Detection of GNAQ/11 Mutations in Uveal Melanoma Patients’ FFPE DNA with Droplet Digital PCR

  Objective  To quantitatively detect GNAQ/11 mutations in uveal melanoma (UM) by droplet digital PCR (ddPCR).

  Methods   Formaldehyde-fixed paraffin-embedded (FFPE) tumor samples were taken from 78 UM patients with enucleation in West China Hospital between 2009 and 2015. None of the patients received radiotherapy or chemotherapy before enucleation. A retrospective study was conducted to detect GNAQ/11 mutation in UM by ddPCR. To compare the consistency of the results of the two detection methods, DNA sequencing was performed on the target gene by Sanger sequencing. 78 patients with UM were studied retrospectively. GNAQ/11 mutations in uveal melanoma was detected by ddPCR. The consistency of the results of the two detection methods was analyzed.

  Results  GNAQ/11 mutations frequency was 91.9%. The consistency test between Sanger sequencing and ddPCR of GNAQ/11 mutations in 74 patients with UM was conducted. Kappa coefficient=0.436, P=0.001. The error rate of Sanger sequencing results was significantly higher in the heterogeneous group than in the homogeneous group (12/37 vs. 3/16, P=0.53), but the difference was not statistically significant.

  Conclusion  The results of ddPCR and Sanger sequencing showed good consistency, and the mutation ratio of GNAQ/11 in UM was significantly different. GNAQ/11 mutation frequency in UM patients detected by ddPCR was close to the reported frequency. It is more recommended to use ddPCR with high sensitivity to detect gene mutations in samples of tumor tissue DNA derived from FFPE. Sanger sequencing is prone to false negative results.