Low Tumor-infiltrating Lymphocytes Research Articles

[Clinicopathological characteristics and immune microenvironment of breast squamous cell carcinoma].

Objective: To investigate the clinicopathological characteristics of breast squamous cell carcinoma and to analyze the relationship between its immune microenvironment tumor infiltrating lymphocytes (TILs) and prognosis. Methods: Forty-four cases of primary squamous cell carcinoma of the breast diagnosed and treated in the First Affiliated Hospital of Air Force Medical University, Xi'an, China from January 2006 to July 2022 were selected. Their clinicopathological characteristics were analyzed. The cell composition of TILs was evaluated using immunohistochemistry (Mainly markers of B lymphocytes, T lymphocytes and plasma cells). The relationship between TILs and prognosis was also analyzed. Results: The 44 patients of breast squamous cell carcinoma were all female and all were invasive carcinoma. Eight cases (8/44, 18.2%) were squamous cell carcinoma, while 36 cases (36/44, 81.8%) were mixed squamous cell carcinoma. The mixed components included non-specific carcinoma and spindle cell metaplastic carcinoma (17 cases each). One case contained ductal carcinoma in situ of the breast and 1 case contained tubular carcinoma. The proportion of squamous cell carcinoma was 10% to 90%. The cases with pure squamous cell carcinoma often had a large cystic cavity, which was lined by atypical squamous epithelium, while infiltrating squamous cell carcinoma nests were seen in the breast tissue around the cystic cavity. Immunohistochemical staining showed that p63 and CK5/6 were expressed in the squamous cell carcinoma component, but ER, PR and HER2 were not, except for one case of HER2 1+. The positive rates of TRPS1 and PDL-1 were 76% and less than 1%, respectively. Fifteen cases were in the high TILs group (TILs≥30%) and 29 cases were in the low TILs group (TILs<30%). Twenty-three patients were followed up for 5 to 118 months. Among them, 12 died within 3 years and 9 were alive at the end of the follow up. There was no significant difference in TNM stage, TILs and prognosis between simple squamous cell carcinoma and mixed squamous cell carcinoma. Conclusions: Breast squamous cell carcinoma can be divided into simple squamous cell carcinoma and mixed squamous cell carcinoma. There are differences in gross findings and histology between the simple and mixed squamous cell carcinoma of the breast. Sufficient samples should be taken to avoid missing the diagnosis of a minor squamous component. The prognosis of patients with high TILs is significantly better than that of patients with low TILs. The expression rate of TRPS1 in primary squamous cell carcinoma of breast is high and helpful to the differential diagnosis from metastatic squamous cell carcinoma.

Abstract LB119: The immune-modulation of HM16390, firing up the poor tumor microenvironment to induce a potent anti-tumor efficacy

Abstract Immunotherapy, encompassing immune checkpoint blockades (ICBs) and immune stimulators, has become a widespread approach in cancer treatment. However, the effectiveness of these strategies relies significantly on the characteristics of the current tumor microenvironment (TME). This reliance underscores the urgent need for a potent immune modulator capable of inducing a favorable TME, particularly in non-immunogenic cold tumors. Here, we demonstrate that HM16390, a long acting IL-2 analog, has the potential to modify the immunogenicity of the TME by activating, expanding, and recruiting cytotoxic effector cells in cold tumor. In the murine B16F10 melanoma model, following the early upregulated proliferative potential, the proportions of CD8+ T as well as NK cells increased in both blood and tumor. While NK cell frequency initially increased and then recovered, CD8 T cells exhibited a dose-dependent upregulation in the late phase in both blood and tumor. Given the IL-2Rα (CD25) affinity of HM16390, there was an increase in Treg proportions in blood; however, intriguingly, Treg frequency decreased in the tumor. This pattern emphasizes the crucial role of HM16390 in balancing the hyper-activation of peripheral effector cells while simultaneously inducing a profoundly immune-favorable TME. Effector functions were also upregulated in both blood and tumor. Early upregulation of IFN-γ and TNF occurred in both blood and tumor, but the upregulation of granzyme B displayed a sustained expression pattern in the tumor. These immune-modulated features correlated with the exposures of HM16390, particularly noting a negative correlation between Treg down-regulation and Its exposure in blood. The immune modulations of HM16390 were validated in terms of efficacy in the Pancreatic Ductal Adenocarcinoma (PDAC) model. HM16390 was administered subcutaneously to panc02 tumor syngeneic mice via once-a-week administration for 6 weeks, with or without PD-1 combination treatment. HM16390 treatment group was well-tolerated and resulted in a dose-dependent anti-tumor effect. Notably, all mice treated with the highest dose exhibited a complete response. Furthermore, in panc02 syngeneic mice, known for a poor response to ICBs, the combination treatment of HM16390 with mAnti-PD-1 demonstrated synergistic anti-tumor effects. In conclusion, HM16390 demonstrates improved expansion and functions of effector tumor-infiltrating lymphocytes (TILs), correlating with exposure, and exhibits a safe Treg modulation pattern. These modulations culminate in a significant anti-tumor effect and synergies with PD-1 blockade in the B16F10 and PDAC model. Given that both B16F10 melanoma and Panc02 PDAC are recognized as poor immunogenic murine models with low TIL frequency, HM16390 shows promise in modifying the immunogenicity of the TME, thereby activating proper immune responses in preclinical models. Citation Format: Seongju Jeong, Yunjae Kim, Jooyun Byun, Jinyoung Kim, Jaehyuk Choi, Sungmin Bae, Daejin Kim, In Young Choi. The immune-modulation of HM16390, firing up the poor tumor microenvironment to induce a potent anti-tumor efficacy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB119.

