KR20160069070A - Method for Predicting Treatment Effects of EGFR Inhibitor in Glioblastoma - Google Patents
Method for Predicting Treatment Effects of EGFR Inhibitor in Glioblastoma Download PDFInfo
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Abstract
Description
본 발명은 교모세포종에서 표피 성장인자 수용체(EGFR) 억제제의 치료 효과를 예측하는 방법에 관한 것이다.
The present invention relates to a method for predicting the therapeutic effect of an epidermal growth factor receptor (EGFR) inhibitor in a glioblastoma.
신경교종(glioma)은 원발성 뇌종양(primary brain tumor)의 60%를 차지하는 종양으로 현재까지도 방사선 치료 외엔 특별한 치료법이 없는 악성 종양이다. 특히 가장 악성으로 분류되는 교모세포종(glioblastoma)은 다른 암과 비교하여 방사선 및 항암제 치료에 대한 저항성이 매우 높아 일단 진단되면 기대 생존기간이 1년에 불과하다. 또한, 뇌혈관 장벽이 있어 약물의 전달이 쉽지 않고, 상대적으로 뇌신경 생물학에 대한 이해의 부족 때문에 치료제 개발에서 소외된 분야이기도 하다.Glioma is a tumor that accounts for 60% of primary brain tumors and is a malignant tumor with no special treatment other than radiation therapy. In particular, glioblastoma, the most malignant, is highly resistant to radiation and chemotherapy compared to other cancers, and once diagnosed, the expected survival is only one year. In addition, it is not easy to deliver drugs due to the presence of a cerebrovascular barrier, and it is also a field that has been left out of the development of therapeutic drugs due to the lack of understanding of brain nerve biology.
최근 시스템 생물학(systems biology) 및 생물정보학 기법(bioinformatics tools)의 비약적 발전에 따라 교모세포종의 발생과 병증의 진행과 관련된 것으로 여겨지는 의미 있는 유전적 결함 및 유전자 발현 패턴들이 보고되고 있으나, 임상성과를 대표할 수 있는 분자 마커(molecular marker)들의 발굴 성과는 전무한 실정이며, 유전자 발현 분석이 전사체(trascriptom) 수준에 머물러 있어 실제 암 조직에서의 단백질 발현 정도를 대변하지 못하는 한계가 있다. Recent advances in systems biology and bioinformatics tools have led to reports of significant genetic defects and gene expression patterns that are believed to be related to the development of glioblastoma and the progression of disease, There are no molecular markers that can be represented, and there is a limitation that the gene expression analysis does not represent the degree of protein expression in actual cancer tissues because it remains at the level of trascriptomes.
이중 표피성장인자 수용체(Epidermal Growth Factor Receptor, EGFR)의 변이는 전체 교모세포종 환자의 약 57%에서 검색되는 가장 중요한 종양유발 인자로 알려져 있으나, EGFR 억제제에 대한 임상시험 결과 반응성이 낮았기 때문에, 치료반응 예측인자 개발을 통한 표적환자군 선별이 요구되고 있다. The EGFR mutation is known to be the most important tumorigenic factor in approximately 57% of patients with Glioma. However, because of the low reactivity of EGFR inhibitors in clinical trials, The selection of the target patient group is required through the development of the response predictor.
이에 본 발명자들은 교모세포종에서 EGFR 억제제의 치료 효과의 예측 마커로 이용할 수 표지자로서 EGFR 카피수(copy number)를 이용할 수 있음을 규명함으로써 본 발명을 완성하게 되었다.
Thus, the present inventors have completed the present invention by confirming that EGFR copy number can be used as a marker to be used as a predictive marker of the therapeutic effect of an EGFR inhibitor in a glioblastoma.
본 명세서 전체에 걸쳐 다수의 논문 및 특허문헌이 참조되고 그 인용이 표시되어 있다. 인용된 논문 및 특허문헌의 개시 내용은 그 전체로서 본 명세서에 참조로 삽입되어 본 발명이 속하는 기술 분야의 수준 및 본 발명의 내용이 보다 명확하게 설명된다.
Numerous papers and patent documents are referenced and cited throughout this specification. The disclosures of the cited papers and patent documents are incorporated herein by reference in their entirety to better understand the state of the art to which the present invention pertains and the content of the present invention.
