Histopathological Assessment of Tumor-Infiltrating Lymphocytes and Their Clinicopathological Associations in Breast Carcinoma- Cross Sectional Retrospective Study

Breast cancer is the most common cancer among women worldwide and is also a major cause of cancer-related deaths in India¹. In recent years, the number of breast cancer cases has been increasing, and many patients in developing countries present at a younger age and in advanced stages of disease. Although there have been improvements in diagnosis and treatment, breast cancer shows wide variation in behavior, even among patients with similar clinical and pathological features. This highlights the need for additional markers that can help in better understanding tumor biology.

Recent studies have shown that the tumor immune microenvironment plays an important role in the development and progression of breast cancer². Tumor-infiltrating lymphocytes (TILs) are immune cells present within and around the tumor tissue and represent the host immune response against cancer cells. These lymphocytes mainly include T cells, B cells, and natural killer cells. Their presence reflects the interaction between the immune system and tumor cells, and therefore TILs have gained importance as potential prognostic and predictive markers in breast carcinoma³.

To ensure uniform assessment, the International Tumor-Infiltrating Lymphocytes Working Group proposed standard guidelines for the evaluation of TILs on routine hematoxylin and eosin–stained sections⁴. Several studies have reported that higher levels of TILs are commonly associated with favorable clinicopathological features such as smaller tumor size, lower histological grade, and absence of lymph node metastasis. These findings have been observed more frequently in aggressive subtypes such as triple-negative and HER2-positive breast cancers5-7.

However, studies evaluating the association of TILs with clinicopathological parameters in Indian breast cancer patients are limited. Differences in genetic background, tumor characteristics, and healthcare practices may influence immune response patterns. Therefore, more data from the Indian population are required to better understand the role of TILs.

The present study aims to evaluate tumor-infiltrating lymphocytes in breast carcinomas and to correlate their grades with various clinicopathological characteristics, thereby assessing their usefulness as a simple and cost-effective parameter in routine histopathological practice.

This retrospective study was carried out on 157 cases of histologically confirmed invasive breast carcinoma diagnosed at a tertiary care teaching hospital. Relevant clinical and pathological details were collected from hospital records and pathology registers. The parameters studied included tumor size, histological grade, lymph node status, presence or absence of necrosis, and histological type of the tumor.

Formalin-fixed, paraffin-embedded tissue blocks were retrieved, and hematoxylin and eosin–stained sections were reviewed. Tumor-infiltrating lymphocytes were evaluated on routine H&E slides according to the recommendations of the International Tumor-Infiltrating Lymphocytes Working Group. Only stromal TILs within the invasive tumor area were assessed, while areas of necrosis, fibrosis, and in situ carcinoma were excluded from evaluation.

Based on the density of lymphocytic infiltration in the tumor stroma, TILs were graded into three categories: low, intermediate, and high. Each case was assessed carefully to ensure consistency in grading. Cases with incomplete clinicopathological data were excluded from specific analyses where required.

Statistical analysis was performed using the Chi-square test to assess the association between TIL grades and various clinicopathological parameters. A p-value of less than 0.05 was considered statistically significant.

Association of TIL Grade with Tumor Size (Table 1)

As shown in Table 1, TIL grade demonstrated a statistically significant association with tumor size (χ² = 17.33, df = 4, p = 0.0017). Higher TIL grades were more frequently observed in tumors measuring 2–5 cm and less commonly in tumors larger than 5 cm, indicating a favorable immune response in smaller tumors.

Table 1: Association of TIL Grade with Tumour Size (n = 157)

Chi-square test: χ² = 17.33, df = 4, p = 0.0017. Statistically significant association.

Association of TIL Grade with Histological Grade

Table 2 illustrates the relationship between TIL grade and histological grade. A highly significant association was observed (χ² = 48.46, df = 4, p < 0.001), with higher TIL grades predominantly seen in lower histological grades, suggesting an inverse relationship between tumor aggressiveness and immune infiltration.

Table 2: Association of TIL Grade with Histological Grade (n = 157)

Chi-square test: χ² = 48.46, df = 4, p = <0.001. Statistically significant association.