Tumor-Infiltrating Lymphocytes (TILs) in Breast Cancer: Prognostic and Predictive Significance across Molecular Subtypes

Tumor-infiltrating lymphocytes (TILs) are pivotal in the immune response against breast cancer (BC), with their prognostic and predictive significance varying across BC subtypes. In triple-negative BC (TNBC), higher TIL levels correlate with improved prognosis and treatment response, guiding therapeutic strategies and potentially offering avenues for treatment de-escalation. In metastatic TNBC, TILs identify patients with enhanced immunotherapy response. HER2+ BC, similar to TNBC, exhibits positive correlations between TILs and treatment response, especially in neoadjuvant settings. Luminal BC generally has low TILs, with limited prognostic impact. Single hormone receptor-positive BCs show distinct TIL associations, emphasizing subtype-specific considerations. TILs in ductal carcinoma in situ (DCIS) display ambiguous prognostic significance, necessitating further investigation. Standardizing TIL assessment methods is crucial for unlocking their full potential as biomarkers, guiding treatment decisions, and enhancing patient care in BC.

Evaluation of tumor response to immune checkpoint inhibitors by a 3D immunotumoroid model.

Only 20 percent of renal and bladder cancer patients will show a significant response to immune checkpoint inhibitor (ICI) therapy, and no test currently available accurately predicts ICI response. We developed an "immunotumoroid" cell model system that recapitulates the tumor, its microenvironment, and necessary immune system components in patient-derived spheroids to enable ex vivo assessment of tumor response to ICI therapy. Immunotumoroids were developed from surgically resected renal cell carcinomas and bladder carcinomas selected for high tumor-infiltrating lymphocytes (TILs) and survived more than a month without media exchange. Immunohistochemistry was used to detect immune and non-immune cells in cryopreserved source tumors and the resulting immunotumoroids. Immunotumoroid response to ICIs (nivolumab, pembrolizumab, and durvalumab) and chemotherapy (cisplatin, gemcitabine, and paclitaxel) was monitored in real-time with Cytotox Red staining in an Incucyte device, and the immunotumoroid response was compared to retrospective clinical drug responses. Six of the 13 cases tested grew viable immunotumoroid models, with failed cases attributed to extensive tumor tissue necrosis or excess lymphocytes preventing spheroid formation. One successfully cultured case was excluded from the study due to low TIL infiltration (<5%) in the primary tumor sample. The five remaining models contained immune cells (CD4+ and CD8+ T cells, and macrophages), non-immune cells (fibroblasts), and tumor cells. Chemotherapy and ICI drugs were tested in immunotumoroids from 5 cases and compared to clinical outcomes where data was available. Four/five models showed cell killing in response to chemotherapy and two/five showed sensitivity to ICI. In three cases, the immunotumoroid model accurately predicted the patient's clinical response or non-response to ICIs or chemotherapy. Our immunotumoroid model replicated the multicellular nature of the tumor microenvironment sufficiently for preclinical ICI screening. This model could enable valuable insights into the complex interactions between cancer cells, the immune system, and the microenvironment. This is a feasibility study on a small number of cases, and additional studies with larger case numbers are required including correlation with clinical response.

Abstract 3437: Stromal tumor-infiltrating lymphocytes in association with body size and composition among Black and White women with breast cancer

