High Accuracy Salivary miRNA Signature for Non-Invasive Diagnosis of Superficial Peritoneal Endometriosis: A Large Prospective Multicenter Study

INTRODUCTION: Endometriosis affects approximately 10% of women of reproductive age and is associated with substantial morbidity, including chronic pelvic pain, dysmenorrhea, and infertility. Superficial peritoneal endometriosis (SUP) is the most common endometriosis phenotype yet remains the most difficult to diagnose, as lesions are frequently small and poorly detected by transvaginal ultrasound or pelvic MRI. This leads to diagnostic uncertainty and delay in symptomatic patients. Laparoscopy with histologic confirmation is invasive, operator dependent and resource-intensive. These limitations highlight the need for accurate, non-invasive diagnostic tools. The salivary miRNA signature for endometriosis offers advantages in terms of accessibility, stability, and reproducibility. OBJECTIVE: To evaluate the diagnostic accuracy of the salivary miRNA signature for endometriosis in women with histologically confirmed SUP. METHODS: We present finding of a subgroup analysis of the prospective, multicenter ENDOmiRNA Saliva Test study (NCT05244668). Women aged 18–43 years with symptoms suggestive of endometriosis were enrolled. All analysed patients underwent transvaginal ultrasound and/or pelvic MRI, followed by laparoscopy with histological confirmation. Saliva samples were collected at enrolment or prior to surgery, irrespective of menstrual cycle phase or ongoing medical treatment. RNA extraction, miRNA sequencing, and quantification followed standardized protocols. Endometriosis status according to the salivary miRNA signature for endometriosis (109-miRNA panel) was determined blinded to imaging and surgical findings. Diagnostic performance was assessed in patients presenting with superficial peritoneal endometriosis, either isolated or associated with other phenotypes classified in the patients with SUP group and in patients with isolated SUP. RESULTS: Among 556 patients with an analysable sample, 340 had histologically confirmed endometriosis, and 216 served as controls. In the patients with SUP group (n=332), the salivary miRNA signature for endometriosis demonstrated an accuracy of 96.0% (95% CI 93.8–97.5%), sensitivity of 97.0% (95% CI 95.2–98.2%), specificity of 94.4% (95% CI 91.7–96.3%), PPV of 0.964 (95% CI 0.946–0.976), and NPV of 0.953 (95% CI 0.925–0.972).Within the subgroup of patients with histologically proven isolated SUP (n=81), diagnostic performance remained high, with an accuracy of 95.3% (95% CI 92.1–96.8%), sensitivity of 96.3% (95% CI 90.4–99.0%), specificity of 94.9% (95% CI 92.7–95.9%), PPV of 0.876 (95% CI 0.823–0.901), and NPV of 0.986 (95% CI 0.963–0.996). CONCLUSIONS: This large, prospective, multicenter analysis demonstrates that the salivary miRNA signature for endometriosis achieves high diagnostic accuracy in patients with superficial peritoneal endometriosis, including the particularly challenging isolated SUP phenotype. The test markedly outperforms conventional imaging in sensitivity while preserving high specificity, and its consistently high negative predictive value supports its use as a robust rule-out tool in clinical practice. Collectively, these performance characteristics translate into meaningful clinical utility. As a non-invasive, standardized, and easily deployable assay, the salivary miRNA signature constitutes a valuable addition to the diagnostic armamentarium for endometriosis, with the potential to reduce diagnostic delay, refine clinical decision-making in imaging-negative patients, and ultimately improve patient care in endometriosis.Table 1Table 2