Prognostic Value Of Tumor-infiltrating Lymphocytes Research Articles

The prognostic value of tumor-infiltrating lymphocytes in head and neck squamous cell carcinoma: A systematic review and meta-analysis

Head and neck squamous cell carcinoma (HNSCC) is experiencing a rising incidence and mortality worldwide, emphasizing the need for reliable prognostic markers. Tumor-infiltrating lymphocytes (TILs) have emerged as a promising biomarker for predicting HNSCC prognosis, yet no systematic reviews have exclusively focused on hematoxylin and eosin (H&E)–stained formalin-fixed paraffin-embedded (FFPE) samples, which are routinely used in clinical practice. This systematic review and meta-analysis followed the PRISMA guidelines to examine the prognostic value of TILs in HNSCC using H&E-stained FFPE samples. Data were pooled from 43 studies, including 26 studies in a meta-analysis, analyzing 5037 HNSCC samples. We found that a high TIL count associated with a significantly improved overall survival (OS) (HR 0.47, 95 % CI 0.41–0.55, p < 0.0001), disease-free survival (DFS) (HR 0.55, 95 % CI 0.41–0.55, p < 0.0001), and disease-specific survival (DSS) (HR 0.58, 95 % CI 0.46–0.73, p < 0.0001). The heterogeneity was moderate for the pooled analysis (OS: I² = 40 %; DFS: I² = 39 %; DSS: I² = 51 %), but low for the subgroup analysis based on tumor site in oral, oropharyngeal, laryngeal, and nasopharyngeal cancer (OS and DFS: I² = 0–14 %). This review is the first to systematically evaluate TILs in HNSCC using H&E-stained samples, confirming their prognostic value. A high TIL count is associated with improved survival outcomes, suggesting their potential as prognostic biomarkers in clinical settings.

Prognostic value of tumor-infiltrating lymphocytes and PD-L1 expression in esophageal squamous cell carcinoma.

Tumor cells (TC) participate in tumor progression by altering the immune responses in the tumor microenvironment. However, the clinical relevance and prognostic effect of PD-L1 expression and tumor-infiltrating lymphocytes (TILs) in esophageal squamous cell carcinoma (ESCC) are unknown. The purpose of this study was to investigate the interactions and clinical significance of PD-L1 expression and TILs in ESCC. Tissue specimens were collected from 126 patients with ESCC who underwent curative esophagectomy. Immunohistochemical analysis and multiplex immunofluorescence for CD4, CD8, CD25, FOXP3, and PD-L1 in the tumor were used to identify multiple tumor-infiltrating immune cells (TIIC), Tregs, and TC. PD-L1 was expressed in tumor cells (PD-L1 TC). PD-L1 TIIC and PD-L1 TC affected the biological behavior of TC. The positive expression rate of PD-L1 TC and CD8+ TILs was 27.8% (35/126) and 31.7% (40/126), respectively. Kaplan-Meier analysis showed that overall survival (OS) was significantly associated with decreased CD8+ TILs and PD-L1 TC-positive expression, which promote ESCC progression and metastasis. Tumor depth, CD8, and PD-L1 TC were independent prognostic factors in ESCC, and a predictive nomogram with these three risk factors improved the accuracy of predicting OS in patients with ESCC after surgical resection. The conjoint analysis of multiple immune-related factors is beneficial for stratifying patient survival risk.

Association of tumor-infiltrating lymphocytes with clinical outcomes in patients with triple-negative breast cancer receiving neoadjuvant chemotherapy: a systematic review and meta-analysis.

Triple-negative breast cancer (TNBC) presents a clinical challenge as an aggressive tumor, correlated with unfavorable prognosis. Tumor-infiltrating lymphocytes (TILs) have garnered interest as a potential prognostic biomarker. However, the disparity in outcomes between varying TILs rates remains inadequately explored. PubMed, Scopus, Web of Science, and Cochrane databases were searched for studies about the prognostic value of TILs in patients with TNBC receiving neoadjuvant chemotherapy. The hazard ratios (HRs) or odds ratios (ORs) were computed for binary endpoints, with 95% confidence intervals (CIs). Twenty-nine studies were included, involving a population of six thousand one hundred sixty-one (80.41%) with TNBC. The cut-off TILs value ranged from 10 to 60%, with 50% being the most related value. Compared with the low-TIL expression group, the disease-free survival (DFS) (HR 0.71; 95% CI 0.61-0.82; p < 0.00001) and overall survival (OS) (HR 0.76; 95% CI 0.63-0.90; p = 0.002) rates showed significant improvement with higher TIL infiltrations. In the subgroup analyses of the lymphocyte subtypes CD4 + and CD8 + , there was statistical significance favoring higher TILs rates in both subtypes, each associated with improved DFS (HR 0.48; 95% CI 0.33-0.71; p = 0.0002) and OS (HR 0.53; 95% CI 0.36-0.78; p = 0.001), regardless of which cell subtype was predominantly infiltrated. The complete pathological response analysis showed better rates for the higher TIL group than the control for both the TIL (OR 1.29; 95% CI 1.13-1.48; p = 0.0003) and Ki-67 (OR 2.74; 95% CI 2.01-3.73; p < 0.00001) analyses. Higher expressions of TILs in patients with TNBC were associated with improved significantly DFS, OS, and pCR outcomes.

