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Digital Arrayを用いたNSCLCにおける血漿からの変異型EGFRのコピー数解析

 本研究では、新しい超高感度検出定量の技術であるdigital PCRを用いて肺がん患者の血液からEGFR遺伝子変異を検出しコピー数の定量を行いました。患者から切除した原発巣および採取した血漿から抽出したDNAを用いて検出・比較を行いました。その結果、原発巣がシークエンス法で変異陽性だった症例の92%において、digital PCRによって血漿から変異を検出することができました。また、従来のシークエンス法では変異が検出できなかった原発巣サンプルについても、digital PCRによって変異を検出することができました。さらに興味深いことに、EGFR-TKI(イレッサ、タルセバ)の投与前後の血漿中の変異型EGFRのコピー数をdigital PCRで測定した結果、EGFR-TKIによる治療効果に応じて変異型EGFRのコピー数が変動したことから、血漿中EGFR遺伝子変異のコピー数が治療効果の指標として利用できることが示唆されました。今後、非侵襲的なサンプルである血液サンプルからdigital PCRにより高感度にEGFR遺伝子変異を検出・コピー数定量することにより、より簡便にかつ高精度に治療の奏功性や予後の予測ができるようになることが期待されます。
(各文献の解釈につきましては、弊社は一切の責任を負いません)

Single-Molecule Detection of Epidermal Growth Factor Receptor Mutations in Plasma by Microfluidics Digital PCR in Non-Small Cell Lung Cancer Patients
Yung TK, Chan KC, Mok TS, Tong J, To KF, Lo YM.
Clinical Cancer Research.  2009;15:2076-2084.

【Abstract】


Pupose:
We aim to develop a digital PCR-based method for the quantitative detection of the two common epidermal growth factor receptor (EGFR) mutations (in-frame deletion at exon 19 and L858R at exon 21) in the plasma and tumor tissues of patients suffering from non-small cell lung cancers. These two mutations account for >85% of clinically important EGFR mutations associated with responsiveness to tyrosine kinase inhibitors.
Experimental design: DNA samples were analyzed using a microfluidics system that simultaneously performed 9,180 PCRs at nanoliter scale. A single-mutant DNA molecule in a clinical specimen could be detected and the quantities of mutant and wild-type sequences were precisely determined.
Results: Exon 19 deletion and L858R mutation were detectable in 6 (17%) and 9 (26%) of 35 pretreatment plasma samples, respectively. When compared with the sequencing results of the tumor samples, the sensitivity and specificity of plasma EGFR mutation analysis were 92% and 100%, respectively. The plasma concentration of the mutant sequences correlated well with the clinical response. Decreased concentration was observed in all patients with partial or complete clinical remission, whereas persistence of mutation was observed in a patient with cancer progression. In one patient, tyrosine kinase inhibitor was stopped after an initial response and the tumor-associated EGFR mutation reemerged 4 weeks after stopping treatment.
Conclusion: The sensitive detection and accurate quantification of low abundance EGFR mutations in tumor tissues and plasma by microfluidics digital PCR would be useful for predicting treatment response, monitoring disease progression and early detection of treatment failure associated with acquired drug resistance.

PMID 19276259
 

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Dube Simant, Qin Jian, Ramakrishnan Ramesh
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Fan H. Christina, Quake Stephen R.
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Development of Potential Pharmacodynamic and Diagnostic Markers for Anti-IFN-I± Monoclonal Antibody Trials in Systemic Lupus Erythematosus
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Circulating Tumor Cells Are Transcriptionally Similar to the Primary Tumor in a Murine Prostate Model
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Enhanced Generation of Induced Pluripotent Stem Cells from a Subpopulation of Human Fibroblasts
Byrne James A., Nguyen Ha N., Reijo Pera Renee A.
PLoS ONE.  2009;4:e7118.

Transplantation directs oocyte maturation from embryonic stem cells and provides a therapeutic strategy for female infertility
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Pharmacogenetic Pathway Analysis for Determination of Sunitinib-Induced Toxicity
van Erp NP, Eechoute K, van der Veldt AA, Haanen JB, Reyners AK, Mathijssen RH, Boven E, van der Straaten T, Baak-Pablo RF, Wessels JA, Guchelaar HJ, Gelderblom H
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Evaluation of current methods used to analyze the expression profiles of ATP-binding cassette transporters yields an improved drug-discovery database

Orina JN, Calcagno AM, Wu CP, Varma S, Shih J, Lin M, Eichler G, Weinstein JN, Pommier Y, Ambudkar SV, Gottesman MM, Gillet JP
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Association of reactive oxygen species levels and radioresistance in cancer stem cells
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