KR100886670B1 - New diagnostic markers and methods of papillary thyroid cancer - Google Patents
New diagnostic markers and methods of papillary thyroid cancerDownload PDFInfo
- Publication number
- KR100886670B1 KR100886670B1KR1020070003463AKR20070003463AKR100886670B1KR 100886670 B1KR100886670 B1KR 100886670B1KR 1020070003463 AKR1020070003463 AKR 1020070003463AKR 20070003463 AKR20070003463 AKR 20070003463AKR 100886670 B1KR100886670 B1KR 100886670B1
- Authority
- KR
- South Korea
- Prior art keywords
- leu
- asp
- mcm3
- ala
- val
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
Images
Classifications
- A—HUMAN NECESSITIES
- A47—FURNITURE; DOMESTIC ARTICLES OR APPLIANCES; COFFEE MILLS; SPICE MILLS; SUCTION CLEANERS IN GENERAL
- A47L—DOMESTIC WASHING OR CLEANING; SUCTION CLEANERS IN GENERAL
- A47L15/00—Washing or rinsing machines for crockery or tableware
- A47L15/42—Details
- A47L15/4285—Water-heater arrangements
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
- G01N33/57407—Specifically defined cancers
- A—HUMAN NECESSITIES
- A47—FURNITURE; DOMESTIC ARTICLES OR APPLIANCES; COFFEE MILLS; SPICE MILLS; SUCTION CLEANERS IN GENERAL
- A47L—DOMESTIC WASHING OR CLEANING; SUCTION CLEANERS IN GENERAL
- A47L2501/00—Output in controlling method of washing or rinsing machines for crockery or tableware, i.e. quantities or components controlled, or actions performed by the controlling device executing the controlling method
- A47L2501/06—Water heaters
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
- G01N2333/46—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans from vertebrates
- G01N2333/47—Assays involving proteins of known structure or function as defined in the subgroups
- G01N2333/4701—Details
- G01N2333/4703—Regulators; Modulating activity
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Urology & Nephrology (AREA)
- Hematology (AREA)
- Biomedical Technology (AREA)
- Chemical & Material Sciences (AREA)
- Molecular Biology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Cell Biology (AREA)
- Hospice & Palliative Care (AREA)
- Biotechnology (AREA)
- Food Science & Technology (AREA)
- Oncology (AREA)
- Microbiology (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Peptides Or Proteins (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
Translated fromKoreanDescription
Translated fromKorean도 1A는 PTC의 고유한 특성과 PTC 내 MCM3의 면역반응성을 보여주는 결과 사진으로서, 왼쪽 사진은 PTC의 특징적인 섬유혈관 심지를 갖는 유두엽 형상을 보여주는 것이고(100배), 오른쪽 사진은 정상 갑상샘 소포 세포(thyroid follicular cells)에 대한 음성 염색 결과를 보여주는 것이며(200배),FIG. 1A is a photograph showing the intrinsic properties of PTC and the immunoreactivity of MCM3 in PTC. The left photograph shows the papillary morphology with PTC's characteristic fibrovascular wick (100-fold), and the right photograph shows normal thyroid vesicle cells. shows negative staining results for thyroid follicular cells (200-fold),
도 1B는 PTC 내 MCM3의 면역반응성을 보여주는 도면으로서, MCM3에 대한 면역조직화학 염색결과 PTC 세포내 핵 염색 패턴을 보여주는 사진이며(100배),Figure 1B is a diagram showing the immunoreactivity of MCM3 in PTC, a photograph showing the nuclear staining pattern in PTC cells by immunohistochemical staining for MCM3 (100 times),
도 2는 증식중인 림프구내 MCM3에 대한 면역염색 결과를 보여주는 도면으로서, 유두갑상선 종양 근처 림프구 집합체내 증식중인 림프구들에서 MCM3 면역염색이 나타난 사진이며(200배),2 is a diagram showing the results of immunostaining for MCM3 in proliferating lymphocytes, a photograph showing MCM3 immunostaining in proliferating lymphocytes in lymphocyte aggregates near the papillary thyroid tumor (200 times).