Association of TIL Grade with Necrosis

The association between TIL grade and necrosis is presented in Table 3. A statistically significant correlation was noted (χ² = 32.57, df = 6, p < 0.001). Higher TIL grades were more frequently observed in tumors without necrosis, while extensive necrosis was associated with lower TIL grades.

Table 3: Association of TIL Grade with Necrosis (n = 157)

Chi-square test: χ² = 32.57, df = 6, p = <0.001. Statistically significant association.

Association of TIL Grade with Lymph Node Status

Table 4 depicts the association between TIL grade and lymph node involvement. A significant relationship was observed (χ² = 27.26, df = 6, p < 0.001). Higher TIL grades were more commonly seen in lymph node-negative cases, whereas extensive nodal involvement correlated with lower TIL grades.

Table 4: Association of TIL Grade with Lymph Node Status (n = 131)

Chi-square test: χ² = 27.26, df = 6, p = <0.001. Statistically significant association.

Association of TIL Grade with Histological Type

As shown in Table 5, TIL grade varied significantly across different histological types (χ² = 100.77, df = 20, p < 0.001). Medullary and mucinous carcinomas demonstrated higher TIL grades compared to invasive ductal carcinoma, indicating a stronger immune response in these subtypes.

Table 5: Association of TIL Grade with Histological Type (n = 157)

Chi-square test: χ² = 100.77, df = 20, p = <0.001. Statistically significant association.

Overall analysis showed that tumor-infiltrating lymphocyte (TIL) grades had a statistically significant association with all the studied clinicopathological parameters. TIL grade was significantly related to tumor size, histological grade, presence of necrosis, lymph node status, and histological type. Higher TIL grades were more commonly observed in smaller tumors, lower histological grades, tumors without necrosis, and lymph node–negative cases. Certain histological subtypes also demonstrated higher TIL levels. These findings indicate that increased lymphocytic infiltration is associated with more favorable clinicopathological features in breast carcinoma.

Figure 1: TIL grade: Low

Figure 2: TIL grade: Intermediate

Figure 3: TIL grade: Hight

Tumor-infiltrating lymphocytes represent the host immune response against tumor cells and have gained increasing importance in breast carcinoma. In the present study, TIL grades showed a significant association with several clinicopathological parameters, indicating their potential role in reflecting tumor behavior.

In this study, higher TIL grades were more commonly seen in smaller tumors. Similar observations have been reported by Salgado et al. and Denkert et al., who found that tumors with higher immune infiltration were often associated with favorable tumor characteristics⁴˒⁵. This suggests that an active immune response may limit tumor growth.

A significant association was also observed between TIL grade and histological grade, with higher TIL grades seen in lower-grade tumors. This finding is in agreement with studies by Loi et al. and Vaid et al., who reported an inverse relationship between tumor grade and lymphocytic infiltration⁶˒⁷. Lower-grade tumors may provide a more favorable environment for immune cell infiltration compared to poorly differentiated tumors.

Lymph node status also showed a significant correlation with TIL grade. Higher TIL grades were more frequently observed in lymph node–negative cases. This observation is consistent with studies by Tsang et al. and Thanna et al., who reported lower rates of nodal metastasis in tumors with higher immune cell infiltration 8,9˒¹⁰.

The present study demonstrated that tumors without necrosis showed higher TIL grades. Similar findings were reported by Marchetti et al., who suggested that extensive necrosis may be associated with aggressive tumor behavior and reduced immune response¹¹.

Regarding histological type, medullary and mucinous carcinomas demonstrated higher TIL grades compared to invasive ductal carcinoma of no special type. This finding supports previous observations that certain histological subtypes are more immunogenic in nature.

Overall, the findings of this study support the usefulness of TIL grading as a simple histopathological parameter that correlates well with established clinicopathological features in breast carcinoma.

Tumor-infiltrating lymphocytes show a significant association with key clinicopathological characteristics of breast carcinoma. Higher TIL grades correlate with favorable prognostic parameters, suggesting their potential role as a simple, cost-effective prognostic biomarker in routine pathology practice.