ReviewTumor microenvironment: Challenges and opportunities in targeting metastasis of triple negative breast cancer
Graphical abstract
Introduction
Triple negative breast cancer (TNBC) is aggressive, metastatic and a remarkable drug-resistant subtype of breast cancers [1]. It is the most challenging subtype due to the dearth of specific targets. Worldwide, TNBC is rising and accounts for nearly 20 % of the total breast cancers [2] and 83 % of disproportionate deaths compared to other subtypes [3]. Chemotherapy is the only option for the treatment of TNBC. The efficacy of currently approved chemotherapeutics including cisplatin, anthracycline, paclitaxel, and tamoxifen [[4], [5], [6]] is limited [7,8] due to heterogeneity with oncogenic drivers [[9], [10], [11]] and development of resistance [[12], [13], [14]].
TNBC has heterogeneous subtypes: basal like-1 (BL1), basal like-2 (BL2), luminal androgen receptor (LAR), immunomodulatory (IM), mesenchymal (M), mesenchymal stem-like (MSL), and unstable (UNS) subtypes [15,16]. BL-subtypes symbolized as BRCA1 and BRCA2-mutants, are characterized by genomic instability with high proliferation and defects in cell-cycle checkpoints. These subtypes are sensitive to platinum-based regimens [17]. MSL subtype is characterized by Epithelial-mesenchymal transition (EMT) and stem cells, which are sensitive to EMT inhibitors [18]. The IM subtype is fortified with immune signalling. M group subtype is metaplastic TNBCs, with PIK3CA mutations, which are susceptible to PI3K inhibitors [19]. LAR subtype is sensitive to AR-targeting agents [15]. In spite of advances in targeted therapies, TNBC has high morbidity and mortality due to the development of lung/bone and brain metastases [[20], [21], [22], [23], [24], [25], [26]]. Therefore, understanding the tumor microenvironment and its regulators will be useful for designing the immune based targeted therapies for TNBC.
The TME is indispensable for the initiation and progression of TNBC. The induction of proliferation, angiogenesis, inhibition of apoptosis, immune system suppression, and evasion of immune surveillance are intrinsically linked to TME [27]. TME can be designated as a hallmark of TNBC. TME is highly complex and heterogeneous [28,29]. Rapid proliferation of cancer cells induces hypoxia, with consequent reprogramming of cancer cells in the TME. The tumor cells and surrounding TME cells constantly adapt to the new conditions and promote tumor growth. TME creates a niche for residing and interacting cancer cells with their surrounding endothelial, and immune cells as well as fibroblasts [30] and can serve as potential targets for the TNBC. The reciprocal communication between cancer cells and stromal cells as well as immune cells induces changes in the cellular components of TME, which predisposes cancer cells to metastasis (Fig. 1) [31]. Further, TME induces the transition of epithelial cells to TNBC stem cells [32]. The interactions between these cells in TME nurture several biological events that underpin cancer growth, invasion, angiogenesis, and metastasis.
TME is made up of cues of ECM components, cancer cells, CSC, immune cells, cancer-associated fibroblasts (CAFs), blood vessels, and differentiated cells. ECM supports tumor progression by furnishing signals for proliferation, escape from tumor growth suppressors, evading apoptosis, empowering replicative immortality, inducing neovascularization and elevating invasion and metastasis (Fig. 2). The CAFs that conjoined with cancer are critical players of cancer progression as well as drug resistance [33]. CAFs are the most abundant stromal cells in the TME [34]. They produce growth factors, and chemokines and also facilitate intrusion of immune cells into the TME for enhancement of growth and survival of tumor cells [34].
CAFs stimulate invasion of cancer cells by generating MMPs that restructure the ECM of the TME [35] and build up the hypoxic environment [36]. Recently, ECM is recognized as one of the potential strategies for targeting TME [37]. The divergency in tumor cells, hypoxia in tissue or enhanced inflammation in the TME induces the changes in the ECM. During cancer progression, the ECM components are disorganized by local modulators in the TME [35]. The enhanced expression of MMPs is also a prerequisite for altering ECM [38]. In hypoxia, CAFs also produces an ECM that differs from ECM under normoxia in terms of stiffness and alignment, which support cancer cell migration [39]. ECM components and their proteolysis products constitute matrisome [40], which may regulates key steps of the cancer-immunity cycle within TME.