본 발명자들은 차세대 유전체 분석법(New Generation Sequencing, NGS) 기반의 교모세포종 신선동결조직에서 채취한 DNA에서 EGFR 카피수를 측정하였을 때, EGFR 카피수에 따라 EGFR 억제제의 치료 효과에 대한 감수성이 유의하게 높음을 규명함으로써 본 발명을 완성하였다. The present inventors measured the EGFR copy number in DNA extracted from fresh frozen tissue of glioblastoma based on New Generation Sequencing (NGS), and found that the susceptibility to the therapeutic effect of EGFR inhibitor was significantly higher according to the EGFR copy number To complete the present invention.
따라서, 본 발명의 목적은 EGFR(epidermal growth factor receptor) 억제제에 대한 대상(subject)의 치료 효과의 유효성(effectiveness) 예측 방법을 제공하는 데 있다.
It is therefore an object of the present invention to provide a method for predicting the effectiveness of a therapeutic effect of a subject on an epidermal growth factor receptor (EGFR) inhibitor.
본 발명의 목적 및 이점은 하기의 발명의 상세한 설명 및 청구범위에 의해 보다 명확하게 된다.
The objects and advantages of the present invention will become more apparent from the following detailed description of the invention and claims.
본 발명의 일 양태에 따르면, 본 발명은 다음 단계를 포함하는 EGFR(epidermal growth factor receptor) 억제제에 대한 대상(subject)의 치료 효과의 유효성(effectiveness) 예측 방법을 제공한다:According to one aspect of the present invention, the present invention provides a method for predicting the effectiveness of a therapeutic effect of a subject on an epidermal growth factor receptor (EGFR) inhibitor comprising the steps of:
(a) 대상으로부터 분리한 생시료(biosample)에서 지놈 DNA를 추출하는 단계; (a) extracting genomic DNA from a biosample separated from a subject;
(b) 상기 추출된 지놈 DNA의 EGFR 유전자의 카피수(copy number)를 분석하는 단계; 및(b) analyzing the copy number of the EGFR gene of the extracted genome DNA; And
(c) 상기 분석된 EGFR 유전자의 카피수가 10 이상이면 상기 대상은 상기 EGFR 억제제에 대하여 유효한 치료 효과를 갖는 대상이고, EGFR 유전자의 카피수가 10 미만이면 EGFR 억제제에 대하여 유효하지 않은 치료 효과를 갖는 대상인 것으로 결정하는 단계.
(c) a subject having an effective therapeutic effect on the EGFR inhibitor when the number of copies of the EGFR gene is 10 or more, and a subject having an ineffective therapeutic effect on the EGFR inhibitor if the copy number of the EGFR gene is less than 10 ≪ / RTI >
본 발명자들은 차세대 유전체 분석법(New Generation Sequencing, NGS) 기반의 교모세포종 신선동결조직에서 채취한 DNA에서 EGFR 카피수를 측정하였을 때, EGFR 카피수에 따라 EGFR 억제제의 치료 효과에 대한 감수성이 유의하게 높음을 규명함으로써 본 발명을 완성하였다.
The present inventors measured the EGFR copy number in DNA extracted from fresh frozen tissue of glioblastoma based on New Generation Sequencing (NGS), and found that the susceptibility to the therapeutic effect of EGFR inhibitor was significantly higher according to the EGFR copy number To complete the present invention.
본 발명을 각 단계 별로 상세하게 설명하면 다음과 같다:The present invention will now be described in detail in the following order:
단계 (a): 지놈 DNA의 추출 Step (a): extraction of genomic DNA
본 발명에 따르면, 우선 대상(예컨대, 인간, 구체적으로 EGFR-관련 질환의 위험이 있거나 EGFR-관련 질환을 가지고 있는 인간)으로부터 분리한 생시료(biosample)에서 지놈 DNA를 추출한다. 본 발명에 적용될 수 있는 생시료는 체외로 분리된 동물의 생체시료로서, 조직, 세포, 혈액, 혈장 또는 혈청을 의미하며, 보다 구체적으로 조직 또는 세포, 보다 구체적으로 교모세포종의 종양세포 또는 종양줄기세포이다.In accordance with the present invention, genomic DNA is first extracted from a biosample isolated from a subject (e.g., a human, specifically a human at risk for an EGFR-related disorder or an EGFR-related disorder). The biological sample that can be applied to the present invention means a tissue sample, a cell, blood, plasma or serum as a biological sample of an animal isolated from the outside of the body, more specifically, a tissue or a cell, more specifically, a germ cell of a glioblastoma, It is a cell.