Prognostic implications of tumor-infiltrating lymphocytes within the tumor microenvironment in gastric cancer

Tumor-infiltrating lymphocytes (TILs) play a regulatory role in the tumor-associated immune response and are important in the prognosis and treatment response of several cancers. However, because of its heterogeneity, the prognostic value of TILs in gastric cancer (GC) is still controversial. Thus, this study aimed to investigate the association between the density of TILs and patients' outcomes in GC. Patients with gastric adenocarcinoma who underwent curative intent gastrectomy were retrospectively investigated. The groups for analysis were determined on the basis of TIL intensity and percentage of CD3+T-cell infiltration by immunohistochemical. Furthermore, Epstein-Barr virus (EBV), microsatellite instability (MSI), T-cell ratio of CD4 to CD8, and programmed death protein ligand 1 (PD-L1) status were evaluated. A total of 345 patients were enrolled: 124 patients with GCs (35.9%) were classified as the low-CD3+TIL group, and 221 patients with GCs (64.1%) were classified as the high-CD3+TIL group. Poorly differentiated histology (P=.014), EBV-positive status (P<.001), PD-L1-positive status (P=.001), and CD4<CD8 (P<.001) were associated with high-CD3+GC. There was no difference regarding MSI status, the degree of tumor invasion (pT), the presence of lymph node metastasis, and pTNM stage between low- and high-CD3+groups. In survival analysis, the high-CD3+group had better disease-free survival and overall survival rates than had the low-CD3+group (P=.055 and P=.041, respectively). In the multivariate analysis, total gastrectomy, lymph node metastasis, advanced pT stage, and low CD3+levels were independent factors related to worse survival. High CD3+TILs levels were significantly associated with improved survival and could serve as prognostic biomarkers in GC. In addition, CD3+T-cell infiltration was related to both EBV-positive and PD-L1-positive GC and may assist in the investigation of targets in immunotherapy.

Clinicopathological and prognostic value of TIL and PD L1 in triple negative breast carcinomas

Triple negative breast cancer (TNBC), a highly aggressive subtype of breast cancer, accounts for 15 % of all diagnosed breast cancers. This group, which has the worst clinical outcome, high recurrence rate and poor prognosis, does not benefit from specific treatment. Therefore, there is a need to develop more effective biomarker and therapeutic strategies especially for this group. A positive level of immunity has been found to be associated with patient survival in various organ cancers. More specifically, tumor infiltrating lymphocytes (TIL) have been documented to have strong prognostic value. The programmed cell death 1 (PD 1) protein on the surface of T lymphocytes is activated by the Programmed cell death ligand 1 (PD-L1) protein on the cancer cell surface. PD- L1 is thought to form a pathway that results in suppression of antitumor responses when activated. Patients with breast cancer (BC) who underwent resection without neoadjuvant chemotherapy between 2010 and 2020 were included in this study. Of the 302 BCs examined, 21 constitute the group with TNBC. In our study, the mean age of the Triple positive breast cancer (TPBC) and TNBC groups was similar (55.67 ± 12.61 vs. 53.23 ± 8.21, p = 0.384). There was no significant correlation between TPBC and TNBC and tumor size, lymph node, histological grade, and PD-L1 positivity in the center of the tumor (all p-value >.05). It was observed that tumor stage was higher in patients with TNBC than in patients with TPBC (19 % vs. 1.1 %, p = .002). The Ki 67 proliferation index was found to be higher in patients with TNBC than in patients with TPBC (90.5 % vs. 41.8 %, p .001). Although not statistically significant, clinically, CD 3 and CD 8 immune scores with high tumor margin were higher in patients with TNBC than in patients with TPBC (90.4 % vs, 9.6 % and 85.7 % vs. 14.3 %, respectively). Positive expression of PD-L1 at the tumor margin was significantly higher in patients with TNBC than patients with TPBC (20.3 % vs, 52.4 %, p = .002). By Kaplan-Meier analysis, the survival distribution of CD 3 and CD 8 immunoscore, tumor central and margin PD-L1 values were compared. Mean follow-up was 136.18 months (range, 1 – 144 months); and the 10-year Overall Survival (OS) estimate for the population was 90.9 % (95 % CI, 85.5 – 96.7). In this study, this difference was not statistically significant according to the log-rank test. In this study, we aimed to evaluate the relationship between CD 3, CD 8 T lymphocyte immune score and PD-L1 expression at the tumor center and margin in TNBC, the prognostic value and clinicopathological significance of this relationship.

Deep learning based quantitative models to distinguish tumoral and stromal tumor-infiltrating lymphocytes and the prognosis patients with high-grade serous ovarian cancer.

