Non-coding RNA-mediated mechanisms underlying tamoxifen resistance in breast cancer

Abstract

Tamoxifen remains a cornerstone therapy for estrogen receptor-positive (ER⁺) breast cancer; however, the emergence of drug resistance significantly limits its therapeutic success. Growing evidence has revealed that non-coding RNAs (ncRNAs), including microRNAs (miRNAs), long non-coding RNAs (lncRNAs), and circular RNAs (circRNAs), act as pivotal regulators of the molecular networks driving tamoxifen resistance. These “silent regulators” modulate multiple signaling pathways, including PI3K/AKT, MAPK, and estrogen receptor signaling, through mechanisms such as post-transcriptional regulation, epigenetic modification, and competitive endogenous RNA (ceRNA) interactions. Dysregulated ncRNAs can alter drug efflux, apoptosis, cell cycle progression, and epithelial–mesenchymal transition (EMT), contributing to reduced tamoxifen sensitivity and tumor recurrence. Our knowledge of BC resistance to tamoxifen will be aided by the new molecular insights into the mechanistic understanding of resistance to endocrine treatment as a whole that come from the growing functions of ncRNAs as miRNA sponges, transcriptional scaffolds, and ceRNAs, among others. The biosynthesis and molecular properties of ncRNAs, their involvement in tamoxifen resistance, and their potential use as therapeutic targets or diagnostic biomarkers are the main topics of this thorough analysis. In order to perhaps serve as a foundation for upcoming RNA-based therapies aimed at overcoming endocrine resistance in ER⁺ BC, we would also consolidate the current body of knowledge.

Introduction

Breast cancer (BC) is the most common cancer in females, as well as the second most frequent cause of cancer death (only lung cancer is responsible for more cancer deaths). In 2022, there were estimated to be 2.3 million newly diagnosed cases worldwide and ~ 660–670 thousand deaths from BC [1, 2]. BC is heterogeneous disease classified into distinct molecular subtype; hormone receptor-positive “luminal” subtypes (luminal A: ~30–40% of cases; luminal B: ~20–30% of cases), HER2-enriched tumors (10–15% of cases), and triple-negative (basal-like) BC (15–20% of cases) [3]. The subtypes have different prognoses and lead to different treatment decisions. The standard management of these malignancies is multimodal: local control with surgical resection and radiotherapy, and systemic therapy such as chemotherapy, endocrine therapy for estrogen receptor-positive (ER⁺) tumors, and targeted agents like anti-HER2 antibodies to prevent recurrence or metastatic disease [3, 4]. Despite the considerable advances in BC therapy, metastatic and treatment-resistant disease remain largely incurable, highlighting the urgent need to better understand the underlying molecular mechanisms driving disease progression and therapeutic resistance.

Non-coding RNAs (ncRNAs) have emerged as critical regulators of gene expression in cancer biology. Although only ~ 3% of human genes encode proteins; the majority of the transcriptome is non-coding. NcRNAs are typically subdivided based on their lengths: small ncRNAs (< 200 nt) and long ncRNAs (> 200 nt) [5, 6]. MicroRNAs (miRNAs) are small ~ 18–24 nt RNAs that repress gene expression by post-transcriptionally binding target mRNAs to inhibit translation or promote degradation. Long non-coding RNAs (lncRNAs) are considered anything longer than 200 nt and can modulate chromatin states, transcription and other processes. Circular RNAs (circRNAs), which form covalently closed loops via back-splicing, exhibit exceptional stability and frequently act as “sponges” to sequester miRNAs or regulate gene expression in other ways [7–9]. Together, miRNAs, lncRNAs and circRNAs represent a pervasive influence on cancer biology, affecting rates of cell proliferation, cell migration/metastasis, apoptosis, and drug response [6]. In BC, dysregulation of ncRNAs has been implicated in tumor progression and resistance to therapy, making them a key focus for understanding disease biology.

Endocrine therapy remains a cornerstone of treatment for ER⁺ BC, with tamoxifen—a selective estrogen receptor modulator (SERM)—being a first-line option [2]. Tamoxifen binds estrogen receptor alpha (Erα) blocking estrogen-driven signaling and reducing the recurrence risk of hormone-sensitive tumors by roughly 50% [1, 10]. However, the development of tamoxifen resistance is prevalent, with about 30–50% of ER⁺ patients having disease that is either intrinsically unresponsive to tamoxifen or acquires resistance during treatment [1]. To understand the mechanism of tamoxifen resistance, it is important to reconstruct a framework of endocrine resistance. Endocrine resistance is complex and may be due to a multitude of biological factors including ESR1 (ERα) gene mutations or loss, altered ER co-regulators, activation of alternative growth pathways (e.g. HER2 or PI3K/AKT/mTOR signaling), and altered metabolism or efflux of the drug itself [11]. Notably, recently documented contributions of ncRNAs to these pathways should be acknowledged. For example, the oncomiR-221/222 family can suppress ERα and PTEN expression, promoting tamoxifen resistance, whereas inhibition of these miRNAs restores ERα levels and re-sensitizes cells to therapy [12]. Additionally, certain lncRNAs (e.g., HOTAIR, MALAT1) and circRNAs have been reported to modulate the signaling networks and gene expression programs that govern endocrine escape [13, 14]. In summary, there is evidence that ncRNAs are potential upstream regulators of the molecular events responsible for both intrinsic and acquired tamoxifen resistance in ER⁺ BC.

In this review, we will first review the conventional mechanisms and pathways of tamoxifen resistance in BC, followed by a focus on the biology of ncRNAs and the myriad of mechanisms related to how miRNAs, lncRNAs, and circRNAs can alter estrogen receptor signaling and associated signaling pathways to promote or mitigate drug resistance. We will also include a discussion on the therapeutic potential of targeting these ncRNAs. By providing a comprehensive overview of ncRNA-mediated tamoxifen resistance we aim to highlight several promising approaches to improving endocrine therapy in BC.

Non coding rna’s biogenesis

Non coding RNAs, comprising a large part of the transcriptome, are functionally diverse regulators of gene expression. They are categorized into different types, including miRNAs, lncRNAs, and circRNAs, which all play important roles in development, cell differentiation, and tumorigenesis, often through epigenetic and post-transcriptional mechanisms (Table 1) [15, 16].

Table 1.

Comparative overview of miRNAs, lncRNAs, and circrnas

Feature	miRNA	lncRNA	circRNA
Length	~ 22 nucleotides	> 200 nucleotides	Variable; often > 100 nt
Biogenesis	Transcribed by RNA Pol II → pri-miRNA → processed by Drosha-DGCR8 → pre-miRNA → exported by Exportin-5 → cleaved by Dicer → mature miRNA	Transcribed by RNA Pol II → splicing, 5’ capping, 3’ polyadenylation (like mRNA)	Back-splicing of pre-mRNA exons or introns; facilitated by complementary sequences or RNA-binding proteins (RBPs)
Structure	Single-stranded, mature miRNA duplex with seed region	Linear, often structured with multiple functional domains	Circular covalently closed loop; lacks 5’ cap and 3’ poly(A) tail
Stability	Moderate; susceptible to exonucleases	Moderate; some nuclear-retained forms are stable	High; resistant to RNase R and exonucleases
Location	Cytoplasm (majority), some nuclear miRNAs	Mostly nuclear; some cytoplasmic	Predominantly cytoplasmic, some nuclear forms (e.g., EIciRNAs)
Functional Mechanism	Binds to 3’ UTR of mRNA in RISC complex → mRNA degradation or translational repression	Acts as guide, scaffold, decoy, or sponge; regulates chromatin modifiers, transcription, or other RNAs	Sponges miRNAs or RBPs; modulates transcription, splicing; some can be translated
Key Molecular Players	Drosha, DGCR8, Exportin-5, Dicer, Argonaute (AGO) proteins	RNA Pol II, PRC2, LSD1, SWI/SNF, hnRNPs	QKI, FUS, MBL, Alu repeats; possible IRES or m6A for translation
Examples	miR-21, miR-155, let-7, miR-34a	HOTAIR, MEG3, MALAT1, NEAT1, GAS5	ciRS-7 (CDR1as), circHIPK3, circFOXO3
Ref	[15, 17, 19]	[15, 22–24]	[16, 23, 25]

miRNAs are ~ 22-nucleotide RNAs that post-transcriptionally regulate gene expression by binding to mRNAs base paired with complementary regions in the 3’ untranslated regions (UTRs), with the end result being either mRNA degradation or translational repression. The canonical pathway of miRNA biogenesis begins when RNA polymerase II transcribes primary miRNA transcripts (pri-miRNAs) that have one or more stem-loop structures and have been capped and polyadenylated [17, 18]. In the nucleus, these pri-miRNAs are processed into ~ 70-nt precursor miRNAs (pre-miRNAs) by the microprocessor complex, which contains RNase III enzyme Drosha and double-stranded RNA-binding protein DGCR8. Pre-miRNAs are transported into the cytoplasm by Exportin-5 in a Ran-GTP-dependent pathway. Once in the cytoplasm, Dicer, a second RNase III enzyme, cleaves the pre-miRNA into a ~ 22-nt miRNA duplex. One strand (the guide strand) will become part of the RNA-induced silencing complex (RISC), which contains Argonaute (AGO) proteins and the passenger strand will typically be degraded. Target repression occurs via base pairing, and base pairing can either be perfect or near perfect leading to mRNA cleavage (common in plants) or partial base pairing resulting in translational repression and subsequently deadenylation (for animals). Additionally, there are also non-canonical pathways of miRNA biogenesis, such as miRtrons, in which miRNAs are derived from spliced introns without undergoing Drosha processing [15, 17, 19].

LncRNAs are considered to be transcripts over 200 nucleotides in length, that structurally resemble mRNAs, but do not possess coding potential. This type of ncRNAs is transcribed by RNA polymerase II, and are often spliced, capped, and polyadenylated, but many of these transcripts stay in the nucleus and can have tissue-specific and context-dependent regulation. LncRNA genes can be intergenic (lincRNAs), intronic, or antisense to protein-coding genes, and some can originate in the enhancer regions (eRNAs) and they can be regulated through many epigenetic modifications (e.g., histone acetylation/methylation and DNA methylation) that can influence their transcriptional activity [20, 21]. LncRNAs can function through four archetypes: (A) Scaffold – lncRNAs such as HOTAIR serve as platforms for chromatin modifiers (e.g., PRC2, LSD1) allowing them to be recruited to genomic loci to remodel chromatin resulting in silencing of underlying genes. (B) Guide – lncRNAs direct chromatin modifiers or transcription factors to specific DNA sequences. (C) Decoy – lncRNAs such as GAS5 sequester transcription factors or miRNAs, thereby regulating downstream signaling. (D) Sponges – lncRNAs such as MALAT1 or NEAT1 sequester miRNAs or RBPs to inhibit their activities on target mRNAs [15, 22–24]. As an example, HOTAIR, one of the most studied oncogenic lncRNAs, represses expression from metastasis suppressor genes, by recruiting PRC2 to the HOXD locus to induce PRC-mediated H3K27me3 and subsequent silencing of transcription. Conversely, MEG3 functions as a tumor suppressor due to its ability to activate p53-dependent transcription and modulate transcriptional activity through histone modifications [23, 25–27]. LncRNAs participate in nucleolar biology as well and various nucleolar lncRNAs affect rDNA transcription, impact ribosomal biogenesis, and influence nucleolar stress responses [23].

CircRNAs are covalently closed-loop RNAs formed from back-splicing, in which a downstream splice donor joins an upstream splice acceptor by forming a covalent bond that establishes the circular RNA. Unlike linear RNAs, circRNAs lack a 5’ cap and 3’ poly(A) tail and this structure provides circRNAs with stability, and its closed-loop structure provides considerable resistance to exonucleases [28, 29]. The biogenesis of circRNAs can be exon-based, exon-intron-based (EIciRNAs), or intron-based (ciRNAs). The back-splicing process is thought to be aided by the presence of inverted repeat sequences in the flanking introns or with the use of RNA-binding proteins (RBPs) such as Quaking (QKI), FUS, and MBL that promote a circularized structure [23]. CircRNAs have various activities: (A) miRNA sponging: ciRS-7 (CDR1as) contains > 70 miR-7 binding sites where it sequesters this tumor suppressive miRNA and promotes oncogenic signaling. (B) Protein decoying: Some circRNAs bind and sequester protein functions. (C) Translation: Despite being non-coding by definition, some circRNAs are able to contain internal ribosome entry sites (IRES) and m6A modifications, allowing these circRNAs to be translated into regulatory peptides in a cap-independent manner. (D) Transcriptional regulation: EIciRNAs and ciRNAs bind U1 snRNP and RNA Pol II and upregulate parental gene expression [16, 23, 25].

