High tumor infiltrating lymphocytes are significantly associated with pathological complete response in triple negative breast cancer treated with neoadjuvant KEYNOTE-522 chemoimmunotherapy.

INTRODUCTION: For patients with locally advanced triple negative breast cancer (TNBC), the standard of care is to administer the KEYNOTE-522 (K522) regimen, including chemotherapy and immunotherapy (pembrolizumab) given in the neoadjuvant setting. Pathological complete response (pCR) is more likely in patients who receive the K522 regimen than in patients who receive standard chemotherapy. Studies have shown that pCR is a strong predictor of long-term disease-free survival. However, factors predicting pCR to K522 are not well understood and require further study in real-world populations. METHODS: We evaluated 76 patients who were treated with the K522 regimen at our institution. Twenty-nine pre-treatment biopsy slides were available for pathology review. Nuclear grade, Nottingham histologic grade, Ki-67, lymphovascular invasion, and tumor infiltrating lymphocytes (TIL) were evaluated in these 29 cases. For the cases that did not have available slides for review from pre-treatment biopsies, these variables were retrieved from available pathology reports. In addition, clinical staging, race, and BMI at the time of biopsy were retrieved from all 76 patients' charts. Binary logistic regression models were used to correlate these variables with pCR. RESULTS: At the current time, 64 of 76 patients have undergone surgery at our institution following completion of K522 and 31 (48.4%) of these achieved pCR. In univariate analysis, only TIL was significantly associated with pCR (p = 0.014) and this finding was also confirmed in multivariate analysis, whereas other variables including age, race, nuclear grade, Nottingham grade, Ki-67, lymphovascular invasion, BMI, pre-treatment tumor size, and lymph node status were not associated with pCR (p > 0.1). CONCLUSION: Our real-world data demonstrates high TIL is significantly associated with pCR rate in the K522 regimen and may potentially serve as a biomarker to select optimal treatment. The pCR rate of 48.4% in our study is lower than that reported in K522, potentially due to the smaller size of our study; however, this may also indicate differences between real-world data and clinical trial results. Larger studies are warranted to further investigate the role of immune cells in TNBC response to K522 and other treatment regimens.

Changes in expression of breast cancer tumor biomarkers between primary tumors and corresponding metastatic sites: common patterns and relationships with survival.

PURPOSE: In metastatic breast cancer, differences in expression patterns of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor-2 (HER2) between the primary tumor (PT) and metastatic site (MET) have been reported. However, there is limited understanding of the relationship of tumor subtype discordance and overall survival (OS). We evaluated patterns of ER/PR/HER2 in PTs and corresponding METs and assessed the relationship between these patterns and OS. METHODS: Patients diagnosed at our center with metastatic breast cancer (2011-2020) were included. ER/PR were stratified as < 1%/1-10%/ > 10% by immunohistochemistry and HER2 as positive/negative by immunohistochemistry/FISH. Tumor subtypes were classified as ER or PR + /HER2-, HER2+ , or triple-negative. Biomarker discordance data from PTs to METs were analyzed for expression patterns. OS was assessed. RESULTS: Of 254 patients, 41 (16.1%) had synchronous and 213 (83.9%) had metachronous METs. Category change of ER/PR/HER2 expression was observed in 56 (22.0%), 117 (40.5%), and 30 (11.8%) patients, respectively. Tumor subtype changed in 56 (22.0%) patients. We identified a difference between PT and MET from ER > 10% to ER < 1% (n = 28,16.2% p < 0.01); PR > 10% to PR < 1% (n = 54,48.2%, p < 0.001); PR > 10% to PR 1-10% (n = 18,16.1%, p < 0.001), and ER or PR+/HER2- to triple-negative (n = 19,13.0%, p = 0.03). In log-rank analysis, change from an ER or PR+/HER2- (5-year OS 88.6%) PT to a HER2+(67.5%) or triple-negative (54.6%) MET was associated with decreased survival (p < 0.01); however, in multivariate analysis, discordant biomarker expression was not associated with decreased survival (p > 0.05). CONCLUSION: Tumor expression of ER/PR/HER2 can differ between the PT and MET. Loss of ER/PR expression is common and may be related to worse survival. Routine assessment of MET tumor markers could inform prognosis and therapeutic decision-making.

Subspecialized breast pathologists have suboptimal interobserver agreement in Ki-67 evaluation using 20% as the cutoff.

