Adenylate kinase 5 (AK5) is a poorly characterized metabolic enzyme with unknown roles in breast cancer. Single-cell transcriptomics revealed AK5 enrichment in malignant epithelial cells, and its low expression correlated with poor patient prognosis, suggesting tumor-suppressive functions. Functionally, AK5 overexpression inhibited, while its knockdown promoted, breast cancer cell proliferation, migration, and invasion. Mechanistically, AK5, dependent on its kinase activity, post-transcriptionally suppressed miR-182-5p maturation, thereby de-repressing-L1 expression. In vivo experiments have proved that AK5 overexpression attenuated tumor growth and synergized with anti-PD-L1 therapy. Our work defines AK5 as a novel tumor suppressor, unveils a kinase-dependent non-canonical role in regulating an immune checkpoint via miR-182-5p, and nominates it as a potential therapeutic target to sensitize tumors to immunotherapy.

Background:Currently, surgical resection remains the first choice for treating early breast cancer. Cryoablation, a minimally invasive ablative technique, can inactivate the target tumor in situ, after which patients typically require further treatments, including surgery. However, reports are limited on the impact of cryoablation on subsequent breast-conserving surgery (BCS). This study aims to clarify the role of preoperative cryoablation in guiding BCS for early breast cancer, specifically whether cryoablation could influence the rate of positive or close resection margin in BCS. Methods:This study retrospectively analyzed the clinical characteristics and treatment data of patients who underwent BCS at our center between December 2024 and December 2025. After propensity score matching (PSM), 111 patients were paired: 37 in the cryoablation group and 74 in the surgery group. The clinical data of both groups were compared to evaluate the feasibility of cryoablation-guided BCS for early-stage breast cancer, its impact on the outcomes of BCS and sentinel lymph node biopsy (SLNB). Results:Ultrasound-guided cryoablation was successfully performed under local anesthesia in all 37 patients, with a complete ablation rate of 94.59% [35/37; 95% confidence interval (CI): 82.3-98.5%]. Compared to the surgery group, the cryoablation group had a lower rate of positive or close margin for BCS (5.41% vs. 32.43%, P=0.002), and a higher rate of accurate surgery (83.78% vs. 59.46%, P=0.01). The mean duration of BCS was shorter in the cryoablation group than in the surgery group (76.65±8.17 vs. 86.50±16.29 min, P<0.001). Additionally, the mean post-operative length of stay (LOS) in the cryoablation group was approximately one day shorter than that in the surgery group (1.59 vs. 2.62 days, P<0.001). Furthermore, the success rate of SLNB was 100% in both groups. Conclusions:This retrospective study demonstrated that cryoablation-guided BCS was feasible and safe. Compared to standard BCS, cryoablation-guided BCS was associated with a higher accuracy in resection, a lower positive or close margin rate, shorter surgical duration and post-operative LOS. Additionally, cryoablation did not negatively impact the success rate of subsequent SLNB.

Chemoresistance remains a major obstacle to effective breast cancer therapy and is driven in part by metabolic reprogramming and dysregulated RNA-mediated signaling. Although tRNA-derived fragments (tRFs) have emerged as important regulators of cellular stress responses, their roles in drug resistance remain incompletely understood. Here, using high-throughput sequencing, we identified tRF-1432 as a markedly upregulated tRF in chemoresistant breast cancer tissues and cell lines, which was further validated in clinical specimens. Functional assays and syngeneic tumor models demonstrated that tRF-1432 enhances resistance to adriamycin by promoting tumor cell survival and suppressing apoptosis under chemotherapeutic stress. Mechanistically, tRF-1432 is a 5'-tRNA^Val-CAC-derived fragment generated by angiogenin cleavage and directly interacts with the RNA-binding protein RBMS1. This interaction attenuates RBMS1-mediated destabilization of IMPDH2 mRNA, leading to increased IMPDH2 expression. As a result, purine metabolic reprogramming is enhanced, intracellular GTP levels are elevated, and proliferative and survival signaling is sustained in the presence of chemotherapy. Importantly, pharmacological inhibition of IMPDH2 using mycophenolic acid (MPA), an FDA-approved drug, effectively reversed chemoresistance both in vitro and in vivo. Collectively, our findings establish a previously unrecognized tRF-1432/RBMS1/IMPDH2 regulatory axis that drives metabolic adaptation and chemoresistance in breast cancer, highlighting this pathway as a potential therapeutic target for overcoming drug resistance.