도 3은 인간 갑상선 조직내 MCM3 단백질의 발현수준에 대한 웨스턴 블롯 분 석 결과를 보여주는 도면으로서, 인간 PTC 조직내 MCM3 단백질의 발현수준(C)을 각각에 대응하는 정상 조직내 MCM3 단백질의 발현수준(N)과 비교한 결과이며, 동일한 시료의 블롯들에 대해 MCM3에 대한 항체와 β-액틴에 대한 항체를 프로브로 사용한 결과를 각각 나타낸 것이다.FIG. 3 shows Western blot analysis of the expression level of MCM3 protein in human thyroid tissue, wherein the expression level (C) of MCM3 protein in human PTC tissue corresponds to the expression level of MCM3 protein in normal tissues. N) and the results of using the antibody against MCM3 and the antibody against β-actin as probes for the blots of the same sample, respectively.
전세계적으로, 갑상선암은 내분비 기관의 악성 종양 중 가장 보편적인 형태이며, 유두갑상선암(PTC)은갑상선암의 가장 일반적인 형태이다. 또한, PTC는 여러가지 암들 중 발병률의 증가속도가 가장 빠른 암으로서, 1991년 10만명당 7.5명으로부터 2004년 8.4명으로 현저히 증가해왔으며, 특히 중년 여성에게서 가장 높은 증가율을 보여주고 있다[보건복지부, 한국중앙암등록 연례보고서, 서울, 2004]. 그리하여, 상기 갑상선암에 대한 종양 발생(tumorigenesis) 및 예후(prognosis)에 대한 관심이 높아져왔다.Worldwide, thyroid cancer is the most common form of malignant tumor of the endocrine organs, and papillary thyroid cancer (PTC) is the most common form of thyroid cancer. In addition, PTC is the fastest growing cancer among various cancers, and has increased significantly from 7.5 in 100,000 people in 1991 to 8.4 in 2004, especially among middle-aged women [Ministry of Health and Welfare, Korea] Central Cancer Registration Annual Report, Seoul, 2004]. Thus, there has been a growing interest in tumorigenesis and prognosis for the thyroid cancer.
나아가, PTC는 보편화된 증식 마커인 Ki-67 및 PCNA와 같은 현저한 증식 마커를 가지지 않은 암이다. 종양 세포의 증식능력은 성장중인 모든 종양에 전형적인 특징이다. 수많은 병리학자들과 임상의들이 세포 증식을 측정하여 예후 정보로 사용해왔다. Ki-67의 면역활성이 폐암, 연부조직 육종(soft tissue sarcoma), 뇌 수막염(meningioma), 전립선암, 비호지킨성 림프종(non-Hodgkin's lymphoma) 등의 몇몇 인간종양에 대한 보편적인 증식표지로서 널리 사용되어왔다[얀센 알엘(Jansen RL) 등, "MIB-1 labelling index is an independent prognostic marker in primary breast cancer",Br J Cancer 1998, 78:460-465 페리 에이(Perry A) 등, "The prognostic significance of MIB-1, p53, and DNA flow cytometry in completely resected primary meningiomas",Cancer (Phila.) 1998, 82:2262-2269; 마샬 알디(Mashal RD), 등 "Expression of cell cycle-regulated proteins in prostate cancer",Cancer Res 1996, 56:4159-4163; 및 달렌바흐 에프(Dallenbach F) 등, "Growth fractions in malignant non-Hodgkin's lymphomas (NHL) as determined in situ with the monoclonal antibody Ki-67",Hematol Oncol 1984,2:365-371]. Ki-67의 이러한 유용성에도불구하고, 몇몇 보고서를 통해 세포주기에 직접 관련이 있다기보다는 세포 증식기 리보솜 생합성에 있어서 어떤 역할을 하는 것일 수도 있다는 보고가 있었으나, 아직까지 Ki-67의 기능에 대해 명백하게 알려지지 않은 상태이다[맥캘럼 디이(MacCallum DE), 등, "The location of pKi67 in the outer dense fibrillary compartment of the nucleolus points to a role in ribosome biogenesis during the cell division cycle",J Pathol 2000, 190:537-544]. 특히, PTC에 대해서는, Ki-67이 신뢰할만한 증식표지인지 여부에 대해서는 의문시 되고 있다[에릭슨 엘에이(Erickson LA) 등, "Expression of p27kip1 and Ki-67 in benign and malignant thyroid tumors", Mod Pathol 1998, 11:169-174; 및 카토 알(Katoh R) 등, "Growth activity in hyperplastic and neoplastic human thyroid determined by an immunohistochemical staining procedure using monoclonal antibody MIB-1",Hum Pathol 1995,26:139-146]. 