CIRCULAR RNAS PROFILING FOR THE IDENTIFICATION OF NOVEL PROGNOSTIC BIOMARKERS AND THERAPEUTIC TARGETS IN BREAST CANCER

Abstract

Breast cancer poses a significant clinical challenge due to its complex molecular landscape, emphsaizing the need for improved prognostic and therapeutic strategies. In this study, we investigated the expression profiles, prognostic significance, and therapeutic relevance of circular RNAs (circRNAs) in a cohort of breast cancer patients from the MENA region. Analysis of the MENA cohort identified fifty circRNAs associated with unfavorable relapse-free survival (RFS). Given the aggressive nature of triple-negative breast cancer (TNBC), we performed differential expression analysis within the MENA TNBC subgroup, revealing 344 upregulated circRNAs. To validate these findings, we analyzed a publicly available dataset of TNBC patients, revealing fifty-four circRNAs commonly upregulated in the MENA TNBC cases which were likewise elevated in the independent dataset. RNase R resistance assays and NGS further confirmed the expression of sixteen circRNAs associated with worse RFS and thirteen TNBC-upregulated circRNAs in TNBC models. Divergent primers and Sanger sequencing confirmed the backsplice junctions of three shortlisted unfavorable prognostic (hsa_circ_0001522, hsa_circ_0001278, and hsa_circ_0001801) and three TNBC-upregulated circRNAs (hsa_circ_0000268, hsa_circ_0006990, and hsa_circ_0001681). Functionally, siRNA-mediated knockdown of these circRNAs suppressed cell proliferation, colony formation, three-dimensional tumor growth, and migration in TNBC models. Mechanistic investigations revealed that depletion of the three unfavorable prognostic circRNAs modulated the expression of hsa-miR-4458, hsa-miR-145-5p, and hsa-miR-760, along with downstream target mRNAs including CCND1, ROBO4, and RHOB. Additionally, the TNBC-upregulated circRNAs regulated hsa-miR-537, hsa-miR-616-3p, hsa-miR-6783-3p, hsa-miR-137-3p, hsa-miR-29c-3p, and hsa-miR-3611, leading to the regulation of key oncogenic targets such as EGFR, TGFBR1, CDK6, and ID1. Analysis using the CircAtlas 3.0 database predicted circRNAs interactions with several RNA-binding proteins, including AGO2, CPSF7, TARDBP, UPF1, LIN28B, CPSF6, CSTF2T, HNRNPC, and NUDT21, suggesting roles in post-transcriptional regulation. In summary, we identified and functionally characterized circRNAs that promote TNBC growth and migration, representing promising biomarkers and therapeutic targets, offering new avenues for improving TNBC prognosis and treatment.

Date of Award	2025
Original language	American English
Awarding Institution	HBKU College of Health & Life Sciences