Purpose To establish a predictive model for the sonication energy required for focused ultrasound surgery (FUS) of breast fibroadenomas.Methods This study retrospectively enrolled 87 patients with 154 benign breast tumors treated by FUS in our hospital. Radiomic analysis included 124 tumors from 69 patients, randomly split into a 3:1 ratio for training (96 cases) and validation (28 cases). Three machine learning algorithms were applied for feature selection. Then, all the selected features were used for the construction of the prediction model via four machine learning algorithms. Residual analysis and Intraclass Correlation Coefficient (ICC) analysis were performed to evaluate the performances of these four models. The importance of each feature is demonstrated by the Root Mean Square Error (RMSE) loss obtained through permutation importance measurement.Results This study collected 11 clinical features and 68 ultrasound radiomics features, totaling 79 independent variables. The Bagging Tree Model, characterized by lower and stable RMSE values and high R2 stability with increasing features, demonstrated superior predictive accuracy and explanatory power compared to other models. At the optimal feature count, identified by the minimum RMSE, 33 features were selected for further modeling. The bagging tree model has the highest ICC value among the four models, at 0.56, with a confidence interval of (0.23, 0.77).Conclusions This study established an interpretable machine learning model that integrates clinical and ultrasound radiomics features to estimate the sonication energy in FUS treatment of breast fibroadenomas.

Background:Due to the undesirable cumulative toxicity of multiple drugs, de-escalated neoadjuvant chemotherapy strategies are needed for human epidermal growth factor receptor 2 (HER2)-positive breast cancer. Pyrotinib, a small-molecule irreversible pan-HER receptor tyrosine kinase inhibitor, shows promising efficacy in the neoadjuvant setting. We aimed to determine the efficacy, safety and predictive biomarkers of the de-escalated neoadjuvant nab-paclitaxel combined with pyrotinib and trastuzumab in intrinsic HER2-enriched breast cancer. Methods:In this multicenter phase 2 study (NCT05659056), patients who were histologically diagnosed with HER2-positive breast cancer (clinical stage ⅡA-ⅢC) were deemed suitable to participate in this study. Participants received pyrotinib (400 mg once), trastuzumab (8 mg/kg loading dose, followed by 6 mg/kg maintenance dose), and nab-paclitaxel (260 mg/m2) on day 1 of each 3-week cycle for six cycles. The primary endpoint was the rate of total pathological complete response (tpCR) among BluePrint HER2-enriched breast cancers, which was defined as complete disappearance of invasive tumor in breast specimen and all sampled axillary lymph nodes (ypT0/is, ypN0). This study has been completed. Findings:Between 3 December 2022 and 6 June 2024, 74 participants were finally enrolled in the study. Of all enrolled participants, 66 had baseline BluePrint and MammaPrint results. Among the 43 participants with BluePrint HER2-enriched breast cancer, 23 achieved tpCR (53%, 95% CI 38%-69%), and 28 achieved breast pathological complete response (bpCR) (65%, 95% CI 49%-79%). Among the 23 participants with non-HER2-enriched subtypes, 7 achieved tpCR (30%, 95% CI 13%-53%), while 10 achieved bpCR (43%, 95% CI 23%-66%). Of 66 participants with MammaPrint risk score index, the tpCR rate in MammaPrint ultra-high group (24/39) was significantly higher than that in high group (6/27, P = 0.0024). With the median follow-up of 19.9 months (IQR, 15.5-25.4), no cases of recurrence, metastasis, or mortality events were observed. Grade 3-4 treatment-related adverse events occurred in 17 (23%) participants. The most common grade 3-4 adverse event was diarrhea (10/74). No treatment-related deaths occurred. Of all enrolled participants, no treatment discontinuations occurred due to disease progression during the neoadjuvant therapy period. Interpretation:De-escalated neoadjuvant cytotoxic chemotherapy regimen is promising for BluePrint HER2-enriched breast cancer. Our results provide critical references for the efficacy and biomarkers of de-escalated neoadjuvant therapy in HER2-positive breast cancer. Funding:National Natural Science Foundation of China and Natural Science Foundation of Jiangsu Province.

