The Clinical Outcomes Regarding to
Tumor-Infıltrating Lymphocytes in the
Breast Cancer Patients Treated with
Ebru Şen1*, Mehmet Emin Güneş1, Cengizhan Özdemir1, Merve Karli1, Ayşegül Gemici2 and Serdar Altinay3
1Department of General Surgery, University of Health Sciences Bakırköy Dr. Sadi Konuk Training and Research Hospital, Turkey
2Department of Radiology, University of Health Sciences Bakırköy Dr. Sadi Konuk Training and Research Hospital, Turkey
3 Department of Pathology, University of Health Sciences Bakırköy Dr. Sadi Konuk Training and Research Hospital, Turkey
Submission:May 15, 2020; Published: July 06, 2020
*Corresponding author:Ebru Şen, University of Health Science, Bakırköy Training and Research Hospital, General Surgery Department Tevfik Sağlam cad. No: 11, Zuhuratbaba mah, Bakırköy/ Istanbul/Turkey
How to cite this article:Ebru Ş, Mehmet E G, Cengizhan Ö, Merve K, Ayşegül G, et al. The Clinical Outcomes Regarding to Tumor-Infıltrating
Lymphocytes in the Breast Cancer Patients Treated with Neoadjuvant Chemotherapy. Open Access J Surg. 2020; 11(5): 555823. DOI:10.19080/OAJS.2020.11.555823.
Objective: Host immune system is a new research area of breast cancer treatment. Tumor-infiltrating lymphocytes (TILs) as an immunologic marker may influence the patients clinical outcome. The aim of this study was to explore the prognostic and predictive roles of TILs in our breast cancer patients treated with neoadjuvant therapy and to determine its association with other clinicopathological parameters.
Materials and Methods: Between 2011-2016, 89 patients were included in this study. The patients were categorized according to molecular subtypes. All patients’ parafin sections were retrospectively evaluated and according to TILs value patients were distributed into two groups in each subtypes.
Results: HER2 positive patients had higher TILs value than others. No relationship was seen between TILs levels and presence of either locoregional recurrence or systemic metastasis. Both in survival outcomes and prediction of neoadjuvant therapy response no association was detected according to TILs value within all subtypes.
Conclusion: The relevance of TILs in small patient population is challenge. Further research is warrented.
Keywords:Tumor-infiltrating lymphocytes; Breast cancer; Response to neoadjuvant therapy
In breast cancer great endeavors are going on to find new markers in determining the prognosis. In addition to well-known prognostic factors such as tumour size, grade, nodal status, molecular subtype, proliferation index, gene-expression based recurrence score, recently there is increasing evidence that tumor-infiltrating lymphocytes (TILs) are of important role as an immune biomarker in breast carcinoma [1-3]. TILs might be a reflection of host immune response which plays an important role in cancer control and patient’s clinical outcome. TILs are mononuclear immune cells that infiltrate tumour tissue and mainly comprised of cytotoxic (CD8+) T cells as well as macrophages, helper (CD4+) T cells, B cells, and NK cells [4,5]. There are two types of TILs: Stromal TILs (sTILs) are defined as the percentage of tumour
stromal field in which lymphocytes are located without direct connection to tumour cells. In contrast, intratumoural TILs reflect immune cells within the tumour cells. Stromal TILs are commonly detected in H&E stained sections by light microscopy, and consist the majority of TILs in breast cancer . Its assessment is the most reproducible parameter by pathologists. Moreover, lymphocytes-predominant breast cancer (LPBC) is defined as those cancer with TILs occupies at least 50 % to 60 % of the stroma .
The clinical importance and amount of TILs appear to vary significantly among different subtypes of breast cancer [2,6]. TILs are more common in triple-negative breast cancer (TNBC) and HER2-overexpressing breast cancer, with LPBC frequency of 15-25 % in these groups. Contrary, luminal tumour is the least immune infiltrated breast cancer subtype [3,6,7]. Large
series suggest that tumour with higher TILs levels are associated
with improvement of prognosis by longer survival rates and
with reduction of locoregional recurrences [8,9]. This positive
prognostic effect is especially striking on TNBC and HER2 positive
patient populations [6,10]. In addition to prognostic value of
TILs, trials also indicate the predictive role of TILs in response
to neoadjuvant chemotherapy [11,12]. The aim of this study
was to determine the association between TILs level and other
clinicopathological parameters and to explore the prognostic and
predictive roles of TILs in our breast cancer patients treated with
The study was conducted to determine the association of
TILs value with the clinical outcomes at neoadjuvant settings.
