Low Tumor-infiltrating Lymphocytes Research Articles

Abstract PS16-06: Tumor-infiltrating lymphocytes and pathologic response to neoadjuvant chemotherapy with the addition of platinum and pembrolizumab in TNBC: A single-center real-world study

Abstract Background Previous studies have presented compelling evidence suggesting that the inclusion of platinum agents alongside anthracyclines and taxanes could potentially enhance treatment outcomes in high-risk triple-negative breast cancer (TNBC). Moreover, pembrolizumab has been integrated into clinical practice as a part of standard-of-care for non-metastatic TNBC with high risk. In light of these findings, we undertook a real-world study to investigate the impact of incorporating platinum agents and pembrolizumab on achieving pathologic complete response (pCR) in TNBC patients undergoing neoadjuvant chemotherapy (NAC). Furthermore, we specifically examined the influence of tumor-infiltrating lymphocytes (TILs) on the treatment outcomes. Patients and Methods In this real-world study conducted at Gangnam Severance Hospital, Seoul, Republic of Korea, we analyzed a cohort of 398 patients with TNBC who underwent surgery following NAC between March 2007 and November 2022. Among them, 247 patients received an anthracycline-taxanes (A-T), 120 received a carboplatin regimen including A-T, and 31 received a pembrolizumab regimen including A-T-carboplatin as part of their neoadjuvant chemotherapy treatment. TIL was evaluated in biopsied samples prior to NAC according to the guideline of TIL international working group. The high TIL was defined with a cutoff of 50%. Results Among the 398 patients analyzed, 87 (21.9%) had high TIL tumors. The pCR rates were 32% in the anthracycline-taxane (A-T) regimen group, 57% in the A-T-carboplatin regimen group, and 68% in the pembrolizumab with A-T-carboplatin regimen group. Within the high TIL group, the pCR rate did not increase with the addition of carboplatin (51.8% in the A-T group and 41.7% in the A-T-carboplatin group), but reached 85.7% with the addition of pembrolizumab and carboplatin. Among the low TIL group, the pCR rate increased from 26.7% to 61.1% with the addition of carboplatin, but there was no difference in the pCR rate between the carboplatin and pembrolizumab groups (61.1% and 60.9%, respectively). In clinically node-positive patients, the pCR rate significantly increased with pembrolizumab in the high TIL group (40.9% versus 100%, p=0.035). However, in low TIL patients, the addition of carboplatin alone significantly increased the pCR rate to 62.3%, whereas the addition of pembrolizumab did not show the same effect. Conclusions Our real-world data consistently demonstrates an increased pCR rate with the addition of carboplatin and pembrolizumab. Among patients with high TIL, the addition of carboplatin did not result in an elevated pCR rate. However, the addition of pembrolizumab tended to maximize the pCR rate, surpassing 80%. On the other hand, among patients with low TIL, the addition of carboplatin significantly increased the pCR rate, while the addition of pembrolizumab did not have the same effect. Efforts should be made to improve the response to pembrolizumab-containing regimens for patients with low baseline TIL levels. Citation Format: Min Ji Kim, Yoonwon Kook, Seung Ho Baek, Junghyun Kim, Sohyun Moon, Seung Eun Lee, Jee Hung Kim, Soong June Bae, Sung Gwe Ahn, Joon Jeong. Tumor-infiltrating lymphocytes and pathologic response to neoadjuvant chemotherapy with the addition of platinum and pembrolizumab in TNBC: A single-center real-world study [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PS16-06.