Background: Breast cancer is one of the most common malignant tumors in women. Mitochondria are critical components within cells. Changes in mitochondrial metabolism play a crucial role in regulating tumor cells and further impacting clinical prognosis. However, their interplay in breast cancer remains a pending issue. This study aimed to reveal the relationship between mitochondrial metabolism-related genes and breast cancer by constructing risk features. Methods: Breast cancer transcriptome chip data were obtained from TCGA and GEO, while mitochondrial gene data were downloaded from the MitoCarta3.01 database. The "ConsensusClusterPlus" package was used for clustering analysis, with subsequent processing for GSEA, GO, and KEGG pathway analyses. A prognosis model for breast cancer was established using Cox regression and LASSO algorithms. The CIBERSORT algorithm and MCPcounter algorithm were utilized to calculate each patient's immune cell infiltration level. Additionally, the expression of pivotal genes in breast cancer patient tissue specimens and cell models was validated to verify their association with biological functions in breast cancer cells. Results: We employed the LASSO regression algorithm to reveal mitochondrial metabolism-related genes in the TCGA BRCA dataset and identified four prognosis-related genes (MYH11, LTF, FADS2, and PSPHP1). These findings were further validated through analysis using the GEO dataset. Based on the four-gene prognosis model derived from mitochondrial metabolism, breast cancer patients with a high-risk score had shorter overall survival compared to the low RS patient group. Immunological analysis revealed that patients with higher risk scores were more likely to be unresponsive to immunotherapy but more sensitive to several conventional chemotherapies, suggesting the potential strategy of combining chemotherapy with immunotherapy to enhance the efficacy of current T cell-based immunotherapy. Univariate and multivariate Cox regression analyses validated the mitochondrial gene model as an independent prognostic indicator for predicting overall survival. They established a nomogram based on the mitochondrial gene model to predict the prognosis of breast cancer. Inpatient tissue validation, pivotal genes exhibited an expression trend consistent with the results of bioinformatic analysis. Finally, through cellular experiments, we verified the expression and function of the key gene FADS2, finding that FADS2 was highly expressed in breast cancer cell lines, and upon its knockout, the cell's invasion, migration, and colony-forming abilities were significantly reduced. Conclusion: This work demonstrates that prognostic features based on mitochondrial-related genes are related to clinical outcomes, tumor progression, and genetic alterations. Our research findings may provide insights for developing new targets for breast cancer treatment, early intervention, and prognosis prediction. Citation Format: Yangyang Cui, Yakun Kang, Yiqin Xia, Yue Huang, Shui Wang, Hui Xie. Mitochondrial Metabolism-Related Features Guiding Precision Subtyping and Prognosis in Breast Cancer, Revealing FADS2 as a Novel Therapeutic Target [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P3-02-05.

Chemoresistance is a significant challenge in the field of oncology and is the leading cause of treatment failure in breast cancer. Cancer cells could adapt to hypoxia microenvironment through metabolic reprogramming, while developing a strong drug resistance phenotype at the same time. However, the mechanism under which is still largely unknown. Efforts need to be done to identify novel targets and treatment modalities to address the complex issue of chemoresistance in breast cancer. In this study, we set our sights on an upregulated tRNA-derived fragment, tRF-1432, while we screened through high-throughput sequencing in breast cancer hypoxia models, especially in drug-resistant breast cancer cells/tissues. tRFs are a new category of small non-coding RNAs that occur when pre-tRNAs or mature tRNAs are selectively sheared by enzymes under certain pressure, and which play crucial roles in tumorigenesis. Further investigation on the biological roles and clinical value of tRF-1432 has been done. Overexpression of tRF-1432 is positively correlated with tumor cell proliferation and anti-apoptosis ability, and significantly increased chemoresistance of breast cancer in vitro and in vivo. Immunoprecipitation experiments confirmed the direct bond between tRF-1432 and RBMS1, knockdown of RBMS1 may deprive the tumor-promoting and chemoresistance properties of tRF-1432. Furthermore, in tRF-1432 overexpression breast cancer cells we observed a significant decrease of CCL3, a gene which could induce the transition in the phenotype of macrophages into a proinflammatory one. As a crucial member of tumor microenvironment (TME), macrophages participate in the mediation of therapeutic responses. Silencing tRF-1432 could facilitate M1 macrophage polarization and enhance the response to doxorubicin chemosensitivity in breast cancer. Mechanismly, tRF-1432 decreased the stability of CCL3-mRNA with the cooperation of RNA binding protein RBMS1, thereby reduced M1 macrophage polarization and promote the progression and chemoresistance breast cancer via CCL3-CCR5 signaling. In conclusion, our study clarified the role of hypoxia-induced tRF-1432 in breast cancer development and chemoresistance, and elucidate the molecular mechanism of which, aiming to provide a novel target for reversing chemoresistance in breast cancer. Citation Format: Yuhan Dai, Shuhan Zhao, Huilin Chen, Yue Huang, Yiqin Xia, Jinhui Peng, Jiangdong Jin, Yifan Wu, Shui Wang, Yangyang Cui, Hui Xie. tRF-1432–a novel target for reversing chemoresistance in breast cancer [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P3-02-17.