84 Background: The better prognostic value of tumor-infiltrating lymphocytes (TILs) has been reported in high-grade serous ovarian cancer (HGSOC). However, the TILs evaluation of previous reports largely depends on manual quantification based on a visual assessment within limited section of pathological slides or tissue microarrays, which is subject to inter-observer variability. Since tumor immune microenvironment (TIME) of HGSOC is highly heterogeneous, there is a need to quantify TILs in large areas of pathological slides in an objective and time-saving manner. Here we present an AI-powered quantification method to distinguish tumoral TILs and stromal TILs using hematoxylin and eosin (H&amp;E)-stained whole-slide images (WSIs) of HGSOC. Methods: 161 H&amp;E stained WSIs of HGSOC patients treated at our hospital between 1998 and 2021 were prepared, and by using these WSIs we generated two AI-powered models. Initially, a TILs detection model (TIL-model) was created using a publicly available dataset of 86104 pan-cancer TILs images of 100 × 100 pixels at magnification of x200 (Saltz et al., 2018, Cell Reports). Secondary, a tumor and stroma segmentation model (T/S seg-model) was developed using 136962 patches of 224 × 224 pixels at magnification of x50 which were manually annotated with tumoral regions and stromal regions in 102 H&amp;E stained WSIs of HGSOC from The Cancer Genome Atlas (TCGA). Deep convolutional neural networks of NASNet-Large for the TIL-model and TransUNet for the T/S seg-model were used. TILs density score (number of TILs positive tiles per tumor area) of each sample was calculated, and association between TILs density score and the prognosis was investigated. Results: The established models achieved high accuracy: the TIL-model showed an area under the curve (AUC) value of 0.896, and the T/S seg-model showed an intersection over union value of 0.967. By combining the two models, we successfully distinguished tumoral TILs and stromal TILs. Using our dataset of 161 WSIs, we split the patients by TILs density score at top quartile into high tumoral TILs subgroup (n=40) and low tumoral TILs subgroup (n=121). High tumoral TILs subgroup showed significantly longer overall survival (OS) and progression free survival (PFS) (OS: median 70 months vs not reached, p &lt; 0.005. PFS: median 34 months vs 21 months, p = 0.02). Conclusions: We developed deep learning based pathological AI models to detect TILs and tumors using H&amp;E stained WSIs, which achieved to quantify tumoral and stromal TILs. The objective assessment of TILs using these models will lead to the development of clinically applicable tools for the evaluation of the immune status in HGSOC.

23P Prognostic value of tumor-infiltrating lymphocytes (TILs) and soluble immune checkpoints in early triple-negative breast cancer (TNBC)

23P Prognostic value of tumor-infiltrating lymphocytes (TILs) and soluble immune checkpoints in early triple-negative breast cancer (TNBC)

Combining the guidelines-method and multiphoton imaging method to improve the prognostic value of tumor-infiltrating lymphocytes in breast cancer.

Multiphoton microscopy (MPM) was introduced to label-freely obtain tumor-infiltrating lymphocytes (TILs) images from a total of 611 patients, and the prognostic value of TILs in breast cancer was assessed by the MPM method (TILs-MPM) and guidelines-method proposed by the International Immuno-Oncology Biomarker Working Group (TILs-WG), respectively. Moreover, the clinical (CLI) model, TILs-WG+TILs-MPM model, and full model (CLI+TILs-WG+TILs-MPM) were developed to investigate the prognostic value of TILs. The results show that TILs-WG performs better in ER-negative subgroup, and TILs-MPM is comparable with TILs-WG in the ER-negative subgroup, but has the best performance in the ER-positives subgroup. Furthermore, the TILs-WG+TILs-MPM model can significantly improve the prognostic power compared with the TILs-WG model, and the full model has excellent performance, with high AUC and HR in both ER-positive, ER-negative subgroups, and the complete cohort. Our results suggest that the combination of TILs-WG with TILs-MPM model can greatly improve the prognostic value of TILs.

The prognostic value of tumour-infiltrating lymphocytes, programmed cell death protein-1 and programmed cell death ligand-1 in Stage I-III triple-negative breast cancer.

Tumour-infiltrating lymphocytes (TILs) represent a robust biological prognostic biomarker in triple-negative breast cancer (TNBC); however, the contribution of different subsets of immune cells is unclear. We investigated the prognostic value of immune markers, including stromal TILs (sTILs), CD8+T and FOPX3+T cells, PD-1 and PD-L1 in non-metastatic TNBC. In total, 259 patients with Stage I-III TNBC were reviewed. The density of sTILs along with the presence of total (t), stromal (s), and intratumoral (i) CD8+T cells and FOPX3+T cells were evaluated by haematoxylin and eosin and immunohistochemical staining. Immunohistochemical staining of PD-1, PD-L1 was also conducted. All immune markers were positively correlated with each other (P < 0.05). In the multivariate analysis, sTILs (P = 0.046), tCD8+T cells (P = 0.024), iCD8+T cells (P = 0.050) and PD-1 (P = 0.039) were identified as independent prognostic factors for disease-free survival (DFS). Further analysis showed that tCD8+T cells (P = 0.026), iCD8+T cells (P = 0.017) and PD-1 (P = 0.037) increased the prognostic value for DFS beyond that of the classic clinicopathological factors and sTILs. In addition to sTILs, inclusion of tCD8+T, iCD8+T cells, or PD-1 may further refine the prognostic model for non-metastatic TNBC beyond that including classical factors alone.

