-off
value of 5% was frequently chosen in many clinical trials focusing on
targeted anti-PD-L1 therapies. PD-L1 expression was associated with
worse prognosis for OS and LRFS when a 5% cut-off of positive cells was
applied28. Previous study aimed to explore the effects
of expression of PD-L1 on survival rates and showed that PD-L1
expression were unrelated to in DFS and OS with TPS using different
cutoffs of 1%, 5% and 10%2, in support of the data
reported by Wirsing et. al22. One
study9, using cutoffs of 5% of TPS, PD-L1 positivity
was significantly associated with a better prognosis in DSS and OS, but
this association was disappeared when confounded other
factors8. PPC is defined as PD-L1-staining cells (TCs
or TILs) divided by the total number of each type of cells, of which
difference from TPS or CPS is that the denominator of the calculation
formula of them is the total number of viable TCs. Therefore, a lower
PPC score than TPS or CPS did not accurately represent the effect of
immune checkpoints on TCs under the same conditions. Unlike with TPS,
CPS reflecting an aggregate score of TC and IC evaluated expression of
PD-L1in TC, as well as the impact of different IC on the tumor
microenvironment. A randomized three-arm phase III KEYNOTE-048 trial was
conducted in head and neck cancers (HNSCC)
immunotherapy59,60, PFS and OS were tested in three
groups including the CPS ≥20, the CPS ≥1, and the total population. In
comparison with EXTREME regimen, patients undergoing the treatment of
pembrolizumab plus chemotherapy or pembrolizumab monotherapy
significantly improved OS and PFS, and the protective effect was
progressively declining in the three groups. The percentage of HNSCC
tumor cells expressing PD-L1 was 85% of when CPS was
≥159, which decreased to 50% when measured using
TPS61. The response rate of pembrolizumab seemed to be
increased in higher levels of PD-L1 expression, and CPS was more
excellent in ability to predict response to anti-PD1 therapy comparing
with TPS62. This prompted the FDA to approve the use
of pembrolizumab monotherapy against HNSCC only in patients with a CPS
score ≥1, which drew interest from two studies focusing on the
prognostic value of PD-L1 expression recorded by TPS or CPS in OSCC. One
study11 revealed the protective effect of PD-L1
expression was found on DSS and OS when positivity based on TPS
(>5%) was used, and was observed on OS with the positivity
defined as CPS >1. In the other study9,
no statistical significance was found in the association between PD-L1
and survival in either CPS or TPS. Patients with CPS>1
showed a trend towards improved survival, while TPS>1%
seemingly represented opposite trend. It need to be emphasized that
tumours were classified into into four groups based on the presence of
PD-L1 positivity in TC and IC, and the total prevalence of PD-L1
expression in both TC and IC or only in TC account for 72% in OSCC and
showed good response to immunotherapy. Consequently, it should consider
the combination of PD-L1 expressing in both TC and IC when observing
PD-L1 of response to anti-PD1 therapy.
Immune cells as components of the immune environment, including CD4+
,CD8+ TILs, have shown to be correlated with PD-L1 expression and play
an important role in the mechanisms of immune response evasion of
OSCC63. Several studies reported PD-L1-expressing
tumor cells correlated positively to increased infiltration of CD4+ and
CD8+TILs2,8,22,40, whereas other studies did not show
any correlation11,25. Our study indicated that the
PD-L1 high expression group was significantly associated with high
infiltration by PD-1, CD8+ TILs, strengthening that the activity of
CD8+TILs was reduced through interaction of PD-L1 expression in TCs
activating expression of PD-1 on CD8+ TILs. In recent years, TILs were
gradually considered as a predictor for the response of solid tumors to
anti-PD1 therapy64,65. However, the prognostic value
of PD-L1 in TILs was not fully understood. Our study on prognosis showed
that the PD-L1 expression on TILs did not reveal a relationship with
survival, which was consistent with a previous
study11. Inversely, one study reported that longer OS
and LRFS were appeared in the group of overexpression of PD-L1 in
TILs35. Subramaniam et al.49reported that low TIL PD-L1 expression was
significantly correlated with reduced LRFS, but this association was
disappeared confounded other factors. This was consistent with what was
reported previously in a study that high expression of PD-L1 in TILs was
associated with better OS of OSCC without the confounding factors of
nodal metastases1. It has been called adaptive immune
resistance that a high TIL infiltration appeared simultaneously with
PD-L1 positive TCs. The strong correlation between TILs and tumor PD-L1
staining implied PD-L1 may have been induced via enhanced T cell
production of IFN-γin the same way as in tumour
cells7.
Our study has several limitations. First, the prognostic values of
several studies were estimated rather than provided directly. The actual
association between PD-L1 expression and prognosis was masked on account
of potential confounding factors. Second, the evaluation of the role of
PD-L1expression in TILs on DSS, DFS, PFS and LRFS were not available to
performed as a result of the lack of relevant data.
Conclusions
Current study indicated the expression of PD-L1 in TCs was relatively
high in female patients, non-smoker, non-drinkers. The PD-L1
upregulation in TCs in cancers of advanced stage, N+ status was
observed. The significant association of high PD-L1 expression in TCs in
prognosis was only seen in LRFS. Although high PD-L1 expression in TCs
had a trend toward significance of worse prognosis in OS, DSS and DFS in
the initial analysis, the associations became significant based on
stratified analysis of hypothesized confounders. It was worth noting
that high expression of PD-L1 in TCs of tongue of OSCC was associated
with worse DFS, OS. Future research on the ability to predict response
to anti-PD1 therapy should focus on methods for PD-L1 immunostaining and
various compartments of the oral cavity. In addition, TILs were shown to
be independent of clinicopathological factors and prognosis, and
relevant studies were expected to further confirmed.