Radiofrequency ablation (RFA) and microwave ablation (MWA) have been investigated as treatments for early-stage breast cancer. However, it is unclear which minimally invasive thermal therapy demonstrates superior local efficacy. Also, the cytolytic functions of peripheral T cells after thermal ablation in solid tumors have not been reported. In this study, 60 patients with breast cancer were enrolled from two clinical trials conducted between March 2020 and December 2021. The local effect of thermal ablation evaluated through pathological examinations or radiological imaging was the primary outcome. The secondary outcome involved systemic T-cell responses. Peripheral blood samples were collected before and after treatments. The ablation-induced immune responses were analyzed using flow cytometry, enzyme-linked-immunosorbent assay (ELISA), quantitative real-time PCR (qRT-PCR) and single-cell RNA sequencing. Both RFA and MWA showed favorable local effects in the treatment of breast cancer. Compared to surgery, RFA increased peripheral CD8 + T-cell proportions but did not enhance their cytolytic functions. Conversely, MWA induced stronger cytolytic functions of peripheral T cells and upregulated memory CD4 + T cells. The distinct immune responses induced by MWA and RFA were associated with variations in antigen presentation pathways, types of antigen-presenting cells (APCs), and cytokine secretion profiles. Single-cell RNA sequencing further revealed that dendritic cells were the APCs activated by MWA, exhibiting upregulated fatty acid metabolism. Both thermal ablation therapies are technically feasible for early-stage breast cancer. However, MWA appears superior in enhancing the cytolytic functions of peripheral T cells compared to RFA. This study provides the first mechanistic insight into the different immune responses induced by MWA and RFA, although future clinical trials are necessary to validate these findings. ChiCTR2000029665. Registered February 09, 2020 ( https://www.chictr.org.cn/showproj.html?proj=48315 ) and ChiCTR2000029155. Registered January 16, 2020 ( https://www.chictr.org.cn/showproj.html?proj=48314 ).

Globally, breast cancer remains one of the most prevalent malignancies and a leading cause of cancer-related death in women, with over 2.3 million new cases reported annually. Despite treatment advances, one breast cancer type in particular, HER2-negative breast cancer, lacks precise therapeutic targets. Given the role of N7-methylguanosine (m7G) in gene regulation and its links to cancer progression, we investigated m7G regulatory gene expression and prognostic potential in HER2-negative breast cancer. We analysed publicly available breast cancer datasets (The Cancer Genome Atlas and the Gene Expression Omnibus (GEO)) to analyse the differential expression of 14 m7G-regulatory genes. Clustering analysis, based on m7G patterns, categorised HER2-negative patients into two subgroups. A prognostic model was established through LASSO and Cox regression; subsequently validated by survival analysis, and further supported by functional assays confirming gene function. Our model identified CCNB1 and EIF4E as high-risk genes, with EIF4E overexpression enhancing cell proliferation, migration and invasion. RNA-sequencing (RNA-seq) and pathway analyses showed that upregulated EIF4E activated Wnt signalling and extracellular matrix (ECM) components, processes required for tumour progression. High-risk patients showed reduced immune cell infiltration and poorer survival outcomes. We highlight m7G regulatory gene potential, particularly EIF4E, as prognostic markers and therapeutic targets for HER2-negative breast cancer. Targeting EIF4E-related pathways could provide new therapeutic strategies to improve breast cancer patient outcomes.