Pathways and molecular drivers of Tamoxifen resistance

Tamoxifen as a SERM is still a mainstay of endocrine therapy for ER⁺ BC. Although it is effective, the development of tamoxifen resistance—either intrinsically or acquired—remains one of the major clinical problems, with 30–40% of patients likely to experience recurrence or tamoxifen treatment failure over time [30]. Therefore, understanding the complex and multifactorial mechanisms of tamoxifen resistance is essential to improve treatment outcomes. Biological mechanisms that contribute to tamoxifen resistance are many and interconnected. One major mechanism of tamoxifen resistance involves alteration in estrogen receptor signaling, such as mutations to the ESR1 gene, conformational changes in the receptor, or post-translational modifications that prevent tamoxifen from exerting its inhibitory activity [30, 31]. In addition, cross-talk between ER signaling and growth factor signaling pathways, such as HER2/EGFR and FGFR1, as well as PI3K/AKT/mTOR and downstream MAPK signaling, can induce a proliferative state that is independent of ER, resulting in reduced tamoxifen sensitivity. In addition to stimulating activated survival signals, this signaling also promotes ligand-independent phosphorylation of ER, which allows continued tumor growth despite the presence of tamoxifen [31, 32]. Another vital axis that has emerged down increasing focus is the evasion of autophagy and apoptosis as new contributors to tamoxifen resistance. In fact, cytoprotective autophagy is often activated by tamoxifen in ER⁺ cells, promoting the reformation of the lysosome and survival through the elimination of damaged organelles and cytotoxic compounds [33].

Taken together, all of these situations are associated with a variety of interacting mechanisms that comprise a complex regulatory network that allows for BC cell survival and proliferation in the face of tamoxifen therapy. In the following will look closer at each of these resistance pathways to explore possible therapeutic targets to deterring endocrine resistance.

Growth factors signaling pathways

One major mechanism of tamoxifen resistance in ER⁺ BC is activation of growth factor signaling pathways. These pathways, specifically those involving the epidermal growth factor receptor (EGFR) and HER2 receptor families, provide tumor cells with a method to evade ER-mediated transcriptional control, offering them an alternative signal to survive and proliferate, which renders tamoxifen ineffective in inhibiting growth despite continued ER expression and pathway activation [34, 35]. Understanding how these pathways interact with ER signaling is critical for improving therapeutic outcomes and informing combinatorial treatment strategies. As an example, in animal models of ER⁺ BC, tamoxifen resistance was highly correlated with increased expression of EGFR and HER2, as well as increased expression and activation of their downstream kinases, p42/44 MAPK and p38 [36]. Although tamoxifen effectively inhibited classical ER genomic activity, tumor growth continued due to activation of nongenomic signaling pathways mediated by EGFR and insulin-like growth factor I-receptor (IGF-IR) [37]. Importantly, pharmacologic inhibition of EGFR using gefitinib significantly delayed the onset of tamoxifen resistance, highlighting EGFR signaling as a key contributor to endocrine escape [38]. Taken together, these studies suggest a potential benefit of combination therapy with tamoxifen and HER family inhibitors, regardless of whether tumors initially overexpress EGFR or HER2 [37].

HER3 is a kinase-deficient member of the ErbB receptor family and is activated by heregulin (HRG), and mediates proliferative signaling through heterodimerization of the HRG receptors. Herstatin (Hst), a secreted antagonist of HER2, inhibits HRG-induced activation of HER2/HER3 heterodimers and downstream MAPK and AKT signaling, resulting in suppressed proliferation in HER2-overexpressing BC cells [39]. In addition, Hst downregulates HER3 and HER4 expression and restores tamoxifen sensitivity specifically in HER2⁺ models [40]. Notably, Hst exhibits additive activity when combined with trastuzumab, as it inhibits HRG receptor signaling more broadly. These findings support the development of Hst as a dual-action therapeutic targeting both growth factor signaling and endocrine resistance [40].

The insulin-like growth factor (IGF) signaling pathway play a critical role in cell survival and proliferation and has been implicated in endocrine resistance [41]. In tamoxifen-resistance MCF-7 and T47D BC cells, upregulated autocrine IGF-II activates IGF-IR, leading to c-SRC-mediated phosphorylation of EGFR at Y845 and Y1068 [42]. This initiates a unidirectional cascade of events that potentiates EGFR/MAPK signaling and promotes estrogen-independent growth. Additionally, targeted inhibition of IGF-IR (AG1024) or c-SRC (SU6656) diminished both EGFR activity and cell growth to a significantly greater extent compared to single-drug treatment [42]. Overall, this study identifies a new resistance mechanism via the IGF-II/IGF-IR–c-SRC–EGFR axis [42]. Another investigation demonstrated an important mechanism of how growth factor receptor pathways, especially IGF-IR and EGFR, allow for tamoxifen resistance due to ligand-independent activation of the ER. Activation of downstream PI3K/AKT and MAPK pathways causes phosphorylation of ER and the ER coactivator AIB1 (SRC-3) to promote transcriptional activation of ER in the absence of estrogen [43]. This mechanism can convert tamoxifen from an antagonist into a partial agonist, particularly in HER2-overexpressing and AIB1-high BCs. Clinically, elevated expression of HER2 and AIB1 strongly correlate with poor response to tamoxifen, underscoring the importance of targeting growth factor signaling or ER coactivators to restore endocrine sensitivity [44, 45].

Fibroblast Growth Factor Receptor 1 (FGFR1) has an important function by activating the PI3K/AKT and MAPK pathways and establishing estrogen-independent ER activity. FGFR1 positive BCs are less sensitive to tamoxifen but sensitive to FGFR1 inhibitors, which may represent an avenue for treatment [46]. For instance, Lv and colleagues demonstrated that FGFR1 overexpression in ER⁺ BC is associated with lower levels of ER, lymph node metastasis, and decreased prognosis [47]. It is proposed that FGFR1 aberrant activation is responsible for inducing PI3K/AKT and MAPK pathways which promotes estrogen-independent proliferation and decreased ER expression [47]. Also, knockdown of FGFR1 restores tamoxifen sensitivity in resistant cells. Furthermore, dual inhibition using tamoxifen and FGFR1/VEGFR2 inhibitor brivanib promotes therapeutic response, as FGFR1 may drive endocrine resistance and may be a therapeutic target in ER⁺ BCs [47]. Along with EGFR/ IGF-II/IGF-IR -mediated mechanisms, the TGF-β signaling pathway is another important element of tamoxifen resistance. TGF-β has pleiotropic effects on BC progression by regulating proliferation, apoptosis, and EMT. Crosstalk between TGF-β signaling, ERα, and BC stem cells has been implicated in antiestrogen resistance. Abnormal TGF-β signaling is a contributor to antiestrogen resistance, thus targeting this pathway may offer new avenues of therapy for hormone receptor positive patients that relapse on tamoxifen treatment [48].

ERα mutations

ERα represents the main driver of estrogen mediated transcriptional activity in hormone receptor positive BC. As previously mentioned, tamoxifen is a SERM whose therapeutic efficacy arises from irreversible antagonism to ERα through binding, but the potential for mutations, posttranslational modifications and altered ERα associated signaling can radically alter the conformation, activity and cofactor interactions with ERα; ultimately facilitating the resistance of tamoxifen therapy [49]. Among these mechanisms, phosphorylation of ERα has been identified as a significant regulator [50]. Extensive work has been undertaken to identify the residues responsible for phosphorylation and altering the interaction of ERα with its co-regulators. Following phosphorylation at specific residues (i.e., Ser118, Ser167 and especially Ser305) ERα experiences alterations in interaction with co-regulators and enhanced ligand-independent activity as well as altering tamoxifen from ERα antagonist to partial agonist; thereby negatively confounding therapeutic outcomes [51]. There is also some evidence of ER positive signaling antagonism from tamoxifen within ER negative tumors; despite the absence of classical estrogen signaling with ERα, there is also evidence of either residual or even non-genomic responses to tamoxifen; however, its effects are substantially diminished without ERα reaffirming an essential role of the receptor in mediating therapeutic effects [52]. Also, dysregulation of ERα’s transcriptional co-regulators also significantly impacts tamoxifen response. For example, increases in the amount of coactivator (e.g. SRC-1) and decreases in the amount of corepressors (e.g. NCoR) will result in the presence of an active transcriptional state, allowing proliferation rather than inhibition of growth [53]. One particularly and interesting method is the K303R mutation of ERα, which gives the receptor greater sensitivity to growth factor signaling. Ultimately this means the receptor would be more available to phosphorylation via kinases (e.g. PKA) at Ser305, which leads to increased receptor activation, greater resistance to tamoxifen, and uptake and subsequent expression of estrogen responsive genes even in the presence of tamoxifen [49]. Chen et al. showed that the development of resistance to tamoxifen could be mediated by a reprogramming of the gene landscapes of estrogen-related receptor alpha (ERRα) and ERα target genes. In their report they described how in tamoxifen resistant cells, ERRα exploits the genomic binding regions of ERα, to generate a transcendental proliferative signal when classical ERα signaling is lacking. This resulted in a transcription landscape change, and also promoted estrogen-independent growth pathways to contribute to tamoxifen resistance [54]. Consistent with this research, another study reported that ERα remained a central driver of proliferation, even in tamoxifen resistant conditions; and that HER family/ERK signaling enabled the continued transcription functions ERα was able, even in these conditions. Together this suggested that ERα remained functionally active but was being utilized in mitogenic cascades which enabled tumor cells suppress the signaling of tamoxifen and maintain growth in an estrogen-independent manner [55]. Further, one minireview suggested that the contributors to resistance of ERα were partially due to ERα corepressors and histone deacetylases (HDACs). It discussed that the pool of coactivator/corepressor recruitment, and chromatin remodeling, were imbalance regarding their role in tamoxifen activity on ERα. The loss, reduction or ineffective recruitment of corepressors like NCoR or SMRT, in conjunction with alterations in HDAC activity can shift tamoxifen from an antagonist to a partial agonist [56]. As illustrated in Fig. 1, growth factor–activated signaling pathways, including EGFR/IGFR-driven MAPK and PI3K signaling, promote ligand-independent phosphorylation and nuclear activation of ERα (Fig. 1). This process facilitates ERα interaction with coactivators rather than corepressors at estrogen response elements, ultimately sustaining transcriptional programs that drive proliferation despite tamoxifen treatment. Finally, Michalides et al. investigated the function of Protein Kinase A (PKA) in tamoxifen resistance and showed that PKA phosphorylation of ERα caused a conformational arrest similar to the type seen when an agonist is bound, thus inhibiting tamoxifen antagonist activity and effectively rendering tamoxifen an ER activator under these conditions [57]. The major growth factor–driven molecular mechanisms contributing to tamoxifen resistance in ER⁺ breast cancer, along with their downstream signaling pathways and associated phenotypic outcomes, are summarized in Table 2.

Fig. 1.

The molecular processes behind tamoxifen resistance: Estradiol (E2) and tamoxifen induce distinct conformational and functional states of ERα. E2 binding promotes ERα conformational activation, receptor dimerization, and nuclear translocation, leading to estrogen response element (ERE)-dependent transcription and proliferation. In contrast, tamoxifen binding induces an altered ERα conformation that impairs receptor dimerization and limits ligand-dependent nuclear translocation. Activation of downstream kinases, including ERK1/2, MAPK, and AKT, leads to ligand-independent phosphorylation of ERα and its co-regulatory proteins, facilitating transcriptional modulation in a context-dependent manner. In tamoxifen-resistant settings, phosphorylated ERα engages in bidirectional crosstalk with growth factor receptors (EGFR and IGFR), whereby growth factor signaling enhances ERα activity and ERα-mediated feedback further amplifies receptor tyrosine kinase signaling. HDAC inhibition attenuates MAPK- and AKT-mediated ERα phosphorylation, while EGFR acetylation enhances receptor tyrosine kinase activation, collectively influencing the balance between tamoxifen’s antagonistic and partial agonistic transcriptional effects. Illustration created with BioRender

Table 2.