따라서, PTC의 증식 활성을 보다 정확히 가늠하기 위해 DNA 복제에 직접적인 관련이 있는 또 다른 증식 표지가 필요로 되고 있다.Furthermore, PTC is a cancer that does not have significant proliferative markers such as Ki-67 and PCNA, which are common proliferative markers. The proliferative capacity of tumor cells is typical for all growing tumors. Numerous pathologists and clinicians have measured cell proliferation and used it as a prognostic information. Ki-67's immune activity is widely used as a universal proliferative marker for several human tumors, including lung cancer, soft tissue sarcoma, meningioma, prostate cancer, and non-Hodgkin's lymphoma. Jansen RL et al., "MIB-1 labeling index is an independent prognostic marker in primary breast cancer",Br J Cancer 1998, 78: 460-465 Perry A et al., "The prognostic significance of MIB-1, p53, and DNA flow cytometry in completely resected primary meningiomas ",Cancer (Phila.) 1998, 82: 2262-2269; Marshall RD, et al., "Expression of cell cycle-regulated proteins in prostate cancer",Cancer Res 1996, 56: 4159-4163; And Dalenbach F et al., "Growth fractions in malignant non-Hodgkin's lymphomas (NHL) as determined in situ with the monoclonal antibody Ki-67",Hematol Oncol 1984,2: 365-371]. Despite the usefulness of Ki-67, some reports indicate that it may play a role in cell proliferative ribosomal biosynthesis rather than directly related to the cell cycle, but it is still clear about the function of Ki-67. It is unknown [MacCallum DE, et al., "The location of pKi67 in the outer dense fibrillary compartment of the nucleolus points to a role in ribosome biogenesis during the cell division cycle",J Pathol 2000, 190: 537 -544]. In particular, for PTC, it is questioned whether Ki-67 is a reliable proliferative label [Erickson LA et al., "Expression of p27kip1 and Ki-67 in benign and malignant thyroid tumors",Mod Pathol 1998, 11: 169-174; And Katoh R et al., "Growth activity in hyperplastic and neoplastic human thyroid determined by an immunohistochemical staining procedure using monoclonal antibody MIB-1",Hum Pathol 1995,26: 139-146]. Therefore, in order to more accurately estimate the proliferative activity of PTC, another proliferation label directly related to DNA replication is needed.
한편, 여섯 개 단백질로 구성된 MCM 단백질 군(MCM2 내지 7)은 DNA-결합 단백질들 중 가장 잘 보존된 그룹이고 진핵세포내 DNA 복제에 필요한 단백질로서 알려져있다[타이 비케이(Tye BK), "MCM proteins in DNA replication"Annu Rev Biochem 1999, 68:649-686]. MCM 단백질은 DNA 복제의 개시와 조절에 관련이 있다. MCM 단백질은 DNA 복제 기원에 결합하고 그에 뒤이어 기원 인식 복합체(origin recognition complex, ORC)와 Cdc 6 단백질과 상호작용을 함으로써 복제전 복합체(prereplicative complex)를 형성한다. S-상을 개시하는 때에, Cdc6 단백질이 복제전 복합체로부터 방출되어 DNA 합성을 조절함으로써 DNA가 각 세포주기동안 단 한번만 복제되도록 한다[쉐발리에 에스(Chevalier S) 등, "Cell cycle control of replication initiation in eukaryotes"Curr Opin Cell Biol 1996, 8:815-821]. MCM 단백질들은 세포주기내내 모든 세포에서 발현되며, 휴지상태의 세포와 분화중인 세포에서는 MCM 단백질 발현이 나타나지 않는 것으로 보고되어 있다[매다인 엠에이(Madine MA) 등, "The roles of the MCM, ORC, and Cdc6 proteins in determining the replication competence of chromatin in quiescent cells"J Struct Biol 2000, 129:198-210.]. 