Introduction: Breast conserving surgery (BCS) with radiation is accepted as a standard local treatment for early-stage breast cancer. Advantages in survival and quality of life (QoL) have led to a higher demand for BCS. Success of BCS is characterized by negative margins and a good cosmetic outcome. The aims of this cohort study were to determine whether wire-guided localization (WGL) combined with multidetector CT (MDCT) guided 3-dimensional (3D) reconstruction could guide precision breast conserving surgery (PBCS), and to access the cosmetic outcome reported by patients. Methods: 31 patients with unifocal breast cancer were enrolled for PBCS guided by WGL combined with MDCT guided 3D reconstruction from 2021 to 2022. Under local anesthesia, surrounded WGL was performed, followed by an immediate contrast enhanced MDCT scan. One day after the localization, PBCS guided by MDCT guided 3D reconstruction was performed. 61 women who underwent palpation guided breast conserving surgery (BCS) were included as control. Two-sided Student t test, Fisher’s exact test and chi-square test was applied. Results: Breast Cancer Treatment Outcome Scale (BCTOS) cosmetic subscale was used to access patient reported cosmetic outcomes in our study. In the PBCS group, the results indicated no significant changes (P=0.168) between the baseline assessment and 1 month post operation. This is later followed by adverse changes (P=0.016) between 1 month post operation and 6 months post radiotherapy (RT). In the control group, the results indicated adverse changes (P 0.001) between the baseline assessment and 1 month post operation. This is later followed by deterioration (P=0.028) between 1 month post operation and 6 months post RT. Between the two groups, there was no significant difference noted for the score of the BCTOS at baseline, 1 month post surgery and 6 months post RT. Conclusion: For patients with unifocal breast cancer, WGL combined with MDCT guided 3D reconstruction could achieve better cosmetic outcomes. Citation Format: Yiqin Xia, Yangyang Cui, Yue Huang, Jinghui Peng, Meng Zhao, Hui Xie, Shui Wang. Localization combined with multidetector CT 3-dimensional image reconstruction guided precision breast conserving surgery versus conventional breast conserving surgery: cosmetic analysis of the single center, prospective, cohort study [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P5-09-03.

Hormone receptor-positive (HR+)/human epidermal growth factor receptor 2-negative (HER2-) is the most common molecular subtype in breast cancer (BC), but drug resistance remains an unsolved problem, particularly in metastatic ones. Cell-cycle related genes play a crucial role in tumorigenesis and progression. However, their relationship with drug resistance and patient prognosis is not yet clear. Here, we introduce a novel and robust HR+/HER2- BC Prognostic Signature (HBPS) based on cell cycle-related gene expression and Cox analysis. 421 h+/HER2- BC patients from the TCGA dataset were used as the training set and 3605 patients from GEO and cBioPortal datasets were used as the validation sets. Subsequently, we explored the underlying biological mechanisms and drug susceptibility associated with the HBPS score. Patients with high HBPS scores exhibited significantly worse prognosis across all sets. The high HBPS score group demonstrated lower levels of immune cell infiltration, downregulation of HALLMARK_KRAS_SIGNALING_DN and HALLMARK_IL2-STAT5 signaling. Moreover, further validation revealed that CDKN2C (one critical gene in the HBPS) deficiency was associated with immuno-cold tumor microenvironment and enhanced HR + HER2- breast cancer cells aggressiveness. Overall, the study's biological insights, crucial for comprehending and tackling drug resistance, hold the potential to inform precise drug treatments in HR+/HER2- BC patients.