본 명세서에서 용어 “EGFR-관련 질환”은 구체적으로 암, 보다 구체적으로 뇌암, 보다 더 구체적으로 교모세포종을 의미한다.
As used herein, the term " EGFR-related disorder " specifically refers to cancer, more specifically brain cancer, more specifically glioblastoma.
단계 (b): EGFR 유전자의 카피수(copy number) 분석 Step (b): Analysis of copy number of EGFR gene
이어, 상기 추출된 지놈 DNA의 EGFR 유전자의 카피수(copy number)를 분석한다.Next, the copy number of the EGFR gene of the extracted genomic DNA is analyzed.
EGFR 유전자의 카피수 분석은 당업계에 공지된 다양한 방법에 의해 실시할 수 있다. 구체적으로, EGFR 유전자의 카피수 분석은 총지놈 서열분석법(whole genome sequencing, 참조: Bentley DR, Whole-genome re-sequencing. Curr. Opin. Genet. Dev. 16(6):545552(December 2006); Ahn SM, et al., . The first Korean genome sequence and analysis: Full genome sequencing for a socio-ethnic group. Genome Research 19(9):16229(2009)) 또는 타겟엑솜 서열분석법(targeted exome sequencing, 참조: Sarah B Ng et al., Exome sequencing identifies the cause of a mendelian disorder, Nature Genetics 42(1):3035(2010); Mertes F et al., Targeted enrichment of genomic DNA regions for next-generation sequencing, Brief Funct Genomics. 10 (6):374386(2011); Worthey EA et al., Making a definitive diagnosis: successful clinical application of whole exome sequencing in a child with intractable inflammatory bowel disease, Genet Med. 13(3):255262(Mar 2011))에 의해 수행된다.
The copy number of the EGFR gene can be analyzed by various methods known in the art. Specifically, the copy number of EGFR gene was analyzed by whole genome sequencing (Bentley DR, Whole-genome re-sequencing. Curr. Opin. Genet. Dev. 16 (6): 545552 (December 2006); Genome Research 19 (9): 16229 (2009)) or targeted exome sequencing (see, eg, Ahn SM, et al., The first Korean genome sequence and analysis: Full genome sequencing for a socio- Sarah B Ng et al., Exome sequencing identifies the cause of a mendelian disorder, Nature Genetics 42 (1): 3035 (2010); Mertes F et al., Targeted enrichment of genomic DNA regions for next-generation sequencing, Brief Funct Genomics 13 (3): 255262 (Mar. 2011); Worthey EA et al., "A definitive diagnosis: successful clinical application of whole-body sequencing in a child with intractable inflammatory bowel disease, Genet Med. )).
단계 (c): EGFR 억제제에 대한 치료 유효성 평가 Step (c): Evaluation of therapeutic efficacy against EGFR inhibitors
상기 분석된 EGFR 유전자의 카피수가 10 이상이면 상기 대상은 상기 EGFR 억제제에 대하여 유효한 치료 효과를 갖는 대상이고, EGFR 유전자의 카피수가 10 미만이면 EGFR 억제제에 대하여 유효하지 않은 치료 효과를 갖는 대상인 것으로 결정한다.It is determined that the subject has an effective therapeutic effect on the EGFR inhibitor when the analyzed copy number of the EGFR gene is 10 or more and the subject has an ineffective therapeutic effect on the EGFR inhibitor when the copy number of the EGFR gene is less than 10 .
본 명세서에서, EGFR 억제제는 당업계에 공지된 다양한 EGFR 억제제를 포함하며, 구체적으로 EGFR 억제제는 제피티닙(Gefitinib), 엘로티닙(Erlotinib) 또는 아파티닙(Afatinib)이다.
