CXCL9 Is a Potential Biomarker of Immune Infiltration Associated With Favorable Prognosis in ER-Negative Breast Cancer

The chemokine CXCL9 (C-X-C motif chemokine ligand 9) has been reported to be required for antitumour immune responses following immune checkpoint blockade. In this study, we sought to investigate the potential value of CXCL9 according to immune responses in patients with breast cancer (BC). A variety of open-source databases and online tools were used to explore the expression features and prognostic significance of CXCL9 in BC and its correlation with immune-related biomarkers followed by subsequent verification with immunohistochemistry experiments. The CXCL9 mRNA level was found to be significantly higher in BC than in normal tissue and was associated with better survival outcomes in patients with ER-negative tumours. Moreover, CXCL9 is significantly correlated with immune cell infiltration and immune-related biomarkers, including CTLA4, GZMB, LAG3, PDCD1 and HAVCR2. Finally, we performed immunohistochemistry with breast cancer tissue samples and observed that CXCL9 is highly expressed in the ER-negative subgroup and positively correlated with the immune-related factors LAG3, PD1, PDL1 and CTLA4 to varying degrees. These findings suggest that CXCL9 is an underlying biomarker for predicting the status of immune infiltration in ER-negative breast cancer

Assessing the role of tumour-associated macrophage subsets in breast cancer subtypes using digital image analysis

Purpose: The number of M1-like and M2-like tumour-associated macrophages (TAMs) and their ratio can play a role in breast cancer development and progression. Early clinical trials using macrophage targeting compounds are currently ongoing. However, the most optimal detection method of M1-like and M2-like macrophage subsets and their clinical relevance in breast cancer is still unclear. We aimed to optimize the assessment of TAM subsets in different breast cancer subtypes, and therefore related TAM subset numbers and ratio to clinicopathological characteristics and clinical outcome. Methods: Tissue microarrays of 347 consecutive primary Luminal-A, Luminal-B, HER2-positive and triple-negative tumours of patients with early-stage breast cancer were serially sectioned and immunohistochemically stained for the pan-macrophage marker CD68 and the M2-like macrophage markers CD163, CSF-1R and CD206. TAM numbers were quantified using a digital image analysis algorithm. M1-like macrophage numbers were calculated by subtracting M2-like TAM numbers from the total TAM number. Results: M2-like markers CD163 and CSF-1R showed a moderate positive association with each other and with CD68 (r ≥ 0.47), but only weakly with CD206 (r ≤ 0.06). CD68 + , CD163 + and CSF-1R + macrophages correlated with tumour grade in Luminal-B tumours (P < 0.001). Total or subset TAM numbers did not correlate with disease outcome in any breast cancer subtype. Conclusion: In conclusion, macrophages and their subsets can be detected by means of a panel of TAM markers and are related to unfavourable clinicopathological characteristics in Luminal-B breast cancer. However, their impact on outcome remains unclear. Preferably, this should be determined in prospective series

A nomogram to predict the probability of axillary lymph node metastasis in early breast cancer patients with positive axillary ultrasound

Among patients with a preoperative positive axillary ultrasound, around 40% of them are pathologically proved to be free from axillary lymph node (ALN) metastasis. We aimed to develop and validate a model to predict the probability of ALN metastasis as a preoperative tool to support clinical decision-making. Clinicopathological features of 322 early breast cancer patients with positive axillary ultrasound findings were analyzed. Multivariate logistic regression analysis was performed to identify independent predictors of ALN metastasis. A model was created from the logistic regression analysis, comprising lymph node transverse diameter, cortex thickness, hilum status, clinical tumour size, histological grade and estrogen receptor, and it was subsequently validated in another 234 patients. Coefficient of determination (R-2) and the area under the ROC curve (AUC) were calculated to be 0.9375 and 0.864, showing good calibration and discrimination of the model, respectively. The false-negative rates of the model were 0% and 5.3% for the predicted probability cut-off points of 7.1% and 13.8%, respectively. This means that omission of axillary surgery may be safe for patients with a predictive probability of less than 13.8%. After further validation in clinical practice, this model may support increasingly limited surgical approaches to the axilla in breast cancer

Considering the biology of late recurrences in selecting patients for extended endocrine therapy in breast cancer

Extended endocrine therapy can reduce recurrences occurring more than 5 years after diagnosis (late recurrences) in estrogen receptor (ER)-positive breast cancer. Given the side effects of endocrine therapy, optimal patient selection for extended treatment is crucial. Enhanced understanding of late recurrence biology could optimize patient selection in this setting. We therefore summarized the current knowledge of late recurrence biology, clinical trials on extended endocrine therapy, and tools for predicting late recurrence and benefit from treatment extension. Extending 5 years of tamoxifen therapy with 5 years of tamoxifen or an aromatase inhibitor (AI) reduces late recurrence risk by 2-5%, but results of extending AI-based therapy are inconsistent. Although several clinicopathological parameters and multigene assays are prognostic for late recurrence, selection tools predicting benefit from extended endocrine therapy are sparse. Therefore, we additionally performed a pooled analysis using 2231 mRNA profiles of patients with ER-positive/human epidermal growth factor receptor 2 negative breast cancer. Gene Set Enrichment Analysis was applied on genes ranked according to their association with early and late recurrence risk. Higher expression of estrogen-responsive genes was associated with a high recurrence risk beyond 5 years after diagnosis when patients had received no systemic therapy. Although 5 years of endocrine therapy reduced this risk, this effect disappeared after treatment cessation. This suggests that late recurrences of tumors with high expression of estrogen-responsive genes are likely ER-driven. Long-term intervention in this pathway by means of extended endocrine therapy might reduce late recurrences in patients with tumors showing high expression of estrogen-responsive genes

Fluorescent image-guided surgery in breast cancer by intravenous application of a quenched fluorescence activity-based probe for cysteine cathepsins in a syngeneic mouse model

PURPOSE: The reoperation rate for breast-conserving surgery is as high as 15-30% due to residual tumor in the surgical cavity after surgery. In vivo tumor-targeted optical molecular imaging may serve as a red-flag technique to improve intraoperative surgical margin assessment and to reduce reoperation rates. Cysteine cathepsins are overexpressed in most solid tumor types, including breast cancer. We developed a cathepsin-targeted, quenched fluorescent activity-based probe, VGT-309, and evaluated whether it could be used for tumor detection and image-guided surgery in syngeneic tumor-bearing mice. METHODS: Binding specificity of the developed probe was evaluated in vitro. Next, fluorescent imaging in BALB/c mice bearing a murine breast tumor was performed at different time points after VGT-309 administration. Biodistribution of VGT-309 after 24 h in tumor-bearing mice was compared to control mice. Image-guided surgery was performed at multiple time points tumors with different clinical fluorescent camera systems and followed by ex vivo analysis. RESULTS: The probe was specifically activated by cathepsins X, B/L, and S. Fluorescent imaging revealed an increased tumor-to-background contrast over time up to 15.1 24 h post probe injection. In addition, VGT-309 delineated tumor tissue during image-guided surgery with different optical fluorescent imaging camera systems. CONCLUSION: These results indicate that optical fluorescent molecular imaging using the cathepsin-targeted probe, VGT-309, may improve intraoperative tumor detection, which could translate to more complete tumor resection when coupled with commercially available surgical tools and techniques