PURPOSE: Ki-67 expression levels in breast cancer have prognostic and predictive significance. Therefore, accurate Ki-67 evaluation is important for optimal patient care. Although an algorithm developed by the International Ki-67 in Breast Cancer Working Group (IKWG) improves interobserver variability, it is tedious and time-consuming. In this study, we simplify IKWG algorithm and evaluate its interobserver agreement among breast pathologists in Ki-67 evaluation. METHODS: Six subspecialized breast pathologists (4 juniors, 2 seniors) assessed the percentage of positive cells in 5% increments in 57 immunostained Ki-67 slides. The time spent on each slide was recorded. Two rounds of ring study (R1, R2) were performed before and after training with the modified IKWG algorithm (eyeballing method at 400× instead of counting 100 tumor nuclei per area). Concordance was assessed using Kendall's and Kappa coefficients. RESULTS: Analysis of ordinal scale ratings for all categories with 5% increments showed almost perfect agreement in R1 (0.821) and substantial in R2 (0.793); Seniors and juniors had substantial agreement in R1 (0.718 vs. 0.649) and R2 (0.756 vs. 0.658). In dichotomous scale analysis using 20% as the cutoff, the overall agreement was moderate in R1 (0.437) and R2 (0.479), among seniors (R1: 0.436; R2: 0.437) and juniors (R1: 0.445; R2: 0.505). Average scoring time per case was higher in R2 (71 vs. 37 s). CONCLUSION: The modified IKWG algorithm does not significantly improve interobserver agreement. A better algorithm or assistance from digital image analysis is needed to improve interobserver variability in Ki-67 evaluation.

Breast Cancer With HER2 Immunohistochemical Score 2 and Average HER2 Signals/Cell 6 or More and HER2/CEP17 Ratio Less Than 2 ('ISH Group 3'): A Multiinstitutional Cohort Analysis Emphasizing Outcome and Molecular Subtype.

Breast cancer (BC) with average human epidermal growth factor receptor 2 (HER2) signals/cell ≥6 and HER2/chromosome enumeration probe 17 (CEP17) ratio <2 (in situ hybridization [ISH] group 3) is very rare, accounting for 0.4% to 3.0% of cases sent for the dual-probe ISH assay. Although such patients are currently eligible for treatment with HER2-targeted therapy, their characteristics and outcomes remain poorly understood. Sixty-two BCs with equivocal HER2 immunohistochemical score (2+) and reflex ISH group 3 results were identified across 4 institutions. Available clinicopathologic characteristics, MammaPrint and BluePrint molecular results, and follow-up information were retrospectively analyzed. Most BCs with HER2 equivocal immunohistochemical and ISH group 3 results were histologic grade 2 or 3 (100%), estrogen receptor (ER) positive (90.3%), with an average HER2 signals/cell of 7.3. Molecular profiles revealed that 80% (16/20) of tumors were luminal subtypes, and HER2 molecular subtype was identified in 10% of tumors (2/20). Twelve (19.4%) out of 62 patients developed local recurrence and/or distant metastasis with a median follow-up of 50 months. One (10%) of 10 patients achieved pathologic complete response after neoadjuvant chemotherapy. Forty-nine (79%) out of 62 patients completed anti-HER2 agents, and exploratory analysis showed no statistically significant difference in disease outcomes between patients who completed anti-HER2 treatment and those who did not. Univariate analysis revealed advanced clinical stage, and ER/progesterone receptor negativity was associated with unfavorable disease outcomes, and exploratory multivariate analysis demonstrated that clinical stage was the most significant factor associated with disease outcomes in the studied population. These findings increase our understanding of this rare, but clinically important HER2 category. Large-scale prospective randomized studies are needed to further evaluate the role of perioperative HER2-targeted therapy in this patient population.

Correlation of HER2 Protein Level With mRNA Level Quantified by RNAscope in Breast Cancer.