Molecular mechanisms of Tamoxifen resistance in Estrogen receptor-positive breast cancer

Section	Pathway / Molecular Driver	Downstream Effectors / Pathways	Proliferation	Invasion/Metastasis	Apoptosis	EMT	Other Effects	Model / Study Type	Ref.
Growth factors signaling pathways	IGF-II/IGF-IR–c-SRC–EGFR	↑EGFR/MAPK	↑	↑	↓		Estrogen-independent growth	Cell line	[43]
	HER2/AIB1	↑PI3K/AKT, ↑MAPK, ↑ER phosphorylation	↑	↑	↓		Tamoxifen resistance	Clinical/Cell line	[44, 45]
	FGFR1	↑PI3K/AKT, ↑MAPK	↑	↑	↓		↓ER expression, ↑Lymph node metastasis	Clinical/Cell line	[47]
	TGF-β	↑EMT, ↑Proliferation, ↓Apoptosis	↑	↑	↓	↑	Anti-estrogen resistance	Review	[48]
ERα mutations	ERα mutations (e.g. K303R)	↑ER phosphorylation (Ser305), ↑PKA	↑		↓		Tamoxifen partial agonist	Cell line	[49, 51]
PI3K/AKT/mTOR and MAPK pathways	PI3K/AKT/mTOR	↑Survival, ↑Metabolism, ↑Hedgehog	↑	↑	↓		Metabolic reprogramming	Cell line	[59–61]
	Akt/mTOR/HIF-1α	↑Survival, ↑Resistance	↑		↓		Reversed by glycolysis inhibition	Cell line	[62]
	mTOR/CSC	↑CSC survival, ↑Mammosphere formation	↑				Tamoxifen resistance	Patient-derived CSCs	[66]
	Luteolin/MLL3	↑MLL3, ↑H3K4me1, ↓PI3K/AKT/mTOR	↓		↑		Apoptosis, Sensitization	Cell line	[68]
	MAPK/ERRγ	↑ERRγ	↑		↓		Tamoxifen resistance	Cell line	[69]
	SPRED2/MAPK	↑MAPK	↑		↓		Tamoxifen resistance	Cell line	[70]
Dysregulation of apoptotic and autophagic processes	LDHA/Beclin-1	↑Autophagy, ↓Apoptosis, ↑EMT	↑	↑	↓	↑	Reversed by LDHA inhibition	Cell line	[77]
	GPR30/HMGB1	↑HMGB1, ↑Autophagy (via MEK/ERK)	↑		↓		Tamoxifen resistance	CAFs/Cell line	[78]
	ATP6AP1/V-ATPase	↑Lysosomal acidification, ↑Autophagy	↑		↓		Poor prognosis	Cell line	[79]
	Bcl-2/c-Myc/p21	↓Apoptosis	↑		↓		HDAC inhibitors restore sensitivity	Cell line	[80]
	PKC/ERK	↓Apoptosis	↑		↓		Tamoxifen resistance	Cell line	[81]
	Survivin	↓Apoptosis	↑		↓		Statins sensitize to tamoxifen	Cell line	[82]

PI3K/AKT/mTOR and MAPK pathways

Cell proliferation, survival and metabolism are all largely dependent on the PI3K/AKT/mTOR pathway, which upon chronic activation demonstrates insulin and anti-apoptotic AKT signals in BC [58]. This activation gives tumors the ability to develop resistance to tamoxifen by avoiding the anti-proliferative signals from the estrogen and Ac-3 H-ester tamoxifen metabolite as a result of continuous stimulation of PI3K/AKT/mTOR pathway. This makes the PI3K/AKT/mTOR pathway a main contributor to resistance against endocrine therapy [59]. Previous work encrypted aberrant or constitutive BC signaling of the PI3K/AKT signaling cascade to the induction of Hedgehog signaling pathway in tamoxifen-resistant BC cells [60]. That is to say, genomic crosstalk exists in which these pathways contribute to effects presumably downstream from and/or adjacent to the PI3K signaling pathway that contribute to tumor growth, survival, and maintenance of residual effects from anti-estrogen therapy. Discovery of Hedgehog signaling as a new therapeutic target provide options to prevent the deleterious effects of growth stimulated by compensatory signaling in tumors that have acquired resistance [60]. In tamoxifen-resistant MCF-7 BC cells, PI3K/AKT/PTEN signaling is associated with significant metabolic reprograming including the metabolism of both glucose and glutamine. These compromises in metabolism, resulting from PI3K/AKT-driven cellular reprogramming reactions, yield larger bioenergetic and biosynthetic pools that support the survival and proliferation of cancer cells under the pressure of drug treatment. This PI3K/AKT-mediated metabolic reprogramming phenomenon appears to be an important feature in the acquisition and maintenance of tamoxifen resistance [61]. Moreover, A study has indicated that blocking aerobic glycolysis leads to the disruption of the Akt/mTOR/HIF-1α axis, a crucial metabolic, survival signaling pathway in tamoxifen resistant BC cells. Furthermore, targeting this altered metabolism reversed resistance to the antiestrogen therapy, leading to restored drug sensitivity. This study demonstrated the critical role of metabolic reprogramming to sustain resistance, and that inhibiting aerobic glycolysis could be an exciting therapeutic means to overcome tamoxifen resistance [62]. Interestingly, although in a different cancer type, tamoxifen has also been shown to appear to also reverse P-glycoprotein (P-gp)-mediated multidrug resistance by inhibiting PI3K/AKT in ER-negative gastric cancer. This highlights the potential role of PI3K/AKT signaling in the regulation of drug efflux pumps involved in chemotherapy resistance, and suggests that the potential for PI3K/AKT inhibition by tamoxifen may mediate resistance mechanisms beyond the modulation of estrogen receptor activity [63]. Another study revealed that drugs such as NVP-BEZ235 and GSK2126458 have different efficacies in the treatment of tamoxifen-resistant BC cell lines. Both agents effectively target these important survival signals, but variations in potency and downstream effects suggest custom applications. These inhibitors restore sensitivity to tamoxifen by blocking aberrant PI3K/mTOR pathway activation, thus highlighting the potential of targeted pathway therapies in resistant tumors [64]. Initial studies have shown that mTOR activity inhibition restored the tamoxifen response in BC cells with aberrant Akt activation, and by directly hitting mTOR, downstream proliferative and survival signals were blocked and, consequently, the cells were re-sensitized to tamoxifen. This approach targets a critical mechanism of resistance associated with overactive PI3K/Akt/mTOR signaling [65]. In this regard, Karthik et al. demonstrated that tamoxifen selectively inhibits proliferation of differentiated ER⁺ BC cells, but paradoxically activates BC stem cells (CSCs) via the mTOR pathway by promoting ribosomal biogenesis and mRNA translation. Patient-derived CSCs demonstrated clear upregulation in mTOR activity following tamoxifen treatment, and safety clearance of these CSCs further maintained their stemness and survival. Inhibition of mTOR using pharmacological inhibitors, including dual PI3K/mTOR inhibitors (PF-04691502), reduced sustained functions of the CSCs and the formation of mammospheres-meaning a combined treatment with tamoxifen and mTOR inhibition, could potentially overcome endocrine resistance by targeting the CSCs [66]. Luteolin is a dietary flavonoid that is consumed in many fruits and vegetables which has promising pharmacological activities, including anti-inflammatory, antioxidant, and anti-cancer activity [67]. Luteolin has been shown to upregulate MLL3 expression in tamoxifen-resistant BC cells leading to H3K4 monomethylation, and inhibition of the PI3K/AKT/mTOR signaling pathway. This epigenetic modulation of gene expression re-establishes apoptosis and prevents the known drug-resistance mechanisms of these BC cells. Such studies warrant a broader use of luteolin as a drug to manage, and potentially cancer-related adverse and therapeutic effects in endocrine-resistant BC [68]. Regarding MAPK pathway, a study shown that the ERK/ MAPK signaling pathway regulates expression and transcriptional activity of ERRγ. ERRγ is a critical mediator of tamoxifen resistance in ER⁺ BC. Upon regulating ERRγ, MAPK signaling can alter gene expression programs that promote continued cell survival and proliferation during endocrine therapy. This crosstalk can generate potential therapeutic targets to re-sensitize resistant tumors to tamoxifen [69]. Likewise, absence of SPRED2 results in hyperactivation of the MAPK pathway that drives tamoxifen resistance in ERα-positive BC. Hyperactivation of MAPK increases cell proliferation and encourages survival signaling, resulting in the loss of endocrine therapy efficacy. The study emphasizes the necessity of SPRED2 as a MAPK antagonist along with its restoring function or blockade of MAPK hyperactivation as a means to overcome resistance [70].

Dysregulation of apoptotic and autophagic processes

The processes of autophagy and apoptosis are tightly regulated events that are essential for cellular homeostasis [71]. Thus, the dysregulation of both processes is especially important in the development of tamoxifen resistance in ER⁺ BC. Autophagy in early disease may initially play a tumor suppressive role; however, in an established tumor setting and during therapeutic stress induced by tamoxifen, autophagy promotes survival and drug resistance [72, 73]. Similarly, the evasion against apoptosis which may arise through mechanisms such as the overexpression of anti-apoptotic proteins such as Bcl-2 allows cancer cells to evade tamoxifen induced cell death. Many studies have highlighted the power of restoring apoptosis or targeting autophagy in removing the resistance mechanism to develop novel therapies targeting resistant BC cell lines [74]. Therapeutically targeting autophagy in ER⁺ BC provides dual benefits, both resensitizing cells to tamoxifen and limiting pro-survival adaptations [75]. Several studies demonstrated the use of autophagy inhibitors to inhibit cell proliferation and also re-sensitize cells to tamoxifen-induced cytotoxicity. Mechanistically, the autophagy inhibitors identified block important autophagy regulators including Beclin-1 and LC3, leading to tumor cell viability inhibition and tumor propagation inhibition in endocrine-resistant models [76]. Tamoxifen resistant ER⁺ BC (MCF-7/TAM-R, T47D/TAM-R) cells manifest pro-survival autophagy, diminished ATP and elevated glycolysis, via upregulated LDHA, LDHA interacts with Beclin-1 to enhance autophagy and avoid apoptosis and EM-like changes. Pharmacological or genetic inhibition of LDHA reinstated apoptosis, reversed EMT markers (increased E-cadherin, reduced vimentin), and re-sensitized cells to tamoxifen. LDHA is a functional mediator of resistance to tamoxifen through metabolic reprogramming and autophagy regulation, suggesting LDHA is a potential therapeutic target [77]. Within BC-associated fibroblasts (CAFs), tamoxifen activates GPR30 and increases HMGB1 secretion via a PI3K/AKT signaling pathway. HMGB1 secretion increases autophagy activation via MEK/ERK, ultimately leading to increased tamoxifen resistance of ERα-positive BC cells. Blocking either GPR30 or the downstream signaling pathways can decrease tamoxifen resistance and shows possible targets to improve the effectiveness of endocrine therapies [78]. A recent study highlights ATP6AP1 as a critical mediator of autophagy-related drug resistance in luminal BC. ATP6AP1 facilitates lysosomal acidification and promotes autophagosome-lysosome fusion through interactions with V-ATPase and the Rab7-HOPS complex. Its overexpression is associated with increased tumor proliferation, resistance to tamoxifen, and a correlation with poor patient outcomes. These findings suggest that ATP6AP1 may serve as an innovative therapeutic target in the context of endocrine-resistant BC [79]. In the study by Raha and colleagues demonstrated that tamoxifen-resistant ER⁺ breast cancer cells (referred to as TAMRM and TAMRT) demonstrate increased ER levels and increased levels of Bcl-2, and decreased levels of progesterone receptor leading to almost complete loss of ER responsiveness, which is mediated by transcriptional and epigenetic mechanisms, and involves increased Bcl-2, increased c-Myc levels, and decreased p21 levels. Treatment with HDAC I inhibitors would restore tamoxifen sensitivity by demethylating the epigenetic alterations that lead to resistance. In fact, this is a process that diminishes Bcl-2 expression, which induces apoptosis and reprograms the cells so that resistance no longer exists [80]. In a research study, it demonstrated that PKC and ERK signaling pathways play an important role in the susceptibility of BC cells to tamoxifen, or resistance to tamoxifen. In ER⁺ MCF-7 cells tamoxifen induced apoptosis via activation of PKCδ and inhibition of ERK phosphorylation. In drug resistant ER-negative MCF-7/ADR cells, phosphorylated ERK was maintained due to elevated levels of PKCα which enabled evasion of tamoxifen-induced apoptosis. Inhibition of PKCα and ERK activity increased apoptosis in ER - MDG drug resistant cells. These results provide the basis for new models of therapeutic resistance based on targeting PKCδ/ERK and PKCα/ERK [81]. Moriai et al., defined survivin, an important anti-apoptotic protein involved in tamoxifen resistance in ER⁺ BC. Survivin overexpression inhibited tamoxifen -mediated apoptosis in MCF-7 cells, while survivin knockdown with small interfering RNA (siRNA) enhanced cell death both alone and in combination with tamoxifen. Lovastatin, an HMG-CoA reductase inhibitor (HRI), downregulated survivin in a dose dependent manner and when combined, more than additively increased tamoxifen mediated apoptosis. These results suggest that pharmacological disruption of survivin - through HRIs like lovastatin or rosuvastatin- may be opportunistic strategies to overcome tamoxifen resistance [82].

