MP46-02: Refinement of the Serum miR-371a-3p Test for the Detection of Chemotherapy-naive Minimal Residual Testicular Cancer
Sunday, May 15, 2022
1:00 PM – 2:15 PM
Location: Room 222
John T Lafin, Armon Amini, Bendu Konneh, Anna Savelyeva, Dallas, TX, Jin Piao, Michelle Nuno, Monrovia, CA, Cinzia G Scarpini, Cambridge, United Kingdom, Jeffrey M Howard, Thomas Gerald, Dallas, TX, Nicholas Coleman, Matthew J Murray, Cambridge, United Kingdom, A Lindsay Frazier, Boston, MA, James F Amatruda, Los Angeles, CA, Aditya Bagrodia*, La Jolla, CA
Introduction: While critical for patient care, existing serum tumor markers exhibit middling performance for the detection of testicular germ cell tumors (GCTs). Serum miRNAs, particularly miR-371a-3p, are reported to have excellent performance characteristics in the pre-orchiectomy setting. However, the ability of miR-371a-3p to detect occult disease is understudied. We previously reported in a small cohort of patients that miR-371a-3p outperforms conventional markers in the context of chemotherapy-naïve minimal residual disease. Here, we further refine the assay and expand upon that experience.
Methods: We examined over 250 distinct runs to compare performance between classification based on Cq, normalized Cq (?Cq), or relative quantification (RQ). We prospectively collected clinical information and pre-surgical serum samples from chemotherapy-naïve GCT patients undergoing primary retroperitoneal lymph node dissection (RPLND) at our institution. Patients were classified as “Control” (pure teratoma or no GCT) or “GCT”. RNA was extracted from these serum samples, and miR-371a-3p (target) and miR-30b-5p (reference gene) were detected by qPCR. Performance was assessed by calculation of sensitivity, specificity, and area under the ROC curve (AUC).
Results: Performance did not change appreciably when thresholding based on Cq, ?Cq, or RQ. We therefore decided to move forward with analysis considering only Cq. We found that any given control sample will return a spurious positive result approximately 25% of the time, and estimated an indeterminate range of Cq 28-35. Repeating any indeterminate sample once improved overall performance. We then compared this updated process to a simple binary threshold in a cohort of patients with occult disease. 32 patients were enrolled in the study (15 Control, 17 GCT). 12 samples were indeterminate on first run, and 8 remained indeterminate on a second run, all of which harbored viable GCT or teratoma. Inclusion of a second run prevented 3 false positives, improving specificity from 86% to 100% and yielding a sensitivity of 92% and AUC of 0.96 (95% CI: 0.89-1).
Conclusions: The use of Cq instead of RQ to classify results for the miR-371a-3p test would help to avoid unnecessary sample runs without injuring assay performance. In the context of chemotherapy-naïve minimal residual testicular cancer, consideration of an indeterminate range may improve the performance of the miR-371a-3p test.
Source of Funding: This work was supported by funding from the Cancer Prevention Institute of Texas, St. Baldrick’s Consortium, the Rally Foundation, the Malignant Germ Cell International Consortium, and the Dedman Family Scholarship in Clinical Care.