Analisis Mutasi Gen EGFR dan KRAS Berbasis PCR-HRM (High Resolution Melting) dan RFLP (Restriction Fragment Length Polymorphisme) terhadap Cairan Pleura Pasien Kanker Paru yang Disimpan pada Kertas Saring

Abstract

Lung cancer is the most frequent cause of cancer-related mortality worldwide, including in Indonesia. Today, a medical treatment for lung cancer patient, such as targeted therapies, has been developed to decrease the mortality. The methods need a rapid and a robust molecular characterisation technique to diagnose any gene mutations such as the KRAS and the EGFR. We developed a PCR-HRM and a PCR-RFLP for screening the EGFR and the KRAS mutations in pleural effusion of lung cancer patients. Sensitivity studies using the dilution assay revealed that the PCR-HRM was able to detect KRAS and EGFR mutations as little as 12.5% of mutant DNA in wild type DNA, while the PCR-RPLF was able to detect as little as 3.125 % of DNA mutant in wild type DNA. We analysed 63 of pleural effusion from lung cancer patient that were collected in filter paper. From our sample, we found that 1.6% (1/63) of them had the KRAS mutation; meanwhile 15.9 % (10/63) of them had the EGFR mutations. We also found that the EGFR mutation was more prevalent in men 21.43% (6/28) compared to women 15.3% (4/26). These results showed that the HRM and the RFLP are useful methods to detect the KRAS and the EGFR mutations in the clinical samples of lung cancer. The filter paper was also very potential material to be used in molecular analysis of lung cancer, especially for collecting and transferring the pleural effusion samples.