Objectives: Neoadjuvant radiotherapy (Neo-Rad) improves locoregional control, down-stages the tumor, and increases the success rate of breast-conserving surgery. However, there remains controversy regarding its effect in long-time survival. The aim of this research was to, combined with various clinicopathologic characteristics, compare the survival outcomes of breast cancer (BC) patients underwent Neo-Rad versus adjuvant radiotherapy (Adj-Rad), and to determine whether the radiation sequence was the independent prognostic factor for BC. Methods: A total of 128113 BC patients from Surveillance, Epidemiology, and End Results (SEER) database were identified in our study, of whom 127707 patients underwent Adj-Rad and 406 patients underwent Neo-Rad. The Kaplan-Meier function and the COX proportional hazards model were applied to compare overall survival (OS), the Nelson-Aalen cumulative hazard method and competing risk model were applied to compare breast cancer specific survival (BCSS) between two groups. Results: The OS and BCSS of Neo-Rad group were significantly prolonged than Adj-Rad group (P<0.001) in general. The radiation sequence was as an independent prognostic factor for OS (HR 0.61, 95% CI 0.49-0.75, p<0.001). Neo-Rad group also showed better survival outcomes than Adj-Rad cohort in stage I-II patients, LN positive/negative patients, patients with different molecular subtypes, and patients underwent partial/ total Mastectomy (p<0.05). By contrast, in stage III patients, the OS and BCSS were comparable between two groups (p>0.05). Conclusions: Neo-Rad may be applicable not only for patients with advanced, inoperable, or aggressive subtype of BC, but also in the setting of early-stage, luminal subtype of patients, and it may significantly improve OS and BCSS in most subgroups of patients. These finding warrants more prospective clinical trials to clarify the optimal radiotherapy timing and radiation technologies for corresponding type of patients, and to improve local control and survival outcomes at the same time. Citation Format: Yue Huang, Xie, Yiqin Xia, Yangyang Cui, Jinhui Peng, Shuhan Zhao, Yuhan Dai, Huilin Chen, Yifan Wu, Yixing Yang, Shui Wang, Mingjie Zheng. Survival Outcomes of Breast Cancer Patients Underwent Neoadjuvant Radiotherapy vs. Adjuvant Radiotherapy: A Population-based Observational Study [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P5-11-14.

Background: Target therapies significantly improved the pathological complete response rate (pCR) of human epidermal growth factor receptor 2 (HER2) positive breast cancer. A tyrosine kinase inhibitor (TKI) plus trastuzumab-based regimen showed inconsistent efficacy in neoadjuvant therapy. As we know, HER2-positive breast cancer consists of four intrinsic molecular subtypes of luminal A, luminal B, HER2-enriched, and basal-like. This trial was aimed to evaluate the efficacy of chemotherapy de-escalated neoadjuvant regimen of pyrotinib, trastuzumab and nab-paclitaxel in HER2-enriched subtype of HER2-positive early breast cancer. Methods: This multicenter, single-arm, phase 2 trial was conducted in nine hospitals in China (ClinicalTrials.gov, NCT05659056). Simon’s two-stage design was adopted, and 65 eligible patients with HER2-positive, stage IIA-IIIC invasive breast cancer regardless of hormone receptor status were scheduled to be enrolled. Patients received pyrotinib (400mg, continuous orally), trastuzumab (loading dose of 8 mg/kg, followed by 6 mg/kg every 3 weeks intravenously), and nab-paclitaxel (260mg/m2 every 3 weeks intravenously) for six cycles. The primary endpoint was the total pCR (tpCR, ypT0/Tis and ypN0) of HER2-enriched subtype patients. Intrinsic molecular subtypes of tumor biopsy samples taken at baseline were determined via the BluePrint predictor. Findings: In the first stage, 21 patients were enrolled and tpCR exceeding 35%, so between Nov 22, 2022, and Jul 1, 2024, we recruited 65 patients, of whom 52 (80%) completed neoadjuvant therapy and received surgery as planned, and BluePrint results were available in 50 of them. Of the 50 patients, 32 (64%) were HER2-enriched subtype, 2 (4%) were luminal A, 9 (18%) were luminal B, and 7 (14%) were basal-like. Twenty-two (42.3%, 95% CI 29.0%-56.7%) of 52 patients had tpCR at the time of surgery. Eighteen (56.3%, 95% CI 37.9%-73.2%) of 32 patients with the HER2-enriched subtype and 4 (22.2%, 95%CI 7.3%-48.1%) of 18 patients with non-HER2-enriched subtypes achieved tpCR. Forty-five (86.5%, 95% CI 0.74-0.94) of 52 patients achieved objective response. The treatment related any grade adverse events was 91.2%, and 6 patients changed treatment regimen due to adverse events. And the most common grade 3 or 4 adverse events were diarrhea (12 [21.1%]), and neutrophil count decreased (3 [5.3%]). Conclusion: The trial reached the pre-specified tpCR, had met the primary end point. Neoadjuvant pyrotinib, trastuzumab and nab-paclitaxel reached a high tpCR rate, with manageable toxicity, for patients with HER2-enriched subtype of HER2-positive breast cancer. Chemotherapy can be de-escalated in patients with HER2-enriched subtype breast cancer with BluePrint intrinsic subtyping. Citation Format: Pan Hong, Hui Xie, Yi Ren, Xiaoan Liu, Yi Zhao, Lin Chen, Xiaoming Zha, Tia song Xia, Linlin Zhen, Zhaoji Guo, Jing Lan, Jieqiong Liu, Changchun Li, Jun Zhou, Yanwu Zhang, Zhao Liu, Jing Tao, Shui Wang, Wenbin Zhou. Neoadjuvant Nab-paclitaxel combined with Trastuzumab and Pyrotinib for HER2-enriched subtype of HER2-positive early or locally advanced breast cancer: A multicenter, single-arm, phase 2 trial [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P3-11-25.