비종양성 증식 및 종양성 이상 모두에서 증식 구획을 한정하는데 MCM 단백질에 대한 항체를 사용할 수 있다. 대부분의 연구에서 상기 여섯 개 MCM 단백질들 모두가 유사한 면역염색(immunostaining) 결과를 갖는 것으로 보고되었다[히라이와 에이(Hiraiwa A) 등, "Immunolocalization of hCDC47 protein in normal and neoplastic human tissues and its relation to growth"Int J Cancer 1997, 74:180-184; 및 프리만 에이(Freeman A) 등, "Minichromosome maintenance proteins as biological markers of dysplasia and malignancy"Clin Cancer Res 1999, 5:2121-2132.].On the other hand, the MCM protein group consisting of six proteins (MCM2-7) is the best conserved group of DNA-binding proteins and is known as a protein required for eukaryotic DNA replication [Tye BK, "MCM proteins in DNA replication "Annu Rev Biochem 1999, 68: 649-686. MCM proteins are involved in the initiation and regulation of DNA replication. The MCM protein forms a prereplicative complex by binding to the origin of DNA replication followed by interaction with the origin recognition complex (ORC) and the
그러나, 다양한 종양내 MCM 단백질 발현에 대한 대부분의 연구는 MCM2와 MCM5에 대해 이루어진 것이 모두이다[램나스 엔(Ramnath N) 등, "MCM2 is an independent predictor of survival in patients with nonsmall-cell lung cancer",J Clin Oncol 2001, 19:4259-4266; 스퇴버 케이(Stoeber K) 등, "Diagnosis of genito-urinary tract cancer by detection of minichromosome maintenance 5 protein in urine sediments"J Natl Cancer Inst (Bethesda) 2002, 94:1071-1079; 곤잘레스 엠에이(Gonzalez MA) 등, "Minichromosome maintenance protein 2 is a strong independent prognostic marker in breast cancer"J Clin Oncol 2003, 21:4306-4313; 및 가토 에이치(Kato H) 등, "A new proliferation marker, minichromosome maintenance protein 2, is associated with tumor aggressiveness in esophageal squamous cell carcinoma"J Surg Oncol 2003, 84:24-30]. 많은 연구에서 MCM 단백질 면역반응성이 증식중인 세포의 표지 로서 사용될 수 있음이 제안되었다[가토 에이치 등, "A new proliferation marker, minichromosome maintenance protein 2, is associated with tumor aggressiveness in esophageal squamous cell carcinoma"J Surg Oncol 2003, 84:24-30; 및 하 에스에이(Ha SA) 등, "Cancer-associated expression of minichromosome maintenance 3 gene in several human cancers and its involvement in tumorigenesis"Clin Cancer Res 2004, 10:8386-8395]. 선행연구에서 사체조직들을 이용한 세포주기진입의 평가에서 MCM2가 Ki-67 보다 훌륭한 표지일 수 있음이 보고된 바 있다[로딘스 케이(Rodins K) 등, "Minichromosome maintenance protein 2 expression in normal kidney and renal cell carcinomas: relationship to tumor dormancy and potential clinical utility"Clin Cancer Res 2002, 8:1075-1081]. 그러나, MCM3에 대한 유용성에 대해서는 아직까지 보고된바 없으며, 특히, MCM 단백질 발현수준과 PTC와의 관계에 대해서 밝혀진 바 없다. 본 발명에서는 PTC에 대한 새롭고 신뢰할만한 진단 마커로서의 MCM3의 유용성을 평가하여 PTC 진단을 위한 효과적인 진단마커의 요구에 부응하고자 한다.However, most studies of MCM protein expression in various tumors have been done for both MCM2 and MCM5 (Ramnath N et al., "MCM2 is an independent predictor of survival in patients with nonsmall-cell lung cancer").J Clin Oncol 2001, 19: 4259-4266; Stoeber K et al., "Diagnosis of genito-urinary tract cancer by detection of
본 발명의 목적은 유두 갑상선 암에 대한 효과적인 진단 마커를 제공하는 것이다.It is an object of the present invention to provide an effective diagnostic marker for papillary thyroid cancer.