The prognostic value of tumor-infiltrating lymphocytes in non-small cell lung cancer patients who received neoadjuvant chemotherapy followed by surgery.

The relationship between tumor-infiltrating lymphocyte (TIL) levels and the prognosis of patients with non-small cell lung cancer (NSCLC) who receive neoadjuvant chemotherapy followed by surgery is a problem that requires more research. To evaluate the prognostic value of TIL levels in patients with NSCLC who received neoadjuvant chemotherapy followed by surgery. Patients with NSCLC who received neoadjuvant chemotherapy followed by surgery in our hospital from December 2014 to December 2020 were selected for a retrospective analysis. Sections were stained with hematoxylin and eosin (H&E) to evaluate TIL levels in surgically-resected tumor tissues. Patients were divided into TIL(low-level infiltration) and TIL+ (mediumto high-level infiltration) groups according to the recommended TIL evaluation criteria. Univariate (Kaplan-Meier) and multivariate (Cox) survival analyses were used to analyze the impact of clinicopathological features and TIL levels on prognosis. The study involved 137 patients, including 45 who were TILand 92 who were TIL+. The median overall survival (OS) and disease-free survival (DFS) of the TIL+ group were higher than those of the TILgroup. The univariate analysis showed that smoking, clinical and pathological stages, and TIL levels, were the factors influencing OS and DFS. The multivariate analysis showed that smoking (OS, hazard ratio (HR): 1.881, 95% confidence interval (95% CI): 1.135-3.115, p = 0.014; DFS, HR: 1.820, 95% CI: 1.181-2.804, p = 0.007) and clinical stage III (DFS, HR: 2.316, 95% CI: 1.350-3.972, p = 0.002) were adverse factors affecting the prognosis of patients with NSCLC who received neoadjuvant chemotherapy followed by surgery. At the same time, TIL+ status was an independent factor for a good prognosis in OS (HR: 0.547, 95% CI: 0.335-0.894, p = 0.016) and DFS (HR: 0.445, 95% CI: 0.284-0.698, p = 0.001). Medium to high levels of TILs were associated with a good prognosis in NSCLC patients who received neoadjuvant chemotherapy followed by surgery. The levels of TILs are of prognostic value in this population of patients.

Comparison of the prognostic value of stromal tumor-infiltrating lymphocytes and CD3 + T cells between schistosomal and non-schistosomal colorectal cancer

AimTo compare the prognostic value of tumor-infiltrating lymphocytes (TILs) and CD3 + cells and CD20 + cells between schistosomal colorectal cancer (SCRC) and non-schistosomal CRC (NSCRC).BackgroundAlthough schistosomiasis has been basically eliminated, it has not been completely extinction in China, and occasional outbreaks occur in Europe recently. The role of immune cells in the immune microenvironment of SCRC and NSCRC is remaining obscure, and the inflammation-based prognostic systems of SCRC has rarely been reported.MethodsHE-stained sections of 349 colorectal cancer (CRC) tumors, which were completely resected, were evaluated for density of TILs. Meanwhile, we evaluated CD3 + T lymphocytes and CD20 + B lymphocytes by immunochemistry. The relationship of these infiltrating immune cells with clinicopathological features, including schistosomiasis, and clinical outcomes was evaluated, and the prognostic roles of TILs in SCRC and NSCRC were explored.ResultsExcept for age (P < 0.0001), there were no significant differences between NSCRC and SCRC patients in clinicopathological features (P > 0.05). Beside, the positive expression pattern of sTILs, iTILs, CD3, and CD20 between NSCRC and SCRC patients was also similar (P > 0.05). In the whole cohort, sTILs and CD3 were defined as independent prognostic factors (P = 0.031 and P = 0.003, respectively). CD3 was an independent prognostic factor both in the NSCRC and SCRC set (P = 0.026 and P = 0.045, respectively). Higher sTILs, CD3, and CD20 were correlated with less aggressive tumor characteristics in the whole cohort and in subgroups.ConclusionAlthough CD3 was an independent prognostic factor for both NSCRC and SCRC set, there were no significant differences between SCRC and NSCRC patients in sTILs, CD3, CD20, and in other clinicopathological features.

Impact of Tumor-Infiltrating Lymphocytes on Disease Progression in Human Papillomavirus-Related Oropharyngeal Carcinoma.