Abstract PS09-06: Tumor infiltrating lymphocytes as a predictor of pathologic complete response to neoadjuvant therapy in HER2 positive breast cancer

Abstract Background The presence of tumor infiltrating lymphocytes (TILs) has shown positive prognostic relevance for certain subtypes of breast cancer, although its value as a predictive biomarker is still uncertain. The purpose of this study was to study the association between TILs expression with the pathologic complete response (pCR) following neoadjuvant therapy in Mexican patients with HER2+ breast cancer. Methods Retrospective cohort of patients with stage I-III HER2+ breast cancer who received neoadjuvant therapy between 2017 and 2020 at Instituto Nacional de Cancerología (INCan). We collected demographic and clinico-pathological characteristics. Stage was assigned using AJCC 7th criteria; hormone receptor (HR) and HER2 status were determined according to ASCO-CAP guidelines. Quantification of TILs was made by microscopy and stained with hematoxylin-eosin. TIL levels were defined as low (1-10%) intermediate (11-40%) and high ( >40%). Complete pathologic response was defined as the absence of invasive disease in breast and axilla (ypT0/is N0). pCR rates were compared using X2 and Fisher's exact tests according to demographic and clinicopathologic characteristics. Univariate logistic regression analysis was performed to estimate the probability of pCR according to relevant characteristics. Those parameters with p ≤0.10 on univariate analysis were included in a multivariate logistic regression model. A p< 0.05 was considered as statistically significant. Results We included 164 patients (mean age 51.0 years, SD 11.4). 86 patients (54.1%) were postmenopausal, 75 patients (47.2%) had T3-T4 tumors, 129 patients (81.1%) had positive lymph nodes, 133 patients (83.6%) presented Ki67 ≥30%, 59 patients (37.1%) had negative hormone receptors, 136 patients (85.5%) were treated with anthracyclines and 20 patients (12.6%) received dual anti-HER2 therapy with trastuzumab/pertuzumab. TIL expression was low in 84 patients (52.8%), intermediate in 47 (29.6%), and high in 28 patients (17.6%). The pathologic complete response (pCR) rate in the overall population was 50.9%. pCR rates according to the demographic and clinico-pathological features are listed in Table 1. In patients with intermediate-high TILs, pCR rates were 73.1% with trastuzumab, and 75.0% with dual HER2 blockade (p=0.91). Among patients with low TILs, pCR rates were 23.6% and 75.0%, respectively (p < 0.001). On univariate analysis, intermediate TILs (OR 3.59; 95% C.I 1.70-7.59; p= < 0.001), high TILs (OR 29.00; 95% C.I 6.40-131.37; p= < 0.001), Ki67 ≥30% (OR 2.73, 95% C.I 1.11-6.73; p=0.02), and dual anti-HER2 therapy (OR 3.31; 95% C.I 1.14-9.63 p= 0.021) were associated with a higher probability of pCR. T3/T4 tumor size (OR 0.43; 95% C.I 0.22-0.81; p = 0.009) was associated with a lower probability of pCR. Positive lymph nodes, HR status , and use of anthracyclines were not significantly associated. On multivariate analysis, intermediate TILs (OR 3.28; 95% C.I 1.46-7.32 p=.004), high TILs (OR 32.37; 95% C.I 6.84-153.04 p= < 0.001), and dual anti-HER2 therapy (OR 6.58; 95% CI 2.01-21.47 p= .002) remained significantly associated with pCR. Conclusions Intermediate/high TIL expression was significantly associated with pCR. Our results suggest that TIL expression could help select patients with locally advanced HER2+ breast cancer for treatment intensification with dual HER2 blockade in resource-limited settings. However, these findings require confirmation in larger, prospective studies before implementation into routine practice. TABLE 1. Pathologic complete response (pCR) rates according to demographic and clinical pathological features (n=164). Citation Format: Alberto Monroy Chargoy, Haydee Cristina Verduzco-Aguirre, Javier Monroy Chargoy, Jose Rodrigo Espinosa-Fernandez, Jesus Antonio Martinez Ojeda, Paula Cabrera-Galeana. Tumor infiltrating lymphocytes as a predictor of pathologic complete response to neoadjuvant therapy in HER2 positive breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PS09-06.