As used herein, an EGFR inhibitor comprises a variety of EGFR inhibitors known in the art. Specifically, the EGFR inhibitor is Gefitinib, Erlotinib, or Afatinib.
본 발명의 특징 및 이점을 요약하면 다음과 같다:The features and advantages of the present invention are summarized as follows:
(a) 본 발명은 EGFR(epidermal growth factor receptor) 억제제에 대한 대상(subject)의 치료 효과의 유효성(effectiveness) 예측 방법을 제공한다. (a) The present invention provides a method for predicting the effectiveness of the therapeutic effect of a subject on epidermal growth factor receptor (EGFR) inhibitors.
(b) 본 발명은 차세대 유전체 분석법(New Generation Sequencing, NGS) 기반의 방법을 통하여 대상에서 채취한 DNA에서 EGFR 카피수를 측정하였을 때, EGFR 카피수에 따라 EGFR 억제제의 치료 효과에 대한 감수성이 유의하게 높음을 규명함으로써, 본 발명은 EGFR(epidermal growth factor receptor) 억제제에 대한 대상(subject)의 치료 효과의 유효성(effectiveness) 예측방법으로서 유용하게 이용될 수 있다.
(b) The present invention is based on the observation that the sensitivity of the EGFR inhibitor to the therapeutic effect of the EGFR copy number is significant when the EGFR copy number is measured in the DNA collected from the subject through a method based on a New Generation Sequencing (NGS) , The present invention can be usefully used as a method for predicting the effectiveness of the therapeutic effect of a subject on an epidermal growth factor receptor (EGFR) inhibitor.
도 1은 교모세포종종양조직에서 DNA를 추출하여 NGS(New Generation Sequencing) 방법으로 총지놈(genome)의 엑솜 시퀀싱(whole exome sequencing)을 수행하는 실험의 모식도를 나타낸 도이다.
도 2는 NGS 방법을 활용하여 종양의 EGFR 카피수(copy number)를 측정한 실험의 모식도를 나타낸 도이다.
도 3은 각 환자의 종양에서 분리한 DNA에서 EGFR 카피수(copy number)와 EGFR 억제제의 치료 효과를 비교하여 양성 상관관계를 확인한 것을 그래프로 나타낸 도이다.
도 4는 EGFR 카피수(copy number)가 10 이상인 종양에서 EGFR 억제제들의 치료효과가 우수하게 측정된다는 것을 확인한 실험결과를 그래프를 나타낸 도이다.FIG. 1 is a schematic diagram of an experiment in which DNA is extracted from a glioblastoma tumor tissue and whole exome sequencing of the genome is performed by the NGS (New Generation Sequencing) method.
FIG. 2 is a schematic diagram of an experiment in which an EGFR copy number of a tumor is measured using the NGS method. FIG.
FIG. 3 is a graph showing positive correlation between EGFR copy number and therapeutic effect of EGFR inhibitor in DNA isolated from each patient's tumor.
FIG. 4 is a graph showing experimental results confirming that the therapeutic effect of EGFR inhibitors is excellent in a tumor having an EGFR copy number of 10 or more. FIG.
이하, 실시예를 통하여 본 발명을 더욱 상세히 설명하고자 한다. 이들 실시예는 오로지 본 발명을 보다 구체적으로 설명하기 위한 것으로, 본 발명의 요지에 따라 본 발명의 범위가 이들 실시예에 의해 제한되지 않는다는 것은 당업계에서 통상의 지식을 가진 자에 있어서 자명할 것이다.
Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are only for describing the present invention in more detail and that the scope of the present invention is not limited by these embodiments in accordance with the gist of the present invention .
실시예Example
실험 재료 및 실험 방법Materials and Experiments
1. 교모세포종 검체 획득 및 세포배양1. Glioblastoma specimen acquisition and cell culture
동의서 취득 후 교모세포종 검체를 적법한 절차에 따라 적제 후 연구목적으로 체취하였다. 약 5 X 5 X 5 mm의 검체를 액체질소를 이용하여 순간 동결하고 핵산을 추출하여 유전체 검사를 실시하였다. 일부 검체는 효소를 이용하여 분해(dissociation)한 후, 암세포를 분리하여 암줄기세포 배양용 배지(Neurobasal media with N2 and B27 supplements 0.5X each, and human recoombinant basic fibroblast growth factor and epidermal growth factor, 20 ng/ml)에서 배양하였다. 상기 실험 방법에 대한 모식도를 도 1에 나타내었다.
