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Page 10 of 20 Singh et al. Cancer Drug Resist. 2025;8:56

with ddPCR [102] . High-sensitivity droplet dPCR provides sensitive absolute quantitation of circRNAs as a
complementary method for potentially longitudinal monitoring and/or resistance contributions .
[103]
The combination of circRNA-specific back-splice junction detection with deconvolution-based methods
improves specificity, which would enable the identification of cancer type- or drug resistance-associated
circRNAs at low abundance. Deconvoluted sequencing in combination with longitudinal RNA-seq also
facilitates early detection of therapeutic resistance [101] . Future integration of deconvoluted sequencing with
artificial intelligence (AI) and single-cell RNA-seq libraries is expected to further improve sensitivity and
predictive power, and to support the use of bulk RNA-seq as a non-invasive monitoring tool [101-104] . As
RNA-seq becomes more affordable, global profiling of circRNAs may be achieved through the use of
rRNA-depleted or RNase R-treated libraries and bioinformatics tools such as CIRCexplorer2, find_circ and
CIRI2 [105] . Exosome-derived circRNAs also display tumor specificity, as drug-resistant tumor cells tend to
preferentially package them, which could provide insight into the persistence and localization of drug
resistance during therapy [106] . The integration of bioinformatics platforms and databases (circBase,
circRNADb, and CircInteractome) will facilitate circRNA annotation, differential expression analysis, and
functional characterization, while incorporating pathway and network analysis platforms will help elucidate
the mechanisms by which circRNAs contribute to cancer drug resistance [Figure 3] .
[107]

The integration of these tools enables the development of dynamic circRNA-based biomarker panels that can
provide real-time guidance for therapeutic decisions . Although these approaches are still evolving,
[95]
challenges remain in standardizing sample handling, accounting for pre-analytical variability, and
differentiating tumor-derived circRNAs from those originating from non-malignant sources . Nevertheless,
[16]
the convergence of effective detection methods, sophisticated analytical methods, and key clinical
investigations is positioning circRNA profiling as a key part of future precision oncology . Looking ahead,
[96]
liquid biopsies leveraging circRNAs to monitor therapeutic resistance are expected to increasingly support
personalized, adaptive cancer treatment plans . A comparison of RNA detection methods is presented in
[2]
Table 5.

CLINICAL TRIALS ON LIQUID BIOPSY-BASED DETECTION OF CIRCULATING CIRCRNAS FOR
TRACKING DRUG RESISTANCE IN CANCER
Various preclinical and translational studies have demonstrated that circRNAs such as circFOXO3,
circHIPK3, and F-circEA are involved in drug resistance pathways, including the evasion of apoptosis,
regulation of autophagy, and drug efflux [Table 6] [105] . Moreover, trials involving liquid biopsies have
concentrated on ctDNA, CTCs, or miRNAs to monitor treatment response or early resistance in cancers
such as breast, lung, and CRC [106] . For example, ctDNA-based clinical trials, including SERENA-6 by
AstraZeneca and the NHS London pilot, have demonstrated the utility of early mutation detection in guiding
therapy switches [107] . In contrast, circRNAs have not yet been integrated into routine clinical workflows .
[52]
Efforts have been made in observational studies and small cohort studies to study circRNAs in biofluids
(especially in exosomes) as indicators of drug resistance in breast, prostate, and NSCLCs [111] . Importantly,
these earliest studies provide proof-of-concept evidence that circRNA profiles can capture the dynamic
evolution of a tumor’s adaptation to therapy . Given the advances in sequencing and computational tools
[24]
available for back-splice junction recognition, detecting circRNAs in clinical samples is technically
feasible [112] . To translate these findings into clinical practice, dedicated trials are needed to evaluate the
sensitivity, specificity, and predictive value of circRNA panels in monitoring drug resistance [113] . Future
clinical investigations should focus on validating circRNA signatures in longitudinal patient samples,
combining ctDNA and circRNA assays, and gradually integrating circRNAs into companion diagnostic
workflows. Conducting such trials would represent an important milestone in personalized oncology,

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