Cell-free DNA From Pleural Effusion Samples: Is It Right for Molecular Testing in Lung Adenocarcinoma?

Abstract

Pathogenic molecular features gained specific significance in therapeutic decisions in lung carcinoma in the past decade. Initial and follow up genetic testing requres appropriate amounts and quality of tumor derived DNA, but tumor sampling, especially for disease monitoring is generally limited. Further to the peripheral blood (PB), samples from pleural fluid, accumulating in diverse lung processes might serve as an alternative source for cell-free DNA (cfDNA) for genetic profiling. In our study, cfDNA isolated from the pleural effusion and from the PB, and genomic DNA (gDNA) obtained from tissue/cellular samples were analyzed and compared from altogether 65 patients with pulmonary disease, including 36 lung adenocarcinomas. The quantity of effusion cfDNA yield appeared to be significantly higher compared to that from simultaneously collected PB plasma (23.2 vs. 4.8 ng/μl, p < 0.05). Gene mutations could be safely demonstrated from the effusion cfDNA fraction obtained from adenocarcinoma patients, 3/36 EGFR, 9/36 KRAS and 1/36 BRAF gene variants were detected. In this series, 9/13 samples showed an effusion+/plasma-mutational status, while only 1/13 samples presented with the opposite findings (effusion-/plasma+). gDNA analysis from sediment cell blocks from the identical effusion sample was surprisingly ineffective for lung adenocarcinoma profiling due to the low DNA yield. In conclusion, the cell free supernatant of pleural effusions appears to concentrate cancer derived cfDNA and seems to be particularly suitable for serial genotyping of pulmonary adenocarcinoma.

Transcending Blood—Opportunities for Alternate Liquid Biopsies in Oncology

“…There is some debate as to whether similar conclusions based on size analysis can be drawn regarding non-blood cfDNA. cfDNA with a similar size profile to blood has been reported in various fluids [ 22 , 32 , 79 , 80 ], and tumour enrichment in short cfDNA fragments has been demonstrated in CSF and urine [ 81 , 82 ]. However, distinct size profiles have also been observed.…”

“…Improved ctDNA detection rates have been observed in cfDNA from non-blood sources compared to blood [ 29 ], with the exception of saliva [ 35 ] and sputum [ 17 , 38 ]. This has been repeatedly demonstrated in pleural effusions secondary to lung cancer, with an increased sensitivity to EGFR mutations in malignant pleural effusions over plasma, as well as over cell pellets from the effusions [ 31 , 32 , 33 , 36 ]. Furthermore, in two independent cohorts of lung adenocarcinoma patients, higher ctDNA abundance was seen in other body fluids, including ascites, pericardial effusions, and CSF, compared to blood plasma [ 39 , 40 ].…”

“…In two case studies of recurrent medulloblastoma, where driver mutations were found to differ between the initial and recurrent disease, cfDNA from CSF was found to accurately detect the changes [ 30 ]. Similarly, cfDNA from pleural effusions has been found to correspond to matched tumour tissue [ 31 , 32 ]. Among 29 patients with lung cancer, 93% of the driver mutations identified in tumour tissue were similarly detected in the cfDNA from pleural effusions [ 33 ].…”

Circulating Tumor DNA Liquid Biopsy in Patients with Non-Small Cell Lung Cancer

“…В качестве высоко репрезентативных источников цоДНК также можно отметить плевральный/перикардиальный выпот и цереброспинальную жидкость [35,63]. Несмотря на определенные преимущества данных подходов (высокая концентрация цоДНК и практически полное отсутствие примесей неопухолевой цоДНК соответственно), они являются узкоспециализированными и данные виды биоматериала используются значительно реже плазмы крови в связи с высокой инвазивностью получения биологического образца.…”

EGFR mutation testing from pleural effusions of non-small cell lung cancer patients at the institute for oncology and radiology of Serbia

“…Cell samples have been shown to be a good source of DNA for molecular diagnostic analyzes in lung cancer, such as determining mutations in the EGFR, KRAS or BRAF genes. However, cell sediment testing had an average sensitivity of around 60% (from 40 to 87%), revealing a distinct limitation of this approach [11] . cfDNA in the supernatant of PE has been proved to be superior to cell sediment in detection of genetic variants.…”