Drug efflux transporters

Multidrug resistance (MDR) in human malignancies is largely attributed to enhanced drug efflux mechanisms. In particular, members of the ATP-binding cassette (ABC) transporter superfamily actively export hydrophobic chemotherapeutic agents from tumor cells, thereby reducing intracellular drug accumulation, diminishing therapeutic efficacy, and ultimately contributing to treatment failure and tumor recurrence [83]. A total of 48 human ABC transporter genes has been classified into seven subfamilies (ABCA to ABCG) based on structural similarities and sequence homology [84]. Multidrug resistance protein 1 (MRP1/ABCC1), P-glycoprotein (P-gp/ABCB1), and breast cancer resistance protein (BCRP/ABCG2) are thought to be the members of the ABC transporter superfamily that are most closely associated with MDR in cancer cells [85, 86]. Emerging evidence indicates that ncRNAs are key regulators of ABC transporter expression and function, thereby contributing to tamoxifen resistance. For example, Chen et al., discovered that mRNA ABCB1 was increased based on the RNA expression profiles in adriamycin-resistant BC cells, but GAS5 was downregulated. GAS5 regulates its target Dickkopf 2 (DKK2) by acting as a molecular sponge of miR-221-3p and preventing activation of the Wnt/β-catenin pathway, according to a detailed investigation of the associated mechanism [87]. The ABCB1 promoter contains TCF4/LEF binding domains, which are targets of β-catenin/TCF4 transcriptional regulators [88]. Thus, downregulating GAS5 will boost the expression of ABCB1, disinhibit the Wnt/β-catenin pathway, and encourage adriamycin to leave intracellular sources. Targeting lncRNAs may become a viable strategy to eradicate or reduce MDR by decreasing drug efflux from tumor cells due to their role in controlling drug efflux metabolism. Moreover, ROR1 (Receptor Orphan tyrosine kinase-like Receptor 1) is an upstream regulator of ABCB1, a subfamily of ABC transporters. Through MAPK/ERK and p53, ROR1 controls ABCB1 transcription and stability [89].

Additionally, it was found that ROR1 overexpression is associated with poor treatment response and tumor recurrence in BC chemoresistant cells MDA-MB-231 and SUM-159PT. BC cells were made more sensitive to DOX and cisplatin treatment by ROR1 inhibition [89]. Although most studies have focused on ABCB1 and ABCC family members in tamoxifen resistance, emerging evidence from other therapeutic contexts supports a broader role for ncRNA-mediated regulation of ABC transporters [90]. For example, exosome-derived miR-187-5p from taxol-resistant breast cancer cells was shown to regulate tumor growth and drug resistance through targeting ABCD2 and activation of Wnt/β-catenin signaling, highlighting the potential for exosomal ncRNAs to modulate transporter-associated resistance pathways across treatment modalities [91]. Collectively, these findings underscore the importance of ncRNA-driven regulation of drug efflux transporters as a critical and underappreciated mechanism of tamoxifen resistance. Targeting ncRNA–ABC transporter regulatory networks may therefore represent a promising strategy to restore intracellular drug accumulation and re-sensitize ER⁺ breast cancer cells to endocrine therapy.

NcRNAs as regulators of multiple mechanistic routes to Tamoxifen resistance

In addition to the mechanisms of tamoxifen resistance previously discussed, there has been a growing emphasis in recent years on the role of ncRNAs in this area. ncRNAs, which include miRNAs, lncRNAs, and circRNAs, have been shown to regulate tamoxifen resistance in ER⁺ BC by targeting various pathways. For example, miRNAs like miR-221/222 promote resistance to tamoxifen by downregulating tumor suppressor genes and promoting cell survival [14]. Other miRNAs, such as miR-18a regulate hypoxia-inducible factors (HIFs) that promote adaption to cellular stress [92]. In the same manner, lncRNAs function as molecular sponges and scaffolds by regulating miRNAs and/or their signaling pathways. For instance, lncRNA UCA1 increases tamoxifen resistance by sequestering miR-18a which in turn increases expression of HIF1α and activates hypoxia pathways [92]. Whereas circRNAs are stable and primarily regulate miRNAs which function as sponges and indirectly influence the expression of genes related to drug resistance [93]. Some circRNAs regulate apoptosis and genes associated with autophagy that influence cancer cell survival under tamoxifen treatment [94]. Furthermore, ncRNAs influence not only intracellular pathways but also the tumor microenvironment and intercellular communication, which adds to the complexity of resistance [92, 94]. A comprehensive understanding of this intricate network is crucial, as targeting specific ncRNAs or their interactions presents promising therapeutic strategies to address tamoxifen resistance. In the following sections, we will provide detailed analyses of the specific roles and molecular mechanisms of these ncRNAs to uncover novel biomarkers and therapeutic targets in tamoxifen-resistant BC.

MiRNAs

The role of miRNAs as contributors to endocrine resistance in BC, specifically tamoxifen resistance, has been intensely studied in the last few years. These non-coding small regulatory RNAs are able to control gene expression at the post-transcriptional level and have both been functionally categorized into oncogenic miRNAs or tumor suppressive miRNAs. Beyond their role in cancer, miRNAs and lncRNAs have been extensively studied as therapeutic targets and therapeutic molecules in other diseases, such as preeclampsia (PE), a pregnancy-associated disorder. In PE, miRNA-based approaches, including miRNA mimics, inhibitors, lncRNA sponges, and exosome-associated miRNAs, have demonstrated the potential to modulate disease pathways [95]. Although clinical validation remains limited, these studies highlight the versatility of non-coding RNAs as both biomarkers and therapeutic agents. In this regard, studies in PE have shown that miRNAs, such as miR-510-3p, can act as therapeutic targets and molecules by modulating key signaling pathways like VEGFA/PI3K/AKT/eNOS/mTOR. These findings highlight the potential of ncRNAs to both mediate disease and serve as therapeutics [96]. Similarly, in breast cancer, miRNAs and lncRNAs may regulate tamoxifen resistance, suggesting that RNA-based strategies, like mimics, inhibitors, or sponges, could offer novel approaches to overcome endocrine resistance. Oncogenic miRNAs generally advertise cancer progression and therapeutic resistance by inhibiting tumor suppressor genes, and tumor suppressive miRNAs work by inhibiting oncogenic processes [14, 97, 98]. As illustrated in Fig. 2, miRNAs play a dual role in modulating tamoxifen resistance and sensitivity in ER⁺ breast cancer. There have been altered expressional profiles of both types of miRNAs associated with tamoxifen resistance, modulating several cell processes including cell proliferation, apoptosis, autophagy, EMT, and survival signaling (Fig. 2). Many of these regulatory processes can occur in the tumor, but also in the surrounding microenvironment, affecting treatment response and cancer progression. The interactions between miRNAs and signaling pathways are complex and those interactions could be biomarkers of endocrine resistance or therapeutic targets to reverse tamoxifen insensitivity [99]. miRNAs have a critical regulatory consequence and understanding their functional role sheds light on how resistance is established and future attempts to improve the efficacy of endocrine therapies in patients with BC.

Fig. 2.

MicroRNAs in tamoxifen resistance and sensitivity in breast cancer. The figure depicts the contribution of oncogenic, tumor-suppressive, and exosomal miRNAs to the tamoxifen response in breast cancer. Tumor oncogenic miRNAs, including miR-519a, miR-155, and miR-92a-3p, enhance PI3K/AKT and STAT3 signaling processes that drive resistance to tamoxifen. Tumor-suppressive miRNAs, such as miR-27b-3p, miR-342, miR-320a, miR-449a, let-7b/i, and miR-873, inhibit proliferation as well as reinstate ER signaling or enhance apoptosis to induce tamoxifen sensitivity. Exosomal miRNAs (miR-9-5p, and miR-221/222) impact apoptosis and proliferation, which are components of the resistant processes in the tumor microenvironment. Illustration created with BioRender

Oncogenic MiRNAs (oncomiRs)

Recent studies have demonstrated that a member of the C19MC cluster of microRNAs, namely miR-519a, is a novel oncomiR associated with the upregulation of tamoxifen-resistant ER⁺ BC cells. miR-519a promotes resistance by co-targeting well-characterized tumor suppressor genes (PTEN, RB1, and CDKN1A) that govern key processes primarily through PI3K signaling and cell cycle regulation. The provision of simultaneous repression of these pathways promotes cell proliferation and provides resistance to tamoxifen. Further associative evidence exists based on clinical findings correlated with miR-519a expression and poor survival, supporting its potential utilization as a therapeutic target or predictive biomarker [100]. Miller and coworkers have identified many differentially expressed miRNAs in tamoxifen-resistant MCF-7 BC cells, characterizing miR-221 and miR-222 as the most up-regulated. These miRNAs can directly bind p27Kip1, a tumor suppressor in BC, to either promote cell cycle progression or attenuate the activity of tamoxifen. Further, miR-221/222 over-expression was seen in HER2-positive breast tumors, implicating a potential clinical connection to resistance of endocrine therapy. Chemotherapy would, in a way, be effectively selected for using either miR-221/222 as a status indicator. Restoring p27Kip1 expression showed increased tamoxifen-induced cell death via apoptosis and possibly autophagy [101], following that another group of scientists showed that exosomal miR-221/222, secreted by tamoxifen-resistant BC cells had transferred into sensitive cells and downregulated p27 and ERα to propagate tamoxifen resistance ; however, a blockade of miR-221/222 could reverse this phenomenon. Overall, the study confirmed exosomal miR-221/222 as key mediators of intercellular communication and resistance propagation and suggested their potential as both diagnostic biomarkers and therapeutic targets in the struggle against tamoxifen resistance in ER⁺ BC [102]. Likewise, a case-control study found that miR-221 expression was significantly higher in patients with luminal-subtype BC who had local or distant recurrence after at least one year of tamoxifen therapy, relative to patients with no recurrence. Associations with progesterone receptor (PR) status, Ki-67, lymphovascular invasion, or the stage of cancer were not significant. The study indicated that high levels of serum miR-221 may be a potential biomarker of tamoxifen resistance in BC [12]. Another study identified miR-155 as significantly upregulated in resistant BC cells and patient samples. Overexpression of miR-155 promoted cell survival and tamoxifen resistance, by directly targeting SOCS6 and inhibiting its activity, while inhibiting miR-155 restored drug sensitivity and enhanced apoptosis. The conclusion from these findings suggested the miR-155/SOCS6-STAT3 axis was more important in tamoxifen resistance, which would be a novel pathway in the pathway, suggesting miR-155 was a feasible therapeutic target in BC [103]. Communication between cells via exosomal microRNAs has a significant impact on drug resistance in BC. In Liu and his coworkers’ research, they show that exosomes from tamoxifen-resistant MCF-7/TAM cells contained higher levels of miR-9-5p, which, when transferred to MCF-7 sensitive cells, inhibited apoptosis, and increased resistance to tamoxifen. They studied how miR-9-5p increased the resistance to tamoxifen and determined that miR-9-5p represses ADIPOQ, a gene that was inversely correlated with drug resistance. Also, when tumors were exposed to exosomal miR-9-5p in vivo, tumors had increased tamoxifen resistance [104]. Utilizing integrative bioinformatics methods, researchers have identified that miR-92a-3p is significantly overexpressed in tamoxifen-resistant BC cells and tissues, and higher expression of miR-92a-3p associates with poor prognosis, which in this context is shown to be dependent on complex regulatory networks involving several key genes and pathways, including IL-6/STAT3 and Wnt/β-catenin. Although experimental validation is currently lacking, miR-92a-3p could become a marker and target for therapeutics in tamoxifen resistance in BC [105] (Table 3).