Trastuzumab deruxtecan (T-DXd) has been approved by the US Food and Drug Administration (FDA) to treat patients with metastatic HER2-positive and HER2-low breast cancer, and clinical trials are examining its efficacy against early-stage breast cancer. Current HER2 immunohistochemical (IHC) assays are suboptimal in evaluating HER2-low breast cancers and identifying which patients would benefit from T-DXd. HER2 expression in 526 breast cancer tissue microarray (TMA) cores was measured using the FDA-approved PATHWAY and HercepTest IHC assays, and the corresponding RNA levels were evaluated by RNAscope. HER2 protein levels by regression analysis using a quantitative immunofluorescence score against cell line arrays with known HER2 protein levels determined by mass spectrometry were available in 48 of the cores. RNAscope was also performed in 32 metastatic biopsies from 23 patients who were subsequently treated with T-DXd, and the results were correlated with response rate. HER2 RNA levels by RNAscope strongly correlated with HER2 protein levels (P < .0001) and with HER2 IHC H-scores from the PATHWAY and HercepTest assays (P < .0001). However, neither protein levels nor RNA levels significantly differed between cases scored 0, ultralow, and 1+ by PATHWAY and HercepTest. The RNA levels were significantly higher (P = .030) in responders (6.4 ± 8.2 dots/cell, n = 12) than those in nonresponders (2.6 ± 2.2, n = 20) to T-DXd. RNAscope is a simple assay that can be objectively quantified and is a promising alternative to current IHC assays in evaluating HER2 expression in breast cancers, especially HER2-low cases, and may identify patients who would benefit from T-DXd.

Reproducible Reporting of the Collection and Evaluation of Annotations for Artificial Intelligence Models.

This work puts forth and demonstrates the utility of a reporting framework for collecting and evaluating annotations of medical images used for training and testing artificial intelligence (AI) models in assisting detection and diagnosis. AI has unique reporting requirements, as shown by the AI extensions to the Consolidated Standards of Reporting Trials (CONSORT) and Standard Protocol Items: Recommendations for Interventional Trials (SPIRIT) checklists and the proposed AI extensions to the Standards for Reporting Diagnostic Accuracy (STARD) and Transparent Reporting of a Multivariable Prediction model for Individual Prognosis or Diagnosis (TRIPOD) checklists. AI for detection and/or diagnostic image analysis requires complete, reproducible, and transparent reporting of the annotations and metadata used in training and testing data sets. In an earlier work by other researchers, an annotation workflow and quality checklist for computational pathology annotations were proposed. In this manuscript, we operationalize this workflow into an evaluable quality checklist that applies to any reader-interpreted medical images, and we demonstrate its use for an annotation effort in digital pathology. We refer to this quality framework as the Collection and Evaluation of Annotations for Reproducible Reporting of Artificial Intelligence (CLEARR-AI).

Training pathologists to assess stromal tumour-infiltrating lymphocytes in breast cancer synergises efforts in clinical care and scientific research.

A growing body of research supports stromal tumour-infiltrating lymphocyte (TIL) density in breast cancer to be a robust prognostic and predicive biomarker. The gold standard for stromal TIL density quantitation in breast cancer is pathologist visual assessment using haematoxylin and eosin-stained slides. Artificial intelligence/machine-learning algorithms are in development to automate the stromal TIL scoring process, and must be validated against a reference standard such as pathologist visual assessment. Visual TIL assessment may suffer from significant interobserver variability. To improve interobserver agreement, regulatory science experts at the US Food and Drug Administration partnered with academic pathologists internationally to create a freely available online continuing medical education (CME) course to train pathologists in assessing breast cancer stromal TILs using an interactive format with expert commentary. Here we describe and provide a user guide to this CME course, whose content was designed to improve pathologist accuracy in scoring breast cancer TILs. We also suggest subsequent steps to translate knowledge into clinical practice with proficiency testing.

Mechanical Damage Caused by Compression and Its Effects on Storage Quality of Mandarin.

Mandarin is vulnerable to a range of external loads during processing and shipping, which can cause interior mechanical damage that can happen right away or over time and cause serious rotting when kept in storage. In this study, mandarin was treated to a certain quantity of compression load that did not result in a noticeable rupture of the peel. The interior pulp structure of mandarin was examined for damage prior to peel damage using CT scanning and image reconstruction. An image segmentation method based on mask processing was then used to calculate the pulp damage rate. We examined the variations in physiological activities and internal components between the test group that underwent compression load and the control group that did not undergo this type of stress during storage. The aim was to investigate the factors that contributed to the faster decay of mandarin following mechanical damage. Regression analysis was also used to establish a quantifiable relationship between the amount of compression deformation and the rates of damage and decay of mandarin during storage. The findings demonstrated that mandarin pulp exhibited visible mechanical damage when compression deformation exceeded 8 mm. This led to the disruption of physiological processes like respiration and polysaccharide breakdown, which in turn decreased the hardness of the fruit and sped up its rotting. This study identifies the critical range of compression deformation that leads to the beginning of pulp damage in mandarins. Additionally, it clarifies the quality deterioration mechanism of mandarins that have been subjected to compression damage during the storage period. Therefore, in practical production, various methods of picking, sorting, and collecting mandarins can be optimized to control the amount of compression deformation within a suitable range. This will reduce the probability of pulp damage. According to the study's conclusions, storage conditions can be optimized to regulate the physiological activities of mandarins in a targeted manner. This can minimize the probability of fruit decay and reduce economic losses.