After obtaining the consent form, the glioblastoma specimens were collected according to the legal procedure for the study purpose after the application. Approximately 5 X 5 X 5 mm specimens were frozen for an instant using liquid nitrogen and nucleic acids were extracted and subjected to genomic tests. Some of the specimens were dissociated using enzymes, and then the cancer cells were separated and cultured in a medium for the culture of human stem cells (Neurobasal media with N2 and B27 supplements, 0.5X each, and human recom- binant basic fibroblast growth factor and epidermal growth factor, 20 ng / ml). A schematic diagram of the above experimental method is shown in Fig.
2. 유전자 분석2. Genetic analysis
추출된 핵산은 애질런트 슈어셀렉트 키트(Agilent Sureselect kit)를 이용하여 엑손 DNA 단편(exonic DNA fragment)을 수득한 후, 일루미나 하이시퀀싱2000(Illumina Hiseq2000)을 이용하여 시퀀싱을 수행하였다. 시퀀싱 리드 FASTQ 파일은 버로우-휠러 얼라이너(Burrows-Wheeler Aligner)를 이용하여 매핑(mapping)한 후 점 돌연변이(point mutation)의 발생 여부를 하였다. ngCGH 파이톤 패키지(python package) 프로그램을 이용하여 환자 매칭 정상 세포의 유전체와 비교 후, 종양의 특정 유전자 카피수(copy number)를 분석하였다. DNA 복제체(copy) R 패키지(package)를 이용하여 CBS 알고리즘을 기반으로 분획화(fractionation)한 후, 모든 엑손(exonic) DNA 분획물의 평균값과 비교하여 종양 유전자의 카피수를 계산하였다. 상기 실험 방법에 대한 모식도를 도 2에 나타내었다.
The extracted nucleic acid was subjected to sequencing using an Illuminous High Sequence 2000 (Illumina Hiseq 2000) after obtaining an exonic DNA fragment using an Agilent SureSelect kit. The sequencing lead FASTQ file was mapped using Burrows-Wheeler Aligner and then point mutation occurred. The ngCGH python package program was used to compare the genomes of patient-matched normal cells and to analyze the specific gene copy number of the tumor. DNA copies were fractionated based on the CBS algorithm using the R package and the number of copies of the oncogene was calculated by comparing to the average value of all exonic DNA fractions. A schematic diagram of the above experimental method is shown in Fig.
3. 세포 기반의 약물 반응성(drug sensitivity) 고속 대량 스크리닝(High-throughput Screening) 3. Cell-based drug sensitivity. High-throughput screening.
교모세포종 일차배양 세포는 384웰(well)에 웰당 500개 세포로 플레이팅(plating)을 수행하였다. 두 시간 후, 43종의 표적 항암제를 4배의 농도에서 7배 농도의 포인트로 야누스 자동화 워크스테이션(Janus Automated Workstation)을 이용하여 처리하였다. 상기 표적 항암제를 하기 표 1에 나타내었다.
The glioblastoma primary cultured cells were plated in 384 wells to 500 cells per well. Two hours later, 43 kinds of target anticancer drugs were treated with 4 times the concentration and 7 times the concentration point using the Janus Automated Workstation. The target anticancer agent is shown in Table 1 below.
약물을 처리한 지 6일 후, 파이어플라이 루시퍼레이즈(firefly luciferase)(ATPLite kit)를 이용하여 측정하였고, 살아있는 세포는 인비전 멀티 리더(EnVision Multilable Reader)를 활용하여 분석되었다. DMSO만 처리한 대조 세포군의 생존률을 100으로 놓고 약물 처리군의 상대 생존률을 계산하여 용량 반응 곡선(Dose-response curve, DRC)을 그린 후, 약물의 반응성 정도를 용량 반응 곡선의 곡선 아래 면적(Area Under Curve, AUC)를 계산하여 분석하였다.