Abstract PO3-13-06: TILs are an independent prognostic factor in early-stage TNBC patients achieving pathologic complete response after neoadjuvant chemotherapy

Abstract Introduction: In patients with triple negative breast cancer (TNBC) the achievement of pathological complete response (pCR) after neoadjuvant chemotherapy (NACT) is associated with better long-term clinical outcomes. However, the attainment of pCR does not guarantee cure, since ~ 10% of patients undergo disease relapse. Higher levels of tumor infiltrating lymphocytes (TILs) are associated with better prognosis in TNBC patients undergoing upfront surgery and in patients with residual disease after NACT. However, the prognostic role of TILs among TNBC patients undergoing pCR during NACT has not been investigated so far. Methods: We included 134 TNBC patients (ER cutoff< 10%) who achieved pCR after NACT. These cases were identified from a cohort of 348 TNBC patients who received NACT at Istituto Oncologico Veneto (Padova) and Istituto Nazionale Tumori (Milano) from April 2004 to April 2021. TILs were quantified in pre-NACT tumor biopsy according to International guidelines. We assessed the association between TILs and relapse-free survival (RFS) or overall survival (OS). Results: Main patients characteristics were: median age 48 years; ductal histotype 92.8%; histologic grade 3 95.2%; stage I 9.7%, stage II 71.6%, stage III 18.7%. Almost all patients received anthracyclines (98.5%) and taxanes (99.3%) as part of NACT for a median number of 4 cycles each; 55.2% of patients received carboplatin. At a median FU of 4.6 years, the 5-yr RFS and OS rates were 90.4% and 94.4%, respectively. As a continuous variable (1% increment), TILs were independently associated with better RFS and OS after adjusting for patient age and tumor stage: HR 0.90 (95%CI 0.83-0.98), p=0.015 and HR 0.89 (95%CI 0.80-1.00), p=0.041, respectively. The best prognostic TILs cut-off in this cohort was 20%. Patients with TILs>20% showed a 100% 5-yr RFS rate (0/63 events) vs 82.6% (12/71 events) for patients with TILs< 20% (log-rank p< 0.001). OS rates at 5 years were 100% and 90.1% for high vs low TILs (log-rank p=0.007). Conclusions: This is the first demonstration of an independent prognostic role of baseline TILs in TNBC patients achieving pCR after NACT. Baseline TILs may discriminate patients who are cured after achieving pCR from patients at risk of disease relapse despite the achievement of pCR. These latter may be candidate to treatment escalation in the context of clinical trials. Our work provides relevant data in the ongoing debate on the role of adjuvant therapy (primarily pembrolizumab) following pCR. Citation Format: Maria Vittoria Dieci, Claudio Vernieri, Davide Massa, Lorenzo Nicolè, Gaia Griguolo, Federica Miglietta, Andrea Vingiani, Riccardo Lobefaro, Fabio Girardi, Grazia Vernaci, Tommaso Giarratano, Carlo Alberto Giorgi, Giancarlo Pruneri, Matteo Fassan, Filippo De Braud, Valentina Guarneri. TILs are an independent prognostic factor in early-stage TNBC patients achieving pathologic complete response after neoadjuvant chemotherapy [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-13-06.

Abstract PO3-06-14: Phase I/II study of pembrolizumab in combination with oral binimetinib in patients with unresectable locally advanced or metastatic triple-negative breast cancer