Table 3.

Summary of MiRNAs associated with Tamoxifen resistance in breast cancer

miRNA	Role	Expression in BC	Molecular Mechanism	Outcome	Ref
miR-519a	OncomiR	UP	Targets PTEN, RB1, CDKN1A; activates PI3K/cell cycle	↑Proliferation, ↑Tamoxifen resistance, Poor survival	[100]
miR-221/222	OncomiR	UP	Targets p27Kip1, ERα; exosomal transfer; HER2 + tumors	↑Proliferation, ↓Apoptosis, ↑Tamoxifen resistance, Biomarker	[12, 101, 102]
miR-155	OncomiR	UP	Targets SOCS6, activates STAT3	↑Survival, ↑Tamoxifen resistance, ↓Apoptosis	[103]
miR-9-5p	OncomiR	UP (exosomal)	Represses ADIPOQ	↓Apoptosis, ↑Tamoxifen resistance	[104]
miR-92a-3p	OncomiR	UP	Regulates IL-6/STAT3, Wnt/β-catenin, complex networks	↑Tamoxifen resistance, Poor prognosis	[105]
miR-27b-3p	Tumor Suppressor	DOWN	Targets NR5A2, CREB1	↑Tamoxifen sensitivity, ↓Proliferation, ↓Aromatase	[106]
miR-342	Tumor Suppressor	DOWN	Regulates apoptosis, cell cycle (targets TXNIP, EVL)	↑Apoptosis, ↓Growth, ↑Tamoxifen sensitivity, ↑Survival	[107, 108]
miR-320a	Tumor Suppressor	DOWN	Targets ARPP-19, ERRγ, c-Myc/Cyclin D1 axis	↓Proliferation, ↓Colony formation, ↑Tamoxifen sensitivity	[109]
miR-449a	Tumor Suppressor	DOWN	Targets ADAM22 (affects PPARG, KRAS, LYN)	↑Tamoxifen sensitivity, ↓Tamoxifen resistance	[110]
let-7b/let-7i	Tumor Suppressor	DOWN	Targets ER-α36, inhibits MAPK/Akt pathways	↑Tamoxifen sensitivity, ↓MAPK/Akt, ↓ER-α36	[111]
miR-375	Tumor Suppressor	DOWN	Targets MTDH, reverses EMT	↑Tamoxifen sensitivity, ↓EMT, ↓Invasion	[112]
miR-873	Tumor Suppressor	DOWN	Targets CDK3, inhibits ERα phosphorylation	↓Proliferation, ↑Tamoxifen sensitivity	[113]

Tumor-suppressive MiRNAs

Zhu et al. reported that miR-27b-3p, a tumor suppressor miRNA, was significantly downregulated in tamoxifen-resistant breast cancer cells and patient tissues compared to controls. Functional experiments indicated that the restoration of expression of miR-27b-3p in the model systems increased sensitivity to tamoxifen in BC cells in both cell culture systems as well as animal models of BC [106]. Mechanistically, miR-27b-3p was found to directly target NR5A2 and CREB1, which are genes known to regulate processes associated with cellular proliferation, anti-apoptosis, as well as aromatase expression. Most interestingly, miR-27b-3p levels were found to be inversely correlated with NR5A2 and CREB1 levels in clinical samples, indicating potential as a biomarker for diagnosis and treatment [106]. As illustrated in Fig. 2, miRNAs play a dual function in modulating tamoxifen resistance and sensitivity in ER⁺ breast cancer. In an extensive study by Kim et al., large clinical datasets from TCGA and GEO were evaluated to determine if the miRNAs related to tamoxifen response might be prognostic in BC. Increased expression of miR-342 was significantly associated with improved overall survival and disease-free survival, particularly in ER⁺ patients, and was correlated with expression of gene signatures of tamoxifen sensitivity. In contrast, miR-221, miR-222 and miR-451 had no prognostic value. Ultimately, these results suggest that miR-342 is a potential biomarker for predicting tamoxifen response and guiding treatment in BC [107]. In line with the same study, another group of researchers found that miR-342 is downregulated in tamoxifen-resistant BC cell lines and primary tumors from patients with limited responses to tamoxifen. Restoring miR-342 expression sensitized resistant cells to tamoxifen-induced apoptosis and suppressed cancer cell growth. Transcriptomic analysis indicated that many targets of miR-342 are involved in apoptosis and cell cycle regulation, including TXNIP. Additionally, a significant correlation between miR-342 and its host gene, EVL, was observed, suggesting both as potential predictive markers [108]. In tamoxifen resistant BC cells, miR-320a was significantly downregulated, and restoring expression of miR-320a as well as its function sensitized resistant cells to tamoxifen by directly targeting ARPP-19 and ERRγ and, therefore, inhibited the downstream regulators of c-Myc and Cyclin D1. Progesterone treatment considerably increased miR-320a levels by inhibiting c-Myc, while estrogen treatment had the opposite effect. Subsequently, functional assays indicated that restoration of miR-320a inhibited cell proliferation and colony-forming ability, along with the ability to initiate tumors in vivo, reinforcing the potentially therapeutic implications of targeting the c-Myc/miR-320a/ARPP-19/ERRγ signaling axis in tamoxifen resistant BC [109]. Another miRNA is miR-449a which functional analysis showed that restoring miR-449a expression re-sensitized the resistant cells to tamoxifen treatment, while inhibiting miR-449a expression promoted resistance. ADAM22 was pinpointed as a direct target of miR-449a, and silencing ADAM22 reversed tamoxifen resistance induced by the loss of miR-449a expression. Bioinformatic analyses suggested that ADAM22 may regulate the resistance pathways through binding with PPARG, KRAS, and LYN proteins, further providing evidence toward targeting the miR-449a/ADAM22 axis in treatment [110]. Analysis of BC tissues and cell lines revealed that let-7b and let-7i miRNAs are downregulated in tamoxifen-resistant cells, associated with ER-α36 upregulation. In functional assays, let-7 mimics inhibited ER-α36, MAPK and Akt signaling, and re-established tamoxifen sensitivity. Ectopic expression of ER-α36 lacking the 3′UTR negated this activity. This suggests that let-7 miRNAs play a functional role in regulating ER-α36-dependent pathways in BC, and that loss of let-7 contributes to tamoxifen resistance [111]. The detection of miR-375 depletion in a tamoxifen-resistant BC cell model led to the discovery that loss of miR375 is implicated in the tamoxifen-resistant phenotype and has been shown to promote an EMT-like, invasive phenotype. The restoration of miR-375 sensitizes cells back to tamoxifen, and also reverse EMT, most likely because miR-375 targets and potently inhibits MTDH. In BC patients, there is a clinical correlation of miR-375 and MTDH expression as well as an association of high MTDH expression with poor outcomes [112]. Through research on miR-873 in BC, it has been determined that miR-873 is downregulated in tumors and tamoxifen resistant cells. Restoration of miR-873 expression inhibited ERα activity and cell proliferation by directly regulating CDK3; a kinase that is overexpressed in BC tissue and promotes ERα phosphorylation. Further, the re-expression of miR-873 reversed tamoxifen resistance in resistant cells [113] (Table 3).

LncRNAs

LncRNAs have garnered increasing enthusiasm as gene regulators and key mediators of cellular function in various contexts, including progression of malignancies and cancer therapy resistance [114, 115] (Fig. 3). Studies have shown increased levels of specific lncRNAs are associated with tamoxifen-resistant BC, which actively perturbed ER function via enhanced pro-ER signaling, mediated apoptotic responses and modulated key survival active molecules involved in survival pathways in BC. Emerging evidence indicates that lncRNAs can have oncogenic roles or act as tumor suppressors dependent on the network and context of their targets. Additionally, lncRNAs can act as molecular sponges for miRNAs and thereby indirectly regulate gene expression, modulate chromatin dynamics or act as scaffolds for protein complexes that drive transcription and/or drivers of signaling cascades [14, 114]. Additionally, evidence suggests that lncRNAs play a crucial role in maintaining a tumor microenvironment in drug-resistant cancers. They may help establish pro-survival conditions that promote either autophagy or the proliferation of ER⁺ BC cells in response to tamoxifen therapy. On the basis of their diverse regulatory activities, lncRNAs would appear to have a critical role to the mediation of endocrine resistance. As detailed understanding is realized regarding the contribution of lncRNAs to the inability to respond to tamoxifen, the realization of RNA targeting and/or routinizing lncRNA in combination will allow for novel methods to improve person’s clinical outcomes with BC [92, 116] (Table 4).

Fig. 3.

lncRNA-mediated regulation of tamoxifen resistance via autophagy and apoptosis pathways. This figure illustrates the mechanisms by which lncRNAs can cause tamoxifen resistance in breast cancer by regulating autophagy and apoptosis. In the autophagy pathway, the lncRNAs, H19 and ROR appear to function by regulating Beclin1 expression or autophagosome-lysosome fusion, both of which alter the cellular drug response. In the apoptosis pathway, lncRNAs such as UCA1, MAFG-AS1, TTN-AS1, LINC00626, and GAS5 can modulate important signaling pathways, such as PI3K/AKT, ER stress response, mitochondrial outer membrane permeabilization (MOMP) or caspase activity. Together, these lncRNAs interfere with both intrinsic and extrinsic apoptotic stimuli and drive tamoxifen resistance. Illustration created with BioRender

Table 4.