Abstract Background: Obesity is a major risk factor for breast cancer and negatively affects disease prognosis. Tumor-infiltrating lymphocytes (TILs) in the stroma of breast tissue play a major role in eliminating cancer cells and modulating immunotherapy efficacy. We aim to study the association between body size and composition with stromal TILs in breast tumor tissue by race. Methods: The study included 920 Black and 395 White women with newly diagnosed invasive breast cancer who have stromal TIL scores from the Women’s Circle of Health Study (WCHS). Body mass index (BMI), waist circumference (WC), and waist-to-hip ratio (WHR) were measured during the home interview. Pre-treatment H&amp;E-stained tumor tissue sections were reviewed by our study pathologist and stromal TILs were scored on a scale of 0-100% at 10% increment according to the recommendations of the International TILs Working Group. Linear regressions adjusting for age at diagnosis, tumor grade, stage, and molecular subtype were used to test the differences in TILs across the categories of body size and composition indicators separately for Black and White women. Results: A higher stromal TILs score was associated with more aggressive breast cancer features, including higher grade, larger tumor size, more advanced stage, hormone receptor (HR) negativity, and human epidermal growth factor receptor 2 (HER2) positivity in both Black and White women (all P&amp;lt;0.05). Among Black women, WHR &amp;gt;0.90 vs ≤0.85 was associated with lower TILs overall (β = -3.5%, P = 0.045) and in triple-negative tumors (β = -12.3%, P = 0.003). Among White women, BMI 25.0 - 29.9 vs &amp;lt;25 was associated with higher TILs in HR-/HER2+ tumors (β = 28.1%, P = 0.031). A pattern suggested that BMI ≥30 vs &amp;lt;25 was associated with lower TILs in both HR-/HER2+ (β = -10.2, P=0.52) and triple-negative (β = -12.8, P=0.14) tumors, although the associations were not significant. No association was observed between TILs and any of the body composition parameters in women with HR+ tumors. Conclusions and Relevance: Body size and composition correlate with stromal TILs mainly in patients with HR- tumors, and this relation may be race-dependent. These findings suggest that body size and composition potentially modulate breast cancer treatment outcomes at least in part through TILs-dependent mechanisms. Funding: This work was in part supported by grants from the US National Institutes of Health (P01CA151135, R01CA100598, R01CA185623, P30CA016056, P30CA072720, K07CA201334, and R37CA248371), US Army Medical Research and Material Command (DAMD-17-01-1-0334), the Breast Cancer Research Foundation (CBA), and the Philip L. Hubbell family. Citation Format: Rand T. Akasheh, Thaer Khoury, Song Yao, Angela Omilian, Chi-Chen Hong, Elisa V. Bandera, Bonnie Qin, Nur Zeinomar, Christine Ambrosone, Ting-Yuan David Cheng. Stromal tumor-infiltrating lymphocytes in association with body size and composition among Black and White women with breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3437.

Clinical, Epidemiologic, and Pathologic Significance of ERBB2-Low Expression in Breast Cancer

It is unclear whether breast cancer (BC) with low ERBB2 expression (ERBB2-low) is a distinct clinical, pathological, and epidemiological entity from BC classified as no ERBB2 expression (ERBB2-negative). To evaluate the clinical, pathological, and epidemiologic features of BC with ERBB2-low expression compared with ERBB2-negative BC in a large population study. This cohort study was conducted as part of the Pathways Study, a prospective, racially and ethnically diverse cohort study of women with BC enrolled between 2006 and 2013 in Kaiser Permanente Northern California (KPNC). The hematoxylin and eosin slides underwent centralized pathology review, including the percentage of tumor infiltrating lymphocytes (TILs). Breast biomarker results were extracted from pathology reports, and women were included if they had a documented ERBB2 value that was not classified ERBB2-positive. Data were analyzed from February 2023 through January 2024. Clinical and tumor characteristics associated with BC and ERBB2-low or ERBB2-negative status. ERBB2-low was defined as immunohistochemistry score of 1+ or 2+ (negative by in situ hybridization); ERBB2-negative was defined as immunohistochemistry score of 0+. Other data were collected by self-report or extraction from electronic health records, including BC risk factors, tumor characteristics, treatment modality, and survival outcomes, with recurrence-free survival (RFS) as the primary outcome and overall survival (OS) and BC-specific mortality (BCSM) as secondary outcomes. The clinical, pathological, and epidemiological variables were compared between ERBB2-low and ERBB2-negative BC. Of 2200 eligible patients (all female; with mean [SD] age, 60.4 [11.9] years), 1295 (57.2%) had tumors that were ERBB2-low. Hormone receptors were positive in 1956 patients (88.9%). The sample included 291 Asian patients (13.2%), 166 Black patients (7.5%), 253 Hispanic patients (11.5%), 1439 White patients (65.4%), and 51 patients (2.3%) who identified as other race or ethnicity (eg, American Indian or Alaska Native and Pacific Islander). Within the hormone receptor-negative group, patients whose tumors had ERBB2-low staining, compared with those with ERBB2-negative tumors, had better OS (hazard ratio [HR], 0.54; 95% CI, 0.33-0.91; P = .02), RFS (HR, 0.53; 95% CI, 0.30-0.95; P = .03), and BCSM (HR, 0.43; 95% CI, 0.22-0.84; P = .01). In multivariable survival analysis stratified by hormone receptor status and adjusted for key covariates, patients with ERBB2-low and hormone receptor-negative tumors had lower overall mortality (HR, 0.48; 95% CI, 0.27-0.83; P = .009), RFS (HR, 0.45; 95% CI, 0.24-0.86; P = .02), and BCSM (subdistribution HR, 0.21; 95% CI, 0.10-0.46; P < .001) compared with patients with ERBB2-negative and hormone receptor-negative tumors. Within the hormone receptor-negative subtype, patients with ERBB2-low and high TILs tumors had better survival across all 3 outcomes compared with patients with ERBB2-negative and low TILs tumors. Additionally, patients with ERBB2-low and low TILs tumors had better BCSM (subdistribution HR, 0.36; 95% CI, 0.14-0.92; P = .03). These findings suggest that there were clinical, pathological, and epidemiological differences between ERBB2-low and ERBB2-negative BC, raising the possibility that ERBB2-low might be a unique biologic entity.