Abstract Background: Activation of the RAS/MAPK pathway is associated with reduced tumor-infiltrating lymphocytes and poor outcomes in triple-negative breast cancer (TNBC). This trial evaluated the efficacy of pembrolizumab in combination with binimetinib, a MEK inhibitor. Methods: Patients with unresectable locally advanced or metastatic TNBC with ≤ 3 prior lines of therapy were enrolled. Treatment includes a 2-week run-in with binimetinib followed by pembrolizumab. There were 2 dose levels (DL) with binimetinib at 45 mg at DL 0 and 30 mg at DL -1. A standard 3+3 design was used in Phase I, and Simon’s two-stage Optimal design was used in Phase II. PD-L1 22C3 was performed in archival samples with CPS ≥ 10 considered as positive (PD-L1+). Tumor-infiltrating lymphocytes (TILs) were quantified into 0, 1, 2, and 3+. Circulating tumor cells (CTC) and circulating cancer-associated macrophage-like cells (CAML) were isolated using CellSieve microfilters and immunofluorescently labeled with PD-L1 and p-ERK. Wilcoxon rank sum test, Chi-square test, Cox regression model, and Spearman correlation were used for analysis. Results: 22 patients were enrolled with a median age of 58 years old. Dose-limiting toxicity (DLT) was observed in 2 out of 4 patients in DL 0, with grade 3 ALT abnormality, flank pain, and nausea. In the next 6 patients in DL -1, there was 1 DLT with grade 3 AST/ALT abnormality. There were 17 patients treated with DL -1 and were evaluable for response. The objective response rate (ORR) was 29.41% (95% CI: 10.31-55.9) with 1 complete response (CR) and 4 partial responses (PR). The clinical benefit rate (CBR ≥ 24 weeks) was 35.29% (95% CI: 14.21-61.67). ORR in patients without liver metastases was 55.56% (95% CI: 21.20 - 86.30) and CBR was 66.67% (95% CI: 29.93-92.51). No response was observed in all 5 patients with liver metastases. There were 40.9% of patients with PD-L1 CPS ≥ 10. ORR in patients with PD-L1+ was 80%. However, 11.1% of patients with PD-L1 negative tumors also had an objective response and 44.4% of patients had clinical benefit ≥ 24 weeks. PD-L1 expression in archival tissue was associated with ORR (p 0.032) but not CBR (p 0.198), progression-free survival (PFS, p 0.373), and overall survival (p 0.348). TILs in archival tissue were also not associated with CBR (p 0.155) and PFS (p 0.157). One patient with TILs 0 had a stable disease ≥ 24 weeks and 2 patients TILs 1+ also had an objective response. We further evaluated blood-based biomarkers with CTC and CAML. Baseline PD-L1 in CAML (p 0.04) and decline in CAML size (p 0.02) after 1 cycle was significantly associated with CBR. However, baseline CTC count, CAML count (p 0.64), CAML size (p 0.46), p-ERK in CAML (p 0.23), and changes in CTC count, CAML count (p 0.83), p-ERK (p 0.07), and PD-L1 (p 0.08) in CAML were not significantly associated with responses. Using Cox regression analysis, a reduction in CAML count (p 0.02), CAML size (p 0.01), and PD-L1 in CAML (p 0.03) were associated with significant improvement in overall survival but not the reduction in p-ERK (p 0.6). However, PD-L1 expression in CAML is not associated with PD-L1 expression in archival tissue (Spearman correlation 0.13). Conclusions: Pembrolizumab and binimetinib at 30 mg are safe with manageable toxicities. Consistent with the preclinical data that MEKi can restore T cell function, promising activity was observed even in patients with low TILs and PD-L1 negative, particularly in patients without liver metastases. PD-L1 expression in peripheral blood CAML rather than archival tumor tissue may serve as a better biomarker to predict the clinical benefit of this combination. Early reductions in CAML count and size, were also significantly associated with responses. Future larger clinical trials are warranted to further evaluate the efficacy of this chemotherapy-free combination. Citation Format: Saranya Chumsri, Joseph Larson, Daniel Adams, Kathleen Tenner, Cha-Mei Tang, Morgan Weidner, Amanda Arnold, Dana Haley, Pooja Advani, Kostandinos Sideras, Alvaro Moreno-Aspitia, Edith Perez, Keith Knutson. Phase I/II study of pembrolizumab in combination with oral binimetinib in patients with unresectable locally advanced or metastatic triple-negative breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-06-14.

Abstract PO4-16-04: Decoding the immune landscape of breast cancer (BC) during pregnancy (PrBC): Impact of hormone receptors (HR) and tumor-infiltrating lymphocytes (TILs) phenotype on gene expression signatures

