Monitoring of somatic mutations in circulating cell-free DNA by digital PCR and next-generation sequencing during afatinib treatment in patients with lung adenocarcinoma positive for EGFR activating mutations.
Analysis of circulating cell-free DNA (cfDNA) is under intensive investigation for its potential to identify tumor somatic mutations. We have now explored the usefulness of such liquid biopsy testing with both the digital polymerase chain reaction (dPCR) and next-generation sequencing (NGS) during treatment of patients with the epidermal growth factor receptor (EGFR) inhibitor afatinib.



Eligible patients had advanced lung adenocarcinoma with EGFR activating mutations and were treated with afatinib. Plasma samples were collected before and during (4 and 24 weeks) afatinib treatment as well as at disease progression. Tumor and plasma DNA were analyzed by dPCR and NGS.



Thirty-five patients were enrolled. The objective response rate and median progression-free survival (PFS) were 77.1% and 13.8 months, respectively. Tumor and plasma DNA were available for 32 patients. dPCR and NGS detected EGFR activating mutations in 81.3% and 71.9% of baseline cfDNA samples, respectively. In 19 patients treated with afatinib for ≥24 weeks, the number of EGFR mutant alleles detected in cfDNA by dPCR declined rapidly and markedly after treatment onset, becoming undetectable or detectable at only a low copy number (<10 copies per milliliter) at 4 weeks. Median PFS was slightly longer for patients with undetectable EGFR mutant alleles in cfDNA at 4 weeks than for those in whom such alleles were detectable (14.3 versus 10.0 months). A total of 45 somatic mutations was identified in baseline tumor DNA, and 30 (66.7%) of these mutations were identified in cfDNA by NGS. Allele frequency for somatic mutations in cfDNA determined by NGS changed concordantly during afatinib treatment with the number of EGFR mutant alleles determined by dPCR.



Monitoring of cfDNA by dPCR is informative for prediction of afatinib efficacy, whereas that by NGS is reliable and has the potential to identify mechanisms of treatment resistance.
Department of Comprehensive Clinical Oncology, Faculty of Medical Sciences, Kyushu University, Fukuoka.|Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka.
E
E
Iwama
Department of Genome Biology, Kinki University Faculty of Medicine, Osaka-Sayama.
K
K
Sakai
Division of Respirology, Neurology, and Rheumatology, Department of Internal Medicine, Kurume University School of Medicine, Kurume.
K
K
Azuma
Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka.
T
T
Harada
Department of Thoracic Oncology, NHO Shikoku Cancer Center, Matsuyama.
D
D
Harada
Department of Thoracic Oncology, National Kyushu Cancer Center, Fukuoka.
K
K
Nosaki
Center for Innovative Clinical Medicine, Okayama University Hospital, Okayama.
K
K
Hotta
Department of Thoracic Medical Oncology, Cancer Institute Hospital of Japanese Foundation for Cancer Research, Tokyo.
F
F
Ohyanagi
Department of Thoracic Oncology, Kansai Medical University Hospital, Hirakata.
T
T
Kurata
Department of Respiratory Medicine, Miyagi Cancer Center, Natori.
T
T
Fukuhara
Third Department of Internal Medicine, Wakayama Medical University, Wakayama.
H
H
Akamatsu
Department of Thoracic Oncology, National Cancer Center Hospital East, Kashiwa.
K
K
Goto
Clinical Research Support Center Kyushu, Fukuoka.
T
T
Shimose
Department of Research and Development of Next Generation Medicine, Kyushu University, Fukuoka, Japan.
J
J
Kishimoto
Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka.
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Y
Nakanishi
Department of Genome Biology, Kinki University Faculty of Medicine, Osaka-Sayama.
K
K
Nishio
Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka.
I
I
Okamoto
D000230:Adenocarcinoma / Q000097:blood / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics*; D000077192:Adenocarcinoma of Lung; D000077716:Afatinib; D000074141:Circulating Tumor DNA / Q000097:blood / Q000235:genetics*; D066246:ErbB Receptors / Q000235:genetics* / Q000378:metabolism; D005260:Female; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D006801:Humans; D000073890:Liquid Biopsy; D008175:Lung Neoplasms / Q000097:blood / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics*; D008297:Male; D009154:Mutation; D009367:Neoplasm Staging; D016133:Polymerase Chain Reaction / Q000379:methods; D011446:Prospective Studies; D011799:Quinazolines / Q000009:adverse effects / Q000627:therapeutic use*
D000074141:Circulating Tumor DNA; D011799:Quinazolines; D000077716:Afatinib; C512478:EGFR protein, human; D066246:ErbB Receptors