Six days after the drug treatment, the cells were measured using a firefly luciferase (ATPLite kit), and living cells were analyzed using an EnVision Multilable Reader. Dose-response curve (DRC) was calculated by calculating the relative survival rate of the drug-treated group by setting the survival rate of the control cell group treated with DMSO only to 100. After that, the degree of reactivity of the drug was calculated as the area under the curve Under Curve, AUC) were calculated and analyzed.
실험 결과Experiment result
1. EGFR 유전자의 카피수(copy number)의 측정1. Measurement of copy number of EGFR gene
18명의 교모세포종 환자 검체에서 핵산을 분리한 후, 상기 분리된 핵산을 전장 서열 분석(Whole Exome Sequencing, WES)을 수행하여 EGFR 유전자의 카피수를 측정하였다. 또한, 같은 환자의 환자 유래의 암세포를 활용하여, 상기 43종의 표적항암제에 대해 약물 감수성 반응성을 고속 대량 스크리닝(High-throughput) 방법으로 검사를 수행하였다.
Nucleic acid was isolated from 18 glioblastoma patient specimens, and the number of copies of the EGFR gene was measured by performing whole-body sequencing (WES) on the separated nucleic acid. In addition, using the cancer cells derived from the patient of the same patient, the 43 kinds of target anticancer drugs were tested for drug sensitivity reactivity by a high-throughput method.
2. EGFR 카피수와 EGFR 억제제(inhibitor)의 상관관계(corelation) 분석2. Corelation analysis of EGFR copy number and EGFR inhibitor (inhibitor)
EGFR 카피수와 EGFR 억제제 3종(에를로티닙(Erlotinib), 제피티닙(Gefitinib) 및 아파티닙(Afatinib))에 대한 반응성 사이의 상관관계를 조사하였다. 상기 상관관계의 분석 결과를 도 3에 나타내었다. The correlation between the number of EGFR copies and the reactivity to three EGFR inhibitors (Erlotinib, Gefitinib and Afatinib) was investigated. The results of the above correlation analysis are shown in Fig.
도 3에 나타낸 바와 같이, EGFR 억제제와 즉, 에를로티닙(Erlotinib), 제피티닙(Gefitinib) 및 아파티닙(Afatinib)와 EGFR 카피수의 상관관계에 대한 그래프는 양의 상관관계를 갖는 것을 보여준다. As shown in FIG. 3, the graph of the correlation of EGFR inhibitors, that is, Erlotinib, Gefitinib, and Afatinib, and EGFR copy number has a positive correlation Show.
따라서, 상기 분석 결과는 EGFR 카피수가 높을수록, 약물 감수성(drug sensitivity)이 높다는 것을 실험적으로 입증한 것이다.
Thus, the above analysis results demonstrate experimentally that the higher the EGFR copy number, the higher the drug sensitivity.
3. EGFR 카피수를 이용한 교모세포종의 치료 효과의 유효성(effectiveness) 분석3. Efficacy analysis of curative effect of glioblastoma using EGFR copy number
EGFR 카피수(copy number) 10을 기준으로, EGFR 카피수가 10 미만인 종양의 환자유래 세포 및 10이상인 종양의 환자유래 세포의 EGFR 억제제 감수성을 비교 분석하였다. 상기 비교 분석 결과를 도 4에 나타내었다.
Based on the
도 4에 나타낸 바와 같이, EGFR 카피수가 10 이상인 그룹은 EGFR 억제제, 3종(에를로티닙(Erlotinib), 제피티닙(Gefitinib) 및 아파티닙(Afatinib))에 대한 약물 감수성이 모두 유의하게 증가한 것을 보여주고 있다. As shown in FIG. 4, the group with an EGFR copy number of 10 or more had significantly increased drug sensitivity to the EGFR inhibitor, three (Erlotinib, Gefitinib and Afatinib) .
따라서, 상기 실험결과 분석은 차세대 유전체 분석법(New Generation Sequencing, NGS) 기반의 교모세포종 신선동결조직에서 채취한 DNA에서 EGFR 카피수를 측정하였을 때, 카피수 10 이상인 그룹에서 EGFR 억제제의 치료 효과에 대한 감수성(sensitivity)이 유의하게 높다는 사실을 규명한 것이다.