Tumor-Infiltrating Lymphocyte Level Consistently Correlates with Lower Stiffness Measured by Shear-Wave Elastography: Subtype-Specific Analysis of Its Implication in Breast Cancer.

Background: We aimed to elucidate the clinical significance of tumor stiffness across breast cancer subtypes and establish its correlation with the tumor-infiltrating lymphocyte (TIL) levels using shear-wave elastography (SWE). Methods: SWE was used to measure tumor stiffness in breast cancer patients from January 2016 to August 2020. The association of tumor stiffness and clinicopathologic parameters, including the TIL levels, was analyzed in three breast cancer subtypes. Results: A total of 803 patients were evaluated. Maximal elasticity (Emax) showed a consistent positive association with an invasive size and the pT stage in all cases, while it negatively correlated with the TIL level. A subgroup-specific analysis revealed that the already known parameters for high stiffness (lymphovascular invasion, lymph node metastasis, Ki67 levels) were significant only in hormone receptor-positive and HER2-negative breast cancer (HR + HER2-BC). In the multivariate logistic regression, an invasive size and low TIL levels were significantly associated with Emax in HR + HER2-BC and HER2 + BC. In triple-negative breast cancer, only TIL levels were significantly associated with low Emax. Linear regression confirmed a consistent negative correlation between TIL and Emax in all subtypes. Conclusions: Breast cancer stiffness presents varying clinical implications dependent on the tumor subtype. Elevated stiffness indicates a more aggressive tumor biology in HR + HER2-BC, but is less significant in other subtypes. High TIL levels consistently correlate with lower tumor stiffness across all subtypes.

Clinical and radiomics integrated nomogram for preoperative prediction of tumor-infiltrating lymphocytes in patients with triple-negative breast cancer.

The present study aimed to develop a radiomics nomogram based on conventional ultrasound (CUS) to preoperatively distinguish high tumor-infiltrating lymphocytes (TILs) and low TILs in triple-negative breast cancer (TNBC) patients. In the present study, 145 TNBC patients were retrospectively included. Pathological evaluation of TILs in the hematoxylin and eosin sections was set as the gold standard. The patients were randomly allocated into training dataset and validation dataset with a ratio of 7:3. Clinical features (age and CUS features) and radiomics features were collected. Then, the Rad-score model was constructed after the radiomics feature selection. The clinical features model and clinical features plus Rad-score (Clin+RS) model were built using logistic regression analysis. Furthermore, the performance of the models was evaluated by analyzing the receiver operating characteristic (ROC) curve, calibration curve, and decision curve analysis (DCA). Univariate analysis and LASSO regression were employed to identify a subset of 25 radiomics features from a pool of 837 radiomics features, followed by the calculation of Rad-score. The Clin+RS integrated model, which combined posterior echo and Rad-score, demonstrated better predictive performance compared to both the Rad-score model and clinical model, achieving AUC values of 0.848 in the training dataset and 0.847 in the validation dataset. The Clin+RS integrated model, incorporating posterior echo and Rad-score, demonstrated an acceptable preoperative evaluation of the TIL level. The Clin+RS integrated nomogram holds tremendous potential for preoperative individualized prediction of the TIL level in TNBC.

Cyclin-dependent kinase inhibitors enhance programmed cell death protein 1 immune checkpoint blockade efficacy in triple-negative breast cancer by affecting the immune microenvironment.

Clinical studies on programmed death-ligand 1 (PD-L1) immune checkpoint inhibitors for treating triple-negative breast cancer (TNBC) have shown unsatisfactory efficacy due to low tumor-infiltrating lymphocyte (TIL) levels. Inhibitors targeting cyclin-dependent kinase (CDK) proteins can affect the immune microenvironment, increase TIL levels, and promote antitumor immunity, thus providing a new direction for TNBC treatment strategies. The authors tested three CDK inhibitors on the TNBC cell lines MDA-MB-231 and 4T1 and validated their antitumor effects and impact on the immune microenvironment using multiple detection methods. They verified the efficacy and immune-related mechanisms of different combination therapy experiments in a 4T1 cell-transplanted BALB/c mouse model. Treatment with CDK inhibitors for 72 hours inhibited cell proliferation, clone formation, migration, and cell-cycle arrest and induced apoptosis in human breast cancer MDA-MB-231 cells and mouse breast cancer 4T1 cells. CDK inhibitors suppressed DNA methylation by downregulating DNMT1, DNMT3a, and DNMT3b expression. These three inhibitors promoted the secretion of various chemokines, enhanced tumor cell antigen presentation, and increased PD-L1 expression. CDK inhibitors improved the efficacy of immunotherapy in animal models and increased TIL levels. Combination therapy with CDK and PD-L1 immune checkpoint inhibitors affects the immune microenvironment, promotes antitumor immunity, and improves the efficacy of immunotherapy for TNBC.

