Serum miR371 in testicular germ cell cancer before and after orchiectomy, assessed by digital-droplet PCR in a prospective study

Abstract

MicroRNA-371a-3p (miR371) has been suggested as a sensitive biomarker in testicular germ cell cancer (TGCC). We aimed to compare miR371 with the classical biomarkers α-fetoprotein (AFP) and β-human chorionic gonadotropin (hCGβ). Overall, 180 patients were prospectively enrolled in the study, with serum samples collected before and after orchiectomy. We compared the use of digital droplet PCR (RT-ddPCR) with the quantitative PCR used by others for detection of miR371. The novel RT-ddPCR protocol showed high performance in detection of miR371 in serum samples. In the study cohort, miR371 was measured using RT-ddPCR. MiR371 detected CS1 of the seminoma and the non-seminoma sub-types with a sensitivity of 87% and 89%, respectively. The total sensitivity was 89%. After orchiectomy, miR371 levels declined in 154 of 159 TGCC cases. The ratio of miR371 pre- and post-orchiectomy was 20.5 in CS1 compared to 6.5 in systemic disease. AFP and hCGβ had sensitivities of 52% and 51% in the non-seminomas. MiR371 is a sensitive marker that performs better than the classical markers in all sub-types and clinical stages. Especially for the seminomas CS1, the high sensitivity of miR371 in detecting TGCC cells may have clinical implications.

Testicular Neoplasms: Primary Tumour Size Is Closely Interrelated with Histology, Clinical Staging, and Tumour Marker Expression Rates—A Comprehensive Statistical Analysis

“…The authors found positive correlations between primary tumour size and M371 serum levels in both seminomas and nonseminomas, and quite a number of patients with subcentimeter primaries had measurable M371 levels. Notably, among the small tumours, a larger number of nonseminomas compared to seminomas had measurable M371 levels [ 59 ]. Overall, the sensitivity of M371 is better for nonseminoma than for seminoma regarding small testicular masses.…”

DigiMir Test: Establishing a Novel Pipeline for MiR-371a Quantification Using Droplet Digital PCR in Liquid Biopsies From Testicular Germ Cell Tumor Patients

“…DdPCR provides absolute quantification, obviating the need for normalization to housekeeping microRNAs and facilitating setting of cutoffs of positivity, and is less influenced by inhibitory substances, due to the partitioning process. However, its application for microRNA quantification is still largely unexplored [with few studies available (37)(38)(39)(40)(41)(42)(43)(44)], and only one very recent study attempted to apply such protocol to hsa-miR-371a-3p determination in TGCTs (45).…”

MicroRNAs in Differentiation of Embryoid Bodies and the Teratoma Subtype of Testicular Cancer

“…These five microRNAs were generally weakly expressed and detected in few patients even with the pre-amplification step included in the RT-qPCR method. miR-371a-3p, which has been previously reported as a circulating biomarker (31,42,(62)(63)(64)(65)(66)(67)(68) was found to be weakly expressed in serum from one patient with primary TGCT described as pure teratoma, but was not expressed in any of the serum samples collected prior to pcRPLND from patients with pure teratoma in the post-chemotherapy RPLND specimen. In tissue, our finding of a gradient in expression of miR-371a-3p from tumour, through normal testicular tissue to non-testicular CANCER GENOMICS & PROTEOMICS 19: 178-193 (2022) tissue is in concordance with the report from Belge et al (69).…”

“…As a control group for serum analyses, we used serum from 30 healthy, age-matched male blood donors (42). We also downloaded datasets with global circulating microRNA expression data from healthy individuals in order to filter out microRNAs that are expected to be highly expressed in serum from healthy individuals (43,44).…”