Mycorrhizal fungi reduce the photosystem damage caused by drought stress on Paris polyphylla var. yunnanensis.

Drought stress (DS) is one of the important abiotic stresses facing cash crops today. Drought can reduce plant growth and development, inhibit photosynthesis, and thus reduce plant yield. In this experiment, we investigated the protective mechanism of AMF on plant photosynthetic system by inoculating Paris polyphylla var. yunnanensis(P.py) with a clumping mycorrhizal fungus (AMF) under drought conditions. The drought environment was maintained by weighing AMF plants and non-AMF plants. The relative water content (RWC) of plant leaves was measured to determine its drought effect. DS decreased the RWC of plants, but AMF was able to increase the RWC of plants. chlorophyll a fluorescence curve measurements revealed that DS increased the OKJIP curve of plants, but AMF was able to reduce this trend, indicating that AMF increased the light absorption capacity of plants. DS also caused a decrease in plant Y(I) and Y(II). ETRI and ETRII, and increased Y(NO) and Y(NA) in plants, indicating that DS caused photosystem damage in plants. For the same host, different AMFs did not help to the same extent, but all AMFs were able to help plants reduce this damage and contribute to the increase of plant photosynthesis under normal water conditions.

Dihydroquercetin alleviates dopamine neuron loss via regulating TREM2 activation.

BACKGROUND: Parkinson's disease (PD) is a prevalent neurodegenerative disorder, marked by the degeneration of dopamine (DA) neurons in the substantia nigra (SN). Current evidence strongly suggests that neuroinflammation, primarily mediated by microglia, contributes to PD pathogenesis. Triggering receptor expressed on myeloid cells 2 (TREM2) might serve as a promising therapeutic target for PD due to its ability to suppress neuroinflammation. Dihydroquercetin (DHQ) is an important natural dihydroflavone and confers apparent anti-inflammatory, antioxidant and anti-fibrotic effects. Recently, DHQ-mediated neuroprotection was exhibited. However, the specific mechanisms of its neuroprotective effects remain incompletely elucidated. METHODS: In this study, rat models were utilized to induce damage to DA neurons using lipopolysaccharide (LPS) and 6-hydroxydopamine (6-OHDA) to assess the impacts of DHQ on the loss of DA neurons. Furthermore, DA neuronal MN9D cells and microglial BV2 cells were employed to investigate the function of TREM2 in DHQ-mediated DA neuroprotection. Finally, TREM2 knockout mice were used to investigate whether the neuroprotective effects mediated by DHQ through a mechanism dependent on TREM2. RESULTS: The main findings demonstrated that DHQ effectively protected DA neurons against neurotoxicity induced by LPS and 6-OHDA and inhibited microglia-elicited neuroinflammation. Meanwhile, DHQ promoted microglial TREM2 signaling activation. Notably, DHQ failed to reduce inflammatory cytokines release and further present neuroprotection from DA neurotoxicity upon TREM2 silencing. Similarly, DHQ didn't exert DA neuroprotection in TREM2 knockout mice. CONCLUSIONS: These findings suggest that DHQ exerted DA neuroprotection by regulating microglia TREM2 activation.

Role of Rubus chingii BBX gene family in anthocyanin accumulation during fruit ripening.

The B-box (BBX) family, which is a class of zinc finger transcription factors, exhibits special roles in plant growth and development as well as in plants' ability to cope with various stresses. Even though Rubus chingii is an important traditional medicinally edible plant in east Asia, there are no comprehensive studies of BBX members in R. chingii. In this study, 32 RcBBX members were identified, and these were divided into five groups. A collinearity analysis showed that gene duplication events were common, and when combined with a motif analysis of the RcBBX genes, it was concluded that group V genes might have undergone deletion of gene fragments or mutations. Analysis of cis-acting elements revealed that each RcBBX gene contained hormone-, light-, and stress-related elements. Expression patterns of the 32 RcBBX genes during fruit ripening revealed that highest expression occurred at the small green fruit stage. Of note, the expression of several RcBBX genes increased rapidly as fruit developed. These findings, combined with the expression profiles of anthocyanin biosynthetic genes during fruit ripening, allowed us to identify the nuclear-targeted RcBBX26, which positively promoted anthocyanin production in R. chingii. The collective findings of this study shed light on the function of RcBBX genes in different tissues, developmental stages, and in response to two abiotic stresses.

