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Table of Contents
Year : 2020  |  Volume : 3  |  Issue : 1  |  Page : 144-145

Authors' reply to Chadha et al., Sambath et al., and Sharma et al.

1 Department of Medical Oncology, Indira Gandhi Institute of Medical Sciences, Patna, Bihar, India
2 Molecular Biology and Cytogenetics, Oncquest Laboratory, New Delhi, India

Date of Submission14-Jan-2020
Date of Decision16-Jan-2020
Date of Acceptance21-Jan-2020
Date of Web Publication24-Feb-2020

Correspondence Address:
Avinash Pandey
Department of Medical Oncology, Indira Gandhi Institute of Medical Sciences, Patna, Bihar
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/CRST.CRST_25_20

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How to cite this article:
Pandey A, Dutt S, Singh A, Kumar A, Singh S. Authors' reply to Chadha et al., Sambath et al., and Sharma et al. Cancer Res Stat Treat 2020;3:144-5

How to cite this URL:
Pandey A, Dutt S, Singh A, Kumar A, Singh S. Authors' reply to Chadha et al., Sambath et al., and Sharma et al. Cancer Res Stat Treat [serial online] 2020 [cited 2021 Aug 5];3:144-5. Available from: https://www.crstonline.com/text.asp?2020/3/1/144/279103

Chadha et al.[1] have cautioned about the importance of the pre-analytical factors – immediate separation and storage of plasma or use of special tubes to stabilize nucleated cells to minimize the contamination of genomic DNA. In our study, we have used PAXgene Blood cfDNA Tube (BD) for the collection and transportation of the samples. The stabilization reagent in the PAXgene Blood cfDNA Tube prevents blood coagulation, lysis of red blood cells, and apoptosis of white blood cells. The stabilization reagent in the tube is free of cross-linking or cross-linker releasing substances and hence does not chemically modify cfDNA. In a study published in 2017, Warton et al. evaluated the PAXgene Blood cfDNA System for the use of cell-free circulating DNA studies in plasma. They found that the total yield of cfDNA remained stabilized in the PAXgene tubes even when the samples were stored for 7 days at room temperature.[2]

For the collection of plasma, we follow the double centrifugation protocol which increases the yield of high-quality plasma.[3] This approach of pre-analytical and processing protocol in our laboratory has resulted in a significantly low percentage (~3%) of unsatisfactory results for circulating cell free DNA for digital droplet Polymerase Chain Reaction (ddPCR) for EGFR testing (unpublished data).

We concur that in the EGFR mutation-negative liquid biopsy cohort, tumor biopsy should be considered to confirm mutation status, as the sensitivity of liquid biopsy is 70%. Our study only had patients for whom tissue-based biopsy was not feasible upfront. At data lock time point with a median follow-up of 14 months, in the EGFR-negative cohort, 3/5 patients who had refused biopsy, 4/5 in whom biopsy was not feasible, and 4/5 in whom tissue was inadequate had succumbed due to early disease progression. Hence, we could not perform invasive tissue biopsies at later time points or at progression for the above cohort.[4]

Sambath et al.[5] have pointed out correctly that our study was retrospective and had exclusively nonsmokers, as we wanted to enrich the yield of mutation-positive liquid biopsies. Among the nonsmoking, biopsy-ineligible, female-preponderant population, our liquid biopsy EGFR mutation positivity was 39%.[4] This may vary if tested prospectively in unselected wider population of metastatic non-small cell lung cancer patients including smokers. We did not pursue patients who progressed on the first-line therapy in either EGFR mutation-negative or mutation-positive cohort, with invasive tissue biopsy or liquid biopsy to detect emerging drug-sensitive targetable mutations, including T790M.

Sharma et al.[6] have commented that the ideal source of liquid biopsy (ctDNA, CTCs, exosomes, platelets, and microRNA) has not been ascertained yet. We would like to point out that laboratory science and clinical pathology are in a state of dynamic growth, and it is true that many different analytes from the plasma samples are being evaluated as a source of tumor DNA. However, to date, ctDNA is considered to be the best analyte as liquid biopsy for EGFR testing. Studies have shown that ctDNA is a feasible sample type for real-world EGFR mutation testing if robust and sensitive DNA extraction and mutation analysis methodologies are employed.[7],[8]

They have further pointed out that there is still a lack of clarity about the method to be used to detect biomarkers in the liquid biopsy [digital droplet Polymerase Chain Reaction (ddPCR), Next Generation Sequencing (NGS) and Reverse Transcriptase Polymerase Chain Reaction (RTPCR)]. Multiple elegant studies have been published that the analytical performance of the ddPCR, with a detection limit of 0.05%–0.1%, is in concordance with deep-sequencing NGS and RT PCR and has high specificity and sensitivity for detecting EGFR mutations.[9],[10]

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Conflicts of interest

There are no conflicts of interest.

  References Top

Chadha K, Vinarkar S, Chheda P. EGFR mutation in non-small cell lung cancer by liquid biopsy when solid not feasible. Cancer Res Stat Treat 2020;3:140-1.  Back to cited text no. 1
  [Full text]  
Warton K, Yuwono NL, Cowley MJ, McCabe MJ, So A, Ford CE. Evaluation of Streck BCT and PAXgene stabilised blood collection tubes for cell-free circulating DNA studies in plasma. Mol Diagn Ther 2017;21:563-70.  Back to cited text no. 2
Risberg B, Tsui DW, Biggs H, Ruiz-Valdepenas Martin de Almagro A, Dawson SJ, Hodgkin C, et al. Effects of collection and processing procedures on plasma circulating cell-free DNA from cancer patients. J Mol Diagn 2018;20:883-92.  Back to cited text no. 3
Pandey A, Dutt S, Singh A, Kumar A, Singh S. Outcomes with liquid biopsy to determine the EGFR mutation status in poor performance status, biopsy-ineligible, advanced NSCLC patients. Cancer Res Stat Treat 2019;2:197.  Back to cited text no. 4
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Sambath J, Deb B, Kumar P. A commentary on outcomes with liquid biopsy to determine the epidermal growth factor receptor mutation status in poor performance status, biopsy-ineligible, advanced non-small cell lung cancer patients. Cancer Res Stat Treat 2020;3:141-2.  Back to cited text no. 5
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Sharma M, Joga S, Batra U. Liquid biopsy in non-small cell lung cancer: Ready for prime time? Cancer Res Stat Treat 2020;3:142-3.  Back to cited text no. 6
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Douillard JY, Ostoros G, Cobo M, Ciuleanu T, McCormack R, Webster A, et al. First-line gefitinib in Caucasian EGFR mutation-positive NSCLC patients: A phase-IV, open-label, single-arm study. Br J Cancer 2014;110:55-62.  Back to cited text no. 7
Choughule A, D'Souza H. Liquid biopsy in lung cancer-hope or hype? Cancer Res Stat Treat 2019;2:221-3.  Back to cited text no. 8
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Guo QM, Wang L, Yu WJ, Qiao LH, Zhao MN, Hu XM, et al. Detection of plasma EGFR mutations in NSCLC patients with a validated ddPCR lung cfDNA assay. J Cancer 2019;10:4341-9.  Back to cited text no. 9
Thress KS, Brant R, Carr TH, Dearden S, Jenkins S, Brown H, et al. EGFR mutation detection in ctDNA from NSCLC patient plasma: A cross-platform comparison of leading technologies to support the clinical development of AZD9291. Lung Cancer 2015;90:509-15.  Back to cited text no. 10


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