We aim to explore the prognostic value of tumor-infiltrating lymphocytes (TILs) in the primary tumor and metastatic lymph nodes of patients with HPV(+)OPSCC. We hypothesize that TILS density at both sites is associated with disease-free survival in HPV(+)OPSCC. Matched case-control study among HPV(+)OPSCC patients who underwent intent-to-cure surgery. Cases developed locoregional or distant recurrence. Controls were matched based on age, sex, pathologic T, N, and overall stage, year of surgery, type of adjuvant treatment received, and the Adult Comorbidity Evaluation-27 (ACE-27) score. Single tertiary care center, May 2007 to December 2016. Tumoral TILs (tTILs) density was defined as % TILs; stromal TILs (sTILs) density was defined as absent/sparse or moderate/dense crowding. Associations between TILs and time to disease progression were assessed using Cox regression models. Forty-four case-control pairs (N = 88) were included: 42 (48%) AJCC pStage I, 39 (44%) pStage II, and 7 (8%) pStage III. tTILs density ≥10% (hazard ratio [HR] 0.41, 95% confidence interval [CI] 0.17-0.99, p = .048) and a moderate/dense sTILs density (HR 0.21, 95% CI 0.06-0.75, p = .016) in the primary tumor were significantly associated with decreased risk of progression. TILs density in the lymph node was associated with decreased risk of progressionbut did not reach statistical significance. The tTILs and sTILs density correlated strongly between the primary tumor and lymph node. Concordance between the pathologists' was moderate (60%-70%). In HPV(+)OPSCC, a higher density of tumoral and stromal TILs in the primary tumor and possibly the lymph node may predict a lower risk of disease progression.

Prognosis-related novel immunostaining pattern for programmed cell death ligand 1 and prognostic value of tumour-infiltrating lymphocytes in triple-negative breast cancer.

This study aims to determine theprognostic significance ofprogrammed cell death ligand 1 (PD-L1) expression and tumour-infiltrating lymphocytes (TILs) in triple- negative breast cancer (TNBC). PD-L1 expression and TIL percentage were determined in TNBCs that did not receive neoadjuvant therapy. Therelationship between PD-L1 expression and thepercentage ofTILs with survival was investigated. Thepresence ofintratumoural PD-L1-positive tumour-infiltrating immune cells (TIICs) in tumours with ≥ 1% PD-L1 expression was identified as anew PD-L1 evaluation parameter. Thepresence ofintratumoural PD-L1-positive TIICs as anew parameter in PD-L1-positive cases increased overall survival. Thepercentage ofTILs increased in both overall and distant metastasis-free survival (p = 0.040 and p = 0.006, respectively). As aresult, it was found that therisk ofdeath was increased 5.18-fold (p = 0.013) in patients without intratumoural PD-L1-positive TIICs. This risk ofdeath was calculated to be 5.40-fold higher in patients with TIL percentage ≤ 10% than in those with > 40% (p = 0.024), and therisk ofdistant metastasis was calculated to be 11.95 times higher. In our study, we discovered that thepercentage ofTILs made astatistically significant difference in TNBC survival. Thepresence ofintratumoural PD-L1-positive TIICs in PD-L1-positive cases significantly increased survival.

42P Tumor-reactive CD8+ T cells in ovarian and colon cancer in tumors and cell products

Several pivotal studies established the high prognostic value of tumor-infiltrating lymphocytes (TILs) in patients with solid tumors, including colorectal (CRC) and ovarian (OvCa) cancer. Albeit the association between clinicopathological outcomes and tumor infiltration supports the intervention with immunotherapeutic strategies, TIL-adoptive cell transfer (ACT) only led to clinical results in a subset of patients with non-melanoma tumors, raising the question of the abundance of tumor-reactive T cells in these cancers.

Prognostic Value of Tumour-Infiltrating Lymphocytes in an Unselected Cohort of Breast Cancer Patients.

Tumour-infiltrating lymphocytes (TILs) are considered to have prognostic and predictive value for patients with early breast cancer. We examined 1166 breast cancer patients from a prospective, multicentre cohort (Prognostic Assessment in Routine Application (PiA), n = 1270, NCT 01592825) following recommendations from the International TILs Working Group. TIL quantification was performed using predefined groups and as a continuous variable in 10% increments. The primary objective was the distribution of TILs in different breast cancer types. The second objective was the association with the recurrence-free interval (RFI) and overall survival (OS). Stromal infiltration with more than 60% TILs appeared in 2% of hormone receptor (HR)-positive and HER2-negative tumours, in 9.8% of HER2-positive tumours (any HR) and 19.4% of triple-negative breast cancers (TNBCs). Each 10% increment was associated with an improvement in the prognosis in HER2-positive samples (RFI, hazard ratio 0.773, 95% CI 0.587–1.017; OS, hazard ratio 0.700, 95% CI 0.523–0.937). When defining exploratory cut-offs for TILs, the use of a 30% threshold for the HR-positive and HER2-negative group, a 20% threshold for the HER2 group and a 60% threshold for the TNBC group appeared to be the most suitable. TILs bore prognostic value, especially in HER2-positive breast cancer. For clinical use, additional research on the components of immune infiltration might be reasonable.