Breast cancer patients treated after neoadjuvant chemotherapy
at Bakırköy Sadi Konuk Training and Research Hospital from
July 2011 to January 2016 were included in this study. All were
followed up in our hospital. Patient’s demographic information,
characteristics, outcomes and follow-up were obtained from
hospital records. The study protocol was approved by the ethics
committee of our institute (No:2017/09-20). Neoadjuvant
therapy were taxane and anthracycline-based protocol, adding
transtuzumab in the HER2 positive subtypes. Surgical treatment
was decided as to treatment response. Radiotherapy and endocrine
therapy were given in accordance with institutinal guidelines.
Inoperable patients following neoadjuvant therapy were excluded
of this study. The patients were categorized according to molecular
subtypes as triple negative (TN), HER2 positive and luminal type.
All patients in each subtypes were distributed into two groups
according to TILs, with 10 % of cut-off value. In order to determine
TILs value all patients’ paraffin sections stained hematoxilyn and
eosin (H&E) from core biopsies were retrospectively assessed by
an experienced pathologist (S.A.), blinded of clinical information.
Briefly, all mononuclear cells (including lymphocytes and plasma
cells) in the stromal compartment within the borders of invasive
tumour were evaluated and reported as a percentage value. pCR
was defined as ypT0/Tis ypN0.
We analysed associations of TILs with other clinicopathological
factors such as menopausal status, tumour molecular type, ypTNM
classification, histology, grade, lymphovascular and perineural
invasion. In order to evaluate the predictive role of TILs, we
investigated pathological responses to neoadjuvant therapy in
high and low TILs groups of each subpopulation. To determine
the prognostic role of TILs we recorded the patients’ locoregional
recurrence and systemic metastasis in all groups and detected
overall survival and disease free survivals. Statistical analysis
was carried out using NCSS 11 package program. For categorical
variables the frequency and percentage values were calculated
and for continuous variables, average and standard derivation
values were calculated. Standard distribution sort of variables
was made with Kolmogorov-Smirnov test. Student t test was used
in the comparison of two groups and single direction variance
analysis was used in the comparison of more than two groups for
variables exhibiting standard distribution. Disease-free survival
(DFS) and overall survival (OS) were obtained using Kaplan-Meier
survival analysis. The relations between categorical variables
were examined with Chi square test, Fisher exact probability
test and Fisher-Freeman-Halton test. P<0,05 was accepted as
The study consisted of 89 breast cancer patients who
underwent neoadjuvant chemotherapy. The mean follow-up
duration of all patients was 33,8 (15-84) months. According
to molecular subtypes, 39 patients had luminal tumour, and 25
patients each triple negative and HER2 positive.
The mean TILs values were 10.8%, 15.4%, 6% in TN, HER2
positive and luminal groups, respectively. Among the patients
of high TILs levels, HER 2 positive group was the most frequent
molecular subtype (50 %), followed by triple negatives (32%)
(Table 1). Sixty percent of patients with TILs less than 10 % had
luminal tumour. There was statistically significant relationship
between TILs value and both molecular type of tumour
(p=0.0001) and presence of lymphovascular invasion (p=0.048).
No association was detected between TILs and other variables
such as menopausal status, ypTNM classification, histology, grade,
and perineural invasion (Table 2).
No association was seen in any subtypes between TILs levels
and presence of either locoregional recurrence or systemic
metastasis (Table 3). Neither OS nor DFS were significantly
different between high and low TILs groups in both TN and HER2
positive groups. Because there was no death in luminal group, we
could not detected survival analyses in this subtype.
In the neoadjuvant setting, no interaction was found between TILs level and complete pathological response according to
each subtypes (Table 4). But, there was non-significantly better
pathological response rates in high TILs group of HER2 (+)
The constant interaction between tumour and host immunity
influences patient’s outcome. Innate and adaptive immune
response in the tumour milieu require functional cytotoxic T-cells
in order to achieve efficent tumour elimination . The impact
of TILs on the prognosis and treatment prediction suggests
its importance on the reflection of individual immunity . Various studies of TN and HER2 (+) breast cancer have pointed
consistent results that the increased TILs value the patients have,
the better OS and DFS in the adjuvant setting [6,14]. Moreover, the
highest TILs also show the highest expression of T-cell checkpoint
receptors which found on T cells . Antitumour immunity is
enhanced by monoclonal antibodies against checkpoint inhibitors.
In addition to standard therapeutic options, immunotherapy in
breast cancer is a new and promising implication especially in
selected patient population. Therefore, TILs evaluation may be
accepted as a biomarker for immunotherapeutics in patients with
breast cancer at near future.
