FC 12-01INTER-LABORATORY AGREEMENT OF THE FAM19A4/miR124-2 METHYLATION TEST – A VALID-SCREEN (H2020) SUB-STUDY

16. Methylation
A. Floore 1, B. Hesselink 1, J. Bonde 2, M. Poljak 3, K. Cuschieri 4, U. Petry 5, M. Del Pino 6, M. Bleeker 7, S. De Sanjose 8, P. Snijders 7, C. Meijer 7, D. Heideman 7.
1Self-screen B.V. (Netherlands), 2Department of Pathology, Copenhagen University Hospital, Hvidovre, Denmark (Netherlands), 3University of Ljubljana, Ljubljana, Slovenia (Netherlands), 4Scottish Human Papillomavirus Reference Laboratory, Edinburgh, Scotland, United Kingdom (Netherlands), 5Department of Gynaecology and Obstetrics, Klinikum Wolfsburg, Wolfsburg (Netherlands), 6University of Barcelona, faculty of medicine, institut clinic of Gynecology, Barcelona, Spain (Netherlands), 7Department of Pathology, VU University Medical Center, Amsterdam, The Netherlands (Netherlands), 8Catalan Institute of Oncology, Barcelona, Spain (Netherlands)

Background / Objectives

Background - The FAM19A4/miR124-2 methylation test (QIAsure Methylation Test, QIAGEN) is a novel assay, designed by Self-screen, that can be used for the triage testing of HPV-positive women or women with ASC-US. The test is a quantitative methylation-specific PCR that detects hypermethylation of the genes FAM19A4 and hsa-mir124-2 in cervical scrapes and self-collected samples. Within the framework of the VALID-SCREEN (H2020) project, the performance of the FAM19A4/miR124-2 methylation test will be validated in different, well characterised clinical cohorts from different European countries.

Objective - Determine the inter-laboratory agreement of the FAM19A4/miR124-2 methylation test on HPV-positive cervical scrapes collected in different European screening cohorts.


Methods

In total 695 HPV-positive cervical scrapes from five different European screening settings were included, i.e. Slovenia (SL, n=97), Spain (SP, n=239), Scotland (SC, n=96), Germany (G, n=159), and Denmark (D, n=104). Cervical scrapes had been collected in PreservCyt (SP, SL, SC, and G) or Surepath (D) and DNA extraction was performed according to local procedures. The samples were tested locally and sent to the reference lab for retesting. Both test and reference lab performed separately bisulfite-conversion followed by FAM19A4/miR124-2 methylation test (QIAsure Methylation Test) according to manufacturers’ instructions. Testing at both sites was performed blinded to the results of the other lab, and compared afterwards.


Results

Overall inter-laboratory agreement was 90.5% (629/695; 95%CI:88-92) with even higher agreement values in women with CIN3+ (i.e. 96.7%; 29/30; 95%CI:80-100). Agreement values for the five screening settings ranged from 83.9% to 96.8%. Overall kappa value was 0.77 (range laboratories: 0.64-0.91) indicating good inter-laboratory agreement. Discordant test results related to samples of women without clinical relevant disease having methylation values around the clinical cut-off of the assay. 


Conclusion

The inter-laboratory agreement of the FAM19A4/miR124-2 methylation test (QIAsure Methylation Test) was consistently high for the different European screening settings. Thus, the FAM19A4/miR124-2 methylation test is a good and reliable molecular triage assay suited for full molecular screening (i.e. HPV-DNA testing in combination with QIAsure Methylation Test). 


References

This work was supported by the SME Instrument in the Horizon 2020 Work Programme of the European Commission (Valid-screen 666800).