145P Prognostic value of tumor-infiltrating lymphocytes in HER2-positive breast cancer treated with neoadjuvant chemotherapy and dual HER2-blockade: A TRAIN-2 sub study

BackgroundPreviously, it has been demonstrated that stromal tumor-infiltrating lymphocytes (sTILs) provide independent prognostic information in early-stage HER2-positive breast cancer. The aim of our study was to validate these findings by studying sTILs and outcome measures in early-stage HER2-positive breast cancer patients who were randomized to receive neoadjuvant chemotherapy with or without anthracyclines plus trastuzumab and pertuzumab in the TRAIN-2 study.MethodsFollowing sTIL scoring on pre-treatment biopsies by two pathologists, the geometric mean was calculated. When the two scores differed by more than 10%, a final score was assigned after revision by one pathologist. The association between sTILs and pathological complete response (pCR) was studied using logistic regression analyses adjusted for age, hormone receptor (HR) status, clinical T-stage, clinical N-stage, tumor grade and treatment arm. Similarly, the relation between sTILs and event-free survival (EFS) was evaluated by Cox regression analyses.ResultsTable: 145PStromal TILs, pathological complete response and 5-year event-free survival ratesGroupNumber of events5-year EFS (%)95% CILow sTILs, no pCR11/7383.875.3 - 93.2High sTILs, no pCR8/4581.771.0 - 94.1Low sTILs, pCR13/12981.569.7 - 95.2High sTILs, pCR3/12397.594.7 - 100 Open table in a new tab ConclusionsWe did not observe a significant association between sTILs and pCR or EFS after adjustment for clinical variables. However, patients with pCR and high sTILs show excellent 5-year EFS rates compared to patients with no pCR or low sTILs. Early-stage HER2-positive breast cancer patients with high sTILs and pCR may be candidates for de-escalated adjuvant treatment, whereas patients with low sTILs may benefit of additional treatment.Clinical trial identificationNCT01996267.Legal entity responsible for the studyBOOG Study Center.FundingRoche.DisclosureA.E. van Leeuwen-Stok: Financial Interests, Institutional, Funding, Roche funding for the TRAIN-2 study: Roche . H. Horlings: Financial Interests, Institutional, Funding: Roche; Non-Financial Interests, Institutional, Advisory Role: SlideScore.com. R.F. Salgado: Non-Financial Interests, Institutional, Other: Merck , Bristol Myers Squibb; Financial Interests, Institutional, Funding: Puma Biotechnology, Merck , Roche; Financial Interests, Personal, Advisory Board: Roche , Exact Sciences, Bristol Myers Squibb . G.S. Sonke: Financial Interests, Institutional, Funding: Roche , Merck , Novartis , Agendia, AstraZeneca , Seagen; Financial Interests, Institutional, Advisory Role: Seagen, Biovica. All other authors have declared no conflicts of interest. BackgroundPreviously, it has been demonstrated that stromal tumor-infiltrating lymphocytes (sTILs) provide independent prognostic information in early-stage HER2-positive breast cancer. The aim of our study was to validate these findings by studying sTILs and outcome measures in early-stage HER2-positive breast cancer patients who were randomized to receive neoadjuvant chemotherapy with or without anthracyclines plus trastuzumab and pertuzumab in the TRAIN-2 study. Previously, it has been demonstrated that stromal tumor-infiltrating lymphocytes (sTILs) provide independent prognostic information in early-stage HER2-positive breast cancer. The aim of our study was to validate these findings by studying sTILs and outcome measures in early-stage HER2-positive breast cancer patients who were randomized to receive neoadjuvant chemotherapy with or without anthracyclines plus trastuzumab and pertuzumab in the TRAIN-2 study. MethodsFollowing sTIL scoring on pre-treatment biopsies by two pathologists, the geometric mean was calculated. When the two scores differed by more than 10%, a final score was assigned after revision by one pathologist. The association between sTILs and pathological complete response (pCR) was studied using logistic regression analyses adjusted for age, hormone receptor (HR) status, clinical T-stage, clinical N-stage, tumor grade and treatment arm. Similarly, the relation between sTILs and event-free survival (EFS) was evaluated by Cox regression analyses. Following sTIL scoring on pre-treatment biopsies by two pathologists, the geometric mean was calculated. When the two scores differed by more than 10%, a final score was assigned after revision by one pathologist. The association between sTILs and pathological complete response (pCR) was studied using logistic regression analyses adjusted for age, hormone receptor (HR) status, clinical T-stage, clinical N-stage, tumor grade and treatment arm. Similarly, the relation between sTILs and event-free survival (EFS) was evaluated by Cox regression analyses. ResultsTable: 145PStromal TILs, pathological complete response and 5-year event-free survival ratesGroupNumber of events5-year EFS (%)95% CILow sTILs, no pCR11/7383.875.3 - 93.2High sTILs, no pCR8/4581.771.0 - 94.1Low sTILs, pCR13/12981.569.7 - 95.2High sTILs, pCR3/12397.594.7 - 100 Open table in a new tab ConclusionsWe did not observe a significant association between sTILs and pCR or EFS after adjustment for clinical variables. However, patients with pCR and high sTILs show excellent 5-year EFS rates compared to patients with no pCR or low sTILs. Early-stage HER2-positive breast cancer patients with high sTILs and pCR may be candidates for de-escalated adjuvant treatment, whereas patients with low sTILs may benefit of additional treatment. We did not observe a significant association between sTILs and pCR or EFS after adjustment for clinical variables. However, patients with pCR and high sTILs show excellent 5-year EFS rates compared to patients with no pCR or low sTILs. Early-stage HER2-positive breast cancer patients with high sTILs and pCR may be candidates for de-escalated adjuvant treatment, whereas patients with low sTILs may benefit of additional treatment.