Key LncRNAs and their mechanisms in Tamoxifen resistance in breast cancer

Functional Category	LncRNA	Expression in Breast Cancer	Molecular Mechanism / Pathway	Outcome	Ref
ceRNA Activity	CYTOR	UP	Sponges miR-125a-5p, ↑SRF, activates Hippo/MAPK	↑Tamoxifen resistance, ↑Proliferation, Poor response	[117]
	ROR	UP	Sponges miR-205, ↑ZEB1/2, promotes EMT	↑Tamoxifen resistance, ↑Proliferation, ↑Invasion, ↑EMT	[118]
	SNHG6	UP	Sponges miR-101, ↑Sox2/Oct4/EZH2, ↑EMT, ↑Stemness	↑Tamoxifen resistance, ↑Proliferation, ↑Invasion, ↑EMT	[119]
	ADAMTS9-AS2	DOWN	Sponges miR-130a-5p, ↑PTEN	↑Tamoxifen sensitivity, ↓Proliferation, ↑Apoptosis	[120]
	ATXN8OS	UP	Sponges miR-16-5p, ↑VASP	↑Tamoxifen resistance, ↑Invasion	[121]
	DSCAM-AS1	UP	Sponges miR-137, ↑EPS8	↑Tamoxifen resistance, ↑Proliferation, ↓Apoptosis	[122]
	UCA1	UP	Sponges miR-18a, ↑HIF1α, feed-forward loop	↑Tamoxifen resistance, ↑Cell cycle progression	[123]
	MAFG-AS1	UP	Sponges miR-339-5p, ↑CDK2, ↑FOXO1 phosphorylation	↑Tamoxifen resistance, ↑Cell cycle, ↓Apoptosis	[124]
Signaling Pathway Regulation	UCA1	UP	Activates PI3K/AKT/CREB, AKT/mTOR, Wnt/β-catenin	↑Tamoxifen resistance, ↑Survival, ↓Apoptosis	[126, 127]
	91 H	UP	Activates mTOR	↑Tamoxifen resistance, ↑Proliferation, ↑Migration	[128]
	SBF2-AS1	UP	Interacts with YBX1, activates PI3K/AKT/mTOR	↑Tamoxifen resistance, ↑Proliferation	[129]
	TTN-AS1	UP	↓miR-107, ↑ZNRF2, activates PI3K/AKT	↑Tamoxifen resistance, ↑Proliferation, ↑Invasion	[130]
	GAS5	DOWN	Inhibits mTOR, ↑PTEN, ↑p-AMPK2	↑Tamoxifen sensitivity, ↓Proliferation	[131]
	HOTAIR	UP	Stabilizes ER, ↑ER chromatin occupancy, ↑HGF/c-Met/NF-κB	↑Tamoxifen resistance, ↑Proliferation, ↑ER signaling	[133]
	LINP1	UP	↓ER expression, modulates ER signaling	↑Tamoxifen resistance, ↑Proliferation, ↑Migration	[134]
	LINC00894-002	DOWN	↓miR-200a/b, ↑TGF-β2/ZEB1, activates EMT	↑Tamoxifen resistance, ↑Invasion, ↑EMT	[135]
	LINC00626	UP	↑UPF1, represses PERK-ATF4-CHOP apoptotic pathway	↑Tamoxifen resistance, ↓Apoptosis	[136]
Epigenetic Modulation	HOTAIRM1	UP	Interacts with EZH2, ↓H3K27me3 at HOXA1, ↑HOXA1	↑Tamoxifen resistance, ↑HOXA1, Biomarker potential	[13]
	H19	UP	↑NAT1 promoter methylation, ↓NAT1, ↑DNMT3B, ↓Beclin1	↑Tamoxifen resistance, ↓Autophagy, Poor prognosis	[141, 142]
	AGPG	UP	Binds PURα, ↑E2F1 signaling	↑Tamoxifen resistance, ↑Proliferation, Poor outcome	[143]
Cellular Fate Determination	DILA1	UP	Binds Cyclin D1, inhibits degradation	↑Tamoxifen resistance, ↑Proliferation	[145]
	SOX2OT	DOWN	Not via SOX2, mechanism unclear	↑Tamoxifen resistance, ↑Aggressiveness, ↓Survival	[146]
	ROR	UP	Inhibits autophagy, ↓LC3/Beclin1, ↓P-gp, ↓GST-π	↑Tamoxifen resistance, ↑Proliferation, ↑Invasion	[148]
	UCA1 (exosomal)	UP	Exosomal transfer, inhibits apoptosis in recipient cells	↑Tamoxifen resistance, ↑Survival	[150]

LncRNAs acting as CeRNAs

Among the studies investigating ceRNA-mediated mechanisms of tamoxifen resistance, one reports that lncRNA CYTOR is significantly upregulated in MCF‑7 tamoxifen-resistant breast cancer cells. When CYTOR is silenced, tamoxifen sensitivity is regained, which confirms the functional role of CYTOR in tamoxifen resistance. CYTOR directly binds and inhibits miR‑125a‑5p, which causes upregulation of serum response factor (SRF), and activation of the Hippo pathway and MAPK signaling. In the clinical study, in BC patient tumors, high CYTOR levels are indicative of poor response to tamoxifen, and indicate high SRF [117]. LncRNA-ROR is upregulated in tamoxifen-resistant BC cell lines and promotes proliferation, invasion, and EMT. ROR can act as a ceRNA and sponge miR-205, thus lowering miR-205 levels and, in turn, upregulate the EMT transcription factors ZEB1 and ZEB2. Knocking down ROR with siRNA reverses these effects, restoring miR-205 expression and drug sensitivity in the resistant cells [118]. According to a new study, lncRNA SNHG6 was identified to be significantly overexpressed in resistant ER⁺ BC cells. SNHG6 acts as a sponge of miR-101, inhibiting its function. This axis induces EMT and increases stemness markers (Sox2, Oct4, EZH2), while also increasing proliferation and invasion. Silencing SNHG6 or restoring miR-101 reverses those endpoints, but re-sensitizes cells to tamoxifen confirming the SNHG6–miR-101 axis as a potential new driver of resistance [119]. Continuing studies on lncRNAs functioning as ceRNAs, ADAMTS9-AS2 was found to sponge miR-130a-5p and induce PTEN upregulation to overcome tamoxifen resistance in BC. ADAMTS9-AS2 expression was reduced in resistant cells in correlation with increased proliferation and decreased apoptosis. Thus, the ADAMTS9-AS2/miR-130a-5p/PTEN axis is an important regulator of tamoxifen sensitivity [120]. Recent study has identified that ATXN8OS is highly expressed in BC and tamoxifen-resistant cells and contributes to the resistance by sponging miR-16-5p and enhancing VASP levels. Silencing of ATXN8OS was shown as a way to improve the tamoxifen response in both in vitro configuration and in vivo through the miR-16-5p/VASP axis [121]. Tamoxifen-resistant BC upregulated DSCAM-AS1, which promoted resistance in the disease through sponging miR-137 and upregulating EPS8, which encouraged proliferation in tumor cells and promoted progression through the cell cycle while inhibiting apoptosis. DSCAM-AS1 transcription levels were inversely correlated with miR-137 transcription levels but positively correlated with EPS8 transcription levels, establishing the DSCAM-AS1/miR-137/EPS8 axis as a key modulator of tamoxifen resistance [122]. Another investigation indicated that tamoxifen treatment induces UCA1 upregulation through HIF1α, with UCA1 acting as a ceRNA by sponging miR-18a, a negative regulator of HIF1α, thereby generating a feed-forward loop that increases expression of UCA1, and promotes tamponade resistance through tight regulation of the cell cycle and diminished sensitivity. Therapies directed toward the UCA1/miR-18a/HIF1α axis could enhance therapeutic efficacy [123]. LncRNA MAFG-AS1 is overexpressed in ER⁺ BC and is associated with negative prognosis. MAFG-AS1 promotes tamoxifen resistance by sponging miR-339-5p which leads to the upregulation of CDK2. The upregulation allows cell cycle progression and inhibits apoptosis via ER signaling pathway in coordination with FOXO1 phosphorylation. In addition, there is positive feedback pathway MAFG-AS1 and ERα promote resistance. Targeting the MAFG-AS1/miR-339-5p/CDK2 axis could provide novel therapeutic strategies for endocrine-resistant BC [124] (Table 4).

LncRNAs modulating signaling pathway

LncRNAs regulate the PI3K/AKT/mTOR signaling pathway and contribute to BC tamoxifen resistance. This regulation contributes to the cellular resistance of tamoxifen-induced apoptosis, and drives ongoing tumor growth, making the PI3K/AKT/mTOR axis an important target for understanding lncRNA-based endocrine resistance [125]. For example, lncRNA UCA1 was shown to be significantly upregulated in tamoxifen-resistant BC cells, where it induces resistance by interacting with EZH2 to repress p21 expression and activate the PI3K/AKT/CREB signaling pathway. Additionally, another study has shown that UCA1 enhances the effectiveness of tamoxifen resistance by regulating the AKT/mTOR axis to inhibit apoptosis and promote cell survival. Importantly, when AKT/mTOR signaling is inhibited or UCA1 is knocked down, tamoxifen sensitivity is restored, indicating an important role for UCA1 in regulating survival pathways and may therefore be a therapeutic target [126, 127] (Fig. 3). Another study found that knockdown of lncRNA 91 H in ER⁺ BC cell line was able to decrease migration and proliferation of cancer cells, and also significantly increased sensitivity to tamoxifen, through inhibition of the mTOR pathway. mTOR inhibition combined with tamoxifen to reduce cell viability produced a synergistic effect. These results suggest that 91 H modulates tamoxifen resistance primarily through activation of the mTOR pathway [128]. The oncogenic lncRNA SBF2-AS1 interacts with YBX1 to generate a positive feedback loop that activates the PI3K/AKT/mTOR signal pathway promoting cell proliferation and tamoxifen resistance in the BC cellular model. Knocking down YBX1 or SBF2-AS1, disrupts this axis which activates the PI3K/AKT/mTOR pathway along with reducing the PI3K/AKT/mTOR and importantly restricted tamoxifen response. Collectively, the researchers validate the YBX1/SBF2-AS1/PI3K/AKT/mTOR axis as an actionable target to overcome tamoxifen resistance [129]. TTN-AS1 is an additional lncRNA that contributes to tamoxifen resistance by downregulating miR-107 and thereby upregulating ZNRF2. The TTN-AS1/miR-107/ZNRF2 axis activates the PI3K/AKT signaling pathway, which promotes cell proliferation, invasion, and survival. Knockdown of TTN-AS1 or ZNRF2 increased apoptosis and attenuated resistance to tamoxifen, underlining that the TTN-AS1/miR-107/ZNRF2/AKT/PI3K is an important mechanism underlying tamoxifen resistance that could be targeted therapeutically [130]. In contrast, lncRNA GAS5 expression is reduced in tamoxifen-resistant MCF-7R BC cells, and its reinstatement sensitized the cells to tamoxifen by inhibiting activation of the mTOR pathway. Metformin treatment increased expression of GAS5 and led to decreased mTOR, p-mTOR, and p-P70S6K, while increasing PTEN and p-AMPK2 levels. Critically, metformin had no anti-proliferative action on GAS5-deficient cells, indicating that GAS5 mediates this action. Thus, GAS5 sensitizes cells to tamoxifen through negative regulation of the mTOR signaling pathway [131].

An additional pivotal mechanism in tamoxifen resistance is the ER signaling pathway since it is essential for the growth of ER⁺ breast tumor cells. In resistant tumors, this pathway may be constitutively active without estrogen, often in ways we yet do not fully understand, such as via ligand-independent situations using components such as lncRNAs that support ER signaling despite undergoing an endocrine therapy [132]. In this context, lncRNA HOTAIR was significantly upregulated in tamoxifen resistant BC tumors compared to their primary forms. While HOTAIR would typically be repressed by ER signaling, it becomes derepressed with ER signaling blockade, by either hormone deprivation or tamoxifen treatment, causing an upregulation of the HOTAIR. Mechanistically, upon deregulation of HOTAIR, it stabilizes ER protein levels, increases ER chromatin occupancy and amplifies downstream ER transcriptional programs even without estrogen. There is a physical interaction between HOTAIR and ER, which may also facilitate nuclear localization and genomic targeting of ER in the absence of estrogen. Overall, these data indicate that HOTAIR may be promoting ligand-independent ER activation and thus contributes to the proliferation of resistant phenotypes [133]. Recent research shows that high LINP1 expression leads to tamoxifen resistance by regulating cell proliferation, migration, invasion, and sensitivity to tamoxifen in BC cells both in vitro and in vivo. From a mechanistic standpoint, LINP1 expression is stimulated by treatment with tamoxifen or by estrogen starvation. This response to tamoxifen decreases ER expression and promotes resistance to tamoxifen. Therefore, at least to some extent, LINP1 is promoting tamoxifen resistance by modulating ER expression and signaling, but further research would be needed to determine the specific mechanisms involved [134]. An additional study discovered that LINC00894-002, which is directly upregulated by ERα signaling, was significantly downregulated in tamoxifen-resistant MCF-7 cells. When depleted, the levels of tumor-suppressive miRNAs were also decreased, specifically miR-200a-3p and miR-200b-3p, which led to hyperexpression of TGF-β2 and ZEB1, two important drivers of EMT. This ultimately leads to activation of the TGF-β2/ZEB1 axis, promoting invasion and contributing to the development of acquired tamoxifen resistance [135]. LINC00626 is transcriptionally regulated by ERα signaling and is elevated in tamoxifen resistant BC cells. It functions as a mediator of resistance through increases in UPF1 expression, which represses the ER stress-mediated PERK-ATF4-CHOP apoptotic pathway. Inhibition of LINC00626 led to activation of this ER stress response, ultimately sensitizing the cells once again to tamoxifen treatment. Thus, LINC00626 connects ER signaling to unfolded protein response modulation, facilitating endocrine therapy resistance [136].