TME is reengineered to alleviate hypoxia, a mediator of cancer metastasis, immune suppression, and drug resistance. Rapidly proliferating tumor cells have increased requirement for oxygen that cannot be met by the surrounding blood vessels, leading to the development of hypoxia [41]. Hypoxia stimulates the hypoxia-inducible factor-1α (HIF-1 α) [42,43] that transactivates genes that regulating the tumor growth, the polarization of macrophage as well as metastasis [44]. Besides, hypoxia also induces the secretion of angiogenic factors by tumor cells [45,46]. The angiogenic factors induce neovascularization within TME, contributing to the potentiation of tumor development.
Induction of anti-tumor immunity is one of the recently identified strategies for therapeutic use of TME since it displays immense diversity in the immune cells [47,48]. They may impede or support tumor progression - depends on the signals within TME [37]. For instance, if the environment surrounding tumor cells is rich in inflammatory molecules, then the immune system fails to recognize and eliminate cancer cells [49]. However, lymphoid lineage B cells and regulatory T cells (Tregs), which regularly infiltrate into TME cause immunosuppression by releasing cytokines and uphold tumor progression [50]. Tumors accomplish immune dispensation via immunoediting and creation of immunosuppressive TME. Co-inhibitory receptors including CTLA4 and PD1, have indispensable, nevertheless diverse functions in modulation of immune responses [51]. Further, upregulation of tumor-associated antigens owing to genetic or epigenetic changes deregulates immune checkpoint proteins leading to inhibition of the immune system [[52], [53], [54]]. Therefore, targets of anti-tumor immunity can be tailored for effective use in combination of immunotherapies and other targeted therapies.
Analysis of the TME presented a clinical correlation between risk of relapse and the landscape of adaptive immune cells in discrete tumor regions [55]. In TNBC, the TME display infiltration of CD8 + T cells with amplified PD-L1 [56]. However, in the absence of CD8 + T cell infiltration, the expression of B7-H4, an immunosuppressive marker, and stromal fibrotic signatures were elevated [57]. This led to new cancer classification [58] based on the simple immune score, defined by quantifiable density and location of CD3+ and cytotoxic CD8 + T cells within the TME [59,60]. The immune score provides a more accurate prediction of prognosis [61] and a reliable estimate of the breast cancer recurrence in patients [62]. The immune score can be calculated by immunohistochemical and flow cytometric analyses of tumor-infiltrating lymphocytes (TIL) in metastatic breast cancer [63]. Lymphocytes frequently infiltrate into TME of lymph node (LN)-positive TNBC [64]. Higher rate of tumor immune infiltration (TII) is associated with a better outcome in TNBC and HER2 +ve breast cancer [65]. Infiltration of T cells into tumors has prognostic impact in HER2-positive and ER-negative cancers [66,67] and TNBC [68], whereas, γδ T cell infiltration is correlated with poor prognosis [69]. In TNBC patients, increased stromal TILs are associated with reduced risk of recurrence. This suggests that specific localization of TILs in TME could contribute to the prognosis and prediction of TNBC in patients.
TME plays a key role in the establishment of different kinds of drug-resistant mechanisms. The response of breast cancer cells to drugs is majorly controlled by drawing signals from TME. Chemotherapy induces TME to alter the phenotype and to develop resistance to drugs [70]. TNBC is associated with six mechanisms of chemoresistance, namely, modulation of ABC transporters, overexpression of class III β-tubulin, mutations in topoisomerase II and DNA mismatch repair enzymes, mutations in apoptotic genes, ALDH1 and glutathione (GSH)/Glutathione-S-transferase (GST) as well as NF-kB signaling [7]. A tumor-initiating, TNBC stem-cell population is further enhanced by overexpression of aldehyde dehydrogenase (ALDH), which is capable of inactivating chemotherapeutic agents, such as cyclophosphamide.
