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Clinical Evaluation of miR-135b-5p and miR-21-5p Expression on Self-Collected Cervical Swab Samples for Triaging
Farhin Sultana1, Ananya De2, Sreeya Bose3
1Department of Oncogene Regulation, Chittaranjan National Cancer Institute, Kolkata, West Bengal, India.
Objective:
This study aimed to evaluate the triage performance of microRNA (miRNA) biomarkers on self-collected, high-risk human papillomavirus (HR-HPV)-positive cervical swabs to discern cervical intraepithelial neoplasia II-III or high-grade squamous intraepithelial lesions (HSIL) in Indian women for precise diagnosis and treatment prioritization.
Methods:
Through in silico analysis, miR-135b-5p and miR-21-5p were identified as upregulated and amplified miRNAs in cervical cancer (CaCx). Quantitative real-time PCR was used to assess miR-135b-5p and miR-21-5p and their targeting, LIM domain-containing protein 1 (LIMD1) and von Hippel-Lindau (VHL) messenger RNA (mRNA) expression, in a discovery cohort (N = 42; normal cervical epithelium [N = 9], CIN [N = 12], and CaCx [N = 21] tissue samples). Thereafter, miRNA expression was validated in an independent set of self-collected cervical swabs (N = 243; HR-HPV-positive [asymptomatic (N = 52), low-grade squamous intraepithelial lesions (LSIL) (N = 50), and HSIL (N = 52), CaCx (N = 35)] and HPV-negative normal [N = 54]). Receiver operating characteristic (ROC) curve analysis was performed to evaluate the diagnostic performance of these two candidate miRNAs.
Results:
In discovery-phase samples, miR-135b-5p showed progressive upregulation from CIN (p = 0.056) to CaCx (p = 0.012), while miR-21-5p showed significant overexpression at CIN (p = 0.011) and CaCx (p = 0.005) compared to normal tissue. Concurrently, significant downregulation of LIMD1/VHL mRNA expression was observed in the same CaCx samples. In the validation cohort, miR-135b-5p and miR-21-5p were significantly overexpressed in HSIL samples compared to lower-grade lesions (≤LSIL), with a corresponding downexpression of LIMD1/VHL. ROC curve analysis demonstrated high sensitivity and specificity for miR-21-5p and miR-135b-5p-both individually and in combination-for distinguishing HR-HPV + HSIL from LSIL, asymptomatic, or HPV-negative normal samples.
Conclusion:
miR-135b-5p and miR-21-5p constitute a feasible triage biomarker on noninvasive, self-collected cervical swab samples for early, precise diagnosis of women with HR-HPV-positive HSIL, requiring treatment prioritization in low-resource settings.