STAT3 as a potential therapeutic target in triple negative breast cancer: a systematic review

Triple negative breast cancer (TNBC) is the most aggressive form of breast cancer and accounts for much higher recurrence and metastasis rates [1]. Due to the absence of the expression of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2), TNBC is unresponsive to endocrine and HER2-targeted therapies, which results in the high mortality of patients with this disease [1]. When patients are diagnosed with TNBC at the early stage, combination chemotherapy (anthracyclines, taxanes, platinum salts, etc.) with or without radiotherapy is used as standard non-surgical therapy and has shown some efficacy in patients with both primary and metastatic diseases [2]. Because of the inter- and the intratumoral heterogeneities of TNBC, the intrinsic chemoresistance as well as severe side effects are often observed and lead to limited success in the clinic [3, 4]. Targeted therapies (e.g., poly (ADP-ribose) polymerase (PARP) inhibitors and epidermal growth factor receptor (EGFR) inhibitors) and immunotherapies have also shown some promise in preliminary clinical studies, but further investigations are critically needed [5‐7]. More recently, many efforts have been made to identify targetable molecules for treating TNBC via genomic profiling and several critical alternations have been discovered, including the overexpression and aberrant activation of signal transducer and activator of transcription 3 (STAT3) [8, 9]. The emerging data suggest that STAT3 may be a potential molecular target and biomarker for TNBC.

The STAT family of transcription factors is comprised of seven members with high structural and functional similarity, including STAT1, STAT2, STAT3, STAT4, STAT5a, STAT5b, and STAT6 [10, 11]. All STAT proteins consist of an amino acid domain (NH₂), a coiled-coil domain (CCD) for binding with interactive proteins, a DNA binding domain (DBD), a linker domain, a SRC homology 2 (SH2) domain for phosphorylation and dimerization, and a C-terminal transactivation domain (TAD) [11]. Most of these domains are highly conserved among STAT proteins and only TAD is divergent and mainly contributes to their structure diversity [12]. STAT3 was initially discovered to bind to DNA in response to interleukin-6 (IL-6) and epidermal growth factor (EGF) in 1994 [13, 14]. Over the past decades, STAT3 has become one of the most investigated oncogenic transcription factors and is highly associated with cancer initiation, progression, metastasis, chemoresistance, and immune evasion [15, 16]. The recent evidence from both preclinical and clinical studies have demonstrated that STAT3 plays a critical role in TNBC and STAT3 inhibitors have shown efficacy in inhibiting TNBC tumor growth and metastasis.

Considering that there is an unmet medical need for TNBC treatment and innovative therapeutic agents are urgently required, an in-depth understanding of the roles of STAT3 in TNBC will facilitate the development of STAT3-targeted therapeutics and pave the way for a novel TNBC treatment approach. In this review, we focus on the recent findings related to STAT3’s role in TNBC as well as STAT3 inhibitors and current targeting strategies. We also discuss other potential strategies for developing new STAT3 inhibitors for TNBC treatment.

The classical STAT3 signaling pathway that is activated through the binding of cytokines or growth factors to their corresponding cell surface receptors has been extensively reviewed [16‐18]. Here, we present a brief overview of the STAT3 signaling pathway, nonreceptor tyrosine kinases of STAT3, and its intrinsic inhibitors and coactivators, which are depicted in Fig. 1. Briefly, the overexpressed cytokine receptors, e.g., interleukin-6 receptor (IL-6R) and interleukin-10 receptor (IL-10R) and the hyperactive growth factor receptors, e.g., epidermal growth factor receptor (EGFR), fibroblast growth factor receptor (FGFR) and insulin-like growth factor receptor (IGFR) always trigger the tyrosine phosphorylation cascade through the binding of ligands to these receptors, leading to the aberrant activation of STAT3 and the transcription of its downstream target genes [17]. Once the ligands bind to their receptors on the cell surface, these receptors further form dimers and successively recruit glycoprotein 130 (gp130) and Janus kinases (JAKs), thus phosphorylating and activating JAKs [19]. Conversely, the cytoplasmic tyrosine residues of these receptors are phosphorylated by the activated JAKs and then interact with the SH2 domain of STAT3, resulting in STAT3 phosphorylation at Tyr705 by JAKs [16]. In addition, STAT3 can be phosphorylated and activated by several nonreceptor tyrosine kinases, e.g., Src and Abl [20]. The phosphorylated STAT3 (pSTAT3) further forms a homodimer through interaction between their phosphorylated Tyr705 site and SH2 domain, triggering the dissociation of STAT3 dimers from the cell surface receptors and its translocation from cytoplasm to the nucleus [21, 22]. With the help of a variety of coactivator proteins, including NCOA/SRC1a, apurinic/apyrimidinic endonuclease-1/redox factor-1 (APE/Ref-1), and CREB-binding protein (CBP)/p300, the nuclear STAT3 binds to specific DNA sequences and activates the transcription of genes that regulate various phenotypes of cancer cells [17, 18].