Prognostic value of tumor-infiltrating lymphocytes in DCIS: a meta-analysis

BackgroundTumor infiltrating lymphocytes (TILs) have been shown to be associated with the prognosis of breast ductal carcinoma in situ (DCIS). In this systematic review and meta-analysis, we investigated the role of TILs and TIL subsets in predicting the recurrence risk of DCIS.MethodPubMed, Medline, Web of Science, Embase and Cochrane were searched to identify publications investigating the prognostic role of TILs in DCIS. After study screening, data extraction and risk of bias assessment, a meta-analysis was performed to assess the association between TILs (total TILs, CD4+, CD8+, FOXP3+, PD-L1+ TILs) and the risk of DCIS recurrence.ResultsA pooled analysis indicated that dense stromal TILs in DCIS were associated with a higher recurrence risk (HR 2.11 (95% CI 1.35–3.28)). Subgroup analysis showed that touching TILs (HR 4.73 (95% CI 2.28–9.80)) was more precise than the TIL ratio (HR 1.49 (95% CI 1.11–1.99)) in estimating DCIS recurrence risk. Moreover, the prognostic value of TILs seemed more suitable for patients who are diagnosed with DCIS and then undergo surgery (HR 2.77, (95% CI 1.26–6.07)) or surgery accompanied by radiotherapy (HR 2.26, (95% CI 1.29–3.95)), than for patients who receive comprehensive adjuvant therapies (HR 1.16, (95% CI 1.35–3.28)). Among subsets of TILs, dense stromal PD-L1+ TILs were valuable in predicting higher recurrence risk of DCIS.ConclusionThis systematic review and meta-analysis suggested a non-favorable prognosis of TILs and stromal PD-L1+ TILs in DCIS and indicated an appropriate assessment method for TILs and an eligible population.

Objective assessment of tumor infiltrating lymphocytes as a prognostic marker in melanoma using machine learning algorithms.

SummaryBackgroundThe prognostic value of tumor-infiltrating lymphocytes (TILs) assessed by machine learning algorithms in melanoma patients has been previously demonstrated but has not been widely adopted in the clinic. We evaluated the prognostic value of objective automated electronic TILs (eTILs) quantification to define a subset of melanoma patients with a low risk of relapse after surgical treatment.MethodsWe analyzed data for 785 patients from 5 independent cohorts from multiple institutions to validate our previous finding that automated TIL score is prognostic in clinically-localized primary melanoma patients. Using serial tissue sections of the Yale TMA-76 melanoma cohort, both immunofluorescence and Hematoxylin-and-Eosin (H&E) staining were performed to understand the molecular characteristics of each TIL phenotype and their associations with survival outcomes.FindingsFive previously-described TIL variables were each significantly associated with overall survival (p<0.0001). Assessing the receiver operating characteristic (ROC) curves by comparing the clinical impact of two models suggests that etTILs (electronic total TILs) (AUC: 0.793, specificity: 0.627, sensitivity: 0.938) outperformed eTILs (AUC: 0.77, specificity: 0.51, sensitivity: 0.938). We also found that the specific molecular subtype of cells representing TILs includes predominantly cells that are CD3+ and CD8+ or CD4+ T cells.InterpretationeTIL% and etTILs scores are robust prognostic markers in patients with primary melanoma and may identify a subgroup of stage II patients at high risk of recurrence who may benefit from adjuvant therapy. We also show the molecular correlates behind these scores. Our data support the need for prospective testing of this algorithm in a clinical trial.FundingThis work was also supported by a sponsored research agreements from Navigate Biopharma and NextCure and by grants from the NIH including the Yale SPORE in in Skin Cancer, P50 CA121974, the Yale SPORE in Lung Cancer, P50 CA196530, NYU SPORE in Skin Cancer P50CA225450 and the Yale Cancer Center Support Grant, P30CA016359.

Clinicopathological characteristics and prognostic analysis of tumor-infiltrating lymphocytes (TILs) in ductal carcinoma in situ (DCIS) and DCIS with microinvasion (DCIS-Mi) of the breast