Therefore, the analysis of the results of the above experiment showed that the EGFR copy number in DNA extracted from fresh frozen tissue of glioblastoma based on the New Generation Sequencing (NGS) assay showed that the EGFR inhibitor And that the sensitivity is significantly higher.
이상으로 본 발명의 특정한 부분을 상세히 기술하였는 바, 당업계의 통상의 지식을 가진 자에게 있어서 이러한 구체적인 기술은 단지 바람직한 구현예일 뿐이며, 이에 본 발명의 범위가 제한되는 것이 아닌 점은 명백하다. 따라서, 본 발명의 실질적인 범위는 첨부된 청구항과 그의 등가물에 의하여 정의된다고 할 것이다.
While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the same is by way of illustration and example only and is not to be construed as limiting the scope of the present invention. Accordingly, the actual scope of the present invention will be defined by the appended claims and their equivalents.
참고자료Resources
1. Wen, P.Y. & Kesari, S. Malignant gliomas in adults. N Engl J Med 359, 492-507 (2008).1. Wen, PY & Kesari, S. Malignant gliomas in adults. N Engl J Med 359 , 492-507 (2008).
2. Joo, K.M., et al. Patient-specific orthotopic glioblastoma xenograft models recapitulate the histopathology and biology of human glioblastomas in situ. Cell Rep 3, 260-273 (2013).2. Joo, KM , et al. Patient-specific orthotopic glioblastoma xenograft models recapitulate the histopathology and biology of human glioblastomas in situ.
3. Tentler, J.J., et al. Patient-derived tumour xenografts as models for oncology drug development. Nat Rev Clin Oncol 9, 338-350 (2012).3. Tentler, JJ , et al. Patient-derived tumor xenografts as models for oncology drug development. Nat Rev Clin Oncol 9 , 338-350 (2012).
4. Lee, J., et al. Tumor stem cells derived from glioblastomas cultured in bFGF and EGF more closely mirror the phenotype and genotype of primary tumors than do serum-cultured cell lines. Cancer Cell 9, 391-403 (2006).4. Lee, J. , et al. Tumor stem cells derived from glioblastomas cultured in bFGF and EGF more closely mirror the phenotype and genotype of primary tumors than do serum-cultured cell lines. Cancer Cell 9 , 391-403 (2006).
Claims (8)
(a) 대상으로부터 분리한 생시료(biosample)에서 지놈 DNA를 추출하는 단계;
(b) 상기 추출된 지놈 DNA의 EGFR 유전자의 카피수(copy number)를 분석하는 단계; 및
(c) 상기 분석된 EGFR 유전자의 카피수가 10 이상이면 상기 대상은 상기 EGFR 억제제에 대하여 유효한 치료 효과를 갖는 대상이고, EGFR 유전자의 카피수가 10 미만이면 EGFR 억제제에 대하여 유효하지 않은 치료 효과를 갖는 대상인 것으로 결정하는 단계.
A method for predicting the effectiveness of a therapeutic effect of a subject on an epidermal growth factor receptor (EGFR) inhibitor comprising the steps of:
(a) extracting genomic DNA from a biosample separated from a subject;
(b) analyzing the copy number of the EGFR gene of the extracted genome DNA; And
(c) a subject having an effective therapeutic effect on the EGFR inhibitor when the number of copies of the EGFR gene is 10 or more, and a subject having an ineffective therapeutic effect on the EGFR inhibitor if the copy number of the EGFR gene is less than 10 ≪ / RTI >
2. The method of claim 1, wherein said subject is a human being at risk for, or having an EGFR-related disorder.
The method according to claim 1, wherein the raw sample is tissue, cell, blood, plasma or serum.
4. The method according to claim 3, wherein the raw sample is a tissue or a cell.
5. The method according to claim 4, wherein the raw sample is a tumor cell of a glioblastoma or a tumor stem cell.
2. The method of claim 1, wherein step (b) is performed by whole genome sequencing or targeted exome sequencing.
The method of claim 1, wherein the epidermal growth factor receptor (EGFR) inhibitor is Gefitinib, Erlotinib, or Afatinib.
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