Breast cancer is the most frequently diagnosed cancer among women globally, and the second leading cause of cancer-related mortality [1]. According to statistics from the World Health Organization (WHO), approximately 2.3 million new cases and nearly 685,000 deaths occurred due to breast cancer in 2020 alone, representing a growing burden in both developed and low-income nations [2]. With the advent of earlier screening and more effective treatment options, many patients are now benefiting, although these services are not yet accessible to everyone. Moreover, breast cancer treatment remains particularly challenging due to the scarcity of effective interventions and the poor clinical prognosis for metastatic or treatment-resistant patients [3, 4].

Tissue biopsy is the gold standard method of oncological diagnosis since the twentieth century [5]. It is an invasive procedure that is associated with procedural risks, including bleeding, infection, and patient discomfort. Although tissue biopsy can capture only a spatially restricted snapshot of the tumor microenvironment, it may fail to represent the holistic landscape of intertumoral heterogeneity of the metastatic site of cancer [6]. Consequently, tracking the molecular pathways that drive tumor evolution, mutation, and resistance remains a significant clinical challenge due to spatial and temporal heterogeneity, though emerging technologies such as liquid biopsies and multi-region sequencing are beginning to address this limitation [7].

To address unmet needs, liquid biopsy offers a dynamic and minimally invasive window into the tumor’s evolving landscape, establishing it as one of the most compelling frontiers in translational oncology [8]. Moreover, in tumor diagnosis, liquid biopsy enables sampling and analysis of biological specimens, including blood, urine, saliva, cerebrospinal fluid, and pleural effusions, to detect and characterize tumor-derived components in circulation [9]. These tumor-associated components most commonly include cell-free circulating tumor DNA (ctDNA), circulating tumor cells (CTCs), small non-coding RNA molecules such as microRNAs (miRNAs), and extracellular vesicles (EVs), which reflect the characteristics of cancer cells (Figure 1) [10, 11]. Recently, significant advancements in electrochemical biosensors, microfluidic lab-on-chip devices, surface plasmon resonance systems, and nanomaterial-based optical platforms have improved sensitivity and selectivity in cancer detection and treatment prognosis [12–14]. In addition, these advances have positioned liquid biopsy as a viable component of clinical oncology practice in low-resource settings where access to sophisticated molecular diagnostics is currently unavailable [15].

The present review aims to synthesize the current state of the liquid biopsy technique as applied to breast cancer, with a specific focus on early detection and treatment prognosis. Followed by examining the conceptual framework of liquid biopsy, we will validate the translational utility of these approaches and conclude with an appraisal of the unresolved challenges consisting of technical, clinical, and ethical perspectives, which should be navigated for widespread clinical implementation.

The term “liquid biopsy” refers to a heterogeneous group of analytes and analytical methodologies that vary considerably in their clinical relevance, biological sources, and technical requirements for biomarker identification and diagnostic application. As a minimally invasive procedure, liquid biopsy unifies diverse analytes under a common umbrella to obtain tumor-relevant biological information from a non-solid specimen [16, 17]. It is based on the principle that tumors continuously release cells and molecular material into the bloodstream as they grow and evolve. These circulating components, derived from both primary and metastatic sites, provide a minimally invasive and real-time snapshot of tumor biology. Because their composition changes with disease progression and treatment, liquid biopsy offers a dynamic approach for monitoring cancer and therapeutic response.

The minimally invasive nature of liquid biopsy has expanded the contexts in which tumor molecular profiling can be performed, enabling serial sampling and broader clinical accessibility [18]. Traditional tissue biopsies, particularly in metastatic disease, often require image-guided procedures that involve procedural risks, patient discomfort, and logistical complexity. As a result, they are typically carried out only at diagnosis or when there is clear evidence of disease progression [19]. Consequently, the molecular information guiding clinical decision-making may be outdated and may not reflect the dynamic changes that occur as tumors evolve under therapeutic pressure [20]. In breast cancer, therapies such as aromatase inhibitors and CDK4/6 inhibitors often lead to the emergence of resistant subclones driven by newly acquired mutations [21]. Among these, alterations in the ESR1 gene, which encodes the estrogen receptor, are especially well recognized [22]. Notably, such resistance-associated mutations can frequently be detected in ctDNA weeks before radiographic progression is evident [23]. This temporal advantage highlights the potential of serial liquid biopsy monitoring to identify molecular signs of resistance at an early stage, enabling more timely therapeutic adjustments rather than waiting for clinical or imaging-based evidence of progression.