ObjectiveOur purpose is to evaluate the correlation of TILs with clinicopathological characteristics and disease free survival (DFS) in DCIS and DCIS-Mi breast cancer (BC) patients.MethodsWe retrospectively reviewed the data of 360 DCIS patients and 125 DCIS-Mi patients treated by a single institution from 2016 to 2019. TILs are regarded as continuous variables and are divided into low (≤ 5%), medium (5–40%) and high (≥ 40%) for statistical analysis.ResultsIn DCIS and DCIS-Mi patients, larger tumor size, higher nuclear grade, hormone receptor (HR) negativity and human epidermal growth factor receptor 2(HER2) overexpression are all related to high TILs (P < 0.05). In addition, compared with DCIS, DCIS-Mi patients were significantly associated with high TILs (P < 0.001). Based on the different results of the subtypes, we further studied the correlation between TILs and DFS in 279 cases of HER2+ patients (204 of DCIS; 75 of DCIS-Mi). In HER2+ group, DCIS-Mi was significantly associated with HR negativity (P = 0.015) and high TILs (P = 0.002) compared with DCIS patients. In the survival analysis, we found that TILs had no effect on the DFS of DCIS (P = 0.938), DCIS-Mi (P = 0.807), and HER2+ (P = 0.379) BC patients. In the univariate and multivariate cox regression analysis, the correlation between TILs and the prognosis of DFS has not been confirmed in the three BC groups (P > 0.05).ConclusionTILs have played an non-negligible role in the progress of DCIS to DCIS-Mi, especially in HER2+ BC. The predictive and prognostic value of TILs still needs further research to confirm.

Abstract P5-06-14: A novel combination of a 2 gene score &amp; TIL as a predictive Biomarker for responders to novel therapies in Indian TNBC - A population with greater proportion of TNBCs

Abstract Introduction: Triple-Negative Breast Cancer (TNBC) comprises approximately 30% of all breast cancers in Indian women. Given their aggressive nature, TNBCs have high rates of systemic metastasis and mortality with only chemotherapy available for treatment. Compelling evidence has demonstrated the prognostic value of tumor-infiltrating lymphocytes (TILs), in many cancers especially in Breast Cancer and play a critical role in tumour progression, response to therapeutics and prognosis. They are typically measured by H&amp;E staining and immunohistochemistry for research purpose and degree of infiltrate is also known to differ among subtypes. Nonetheless, the literature regarding the types of immune cells characterizing TILs and their prognostic utility in TNBC has been conflicting for lack of accurate “functional” TIL assessments. Herein we have used a combination signature of TILs and a 2 gene immune function expression based signature to develop an unique classifier, to identify a subgroup within TNBC that has better clinical outcome, such as survival and response to treatment. Our findings also suggest a possible use of the score as a predictive biomarker for response to immune checkpoint therapy Patients and Methods: Surgically excised TNBC tumor specimens from 44 women from a single treating hospital in Bangalore, India were accessed under IERB approved protocols and assayed on nCounter® PanCancer Immune Profiling Panel which comprised of 740 genes and characterises 14 different immune cell types. Analysis using Non-negative Matrix factorization (NMF) based unsupervised clustering was done to arrive at stable subtypes characterized by different tumor microenvironments within TNBC. SSGSEA analysis was done to identify immune cell types. TILs were characterised by a pathologist on H&amp;E sections according to guidelines from TIL working group. The Immune classifier was validated on Breast Cancer data sets from TCGA. Results: NMF followed by SAM and PAM analysis yielded 2 stable subtypes (ST1 and ST2) of proportions 59% and 41% respectively. TIL groups also were divided into Dense and Mild groups which were 52% and 48% respectively. Despite similar distributions of the groups, ST1 and ST2 had distinct survival with ST2 having poorer outcome (p=0.023) while the dense and mild TIL groups did not separate as distinct groups. On a closer examination using SSGSEA analysis with Rooney et al signature, ST1 was enriched with T-cells, Dendritic cells, MHC-class 1 and very significantly high cytolytic score (CYT), which was defined by Granzyme A and Perforin expressions, proteins secreted by cytotoxic T cells (p=0.0074). The CYT Score ranged from -35 to 170 and cutoff was based on 75th percentile. We next hypothesized that a combined score of TIL and CYT would represent a functional TIL group with high immune activity. We arrived at a CYT+TIL score based on 75th percentile cut off of CYT and 2 groups of TIL. Of the resultant four groups, TIL High-CYT high that constituted about 20% of all TNBCs indicated an elevated immune response because of their microenvironment constitution and this can be identified using our simple immune classifier. On the other hand, in the TIL-high and CYT-low group, despite lymphocyte migration, outcome was not favorable. The classifier was applied to TNBC from TCGA (n=96) and similar results seen, with ST1 and ST2 which had distinctly separate survival and had similar patterns on CYT high groups. Conclusion: We developed a simple immune classifier with 2 gene signature c and H&amp;E slide TIL assessment for identifying a group with TNBC that can be better targeted with therapies. Our signature is predictive for selecting TNBCs which are potential responders to Immune therapies and has the potential for quick clinical adoption though it requires validation on a larger set. Citation Format: Aruna Korlimarla, Hari PS, Jyothi S Prabhu, Chantirika Ragulan, Ravi B Diwakar, Sandhya Apachu, Maggie Cheang, Rekha V Kumar, BS Srinath, TS Sridhar, Savitha Rajarajan, Annie Alexander, Anguraj Sadanandam. A novel combination of a 2 gene score &amp; TIL as a predictive Biomarker for responders to novel therapies in Indian TNBC - A population with greater proportion of TNBCs [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-06-14.