Integrative spatial omics reveals distinct tumor-promoting multicellular niches and immunosuppressive mechanisms in African American and European American patients with TNBC.

Racial disparities in triple-negative breast cancer (TNBC) outcomes have been reported. However, the biological mechanisms underlying these disparities remain unclear. We integrated imaging mass cytometry and spatial transcriptomics, to characterize the tumor microenvironment (TME) of African American (AA) and European American (EA) patients with TNBC. The TME in AA patients was characterized by interactions between endothelial cells, macrophages, and mesenchymal-like cells, which were associated with poor patient survival. In contrast, the EA TNBC-associated niche is enriched in T-cells and neutrophils suggestive of an exhaustion and suppression of otherwise active T cell responses. Ligand-receptor and pathway analyses of race-associated niches found AA TNBC to be immune cold and hence immunotherapy resistant tumors, and EA TNBC as inflamed tumors that evolved a distinctive immunosuppressive mechanism. Our study revealed the presence of racially distinct tumor-promoting and immunosuppressive microenvironments in AA and EA patients with TNBC, which may explain the poor clinical outcomes.

Validation Study of the Newly Proposed Refined Diagnostic Criteria for Malignant Phyllodes Tumor With 136 Borderline and Malignant Phyllodes Tumor Cases.

The World Health Organization (WHO) diagnostic criteria for malignant phyllodes tumor (MPT) may miss a significant number of MPTs with metastatic potential. New refined diagnostic criteria (Refined Criteria) for MPT were recently proposed. The aim of this study is to validate the Refined Criteria. This validation study included 136 borderline (borderline phyllodes tumor [BoPT]) and MPT cases that were not included in the initial study. We evaluated tumor classifications based on both the Refined Criteria and the WHO criteria. The Refined Criteria defines MPT when these criteria are met (1) stromal overgrowth with ≥ 1 feature(s) of marked stromal cellularity, marked stromal cytologic atypia, or ≥10 mitoses per 10 high-power fields (10 mitoses/10 HPFs) or (2) marked stromal cellularity with ≥1 feature(s) of marked stromal cytologic atypia, ≥10 mitoses/10 HPFs or permeative border. The WHO criteria require all 5 morphologic features (stromal overgrowth, permeative border, marked stromal cellularity, marked stromal cytologic atypia, and ≥10 mitoses/10 HPFs) for an MPT diagnosis. Using the Refined Criteria, none of the 61 BoPTs developed metastasis and 40.0% of the 75 MPTs developed metastases; local recurrence was seen in 11.5% BoPTs and 25.3% MPTs. Using the WHO criteria, 9.6% of the 94 BoPTs developed metastases and 50.0% of the 42 MPTs developed metastases; 14.9% of the BoPTs had local recurrence and 28.6% of the MPTs had local recurrence. Nine (30.0%) of the 30 tumors that developed distant metastases were diagnosed as BoPTs by the WHO criteria. When we combined the 75 MPTs from this validation cohort with the 65 MPT cases from the published data using the Refined Criteria, 50 (35.7%) of the 140 MPTs developed metastases, whereas 8 cases with metastases were <5 cm. In the univariate analysis with log-rank test, stromal overgrowth, marked stromal cellularity, marked stromal cytologic atypia, ≥10 mitoses/10 HPFs, presence of heterologous components other than liposarcomatous component, and presence of stromal necrosis were significantly associated with the risk of metastasis (all with P < 0.05). In multivariate analysis with Cox proportional hazard regression, stromal overgrowth and marked stromal cellularity were significantly associated with metastasis (both with P < 0.001). The Refined Criteria are superior to the WHO criteria in predicting the clinical outcomes of BoPTs and MPTs. Using the Refined Criteria, 35.7% of 140 patients with MPT developed metastases, whereas none (0%) of the patients with BoPT developed metastases. Patients with MPT have a high metastatic rate; these patients may benefit from systemic chemotherapy or targeted therapies. In contrast, patients with BoPT may be managed with complete local excision alone without chemotherapy.

NOS2 and COX2 Provide Key Spatial Targets that Determine Outcome in ER- Breast Cancer.