Background Patients with liver metastases of triple-negative breast cancer (TNBC) show poor prognosis compared with other metastases. Chemotherapy is the primary treatment for advanced TNBC. Tumor cell diversity and the tumor microenvironment could affect therapeutic effect. However, whether liver metastases of TNBC exhibit differential chemotherapy efficacy compared with the primary tumors remains inadequately understood. The specific mechanisms that modulate chemotherapy efficacy in liver metastases need further investigation.Methods Single-cell RNA sequencing data from public databases were leveraged to contrast the immune profiles of liver metastases and primary tumors in TNBC. Murine models bearing liver tumors or primary tumors of TNBC were used to evaluate chemotherapy efficacy. Techniques such as immunohistochemistry, wound healing assays, and colony formation assays were employed to account for tumor heterogeneity. Intratumoral T lymphocytes and macrophages were quantified and characterized using RNA sequencing, immunohistochemistry, and flow cytometry. Antibody-mediated depletion of CD8+T cells or macrophages in mice substantiated their impact on chemotherapy responses.Results Single-cell RNA sequencing data showed the immune microenvironments of liver metastases and primary tumors exhibited significant differences, which may critically influence chemotherapy outcomes. Mouse models confirmed that chemotherapy was less effective against liver tumors compared with subcutaneous tumors. After excluding the influence of tumor cell heterogeneity, the weaker responsiveness in liver tumors was mediated by the impeded infiltration of CD8+T cells, attributed to the decreased activation of macrophages. Augmenting macrophage activation can improve the chemotherapeutic efficacy in liver tumors. Moreover, chemotherapy drove the immune microenvironment towards increased suppression through distinct mechanisms, with neutrophil extracellular traps (NETs) accumulating in liver tumors and impaired functionality of macrophages at the primary site. The combination of NET inhibitors or macrophage activators with chemotherapy enhanced treatment effectiveness.Conclusions These findings disclose the compromised chemotherapeutic efficacy in liver tumors of TNBC and elucidate the underlying immune-related mechanisms within the tumor microenvironment. Targeting the specific underpinnings of immune suppression at different tumor sites with selective drugs could optimize chemotherapeutic efficacy.

Background: The tumor stroma has been reported to be associated with worse prognosis in several solid tumors, but its prognostic value in breast cancer (BRCA) is still undefined. Methods: In this research, multiple public and in-house patient cohorts were collected to demonstrate the clinical and immune correlations of tumor-stroma proportion (TSP) in BRCA. In addition, in vitro assays uncovered the oncogenic role of TSP-related collagen in BRCA. Results: High TSP status based on hematoxylin and eosin (HE) staining was associated with positive hormone receptor status, advanced clinical stages, and poor immune checkpoint blockade (ICB) response. In addition, we developed a RNA-sequencing (RNA-seq)-based stromal score based on four critical genes expression (AEBP1, COL6A3, CTSK, and PLAC9). Both TSP status and stromal score were positively associated with increased M2 macrophage abundance in BRCA. Moreover, tumor collagen has been found to be enriched in samples with the high TSP status, and collagen promoted BRCA cells aggressiveness and macrophage M2 polarization. Conclusions: The tumor stroma was found to be notably related to poor ICB response in patients with BRCA as a result of tumor stroma-macrophage interactions. Thus, the TSP status could predict the clinical outcomes of BRCA patients receiving ICB therapy.