Abstract Introduction: PrBC is an uncommon malignancy with aggressive behavior. Its pathogenesis involves distinct immune mechanisms associated with maternal-fetal tolerance and tumor-host immunoediting. PrBC displays specific patterns of TILs with increased CD8+ cells. Gaining a comprehensive understanding of the molecular processes underlying this immune synergy is crucial for enhancing PrBC patients’ clinical management. Here, we sought to identify dysregulated immune-related genes in PrBC and explore their association with HR status and TILs. Methods: A total of n=75 PrBC (age range 26-43 years) and n=67 age-matched early-onset breast cancer (EOBC) in non-pregnant women (controls; age range 28-43 years) were selected from our Institutional registry. For all cases TILs were quantified according to the International TILs Working Group recommendations and profiled by IHC for CD4 and CD8. RNA was extracted from representative FFPE tissue blocks to perform the expression analysis of 395 genes involved in tumor-immune interactions using a targeted NGS panel (Oncomine™ Immune Response Research Assay, Thermofisher). Samples with >1,000,000 mapped reads and >800,000 valid reads were considered adequate. R package DESeq2 software 1.38.3 was used for sequencing depth differences normalization and differential gene expression analysis. Differentially expressed genes (DEGs) were identified based on a significant p-value (p< 0.05). Results: The comparison between PrBC and EOBC revealed a total of n=7 DEGs. All of these genes were upregulated and belonged to distinct superfamilies, including Cancer/Testis (CT) Antigen (MAGEA1/3, XAGE1B), Interferons/Cytokines (IFNA17, IFNB1), Chemokines (CXCL13), and Immunoglobulin (PECAM1/CD31). Notably, the upregulation of Chemokines in respect to EOBC was observed exclusively in HR+ PrBC, whereas triple-negative (TN) PrBC did not exhibit this pattern compared to the control group. Hence, the upregulation of ALOX15B, an enzyme involved in fatty acid peroxidation, was specific to TN PrBC. The immune signatures showed significant variations between PrBC and EOBC also based on TILs density and subpopulations. Indeed, the upregulation of CT genes (MAGEA1, XAGE1B) was exclusively observed in PrBC cases characterized by low TILs levels and prevalence of CD8+ or CD4+ cells. Finally, CD4+ TILs were absent or low in PrBC with upregulated Interferons, Cytokines, Chemokines, and Immunoglobulin genes, but present in cases with increased expression of CT and KLRF1 (NK cells). Conclusion: These findings highlight the heterogeneity and distinct molecular characteristics of PrBC and EOBC. The upregulation of specific immune-related gene families, such as CT and Chemokines, in different PrBC subtypes may suggest their potential role as actionable biomarkers (e.g. MAGEA, a well-known oncogene and potential immunotherapy target). The differences in immune signatures and TILs subpopulations further emphasize the importance of the immune microenvironment in PrBC biology and behavior. Future studies could delve deeper into the clinical implications of these gene expression patterns and explore their relevance in personalized treatment strategies for PrBC. Citation Format: Konstantinos Venetis, Elham Sajjadi, Chiara Frascarelli, Mariia Ivanova, Marianna D'Ercole, Concetta Blundo, Massimo Giroda, Eugenia Di Loreto, Giovanna Scarfone, Stefano Ferrero, Paolo Veronesi, Viviana Galimberti, Fedro Alessandro Peccatori, Nicola Fusco, Elena Guerini-Rocco. Decoding the immune landscape of breast cancer (BC) during pregnancy (PrBC): Impact of hormone receptors (HR) and tumor-infiltrating lymphocytes (TILs) phenotype on gene expression signatures [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-16-04.

Abstract PO3-14-06: Immunologic features and association with prognosis in hormone receptor-positive/HER2-negative (HR+/HER2-) metastatic breast cancer (MBC) treated with chemotherapy (CT) or CDK4/6-inhibitors (CDK4/6i) + endocrine therapy (ET)