The assessment of tumor-infiltrating lymphocytes in invasive apocrine carcinoma of the breast in relation to the HER2 status.

In the current study, we assessed the prevalence and molecular features of HER2-low phenotype in the apocrine carcinomas of the breast (ApoCa) and its relationship with tumor-infiltrating lymphocytes (TILs). A cohort of 64 well-characterized therapy-naïve ApoCa was used. The TIL distribution was assessed using the hematoxylin and eosin whole slide/scanned images following the international TILs working group recommendations. Next-generation sequencing (NGS) was performed in a subset of HER2-low ApoCa. All patients were women, with a mean age of 62 years. Forty-three carcinomas were pure apocrine carcinoma (PAC; ER-/AR+), and the remaining 21 were classified as apocrine-like carcinomas (ALCs; ER+/-, AR+/-). HER2/neu was positive (score 3+ by IHC and/or amplified by FISH) in 20/43 (47%) PAC and 4/21 (19%) ALC. The prevalence of HER2-low expression (scores 1+ or 2+ without HER2 amplification) in ApoCa was 39% without significant differences between PAC and ALC (P = 0.14); however, the HER2-low phenotype was more prevalent in triple-negative PAC than in ALC (P < 0.001). Levels of TILs were low (≤10%) in 74% of ApoCa (median 5%, range 0%-50%). TIL levels were significantly higher in ALC than in PAC (P = 0.02). HER2 status had no impact on TIL distribution (P = 0.45). The genomic profile of HER2-low ApoCa was similar to other subtypes of ApoCa. ApoCa has predominantly low TIL, particularly PAC.The prevalence of the HER2-low phenotype in ApoCa is high, which should have therapeutic and clinical implications given the recently approved therapies with antibody-drug conjugates (ADCs) for HER2-low breast cancers.

Machine learning-based MRI radiomics for assessing the level of tumor infiltrating lymphocytes in oral tongue squamous cell carcinoma: a pilot study

BackgroundTo investigate the value of machine learning (ML)-based magnetic resonance imaging (MRI) radiomics in assessing tumor-infiltrating lymphocyte (TIL) levels in patients with oral tongue squamous cell carcinoma (OTSCC).MethodsThe study included 68 patients with pathologically diagnosed OTSCC (30 with high TILs and 38 with low TILs) who underwent pretreatment MRI. Based on the regions of interest encompassing the entire tumor, a total of 750 radiomics features were extracted from T2-weighted (T2WI) and contrast-enhanced T1-weighted (ceT1WI) imaging. To reduce dimensionality, reproducibility analysis by two radiologists and collinearity analysis were performed. The top six features were selected from each sequence alone, as well as their combination, using the minimum-redundancy maximum-relevance algorithm. Random forest, logistic regression, and support vector machine models were used to predict TIL levels in OTSCC, and 10-fold cross-validation was employed to assess the performance of the classifiers.ResultsBased on the features selected from each sequence alone, the ceT1WI models outperformed the T2WI models, with a maximum area under the curve (AUC) of 0.820 versus 0.754. When combining the two sequences, the optimal features consisted of one T2WI and five ceT1WI features, all of which exhibited significant differences between patients with low and high TILs (all P < 0.05). The logistic regression model constructed using these features demonstrated the best predictive performance, with an AUC of 0.846 and an accuracy of 80.9%.ConclusionsML-based T2WI and ceT1WI radiomics can serve as valuable tools for determining the level of TILs in patients with OTSCC.

The association of PD-L1 expression and CD8-positive T cell infiltration rate with the pathological complete response after neoadjuvant treatment in HER2-positive breast cancer.

Achieving a pathological complete response (pCR) after neoadjuvant therapy in HER2-positive breast cancer patients is the most significant prognostic indicator, suggesting a low risk of recurrence and a survival advantage. This study aims to investigate clinicopathological parameters that can predict the response to neoadjuvant treatment in HER2 + breast cancers and to explore the roles of tumour-infiltrating lymphocytes (TILs), CD8 + T lymphocytes and PD-L1 expression. This single-centre retrospective study was conducted with 85 HER2-positive breast cancer patients who underwent surgery after receiving neoadjuvant therapy between January 2017 and January 2020. Paraffin blocks from these patients were selected for immunohistochemical studies. A complete pathological response to neoadjuvant treatment was determined in 39 (45.9%) patients. High Ki-67 index (> 30%), moderate to high TIL infiltration, PD-L1 positivity and high CD8 cell count (≥ 25) were significantly associated with pCR in univariate analyses (p: 0.023, 0.025, 0.017 and 0.003, respectively). Multivariate regression analysis identified high Ki-67 index (> 30%) and CD8 cell infiltration as independent predictors for pCR in HER2-positive breast cancer. High Ki-67 index, and high CD8 cell count are strong predictors for pCR in HER2-positive breast cancer. Tumours with high Ki-67 index, high TILs and CD8 infiltration may represent a subgroup where standard therapies are adequate. Conversely, those with low TILs and CD8 infiltration may identify a subgroup where use of novel strategies, including those that increase CD8 infiltration could be applied.