Breast cancer is the most common female malignancy all over the word, at a risen rate of over 1.7 million cases per year globally, where metastasis is the leading cause of death. Accumulating epidemiological evidence indicates that abnormal fat status such as obesity is an important risk and poor prognostic factor for breast cancer. Adipose, a significant component of the stromal tissues, also a type of metabolically active endocrine organ, is the most abundant stromal constituent in the mammary gland surrounding breast cancer. In addition, a growing number of studies have confirmed that adipocytes adjacent to invasive cancer cells, known as cancer-associated adipocytes (CAAs), participate in the regulation of breast cancer progression. Breast tumor adjacent adipocytes have the potential to drive the malignancy of tumor cells by secreting hormones, proinflammatory cytokines, and accelerating tumor metabolic reprogramming. Despite of the diversity effects of adipokines on breast cancer, adipocytes secreted protein derived peptidome remains a black hole to our knowledge, since degradome has been expanding its roles in biology and pathology. In addition to glucose and lipid metabolic products, the fragments/peptides cleaved from extracellular matrix and membrane proteins degradation by proteinase or hydrolase at the level of cancer-tissue microenvironment, are supposed to carry cancer-specific information and further reveal the reciprocal interconnected network of tumor and surrounding stroma tissues, which could be a potential anti-tumor therapeutic target. In addition, little evidence proved that adipocyte-derived peptide could participate in cancer metastasis progression. In this study, we aim to assess the biological effects of breast tumor adjacent adipocyte secreted protein derived peptides on breast cancer cells from multiple angles, and systematically analyze the degradome characters, expecting to realize a basic understanding of its role in the development and progression of breast cancer. Here we found a novel adipocyte-derived peptide MASP6 that may affect the invasive and migrative ability of breast tumor, which is of great significance for exploring new targets for breast cancer. In order to explore the underlying mechanism of communication between breast cancer and adipose tissue, we isolated adipocytes from adipose tissues adjacent to breast tumor (TAA) and breast benign lesions (BAA) and collected the supernatant. The results showed that peptides of adipocyte secreted degradome could increase breast cancer malignancies dominantly. LC-MS/MS and bioinformatic analysis data showed there are about 100 identified peptides contribute to aberrant cell adhesion of tumor cells through regulating focal adhesion. Sparklingly, 2 lipophilic peptides are identified stable, creating sustained effect on tumor cells.What’s more, wound healing test showed that MASP6, one of the adipocyte-derived peptide, promoted the invasive and migrative ability of breast tumor cells. Dual-luciferase assay, quantitative real-time polymerase chain reaction (qRT-PCR) and western blots verified that the expression of Matrix Metalloproteinase (MMP-2/9) was increased after MASP6 treated in breast cancer cells. Mouse xenograft model demonstrated that MASP6 enhanced tumor metastasis in vivo. The PI3K/Akt signal pathway was stimulated after MASP6 treatment. Nuclear and cellular protein extraction, indirect cellular immunofluorescence revealed that MASP6 could enter into cytoplasm and activate nuclear factor kappa B (NF-κB-p65) via promoting its nuclear translocation. In conclusion, our results confirmed mammary gland surrounding adipocytes induce malignancies of breast tumor cells, and delineated the interaction network between secreted degradome of adipocytes and tumor cells, which are believed to afford novel insights into breast cancer therapy. Citation Format: Shuhan Zhao, Jinghui Peng, Yue Huang, Yiqin Xia, Yuhan Dai, Huilin Chen, Jiangdong Jin, Yifan Wu, Yangyang Cui, Ziyi Fu, Yongmei Yin, Shui Wang, Hui Xie. Proteomics-based degradome analysis of mammary tumor adjacent adipocyte reveals adipocyte-derived peptide MASP6 promotes breast cancer cell invasion and migration [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P3-01-19.