Abstract Purpose: Little is known regarding the prognostic and predictive role of tumor immunological features in HR+/HER2- MBC treated with CT or CDK4/6i+ET. Methods: The Italian KENDO phase 2 trial randomized CT-naïve patients with HR+/HER2-neg. MBC with aggressive characteristics to receive CDK4/6i+ET (arm A) or CT+/-ET (arm B). Primary endpoint was progression-free survival (PFS). A tumor sample from the primary or metastatic tumor (archived or newly-obtained) was mandatory for inclusion. In this correlative biomarker analysis, tumor-infiltrating lymphocytes (TILs) and tertiary lymphoid structures (TLS) were detected on baseline tumor samples by hematoxylin&eosin staining, while immune genomic signatures were assessed with the nCounter BC360 panel. Immune pattern at immunohistochemistry was defined as inflamed (IF), excluded (IE) or desert (ID). TILs and signatures were dichotomized (high vs. low) with the maximally-selected rank statistics (MSRS) method. Survival analyses were conducted with the Kaplan-Meier (KM) method and differences were tested with log-rank test. Hazard ratios (HR) with 95% confidence intervals (CI) were estimated with Cox regression. Significance was set at p≤0.05. Since the trial was stopped earlier due to slow accrual all analyses were exploratory. Results: Forty-nine patients were randomized, 17 (34.7%) in arm A and 32 (65.3%) in arm B. No significant clinicopathological differences were observed at baseline between the two arms, except for tumors in arm B showing more TLS (50.0% vs. 12.5%, p=0.013). Median PFS (mPFS) with CT+/-ET was numerically shorter than mPFS with CDK4/6i+ET (11.2 vs. 19.9 months, HR: 1.41, 95%CI: 0.75-2.64, p=0.289). The median OS (mOS) for arm A was not estimable (NE) vs. 30.6 months in arm B (p=0.283). In arm A, median TILs levels were 3% (interquartile range [IQR]: 1-5%) and TLS, with/without germinal centers, were present in 2 cases (12.5%). These were also the only IF tumors in the arm, whereas the rest (87.5%) were ID. In arm B, 7 (25.0%) tumors were IF, 19 (87.5%) ID and 2 (7.1%) IE. Median TILs were 4.5% (IQR: 2-12.8%) and TLS were present in 50% cases. In arm A, ID tumors showed lower mPFS than IF (15.8 vs. 27.5 months), with concordant trend at the OS KM curve. In arm B there was no clear difference in PFS, but IF tumors did not reach mOS, differently from ID (28.7 months) and IE (28.1 months). High vs. low TILs levels were significantly associated with better PFS (p=0.003) and OS (p=0.005) in arm A. High levels of a TGFβ gene expression signature were significantly associated with better PFS (p=0.020) and OS (p=0.005) in arm A and PFS (p=0.03) in arm B; higher levels of a cytokine/chemokine signature were associatedwith better PFS (p=0.02) in arm A, and higher levels of a mastcells signature was associated with worse OS in both arms (p=0.02 and p=0.03). In arm B, higher levels of a macrophage (p < 0.001) and antigen presentation signatures (p=0.04) were associated with worse and better OS, respectively, whereas higher levels of an immune infiltration and a cytotoxic cells signature were associated with better PFS (p=0.03 both). TLS presence was numerically associated to longer mPFS (23.5 vs. 15.8 months) in arm A, with consistent OS KM curve’s trend, and longer mOS (44.5 vs. 28.1 months) in arm B. Conclusions: The KENDO trial further supports CDK4/6i+ET use in aggressive CT-naïve HR+/HER2- MBC. Biomarkers of immune activation such as IF tumors, higher TILs and presence of TLS pointed towards better survival outcomes. Genomic immunological features also showed prognostic effect, with differences according to treatment arm and/or the immune process or cell line tracked. Further research on larger cohorts is needed to confirm these preliminary findings. Citation Format: Francesco Schettini, Michela Palleschi, Francesca Mannozzi, Fara Brasó-Maristany, Lorenzo Cecconetto, Patricia Galván, Marita Mariotti, Alessia Ferrari, Emanuela Scarpi, Anna Miserocchi, Oriana Nanni, Esther Sanfeliu, Aleix Prat, Andrea Rocca, Ugo De Giorgi. Immunologic features and association with prognosis in hormone receptor-positive/HER2-negative (HR+/HER2-) metastatic breast cancer (MBC) treated with chemotherapy (CT) or CDK4/6-inhibitors (CDK4/6i) + endocrine therapy (ET) [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-14-06.

[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.