Besides the PI3K/AKT/mTOR and ER signaling pathway, lncRNAs may act on even more pathways that play into tamoxifen resistance, including the Wnt/β-catenin pathway. Liu and collaborators reported that UCA1 overexpression has a pro-survival and tamoxifen-resistance effect on BC cells, while inhibiting UCA1 activity restores sensitivity to tamoxifen and promotes apoptosis. Mechanistically, UCA1 activated the Wnt/β-catenin signaling pathway, which was evident by increases in β-catenin levels and Wnt reporter activity. Therefore, targeting the UCA1-Wnt/β-catenin axis may enhance the efficacy of tamoxifen in resistant BC settings [137]. c-Met is a receptor tyrosine kinase that, when bound with its ligand hepatocyte growth factor (HGF), will activate downstream pathways with a variety of cellular functions important for organ development and cancer progression [138]. Recent research has shown that activation of the HGF/c-Met pathway plays a major role in the maintenance of tamoxifen resistance in BC. Specifically, HOTAIR, along with EZH2, promotes HGF expression by repressing miR-141/200a and HGF/c-Met further activates NF-κB to establish positive feedback loop. The data presented indicate that the HOTAIR/EZH2-miR-141/200a-HGF/c-Met-NF-κB axis could be a potential target for overcoming tamoxifen resistance [139].

LncRNA-mediated epigenetic modulation

Epigenetic mechanisms within lncRNAs can alter tamoxifen resistance through chromatin structure, histone modification and transcriptional control. lncRNA can function as either a scaffolding molecule or a decoy for chromatin modifiers, resulting in altered expression of important genes in the ER signaling pathway for survival and apoptosis [80, 140]. Kim et al., examines the lncRNA HOTAIRM1 and its role in resistance to tamoxifen in ER⁺ BC. The data suggest that HOTAIRM1 is an oncogenic driver of tamoxifen resistance as it interacts with EZH2 to block polycomb repressive complex 2 (PRC2)-mediated repression. HOTAIRM1 also decreases levels of H3K27me3 at the HOXA1 promoter, which results in increased expression of HOXA1. Silencing either HOTAIRM1 or HOXA1 restores tamoxifen sensitivity in cell line models. Furthermore, the up-regulation of both HOTAIRM1 and HOXA1 in relapse patients indicates the genes may serve as biomarkers and/or therapeutic targets for endocrine resistant BC [13]. Alternate research demonstrates that lncRNA H19 is highly expressed in tamoxifen-resistant MCF-7 BC cells and promotes resistance by increasing the hypermethylation of the NAT1 promoter, leading to NAT1 downregulation. Upon H19 knockdown, NAT1 levels and sensitivity to tamoxifen response are reversed back to wildtype levels. Increased H19 and decreased NAT1 expression were also associated with poorer prognosis in human patient samples treated with tamoxifen, indicating H19 also promotes resistance through epigenetic silencing of NAT1 in clinical tumors and could be employed as a predictive biomarker and a therapeutic target for MCF-7 resistance [141] (Fig. 4) (Table 4). In addition to previous research, Wang et al. showed that lncRNA H19 can mediate tamoxifen resistance in ER⁺ BC in vivo and in vitro in models by modulating autophagy. Mechanistically, H19 inhibits SAHH causing accumulation of S-adenosylhomocysteine while also enhancing DNMT3B activity. DNMT3B binds and hypermethylates the Beclin1 promoter which suppresses Beclin1 expression and reduces autophagic function. Because this autophagy inhibition supports cellular survival under tamoxifen treatment, it confirms the importance of H19 in developing tamoxifen resistance. H19 overexpressed in tamoxifen-sensitive cells curated resistance which confirms a causal role. Therefore, H19/SAHH/DNMT3B/Beclin1 is a new epigenetic mechanism of tamoxifen resistance [142]. Yu et al. demonstrated that lncRNA AGPG drives endocrine resistance in ERα⁺ BC through enhancing E2F1 signaling. AGPG is elevated in ERα⁺ tumors through epigenetic reprogramming and enhancer activation. Importantly, AGPG expression is very high in luminal B tumors and is correlated with poor patient outcomes. In detail, AGPG has high affinity for PURα, a known repressor of E2F1, and disrupts the PURα-E2F1 complex, promoting de-repression and target gene activation of E2F1, which drives tumor progression and resistance to both tamoxifen and CDK4/6 inhibitors. Treatment with AGPG siRNA and tamoxifen together had significant growth inhibition of tumors in xenograft models [143].

Fig. 4.

lncRNAs regulate epithelial-mesenchymal transition, epigenetic reprogramming, and cell cycle control in breast cancer. This diagram depicts the complex modes of action of lncRNAs in breast cancer development. With regard to the epithelial-mesenchymal transition (EMT) pathway, lncRNAs such as ROR, SNHG6, DSCAM-AS1, LINC00894-002, and H19 drive EMT by inhibiting inhibitory tumor-suppressive miRNAs and activating downstream effects through ZEB1/2, Sox2/Oct4, EZH2, EPS8, and TGF-β2 to elicit tumor invasion, intravasation, extravasation, and ultimately colonization at distal sites in metastasis. In terms of epigenetic reprogramming, lncRNAs such as HOTAIRM1, H19, and AGPG are able to regulate DNA and histone methylation to modify chromatin remodeling and subsequently gene expression (HOXA1, EZH2, Beclin1) and transcription factor activity. In relation to the cell cycle and proliferation, lncRNA’s such as CYTOR, MAFG-AS1, LINP1 and SBF2-AS1, are able to modulate critical cell cycle regulators (SRF, CDK2, ER, YBX1) and PI3K/AKT signaling to drive cell cycle progression and proliferation amid tamoxifen selection pressure. These processes highlight the overarching nature of lncRNAs in the regulation of tumor aggressiveness and fidelity of therapy resistance. Illustration created with BioRender

LncRNA-mediated regulation of cellular fate

LncRNAs are now appreciated as crucial modulators of cell fate in BC via processes such as apoptosis, autophagy, EMT, proliferation, and cell cycle. They affect signaling through modulation of gene expression and stimulation of cellular responses to stress and therapy by interacting with a range of molecular partners to achieve tumor progression or therapeutic resistance [144] (Fig. 4). In this regard, lncRNA DILA1 is delineated as a discernible modulator of tamoxifen resistance in ER⁺ BC via direct interaction with Cyclin D1. A conspicuous mechanism by which DILA1, via biding the Thr286 residue, inhibits phosphorylation and degradation, thus lead to the pathological stabilization with Cyclin D1. This sustained build-up activates cell proliferation and neutralizes tamoxifen effect. Functionally silencing DILA1 in these resistant cells re-sensitize these cells, and inhibit the tumor in mice, revealing its feasibility as a novel strategy to restore endocrine responsiveness through Cyclin D1 destabilization [145]. In tamoxifen-resistant BC, lncRNA SOX2OT is significantly downregulated and its expression also negatively correlates with patient survival in both luminal A and luminal B subtypes. Upon dox-induced SOX2OT overexpression, proliferation, migration, and invasion assays also were performed. Surprisingly, SOX2OT overexpression did not stimulate SOX2 expression implying a SOX2-independent mechanism. These data implicate SOX2OT as a meaningful regulator of tumor aggressiveness in tamoxifen resistant BC and potential actor in cellular reprogramming associated with resistance mechanisms [146]. Other observations regarding lncRNA H19 and tamoxifen resistance, demonstrating that H19 is a key mediator of EMT, invasion, and migration in tamoxifen-resistant BC cells via the up-regulation Snail. Importantly, curcumin downregulates H19 with concomitant reversal of EMT markers (decreasing N-cadherin while increasing E-cadherin) and metastatic behaviors in resistant cells. These findings underscore the therapeutic promise of curcumin as an approach to H19-mediated tamoxifen-resistant BC [147]. Using autophagy regulation, lncRNA H19 was highlighted in another study as being implicated in tamoxifen resistance, as discussed in pervious section. H19 increased autophagy by epigenetically modulating Beclin1 expression through the SAHH/DNMT3B axis, leading to tamoxifen-resistance in BC cells [142]. LncRNA ROR is upregulated in BC tissues and contributes to tamoxifen resistance by inhibiting autophagy. Knockdown of ROR in BT474 cells increased autophagy markers (i.e., LC3 and Beclin 1), decreased multidrug resistance proteins (P-glycoprotein, GST-π), and decreased cell proliferation, invasion, and migration. These results suggest that lncRNA ROR promotes tamoxifen resistance by inhibiting autophagy and that inhibition of lncRNA ROR restores drug sensitivity [148] (Fig. 3). Moreover, recent studies have shown that lncRNA OIP5-AS1 is significantly upregulated in advanced stages of oral squamous cell carcinoma (OSCC) compared to non-cancerous tissues. OIP5-AS1 promotes oncogenic pathways and may contribute to therapeutic resistance through acting as a molecular sponge for tumor-suppressive microRNAs. These findings highlight its potential as both a biomarker for early detection and a target for personalized treatment strategies, although further mechanistic studies are needed to fully elucidate its functional roles in OSCC progression [149]. Exosomes are critical mediators of intercellular interactions, transferring molecules such as lncRNAs that subsequently influence behavior of the target cell. In Xu et al., study, they showed that tamoxifen resistant LCC2 BC cells release exosomes that are disproportionately enriched for lncRNA UCA1. This case study showed that exosomes from resistant LCC2 cells treated with tamoxifen and isolated were transferred to tamoxifen sensitive MCF-7 cells. Untreated MCF-7 cells subsequently had increased viability assessed with tryphan blue, lower levels of cleaved caspase-3, and decreased apoptosis rates upon treatment with tamoxifen. Importantly, exosomes incubated with a competitive inhibitor that blocked UCA1 loading to exosomes did not confer resistance, demonstrating that UCA1 plays a critical role in conferring tamoxifen resistance through exosomal communication [150] (Table 4).

CircRNAs

CircRNAs are a distinct and endogenous non-coding RNAs with a closed-loop structure that are important for cancer biology and drug resistance. CircRNAs regulate expression for genes contributing to tamoxifen resistant BC as an output, by sponging miRNAs, binding with RBPs, and modifying transcription [93, 151]. In this manner, circRNAs can regulate tumor-suppressive miRNAs or stabilize oncogenic transcripts that control genes involved in estrogen receptor signaling, proliferation, apoptosis, and cell survival. CircRNAs enable the crosstalk between autophagy and apoptosis, promoting the adaptive response to tamoxifen-induced. The variability in expression in tamoxifen sensitive cells compared to the resistant cells point to a significant role for circRNAs in endocrine resistance. As circRNAs have tissue specific expression and are highly conserved, they are potentially promising biomarkers for responding to treatment and for novel targeting as therapeutics [94, 152]. A greater understanding of circRNAs role in tamoxifen resistance is necessary for identifying post-transcriptional resistance mechanisms, and directing RNA-based therapies.