Recently, cancer stem cells (CSCs) are proven players of tumor initiation as well as drug resistance. Further, studies have reported that stem cells are responsible for recurrence within TME [[71], [72], [73]]. TME also mediates the maintenance and retention of stem cells by controlling stemness via bone marrow niche generated signaling pathways [74]. CSCs re-architect their TME and maintain a supportive niche by promoting interactions with immune cells and other cells [75,76]. To realize specific target therapy, current research is aimed at identifying strategies that target TNBC stem cells [[77], [78], [79]]. Subpopulations within TNBC subtypes that exhibit self-renewable and differentiation capacity, are responsible for initiation, progression, invasion, and drug resistance [80]. TNBC stem cells are more resistant to conventional therapies relative to differentiated cells [81]. Mammosphere cultures of TNBC cells are known to express higher levels of stem cell markers such as CD44+/CD24− and ALDH1+ compared to adherent cells [82] and have a critical role in tumorigenicity and metastasis [[83], [84], [85]]. Sensitization studies using phytochemicals have demonstrated the induction of self-renewal and differentiation within TME [86,87]. Studies have examined the several biochemical factors that affect TME associated metastasis in TNBC. For example, FAK is required for TME associated cancer metastasis in TNBC [88]. Therefore, the therapeutics that targets cancer stem cells can control the TME mediated cancer metastasis.
Section snippets
Regulators of TME in TNBC
TME is regulated by genetic modifiers, developmental and other pathways, growth factors and chemokines, exosomes, epigenetic regulators and microRNAs in TNBC (Fig. 3). Current literature provides an evidence for the interaction of TME modifier candidates with molecular pathways. They include FGFR2, TGFR2 and MKL1, which have function in one of TME cell type and eNOS and TLRs, which exclusively function in TME [[89], [90], [91]]. The pathways regulating TME are critical for cellular mechanisms
Approaches for targeting TME
TME is being increasingly recognized as a key target of metastasis in TNBC (Fig. 4). Studies on cancer-immune interactions have increased benefits of immunotherapy in cancer treatment. However, onco-immunotherapy is showing remarkable antitumor response in patients compared to monotherapy. Several onco-immunotherapeutic strategies have been proposed. Paclitaxel along with proton pump inhibitor, lansoprazole has been evaluated against acidic tumor microenvironment in breast cancer [199].
Phytochemical therapeutics
Recent studies showed that phytochemicals can potentially induce genetic, transcriptomic and proteomic alterations while acting on the breast cancer cells. Phytochemicals like EGCG, caffeic acid, genistein, morin and kaempferol were found to have increased activity in the modulation of multiple coding and non-coding genes associated with TME [229]. Green tea polyphenols were shown to modify the expression of miRNAs in different cancers. Breast cancer cell line, MCF 7, treated with polyphenon-60
TME in early diagnosis of TNBC
TME also serve as an interface that affects the human body's reaction to cancer cells. Worldwide, on the basis of gene expression analysis, there is a socioeconomic difference between the different races of TNBC patients. Emerging data advocated that intrinsic variations in the TME of African-American (AA) and Caucasian (CA) breast cancer patients especially in resistin and IL-6 [249]. Review of Deshmukh et al., provided evidence for racial differences in TME and its underlying genetic and
Current status and future implications
TNBC continues is a most inoperable women cancers worldwide. TME is of prime importance in clinical settings due to development of drug resistance and recurrence of TNBC by inducing genes involved in self-renewal. The therapeutic targeting of immune checkpoints that showed favorable results in various cancers in clinical trials, were unsuccessful in patients with TNBC. Recently, several attempts have been performed to gain new insights into TNBC to maximize the efficacy of combination
Conclusions
TNBC is highly aggressive, metastatic and highly challenging breast cancer subtype due to lack of specific targets and targeted therapeutics. TME is associated with hallmarks of TNBC as well as immune system suppression, escaping immune detection and drug resistance. To improve the clinical benefit of TME inhibitors, the impact of exosomes on components of the TME needs to be described using suitable cancer models, which help in the breakthrough of novel therapeutic strategies for TNBC.
Author contributions
All authors are involved in the critical review and final acceptance of the submission. The authors thank Mr. Manas Malla for providing language help.
Declaration of Competing Interest
The authors declare that there are no conflicts of interest.
Acknowledgment
This review was supported by DST-EMR (EMR/2016/002694, dt. 21st August 2017) (RRM) and CSIR (NO.37(1683)/17/EMR-II, dt. 5th May 2017) (RRM), New Delhi, India.
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