In the light of these rapidly evolving field, in 2014 international
TILs working group has recommended TILs evaluation as a
immune parameter along with standard histopathological practise
to facilitate its use in research setting and clinical trials . They
reported TILs measurement as a percentage of stromal TILs on
a continuous scale by visual assessment of H&E-stained sections
of primary tumour specimens similar to any other quantitative
histological biomarker. Several studies have evaluated lymphocyte
subtypes by using immunohistochemical analysis, yet added value
of immunohistochemistry on to TILs evaluation remains to be
demonstrated [4,15]. Moreover, digital image analysis of TILs can
provide more exact measurement and eliminate inter-pathologists
differences in TILs assessment . Visual measurement of TILs
is prefered at many trials like our study [4,6]. Because this route is
stated as rapid, easly available, less expensive and adequate tool
in TILs assessment. There is no established threshold for high
TILs. Various studies used different cut-off values [7,9,16,17].
Moreover, 50-60 % of TILs value for lymphocytes-predominant
breast cancer had been arbitrarly chosen . In this study, ROC
Curve was applied for the estimation of pathological response
of TILs values, but a significant breakpoint could not be found
(unreported data). Therefore, we accepted 10 % of TILs level as a
cut-off value similar to many studies [6,18].
In our study, we investigated the relation of TILs value with
other clinicopathological parameters and its impact on patients
with all molecular subtypes of breast carcinoma who were
operable following neoadjuvant treatment. The positive relation
of higher quantities of TILs and both negative steroid receptor
status and presence of HER2 positivity is striking in the literature
like in our study . Also LPBC is commonly found in HER2
positive and triple negative subtypes [4,19]. Similarly, in our study
HER2 positive group was the main molecular subtype (50%) that
harbours the high percentage of TILs. But, interestingly unique
patient with LPBC was seen in our series. In this study, patients
with luminal type breast cancer had lower TILs value, with the
median of 6%. Similar to litreture, no significant prognostic role
of TILs was found in patients of our luminal group . According
to the trial by Huszno  higher grades of TILs were present more
frequently in younger patients than older women (47% vs. 24%).
Moreover, higher TILs levels was appeared to be associated with
higher histological grades. On the other hand, TILs value also
was significantly correlated with lymphovascular invasion in our
The positive association between TILs levels and favourable
clinical outcomes was based on Sistrunk’s cohort trial at 1922 .
Eastern Cooperative Oncology Group’s phase III trials showed that
each 10 % increase in TILs levels significantly predicted better
DFS and OS in TNBC with a median follow-up of over 10 years .
A recent meta-analysis of twenty-five trials including more than
twenty thousands patients confirmed the survival benefit of TILs
in TN and HER2 positive breast cancer patients . Moreover,
according to secondary analysis of the NeoALTTO trial reported
by Salgado  patients with high TILs levels at baseline had
better outcomes for event free survival independent of whether
they achieved pCR in the neoadjuvant phase. The association
between higher levels of TILs and decreased distant recurrence
rates in TN breast cancer was found in FinHER trial .
Contrary to obvious results as to prognostic impact of higher
TILs levels on the adjuvant and neoadjuvant settings, we could not
detected any significant relationship between TILs and survival
analysis. Another issue related to TILs is its utility in prediction of
adjuvant and neoadjuvant therapy response . At the adjuvant
setting, the association between higher levels of TILs and increased
transtuzumab benefit in HER2 (+) subtype was detected in phase
III trial including 1010 early stage BC patients . Positive
correlation between TILs levels and pCR rates was prominent
in HER2 (+) tumours. According to the study of Carbognin 
pCR rates were increased by nearly 30 % in the presence of TILs
reported as LPBC. Another trial has also indicated that the level of
TILs greater than 5% was associated with higher pCR rates .
Although we could not detected significant correlation between
TILs value and pCR in our study, there were non-significantly
higher pCR rates in HER2 (+) group. The minority of our patient
population explains the limited power of the statistical analysis.
Host immunity controls cancer and influences patient’s
outcome. How can a host enhance anti-tumor immune response
and which biomarker provides better treatment approach are the
main questions. Although which level of TILs should be used to
determine optimal treatment strategy for breast cancer patients
has not been certain yet, many trials indicate that TILs is of
prognostic effect in long-term disease control and predictive effect
of a better local response to treatment. But, further resaerchs are
warrented before the utility of TILs as an immune biomarker for
routine clinical practise of breast cancer patients.