Estrogen receptor-negative (ER-) breast cancer is an aggressive breast cancer subtype with limited therapeutic options. Upregulated expression of both inducible nitric oxide synthase (NOS2) and cyclo-oxygenase (COX2) in breast tumors predicts poor clinical outcomes. Signaling molecules released by these enzymes activate oncogenic pathways, driving cancer stemness, metastasis, and immune suppression. The influence of tumor NOS2/COX2 expression on the landscape of immune markers using multiplex fluorescence imaging of 21 ER- breast tumors were stratified for survival. A powerful relationship between tumor NOS2/COX2 expression and distinct CD8+ T cell phenotypes was observed at 5 years post-diagnosis. These results were confirmed in a validation cohort using gene expression data showing that ratios of NOS2 to CD8 and COX2 to CD8 are strongly associated with poor outcomes in high NOS2/COX2-expressing tumors. Importantly, multiplex imaging identified distinct CD8+ T cell phenotypes relative to tumor NOS2/COX2 expression in Deceased vs Alive patient tumors at 5-year survival. CD8+NOS2-COX2- phenotypes defined fully inflamed tumors with significantly elevated CD8+ T cell infiltration in Alive tumors expressing low NOS2/COX2. In contrast, two distinct phenotypes including inflamed CD8+NOS2+COX2+ regions with stroma-restricted CD8+ T cells and CD8-NOS2-COX2+ immune desert regions with abated CD8+ T cell penetration, were significantly elevated in Deceased tumors with high NOS2/COX2 expression. These results were supported by applying an unsupervised nonlinear dimensionality-reduction technique, UMAP, correlating specific spatial CD8/NOS2/COX2 expression patterns with patient survival. Moreover, spatial analysis of the CD44v6 and EpCAM cancer stem cell (CSC) markers within the CD8/NOS2/COX2 expression landscape revealed positive correlations between EpCAM and inflamed stroma-restricted CD8+NOS2+COX2+ phenotypes at the tumor/stroma interface in deceased patients. Also, positive correlations between CD44v6 and COX2 were identified in immune desert regions in deceased patients. Furthermore, migrating tumor cells were shown to occur only in the CD8-NOS2+COX2+ regions, identifying a metastatic hot spot. Taken together, this study shows the strength of spatial localization analyses of the CD8/NOS2/COX2 landscape, how it shapes the tumor immune microenvironment and the selection of aggressive tumor phenotypes in distinct regions that lead to poor clinical outcomes. This technique could be beneficial for describing tumor niches with increased aggressiveness that may respond to clinically available NOS2/COX2 inhibitors or immune-modulatory agents.

Spatial analysis of NOS2 and COX2 interaction with T-effector cells reveals immunosuppressive landscapes associated with poor outcome in ER- breast cancer patients.

Multiple immunosuppressive mechanisms exist in the tumor microenvironment that drive poor outcomes and decrease treatment efficacy. The co-expression of NOS2 and COX2 is a strong predictor of poor prognosis in ER- breast cancer and other malignancies. Together, they generate pro-oncogenic signals that drive metastasis, drug resistance, cancer stemness, and immune suppression. Using an ER- breast cancer patient cohort, we found that the spatial expression patterns of NOS2 and COX2 with CD3+CD8+PD1- T effector (Teff) cells formed a tumor immune landscape that correlated with poor outcome. NOS2 was primarily associated with the tumor-immune interface, whereas COX2 was associated with immune desert regions of the tumor lacking Teff cells. A higher ratio of NOS2 or COX2 to Teff was highly correlated with poor outcomes. Spatial analysis revealed that regional clustering of NOS2 and COX2 was associated with stromal-restricted Teff, while only COX2 was predominant in immune deserts. Examination of other immunosuppressive elements, such as PDL1/PD1, Treg, B7H4, and IDO1, revealed that PDL1/PD1, Treg, and IDO1 were primarily associated with restricted Teff, whereas B7H4 and COX2 were found in tumor immune deserts. Regardless of the survival outcome, other leukocytes, such as CD4 T cells and macrophages, were primarily in stromal lymphoid aggregates. Finally, in a 4T1 model, COX2 inhibition led to a massive cell infiltration, thus validating the hypothesis that COX2 is an essential component of the Teff exclusion process and, thus, tumor evasion. Our study indicates that NOS2/COX2 expression plays a central role in tumor immunosuppression. Our findings indicate that new strategies combining clinically available NOS2/COX2 inhibitors with various forms of immune therapy may open a new avenue for the treatment of aggressive ER-breast cancers.