Role of stromal PD-L1 expression in colorectal liver metastasis

Background and AimThe outcomes of immune checkpoint blockade for colorectal cancer (CRC) treatment are unsatisfactory. Furthermore, the efficacy of immune checkpoint blockade for liver metastasis of various cancer is poor. Here, we investigated the relationship between stromal programmed death-ligand 1 (PD-L1) expression and the prognosis of patients with colorectal cancer liver metastasis (CRLM).MethodsThe present study enrolled 84 CRLM patients who underwent surgery (R0) for CRC. Immunohistochemistry was performed to analyze stromal PD-L1 expression in CRLM.ResultsStromal PD-L1 was expressed in 52.3% of CRLM samples, which was associated with fewer not optimally resectable metastases (p = 0.04). Stromal PD-L1 also tended to associate with a lower tumor grade (p = 0.08). Stromal PD-L1-positive patients had longer overall survival (p = 0.003). Multivariate analysis identified stromal PD-L1 expression (p = 0.008) and poorer differentiation (p < 0.001) as independent prognostic indicators. Furthermore, stromal PD-L1 expression was correlated to a high number of tumor-infiltrating lymphocytes (TILs). Stromal PD-L1– and low TIL groups had shorter OS than stromal PD-L1 + and high TIL groups (46.6% vs. 81.8%, p = 0.05) Stromal PD-L1-positive patients had longer disease-free survival (DFS) (p = 0.03) and time to surgical failure (p = 0.001). Interestingly, stromal PD-L1 expression was positively related to the desmoplastic subtype (p = 0.0002) and inversely related to the replacement subtype of the histological growth pattern (p = 0.008).ConclusionsStromal PD-L1 expression may be a significant prognostic marker for CRLM.

Abstract A044: Tumor-infiltrating lymphocyte density correlates with survival in pancreatic adenosquamous carcinoma

Abstract Background: Pancreatic adenosquamous carcinoma (PASC) is a rare and aggressive form of pancreatic cancer, representing only 1-5% of cases. Histologically, it is defined by the presence of at least 30% squamous differentiation mixed with ductal adenocarcinoma. No standard therapies or prognostic biomarkers have been validated to date in this understudied disease. Methods: All patients diagnosed with pancreatic cancer in the Johns Hopkins Cancer Registry from 2013 to 2018 were queried for adenosquamous histology. Archived slides of available tumor specimens were examined to quantify the percentage of squamous differentiation and tumor-infiltrating lymphocyte (TIL) density. Descriptive statistics were used to summarize clinical, pathologic, and molecular characteristics. Kaplan-Meier estimates and log-rank tests were used for survival analyses. Results: Among a total of 8,908 patients queried, 155 patients with adenosquamous histology were identified. Median overall survival (OS) was 14.2 months and 7.5 months for resectable and unresectable/metastatic disease, respectively. Molecular tumor testing revealed frequent somatic mutations in KRAS, TP53, and SMAD4 in addition to multiple DNA damage repair genes. The percentage of squamous differentiation and TIL density was correlated with survival outcomes in a subset of 28 patients with evaluable tumor specimens. No association between the percentage of squamous differentiation and OS was identified. Of note, compared to those with low TIL density (0-10%), patients with an intermediate/high TIL density (&amp;gt;10%) exhibited significantly improved survival (median OS 36.3 vs 14.2 months, p = 0.011). Conclusions: For the first time, intermediate/high TIL density is associated with prolonged OS in a single-institution cohort of PASC patients. Prospective validation in larger cohorts is needed to establish prognostic biomarkers that can guide therapeutic decision-making in this lethal and under-examined disease subtype. Citation Format: Saurav D. Haldar, Christopher D. Jakubowski, Su Jin Lim, Hao Wang, Richard A. Burkhart, Kelly Lafaro, William R. Burns III, Jin He, Daniel A. Laheru, Elizabeth D. Thompson, Nilofer S. Azad. Tumor-infiltrating lymphocyte density correlates with survival in pancreatic adenosquamous carcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr A044.

