Introduction
Triple-negative breast cancer (TNBC), an aggressive breast cancer subtype, lacks the expression of the progesterone receptor, human epidermal growth factor receptor 2 (HER2), and estrogen receptor ( Shang & Xu, 2022 ). As a highly heterogeneous tumor, TNBC shows distinct biological features of aggressiveness, high recurrence rate, and distant metastasis ( Gluz et al., 2009 ; Foulkes, Smith & Reis-Filho, 2010 ), which can be attributed to germline alterations (BRCA1 mutations), differences in genetic characteristics, epigenetic alterations, DNA repair defects, morphological features, and gene expression profiles ( Nik-Zainal et al., 2016 ; Jiang et al., 2019 ; Bareche et al., 2018 ; Pareja & Reis-Filho, 2018 ). Due to this heterogeneity, large tumors may contain multiple cells with different molecular characteristics and displaying different sensitivity to treatment ( Dagogo-Jack & Shaw, 2018 ), which has been demonstrated to be the main reason for drug resistance or lower efficiency of immunotherapy in breast cancer therapy ( Li et al., 2021 ). Relative to other subtypes, TNBC has a poor prognosis with a low 10-year survival rate of less than 50% ( Malorni et al., 2012 ). Effective therapeutic targets for TNBC are scarce ( Huang et al., 2021 ; Zhu, Yang & Xu, 2022 ). Therefore, it is particularly crucial to assess the mechanisms underlying the progression and onset of TNBC for immunotherapy and its diagnosis in clinical settings.

The main feature of cellular senescence is a complex secretory phenotype, known as the senescence-associated secretory proteome (SASP), that by facilitate immune-mediated clearance of senescent cells to keep tissue homeostasis and suppresses tumorigenesis, as a powerful tumor-suppressing mechanism ( Bartkova et al., 2006 ; Xue et al., 2007 ). For example, excessive reactiveoxygenspecies caused by metabolic perturbation could trigger mitogen-activated protein kinase (MAPK)/p38 pathways to facilitate cellular senescence ( Borodkina et al., 2016 ). Researches also uncovered that the MEK/MAPK signaling pathway was involved in inducing senescence in human fibroblasts ( Takasugi, Yoshida & Ohtani, 2022 ). Previous studies suggested that cancer cells have devised various ways to evade this mechanism ( Hollstein et al., 1991 ; Jarrard et al., 1999 ; Foster et al., 1998 ). When co-transplanted with completely malignant cells, the growth of senescent cells increases along with the tumor formation rates in the xenografts ( Bartholomew, Volonte & Galbiati, 2009 ; Bhatia et al., 2008 ; Liu & Hornsby, 2007 ). The impact of senescent cells on the evolution of precancerous tumor progression has been confirmed ( Campisi et al., 2011 ; Collado, Blasco & Serrano, 2007 ). Proliferation is induced in several precancerous cells in the presence of senescent fibroblasts, ultimately resulting in tumor formation ( Krtolica et al., 2001 ; Bavik et al., 2006 ; Castro-Vega et al., 2015 ). On the one hand, cellular senescence, a fail-safe mechanism, inhibits tumorigenesis in a cell-autonomous environment by preventing harmful cells from further proliferation. On the other hand, senescent cells secrete excess growth factors and cytokines ( Coppé et al., 2010 ), including interleukin 8 (IL-6) and IL-6 (collectively referred to as SASP) ( Lasry & Ben-Neriah, 2015 ), thus establishing a microenvironment of immunosuppression, inflammation, and catabolism to promote chemotherapeutic resistance and tumor growth ( Coppé et al., 2008 ). Intriguingly, within multiple element causing TNBC chemoresistance, therapy-induced senescence is well-accepted ( Chakrabarty et al., 2021 ). Despite these extensive effects of cellular senescence on cancer, our understanding of cellular senescence-related characteristics of TNBC patients remains rudimentary. In particular, whether these characteristics of cancer patients can be used as biomarkers to guide clinical prognosis and treatment of TNBC remains elusive.

Given that cellular senescence signature is highly correlated with the progression of cancer, an increasing number of studies have investigated it and demonstrated that senescence related signature is significantly associated with the prognosis of tumor patients. For example, in lung adenocarcinoma (LUAD), a comprehensive analysis constructed a senescence-related signature score (SRS), which was elucidated as an independent prognostic factor for LUAD patients and revealed that it was positively associated with cancer-associated fibroblasts, NK cell infiltration, and cytokine release ( Lin et al., 2021 ). Among them, cancer-associated fibroblasts often present a senescence-associated secretory phenotype (SASP) under stress conditions such as inflammatory factors, which promotes tumor formation and chemoresistance through the release of cytokine IL-6, which also reveals the relevant mechanism of cellular senescence-associated cancer progression from the level of regulation of the immune microenvironment ( Yasuda et al., 2021 ; Wu et al., 2017 ). Another study revealed that SRS in colorectal cancer can be used to assess the genomic mutation status of the tissues, especially the high SRS group showed significant RPN2 mutations ( Yue et al., 2021 ). RPN2 is commonly associated with cell proliferation, migration, and epithelial-mesenchymal transition phenotypes, exacerbates malignant cancer progression through the release of factors such as MMP-9, and inhibits cellular autophagy through STAT3 and NF-κB pathways ( Huang et al., 2019 ; Zhang et al., 2019 ; Han et al., 2023 ). Although the clinical value of cellular senescence-related features has not been explored in TNBC, the above reports still inform our study, and we hypothesize that markers associated with senescence features and prognosis in TNBC could influence cancer progression and prognosis in terms of tumor infiltration microenvironment homeostasis, cytokine release, and genomic mutations.

Based on scRNA-seq and bulk RNA-seq data, we constructed a senescence classifier for TNBC based on cellular senescence-related pathways. Significant differences in prognosis, tumor microenvironment (TME), genome mutations and enriched metabolic pathways were detected from the three subtypes. Next, the heterogeneity of malignant cells among senescent and normal cells was further analyzed in TNBC. A cellular senescence-related risk model for TNBC was constructed. In vitro experiments were also conducted to confirm the expression levels of hub genes selected for the risk model and the functions of certain gene on TNBC progression. Additionally, we also verified the reliability of the signature in assessing the regulatory role of the TNBC immune microenvironment and the patient’s response to immunotherapy. Finally, the possibility of cellular senescence as a clinicopathological feature to improve the prognosis and survival prediction of patients with TNBC was explored. Collectively, our study provides a new direction to the diagnosis and treatment of patients with TNBC.