CircRNA-mediated CeRNA networks

The ceRNA regulatory mechanism is in agreement with hsa_circ_0025202 being highly downregulated in tamoxifen-resistant BC. There have been two separate studies looking at the role of hsa_circ_0025202 in the development of tamoxifen resistance in hormone receptor-positive BC. In the first study, RNA-seq of MCF-7 tamoxifen resistant cells showed marked downregulation of hsa_circ_0025202. Its re-overexpression led to inhibition of proliferation, colony formation and migration, induction of apoptosis, and restoration of tamoxifen sensitivity via sponging of miR-182-5p to upregulate FOXO3a protein levels. Also, with regard to in vivo studies, re-overexpression of hsa_circ_0025202 did inhibit tumor growth and promote tamoxifen response [153]. In the second study, its knockdown increased IC50 to tamoxifen and this was due to those cells seeking to further enter the cell cycle and promote invasion, migration, and inhibit apoptosis. In terms of mechanisms, it sponges miR-197-3p to then upregulate HIPK3, a known tumor-suppressive kinase. The researchers had proven the hsa_circ_0025202/miR-197-3p/HIPK3 axis was in control of tamoxifen resistance beyond just cellular experiments to even xenograft experiments [154]. In contrast, some circRNAs have an oncogenic effect; for example, emerging evidence highlights circTRIM28 is an important modulator of aggressiveness and treatment response in BC. Enhanced circTRIM28 expression, along with the enhances expression of HMGA2 and the decreased levels of miR-409-3p, was found in BC. Silencing circTRIM28 in functional assays decreased malignant actions like cell proliferation and tumorigenicity, as well as enable deficient apoptotic responses to tamoxifen. In this case, circTRIM28 would work as a ceRNA for miR-409-3p, effectively de-repressing HMGA2, which is a downstream effector of oncogenic signaling [155]. Liang et al. emphasize the critical role of hsa_circ_0097922 in BC tamoxifen resistance. This particular circRNA and downstream target ACTN4 are both found to be upregulated, and miR-876-3p is downregulated in resistant cells. Mechanistically, hsa_circ_0097922 acts as a sponge for miR-876-3p, which promotes malignancy and drug resistance by increasing ACTN4. Knockdown of hsa_circ_0097922 itself inhibits tumor growth, restores sensitivity against tamoxifen in vitro and in vivo, suggesting a potential therapeutic axis via miR-876-3p/ACTN4 regulation [156]. Higher levels of hsa-circVIM expression in hormone receptor-positive BC accelerate tumor progression and tamoxifen resistance through its target miR-1294. Silencing hsa-circVIM resulted in decreased cell viability and migration, increased apoptosis, and also restored sensitivity to tamoxifen. Thus, hsa-circVIM/miR-1294 signaling axis is critical in endocrine resistance and may act as a target for therapy and prognostication [157]. Preliminary data indicates that circ_UBE2D2 is significantly upregulated in tamoxifen-resistant BC tissue and cells, and is also secreted as part of exosomes from these resistant cells. Moreover, these exosomes transfer circ_UBE2D2 to sensitive cells, where it sponges miR-200a-3p and enhances cell viability, metastasis, and changes ERα expression. Therefore, targeting circ_UBE2D2 might be a promising way to combat tamoxifen resistance and make tamoxifen more effective as a treatment modality [158] (Table 5) (Fig. 5).

Table 5.

Overview of circrnas mediating Tamoxifen resistance in breast cancer

circRNA	Expression in BC	ceRNA-dependent / Independent	Molecular Mechanism / Pathway	Functional Outcome	Ref
hsa_circ_0025202	DOWN	ceRNA-dependent	Sponging miR-182-5p/FOXO3a, miR-197-3p/HIPK3	↑Tamoxifen sensitivity, ↓Proliferation, ↑Apoptosis	[153, 154]
circTRIM28	UP	ceRNA-dependent	Sponging miR-409-3p/HMGA2	↑Tamoxifen resistance, ↑Proliferation, ↓Apoptosis	[155]
hsa_circ_0097922	UP	ceRNA-dependent	Sponging miR-876-3p/ACTN4	↑Tamoxifen resistance, ↑Invasion, ↑Tumor growth	[156]
hsa-circVIM	UP	ceRNA-dependent	Sponging miR-1294	↑Tamoxifen resistance, ↑Migration, ↓Apoptosis	[157]
circ_UBE2D2	UP	ceRNA-dependent	Exosomal transfer, sponging miR-200a-3p/ERα	↑Tamoxifen resistance, ↑Metastasis, ↑Viability	[158]
circRNA-SFMBT2	UP	ceRNA-independent	Protein binding, ERα stabilization	↑Tamoxifen resistance, ↑Proliferation, ↑ER signaling	[159]
circFOXK2	UP	ceRNA-independent	mRNA stabilization, protein recruitment	↑Tamoxifen resistance, ↑Proliferation, ↑Cell cycle	[162]
circMET	UP	ceRNA-dependent	Sponging miR-204-5p/AHR	↑Tamoxifen resistance, ↑Proliferation, ↑Survival	[165]

Fig. 5.

circRNA-mediated ceRNA networks and their role in tamoxifen resistance in breast cancer. Diagrammatic overview of tumor suppressive and oncogenic circRNAs investigating breast cancer progression and tamoxifen resistance. Tumor suppressive circRNAs (green panel) (e.g., circ_0025202) with regulatory miRNAs (miR-182-5p, miR-197-3p) regulated FOXO3a and HIPK3. Oncogenic circRNAs (pink panel), circTRIM28, circCDK1, circ_0097922, circVIM, and circUBE2D2 which function as competing endogenous RNAs (ceRNA) sponged certain miRNAs (miR-409-3p, miR-489-3p, miR-876-3p, miR-1294 and miR-200a-3p) and stimulated HMGA2, CDK1, ACTN4, malignant progression, and changes to ERα. Other circRNAs such as circMET, circFOXX2 and circRNA-SFMBT2 also stimulated tamoxifen resistance (right panel) by regulating signaling through the upregulation of AHR signaling using miR-204-5p, stabilization of CCND1 mRNA via binding to ELAVL1 protein, and stabilization of ERα protein which allowed continued activity of the oestrogen receptor (ER) and CCND1–CDK4/6–pRB–E2F axis. Illustration created with BioRender

Impact of circrnas on cellular signaling

As an important regulator of ER⁺ BC, circRNA-SFMBT2 is highly expressed in ER⁺ BC cells and is associated with bigger tumor size and worse prognosis. High levels of circRNA-SFMBT2 stabilize the ERα protein by directly binding to its AF2 and DBD domains while RNF181 is recruited to the AF1 domain [159]. Using these interactions, circRNA-SFMBT2 alters both K48- and K63-linked ubiquitination of the ER protein that sustains a prolonged active state of ER signaling and downstream target genes. Functionally, circRNA-SFMBT2 drives cell growth and tamoxifen resistance in vitro and in vivo, establishing its important contribution to endocrine resistance, as a result of altered ER signaling [159]. CircTP63 is upregulated in estrogen receptor‑positive breast cancer and promotes malignant behaviors through the miR‑873‑3p/FOXM1 axis. By sponging miR‑873‑3p, circTP63 prevents repression of the transcription factor FOXM1, enhancing proliferation, invasion, and migration. Silencing circTP63 reduces FOXM1 levels and tumor aggressiveness, highlighting its role in ER⁺ breast cancer progression and its potential as a therapeutic target. This example illustrates that circRNAs can drive cancer progression not only via miRNA sponging but also by modulating oncogenic transcriptional programs [160].

Recent evidence has identified hsa_circ_0062522 as a circRNA that contributes to both tumor progression and tamoxifen resistance in ER⁺ breast cancer. In clinical samples and ER⁺ cell lines, hsa_circ_0062522 expression was significantly upregulated, while its predicted target miR‑3163 was downregulated, suggesting a regulatory interaction. Functional assays demonstrated that overexpression of hsa_circ_0062522 enhanced cell viability and migration, inhibited apoptosis, and increased resistance to tamoxifen, whereas knockdown of hsa_circ_0062522 produced the opposite effects [161]. Dual luciferase reporter and correlation analyses confirmed that hsa_circ_0062522 modulates ER⁺ breast cancer cell behavior and tamoxifen responsiveness by regulating miR‑3163 levels. In vivo xenograft models further corroborated these findings, showing that silencing hsa_circ_0062522 reduced tumor growth and restored tamoxifen sensitivity, highlighting its potential role as both a biomarker and therapeutic target in tamoxifen resistance [161]. In ER⁺ BC, circFOXK2 was determined to be significantly upregulated and was associated with poor prognosis. In detail, circFOXK2 directly interacts with CCND1 mRNA, recruits ELAVL1 and stabilizes CCND1 RNA and expression of protein, leading to activation of the CCND1–CDK4/6–pRB–E2F axis, G1/S transition and tamoxifen resistance. In addition to the evidence, they saw that circFOXK2 knockdown inhibited E2F1 target genes from RNA-seq studies and rescue experiments, the authors demonstrate a circFOXK2 role in ERα-related transcriptional programs. Targeting circFOXK2 inhibited tumor growth and restored tamoxifen sensitivity, representing a key role for circFOXK2 in modulating endocrine resistance through cell cycle and ERα signaling pathways [162]. Moreover, circCDK1 is another circRNA in which its knockdown increased tamoxifen sensitivity by decreasing the IC50 and inhibiting proliferation while increasing apoptosis. CircCDK1 acts as a miR-489-3p sponge, relieving the inhibitory effects on its host gene CDK1. When CDK1 is restored or miR-489-3p is inhibited, circCDK1 knockdown effects are reversed demonstrating that the circCDK1/miR-489-3p/CDK1 axis is a major regulator of tamoxifen resistance and tumor growth in vitro and in vivo [163]. Interestingly, recent work has shown that therapeutic targeting of circTNK2 can reverse tamoxifen resistance and enhance antitumor immunity in ER‑positive breast cancer. In tamoxifen‑resistant models, nanoparticles delivering circTNK2‑specific inhibitors restored drug sensitivity and simultaneously increased natural killer (NK) cell‑mediated immune responses, indicating a dual benefit of this approach. These findings suggest that circTNK2 contributes to endocrine resistance not only through intrinsic tumor cell mechanisms but also by shaping the tumor immune microenvironment, and that nanoparticle‑mediated circRNA targeting may represent a promising therapeutic strategy to improve both endocrine responsiveness and antitumor immunity [164]. Finally, based on Liu et al. study, circMET is over-expressed in tamoxifen resistance cells and drives proliferation and survival through the miR-204-5p/AHR signaling axis. CircMET acts as a molecular sponge to inhibit miR-204-5p, which results in the increased expression of AHR. Reduced circMET activity results in the reactivation of miR-204-5p and downregulation of AHR, which restores sensitivity to tamoxifen [165]. In this regard, Table 5 summarizes circRNAs implicated in tamoxifen resistance and their associated molecular mechanisms in breast cancer.

Conclusion and future perspectives

Tamoxifen resistance in the treatment of ER⁺ BC is a major obstacle in ensuring the long-term efficacy of endocrine therapy. While classic mechanisms including mutations in ER, altered signaling pathways, and the evasion of apoptosis in tumor cells have been well-studied in their contributions to resistance, there is now growing evidence of the important role of ncRNAs in modulating resistance. ncRNAs, including miRNAs, lncRNAs, and circRNAs, play important roles in different biological processes, such as estrogen signaling, autophagy, metabolic reprogramming, and drug efflux, mostly through regulatory networks (like ceRNA interactions) and exosomal transfer. Targeting dysregulated ncRNAs or their molecular partners offers potential to re-sensitize ER⁺ BC to tamoxifen and improve patient outcomes. However, translating these findings into clinically viable therapies faces significant challenges, including efficient and tumor-specific delivery, off-target effects, RNA instability, immune responses, and tumor heterogeneity. Overcoming these barriers will require the development of stable and targeted RNA delivery systems, strategies to enhance specificity and minimize unintended effects, and preclinical studies that account for dynamic tumor resistance mechanisms. Emerging technologies provide opportunities to advance this field. AI-driven multi-omics integration and spatial transcriptomics can facilitate the identification of key ncRNA–mRNA–protein networks, while integrative analyses of miRNA–lncRNA–circRNA cross-regulation can reveal mechanisms underlying chromatin remodeling, DNA methylation, and transcriptional plasticity [95]. These approaches may improve understanding of therapy resistance and tumor microenvironment modulation, enabling more precise patient stratification and personalized interventions. By combining ncRNA-targeted strategies with conventional therapies, and leveraging multi-omics and AI-based modeling, there is potential to accelerate translation from experimental findings to clinically effective interventions, ultimately improving outcomes for patients with tamoxifen-resistant ER⁺ breast cancer.

Author contributions

Conceptualization: M.K.J, R.R, and B.H, Writing-Original Draft: M.K.J, I.A, M.G, M.H.A, and R.E.A, Writing—Review & Editing : M.R.K and I.A, Visualization: M.R.K, Supervision: B.H, All authors reviewed the manuscript and prepared for submission.

Funding

None.

Data availability

No datasets were generated or analysed during the current study.

Declarations

Ethics approval and consent to participate

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Not applicable.

Competing interests

The authors declare no competing interests.