Evaluation of tumor-infiltrating lymphocytes and mammographic density as predictors of response to neoadjuvant systemic therapy in breast cancer

Background Response rates vary among breast cancer patients treated with neoadjuvant systemic therapy (NAST). Thus, there is a need for reliable treatment predictors. Evidence suggests tumor-infiltrating lymphocytes (TILs) predict NAST response. Still, TILs are seldom used clinically as a treatment determinant. Mammographic density (MD) is another potential marker for NAST benefit and its relationship with TILs is unknown. Our aims were to investigate TILs and MD as predictors of NAST response and to study the unexplored relationship between TILs and MD. Material and methods We studied 315 invasive breast carcinomas treated with NAST between 2013 and 2020. Clinicopathological data were retrieved from medical records. The endpoint was defined as pathological complete response (pCR) in the breast. TILs were evaluated in pre-treatment core biopsies and categorized as high (≥10%) or low (<10%). MD was scored (a–d) according to the breast imaging reporting and data system (BI-RADS) fifth edition. Binary logistic regression and Spearman’s test of correlation were performed using SPSS. Results Out of 315 carcinomas, 136 achieved pCR. 94 carcinomas had high TILs and 215 had low TILs. Six carcinomas had no available TIL data. The number of carcinomas in each BI-RADS category were 37, 122, 112, and 44 for a, b, c, and d, respectively. High TILs were independently associated with pCR (OR: 2.95; 95% CI: 1.59–5.46) compared to low TILs. In the univariable analysis, MD (BI-RADS d vs. a) showed a tendency of higher likelihood for pCR (OR: 2.43; 95% CI: 0.99–5.98). However, the association was non-significant, which is consistent with the result of the multivariable analysis (OR: 2.51; 95% CI: 0.78–8.04). We found no correlation between TILs and MD (0.02; p = .80). Conclusion TILs significantly predicted NAST response. We could not define MD as a significant predictor of NAST response. These findings should be further replicated.

Association of tumor-infiltrating lymphocytes with recurrence score in hormone receptor-positive/HER2-negative breast cancer: Analysis of four prospective studies

BackgroundThe clinical value of tumor infiltrating lymphocytes (TILs) in hormone receptor-positive (HR+)/HER2- breast cancer (BC) may be unearthed by focusing on more biologically aggressive tumors. Here we deepen and describe the correlation between RS and TILs, proposing an immuno-genomic model for HR+ /HER2- BC. MethodsWe enrolled T1-T3, N0-N1 BC patients with available RS® and TILs in the context of four multicenter, prospective studies. RS® and TILs were considered as continuous and categorical variables. RS® was categorized into: 0–10 (low risk), 11–25 (intermediate risk) and 26–100 (high risk); TILs were categorized into: low TILs (0–10%), intermediate TILs (11–59%) and high TILs (60–100%). Results811 patients were included. RS distribution was (n = 810): low risk 22.0%, intermediate risk 61.2%, high risk 16.8%. TIL distribution was (n = 455): low TILs 84.6%, intermediate TILs 13.6% and high TILs 1.8%. A significant, weak positive, linear correlation was found between continuous TILs and RS (Pearson coefficient=0.223, p < 0.001). When considering RS and TILs categories, tumors with intermediate/high TIL levels significantly enriched the high RS subgroup (p = 0.006). This was confirmed both within Luminal A and Luminal B cohorts. Among high-RS patients, 16.7% of Luminal A and 26.7% of Luminal B tumors had intermediate/high TILs. ConclusionsWe observed that RS® and TILs capture only slightly overlapping information on the biology of HR+ /HER2- tumor microenvironment. We demonstrated the feasibility of combining RS and TILs into a composite immuno-genomic model, which may serve the purpose of guiding and focalizing patient selection in the further development of immunotherapy strategies for Luminal-like disease.

Clinicopathological features and prognostic analysis of HER2 low and fibrotic focus in HER2-negative breast cancer.

The aim of this study is to evaluate the clinicopathological features and prognostic significance of HER2 low, fibrotic focus (FF), and tumor-infiltrating lymphocytes (TILs) in patients with HER2-negative breast cancer. We retrospectively reviewed the data of 293 patients with HER2-negative, stage I-II, invasive breast cancer of non-specific types. The HER2-negative cases were classified into HER2 low and HER2 0. Digital analysis of hematoxylin-eosin stained whole slide images was used to evaluate the FF expression. TILs were also evaluated using the Whole Slide Image. Furthermore, the association between HER2 low, FF, and TILs as well as their prognostic significance were analyzed. The study cohort included 178 cases (60.8%) with HER2 low and 115 cases (39.2%) with HER2 0. Older age, lower Nottingham histological grade (NHG), estrogen receptor (ER) positivity, progesterone receptor (PR) positivity, and hormone receptor (HR) positivity were all associated with HER2 low. FF was correlated with older age, intermediate and low NHG, vascular invasion, HR positivity, HER2 low status, high Ki67 expression, and low TILs. Univariate survival analysis showed that FF was significantly associated with shorter progression-free survival (PFS). Stratified analysis indicated that in the HR-negative and HR-positive groups, HER2 status and TILs did not affect PFS. DFS was longer in patients without FF compared to those with FF in the HR-positive (hazard ratio [HR] = 0.313) and HER2 low (HR = 0.272) groups. DFS was also significantly longer in patients without FF compared to those with FF in the HR-negative (HR = 0.069) and HER2 0 groups (HR = 0.129). The results indicated that the HER2 low status and the TILs expression did not impact prognosis. However, patients with FF exhibited distinct biological characteristics and prognostic significance, particularly in the HR-negative and HER2 0 groups. This provides a rationale for accurate diagnosis and treatment of HER2-negative breast cancer.