STAT3 is also highly expressed in some normal tissues and organs, including the bone marrow, peripheral nervous system, and digestive tract and plays a physiological role [23‐25]. In the normal physiological conditions, STAT3 phosphorylation and activation are tightly controlled by several intrinsic inhibitors, including protein tyrosine phosphatases (PTPs), the suppressors of cytokine signaling (SOCS), and the protein inhibitor of activated STAT (PIAS) [26]. The Src homology domain-containing tyrosine phosphatases 1/2 (SHP-1/2) directly interact and dephosphorylate JAK and STAT3, resulting in their inactivation [27, 28]. The nuclear PTPs, including TC45 and T-cell protein-tyrosine phosphatase (TC-PTP) induce the inactivation of STAT3 through its dephosphorylation and translocation from nucleus to the cytoplasm [29, 30]. Other PTPs, such as PTP1B and PTPeC have also been reported to regulate STAT3 dephosphorylation and inactivation [31]. Moreover, SOCS directly interacts with JAK and STAT3 and inhibits their phosphorylation and activation via forming a negative feedback loop with JAK-STAT3 signaling pathway [32]. PIAS inhibits the binding of nuclear STAT3 to DNA and induces STAT3 dephosphorylation via protein tyrosine phosphatase receptor T (PTPRT), leading to the reduced expression of its downstream target genes [33]. In addition, the stability of STAT3 protein is also regulated by the ubiquitin-proteasome system via the ubiquitin ligase TRAF6 (tumor necrosis factor receptor-associated factor 6) [34]. Recent studies have also reported that miR-544 directly targets the 3′-untranslated region (UTR) on STAT3 mRNA, thus down-regulating STAT3 expression in TNBC cells [35]. Due to the presence of these endogenous inhibitors, STAT3 is strictly governed to exert its physiological functions in normal cells [36]. Herein, both direct inhibition of STAT3 and activation of the endogenous inhibitors may be considered as potential STAT3-inhibiting strategies for developing novel cancer therapeutics.

The oncogenic potential of STAT3 has been widely recognized through its involvement in regulating the expression of genes related to cancer cell proliferation, anti-apoptosis, migration, invasion, angiogenesis, chemoresistance, immune suppression, stem cell self-renewal and maintenance, and autophagy (as shown in Fig. 2) [17, 18]. Importantly, STAT3 is overexpressed and constitutively activated in TNBC, which is highly related to TNBC initiation, progression, metastasis, resistance to chemotherapy, and the poor survival outcomes [8]. STAT3 is not only capable of eliciting the expression of cancer-related genes, but also physically interacts and functionally cooperates with other oncogenic transcription factors, e.g., GLI1, promoting the aggressiveness of TNBC [8]. A recent study has also found a reduction of the gene associated with retinoic-interferon-induced mortality 19 (GRIM-19), an intrinsic inhibitor of STAT3 transcription accompanied by STAT3 overexpression in TNBC [37]. In addition, TCPTP, including two splice variants TC45 and TC48 are down-regulated in TNBC cells in vitro and in vivo, which also contributes to the activation of STAT3 signaling [38]. Indeed, STAT3 has also been found to localize in the mitochondria, where it is termed mitoSTAT3 and regulates the mitochondrial functions, including electron transport chain, ATP synthesis, calcium homeostasis, and reactive oxygen species (ROS) accumulation [39, 40]. Moreover, mitoSTAT3 has been shown to promote breast cancer cell growth, in which the phosphorylation of Serine 727 plays a critical role [41].

A recent study has shown that acetylated STAT3 is highly elevated in TNBC, causing the methylation and inactivation of tumor-suppressor gene promoters [42]. Importantly, mutation of STAT3 at Lys685 or reducing STAT3 acetylation by resveratrol could induce demethylation and activation of the estrogen receptor-α gene and sensitize TNBC cells to antiestrogens. Considering the emerging data that demonstrate the critical role of STAT3 in TNBC, we herein present a comprehensive overview of its oncogenic functions in this section.

Abundant evidence has suggested that STAT3 may be a promising molecular target for TNBC therapy [86]. Various STAT3 inhibitors have been developed and shown some efficacy in TNBC models in vitro and in vivo, which have been summarized in Table 1. In this section, we discuss the current STAT3-targeting strategies (as shown in Fig. 3) for treating and preventing TNBC, as well as the challenges in developing more specific and effective STAT3 inhibitors.

TNBC is still a treatable but incurable disease with complex genetic heterogeneity. The STAT3 oncogene is overexpressed and constitutively activated in TNBC and is associated with the high metastatic risk and poor survival outcomes. Moreover, STAT3 not only acts as a transcription factor to activate the expression of its downstream target genes but also localizes to mitochondria and regulates its functions, then regulating the various aspects of TNBC cells. Many STAT3-targeted therapies have been successfully developed and shown efficacy in preclinical models of TNBC in vitro and in vivo; several STAT3 inhibitors even enter clinical trials and are currently under investigation in various human cancers, including TNBC. In addition to its role in cancer cells, STAT3 also plays a pivotal role in the immune system. Indeed, STAT3 inhibitors have been found to suppress tumor cells but also boost immune cell responses. Therefore, the STAT3 oncogene is a promising target for TNBC prevention and therapy.

Of note, targeting STAT3 alone has shown excellent anti-TNBC activities in preclinical settings. However, TNBC has been reported to harbor multiple genetic alterations, including STAT3 overexpression and constitutive activation which contribute to the initiation, progression, metastasis, and drug resistance of this disease. Therefore, STAT3 inhibition combined with other targeted therapies may be more effective in treating TNBC. Considering that STAT3 plays a crucial role in chemoresistance, the combination of STAT3 inhibitors with other chemotherapies may exert synergistic effects in treating TNBC. Therefore, further studies are warranted to demonstrate the preventive and therapeutic efficacy of STAT3 inhibitors alone or in combination with chemotherapy and/or other targeted therapies in clinical studies. Moreover, new targeting strategies, i.e. inducing the degradation of STAT3 protein through PROTAC or inhibiting the binding of STAT3 to its co-activators and other interactive proteins can be examined, which may lead to more specific and effective inhibitors for TNBC prevention and therapy.