The prognostic value of tumor mutation burden (TMB) and its relationship with immune infiltration in breast cancer patients

Breast cancer is the most common gynecological cancer worldwide, and it ranks second in female cancer deaths [1]. The incidence rate of breast cancer has still increased year by year in China (27.24 million cases in 2015 but 367,900 cases in 2018) [2, 3]. Although improvements in early detection and treatment have reduced breast cancer mortality by 38%, metastasis of breast cancer at is the major reason of its high lethality [4‐6]. These reports demonstrate that development of innovative treatments for reducing recurrence and death in breast cancer are urgently needed. In recent years, immune checkpoint blockade (ICB) therapy has successfully improved overall survival (OS) in a variety of human cancers, including renal cell carcinoma [7], advanced melanoma [8], non-small-cell lung cancer [9] and bladder urothelial carcinoma [10]. The success of ICB therapy has rekindled the hope for immune-based therapy in breast cancer. Increasing data suggested that the immune system plays a decisive role in breast cancer patients’ response to standard therapy and the long-term survival rate [11].

Tumor mutation burden (TMB), which was reported to be used to predict the efficacy of ICB, is associated with a high neoantigen burden, high T cell infiltration and a high response rate to immune checkpoint inhibitors (ICIs) in different tumor types. TMB has emerged as a useful biomarker for evaluation of immunotherapy effectiveness in several cancer types, but its value is not well understood in breast cancer [12‐17]. Herein, this study aimed to investigate the prognostic value of TMB and its relationship with immune cell infiltration in breast cancer.

In this study, the landscape of different genetic mutations was explored in breast cancer. The results showed that TP53, PIK3CA, TTN and CDH1 were the most prominent mutated genes. TP53 plays a central role in human cancer pathogenesis and is hypermutated in almost all human cancers [18]. There is strong evidence that TP53 mutation is correlated with poorer overall and disease-free survival in breast cancer patients [19‐21]. Recently, the comprehensive cancer genome analyses described that phosphoinositide-3 kinase (PI3K) pathway are frequently altered in human cancers [22, 23]. Mutations of PIK3CA lead to tumorigenesis and hyperactivity of the PI3K pathway [24]. TTN is another frequently mutated gene in a variety of human cancers, such as lung adenocarcinoma, lung squamous cell carcinoma and colon adenocarcinoma [25]. Double mutations of TTN and TP53 may induce breast cancer through regulating a common downstream pathway [26]. The CDH1 gene, located on chromosome 16q22.1, is a well-known tumor suppressor gene. The dysregulation, mutation or transcriptional silence of CDH1 gene probably cause breast cancer development [27]. These reports were consistent with our findings. We found that the low-TMB group had favorable overall survival (OS), being consistent with the results of a survival study in breast cancer conducted by Chen et al. [28]. TMB was significantly correlated with regional lymph node metastasis (N). However, there was no significant association between TMB and other clinical pathological characteristics in breast cancer.

A total of 598 DEGs were identified in breast cancer. KRT gene family members play an important role in breast cancer. We found that KRT83 and KRT1 were upregulated in the low-TMB group, while KRT4 was downregulated. Several studies have shown that the invasion and migration of cancer cells are related to abnormal expression of KRT genes [29‐31]. We observed that DEGs were mainly involved in neuroactive ligand-interaction receptors according to KEGG enrichment analysis. The DEGs associated with the low-TMB group were mainly involved in the MAPK, mTOR, GnRH, TGF-beta and Hedgehog signaling pathways. The MAPK pathway is overactivated in various tumors. Many proteins that involve in MAPK pathway have been identified as oncogenic proteins [32]. Triple-negative breast cancer patients with MAPK/ERK signaling pathway upregulation have a poor survival prognosis [33]. The hedgehog pathway is associated with hormone receptor (HR +)-positive and triple-negative breast cancer patients [34]. Aberrant regulation of other signaling pathways, including mTOR, GnRH, and TGF-bate, could result in breast cancer development [35].

The lower expression of CCL18 and TRGC1 was significantly correlated with favorable survival outcomes in breast cancer, while their higher expression was associated with poor prognosis. CCL18 is produced abundantly by breast tumor-associated macrophages (TAMs), and its expression is also associated with tumor metastasis [36]. CCL18 promotes breast cancer cell invasion and metastasis through [activating/increasing?] Annexin A2 [37]. When CCL18 varied in arm-level gain, the infiltration levels of B cells, CD8 + cells, CD4 + T cells, macrophages and neutrophils decreased significantly in breast cancer samples. Breast cancer patients with low levels of B cells infiltration had a poor survival prognosis. The role of B cells in cancer is controversial in terms of immune cell infiltration. They have been reported to play both positive and negative roles in tumor immunity [38]. Recent years, B cells are considered to be a novel biomarker for ICB therapy and have been demonstrated to be important in ICB-driven antitumor responses because of their secretion of antibodies and helping T cell responses [39]. Daniel P et al. reported that the suppression effect of B cell on T cells may be due to the depletion of T cell subsets caused by antigen presentation, which in turn reduce the efficacy of ICB. The antibodies produced by B cells also play a key role in the ICI response [39]. IgG-secreted B cells can induce cytotoxicity through multiple mechanisms [40]. Daniel P et al. also observed that the efficacy of ICI disappeared when using mice with deficient Ig secretion [39]. These studies indicated that B cells mediate antitumor responses by activating T cells and producing antibodies.

This study revealed that TMB could be a biomarker for predicting overall survival in immunotherapy in breast cancer patients. It was also correlated with immune cell infiltration. Of note, the high-TMB scores were correlated with a reduced survival rate in breast cancer patients. CCL18 is an important gene in breast cancer, and the CNV of CCL18 may reduce immune cell infiltration. Moreover, low B cell infiltration is correlated with poor survival prognosis in breast cancer patients.