Tumor-infiltrating lymphocytes in oral cavity squamous cell carcinoma and its association with clinicopathological parameters

BackgroundOral squamous cell carcinoma is one of the most common cancers of the head and neck region. Tumor-infiltrating lymphocytes are naturally occurring mononuclear cells that infiltrate the tumor microenvironment. The aim of the present study is the evaluation of stromal tumor-infiltrating lymphocytes (TILs) in oral squamous cell carcinoma and its association with clinicopathological parameters like age, gender, site, tumor stage, nodal status, grade, lymphovascular and perineural invasion. MaterialsA total of 51 patients of oral cavity squamous cell carcinomas who underwent resection were included in the study. Assessment of the stromal TILs was done on sections stained with Hematoxylin and eosin using the scoring criteria proposed by the International Immuno-Oncology Biomarker Working Group. TILs were assessed in percentages as continuous scores and subsequently categorized into low TILs (<20%) and high TILs (≥20%). The association between TILs and clinicopathological parameters was evaluated using the Fischer exact/Chi-square test. ResultsAmong the 51 cases evaluated in the present study, low and high TILs were found in 25.5% and 74.5% of cases respectively. TILs were found to show a significant association with tumor size and lymphovascular invasion. However, no significant association was found with age, gender, nodal status, grade, perineural invasion, and site in the oral cavity. ConclusionThe evaluation of TILs as proposed by the International Immuno-Oncology Biomarker Working Group is a simple, inexpensive test. Morphological evaluation of TIL can provide valuable prognostic information in oral cavity squamous cell carcinoma cases and can be incorporated as a part of routine histopathological reporting.

A simple and convenient model combining multiparametric MRI and clinical features to predict tumour-infiltrating lymphocytes in breast cancer

To develop a simple and convenient method based on multiparametric magnetic resonance imaging (MRI) and clinical features to non-invasively predict tumour-infiltrating lymphocytes (TILs) in breast cancer (BC) and to explore the relationship between TIL levels and disease-free survival (DFS). A total of 172 BC patients were enrolled between November 2017 and June 2021 in this retrospective study. The patients were divided into high (≥10%) and low (<10%) TIL groups. Clinicopathological data were collected. MRI features were reviewed by two radiologists. Predictors associated with TILs were determined by using multivariable logistic regression analyses. Kaplan-Meier survival curves based on TIL levels were used to estimate DFS. A total of 102 patients with low TILs and 70 patients with high TILs were included in the study. Tumour size (odds ratio [OR], 1.040; 95% confidence interval [CI]: 1.006, 1.075; p=0.020), apparent diffusion coefficient (ADC; OR, 1.003; 95% CI: 1.001, 1.005; p=0.015), clinical axillary lymph node status (CALNS; OR, 3.222; 95% CI: 1.372,7.568; p=0.007), and enhancement pattern (OR, 0.284; 95% CI: 0.143, 0.563; p<0.001) were independently associated with TIL levels. These features were used in the ALSE model (where A is ADC, L is CALNS, S is size, and E is enhancement pattern). High TILs were associated with better DFS (p=0.016). The ALSE model derived from multiparametric MRI and clinical features could non-invasively predict TIL levels in BC, and high TILs were associated with longer DFS, especially in human epidermal growth factor receptor 2 (HER2)-positive BC and triple-negative BC (TNBC).

Effect of Lipid Levels on Tumor-Infiltrating Lymphocytes and Prognosis in Patients with Triple-Negative Breast Cancer

Objective: Dyslipidemia can promote cell proliferation, malignant transformation, metastasis, and cancer recurrence. Moreover, it could also affect immune infiltration in the tumor microenvironment. Therefore, we aimed to explore the effects of lipid levels on tumor-infiltrating lymphocytes (TILs) and prognosis in patients with triple-negative breast cancer (TNBC). Methods: Samples from 222 patients with TNBC from July 2007 to December 2019 were obtained from the tissue specimen banks in 3 hospitals. The blood samples were used to detect the levels of lipid levels such as apolipoprotein B (Apo B), triglyceride (TG), and low-density lipoprotein cholesterol (LDL-C). The TILs in the 222 TNBC tissues were detected using hematoxylin and eosin (H&E) staining, and the relationship between the lipid levels, clinical characteristics, and prognosis was analyzed. Results: Among TNBC patients, the overall survival (OS) time and disease-free survival (DFS) time were lower in patients with high LDL-C levels than those with low LDL-C levels (p < 0.01, respectively). The DFS was shorter in patients with low stromal TIL (STIL) levels than those with moderate or high STIL levels (p = 0.023). Multifactor Cox regression analysis showed that LDL-C level, Apo B level, and lymphocyte-predominant breast cancer were independent risk factors for OS in TNBC patients. The number of positive lymph nodes, postoperative staging, and total amount of TILs were independent risk factors for DFS in TNBC patients. Conclusion: The LDL-C and STIL levels were correlated with survival and prognosis in patients with TNBC.