Despite its conceptual elegance, the clinical utility of liquid biopsy in early-stage disease is frequently compromised by a signal-to-noise deficit. In patients with localized, low-volume tumors, the stochastic presence of ctDNA and CTCs often falls below the analytical sensitivity of current next-generation sequencing (NGS)-based assays, as these tumor-derived analytes are heavily diluted within the systemic circulation [24–26]. In metastatic breast cancer, ctDNA and CTCs are often present at readily detectable concentrations. In early-stage disease, these analytes are so diluted within the systemic circulation that their detection demands far more sensitive analytical platforms [27–29]. Although analytical sensitivities have been sharpened by sophisticated techniques such as NGS and droplet digital PCR (ddPCR), the diagnosis of early-stage breast cancer remains hindered by frequent false-negative results. To bridge this gap, EV-derived multi-miRNA panels are being explored as promising diagnostic biomarkers for early-stage breast cancer, complementing other liquid biopsy analytes such as ctDNA [30]. For example, combining ctDNA mutation detection with miRNA expression profiling (e.g., miR-21 and miR-155 panels) and EV surface protein markers (e.g., EpCAM-positive EVs) has demonstrated improved sensitivity for early-stage breast cancer detection compared to any single analyte alone [30]. This multi-analyte approach is particularly valuable in subtypes such as triple-negative breast cancer (TNBC) and human epidermal growth factor receptor 2 (HER2)-positive breast cancer, which differ substantially in their molecular profiles and shedding patterns. TNBC tends to shed higher levels of ctDNA due to its aggressive proliferation and genomic instability, whereas HER2-positive tumors may be better detected through protein-based markers and EV profiling [31].

In clinical applications, liquid biopsy serves as a continuous molecular “navigation system” throughout the course of breast cancer management, complementing the gaps left by traditional imaging and invasive tissue sampling [31, 32]. By characterizing the tumor’s genetic profile from a simple blood draw, it enables earlier detection in difficult-to-image dense breast tissue and identifies actionable mutations, such as PIK3CA or ESR1, to guide personalized therapy [33, 34].

Beyond initial diagnosis, the real-time tracking of ctDNA levels allows clinicians to monitor treatment response months before a physical tumor appears on a scan [35]. Most significantly, detecting residual molecular disease after surgery provides a critical window for intervention, potentially preventing recurrence before it becomes clinically established. Figure 2 provides a comparative overview of liquid biopsy and tissue biopsy approaches.

It is important to emphasize that liquid biopsy should be viewed as a complement to, rather than a replacement for, conventional histopathology. Analysis of resected tumor tissue allows for a comprehensive assessment of tumor architecture, morphology, grade, invasive versus in situ components, lymphovascular invasion, receptor status, and spatial heterogeneity. These aspects remain a major advantage of tissue-based pathology and cannot currently be replaced by most liquid biopsy approaches. While liquid biopsy excels at capturing temporal molecular changes and monitoring treatment response, tissue biopsy remains indispensable for initial diagnosis, histological classification, and comprehensive tumor characterization. The integration of both modalities offers the most complete picture of tumor biology.

Liquid biopsy has emerged as an impactful approach in breast cancer diagnosis and real-time therapeutic monitoring [56]. This technique offers advantages over conventional cross-sectional imaging by enabling clinicians to track tumor evolution longitudinally, identify resistance mechanisms, assess early treatment response, and detect minimal residual disease at a stage when therapeutic intervention is most likely to be clinically impactful. Liquid biopsy has reduced reliance on surgical biopsies by analyzing ctDNA, CTCs, and cell-free RNA in the blood. The following discussion presents several significant studies demonstrating the clinical significance of liquid biopsy in breast cancer, with a critical appraisal of their strengths and limitations.

Liquid biopsy provides a minimally invasive approach for real-time molecular monitoring in breast cancer management. While conventional tissue biopsy offers a spatially and temporally controlled view of tumor biology, liquid biopsy provides a complementary and dynamic perspective on heterogeneous tumors by assessing tumor-derived analytes, including CTCs, ctDNA, miRNAs, and EVs.

Several clinical trials, including PADA-1 and SERENA-6, have demonstrated that ctDNA-guided therapeutic strategies can significantly improve progression-free survival in HR+/HER2-negative metastatic breast cancer. In early-stage and pre-invasive disease settings, where ctDNA-based methods have limited sensitivity, CTC-based approaches such as those evaluated in the STIC CTC METABREAST trial offer an alternative detection strategy. Nevertheless, no single biomarker is sufficient for precise diagnosis. Combinatorial approaches incorporating miRNA panels, EV profiling, and integrated multi-analyte strategies represent the most promising path forward for accurate disease assessment.

Several challenges remain before liquid biopsy can achieve widespread clinical adoption. Analytical sensitivity continues to limit detection in early-stage disease, and the lack of standardized pre-analytical and bioinformatic protocols leads to variability across studies and hinders regulatory acceptance. Ambiguous findings can raise ethical concerns regarding overdiagnosis and unnecessary interventions. Furthermore, unequal access to advanced diagnostic technologies threatens to exacerbate existing health disparities. Addressing these challenges will require investment in standardized protocols, robust clinical trial infrastructure, and efforts to make these technologies accessible to patients globally. Liquid biopsy should be viewed as a powerful complement to established diagnostic standards, and its integration into routine clinical practice will depend on resolving these technical, clinical, and ethical hurdles.