본 발명의 다른 목적은 미니염색체 유지 단백질 3(MCM3)의 유두 갑상선 암의 진단마커로서의 새로운 용도를 제공하는 것이다.Another object of the present invention is to provide a novel use of minichromosome maintenance protein 3 (MCM3) as a diagnostic marker for papillary thyroid cancer.
본 발명의 또 다른 목적은 MCM3에 특이적인 항체를 사용하여 유두 갑상선 암을 진단하는 방법을 제공하는 것이다.Another object of the present invention is to provide a method for diagnosing papillary thyroid cancer using an antibody specific for MCM3.
또한, 본 발명의 목적은 MCM3에 특이적인 항체를 포함하는 유두 갑상선 암 진단용 진단키트를 제공하는 것이다.It is also an object of the present invention to provide a diagnostic kit for diagnosing papillary thyroid cancer comprising an antibody specific for MCM3.
본 발명은 유두 갑상선 암(PTC)에 대한 새롭고 효과적인 진단마커를 제공한다. 구체적으로, 본 발명은 미니염색체 유지 단백질 3(MCM3)과 PTC의 임상병리학적 파라미터들과의 관련성을 밝히고 상기 단백질을 PTC에 대한 진단마커로서 제공한다.The present invention provides a new and effective diagnostic marker for papillary thyroid cancer (PTC). Specifically, the present invention reveals the relationship between minichromosomal maintenance protein 3 (MCM3) and the clinical pathologic parameters of PTC and provides the protein as a diagnostic marker for PTC.
본 발명자는 상기 MCM3 단백질의 발현양이 유두 갑상선 암의 조직내에서 현저히높고 정상의 갑상선 조직에서 현저히 낮거나 거의 없는 것으로 확인하였다. 따라서, MCM3 단백질의 발현양을 검출하면 유두 갑상선 암의 진단이 가능하다.The present inventors confirmed that the expression level of the MCM3 protein was significantly high in the tissues of papillary thyroid cancer and was significantly low or little in normal thyroid tissue. Therefore, detecting the expression level of MCM3 protein enables diagnosis of papillary thyroid cancer.
본 발명의 진단방법은 a) 개체로부터 얻은 조직시료를 제공하고, b) 상기 시료를 MCM3와 특이적 결합을 형성하는 진단시약과 접촉시킨 다음, c) (b)에서 형성된 MCM3와 이에 대한 진단시약 간의 복합체 형성을 확인하여 유두 갑상선 암(PTC)를 진단하는 것으로 이루어진다.The diagnostic method of the present invention comprises a) providing a tissue sample obtained from an individual, b) contacting the sample with a diagnostic reagent that forms a specific binding with MCM3, and c) MCM3 formed in (b) and a diagnostic reagent therefor. Diagnosis of papillary thyroid cancer (PTC) consists of confirming complex formation of the liver.
상기 진단방법에 사용되는 진단시약은 MCM3와 특이적인 결합을 형성할 수 있는 단백질, 항원 등으로부터 선택되는 하나를 함유할 수 있고, 바람직하게는 상기 진단시약은 MCM3에 특이적인 항체를 함유하며, 상기 항체는 모노클로날또는 폴리클로날 항체일 수 있다. 또한, 상기 진단시약은 상기 항체의 일부 절편 또는 MCM3와의 결합을 위한 도메인을 필수로 포함하는 절편을 성분으로 할 수 있다.The diagnostic reagent used in the diagnostic method may contain one selected from proteins, antigens, and the like, which may form a specific bond with MCM3. Preferably, the diagnostic reagent contains an antibody specific for MCM3. The antibody may be a monoclonal or polyclonal antibody. In addition, the diagnostic reagent may be composed of some fragments of the antibody or fragments that essentially include a domain for binding to MCM3.
MCM3에 특이적인 항체는 당해 기술분야에 공지된 어떠한 방법에 의해서든 제작될 수 있다. 예들 들면, 윤승규 등, "The human cervical cancer oncogene protein is a biomarker for human hepatocellular carcinoma"Cancer Research, 2004; 64: 5434-5441에 기재된 방법으로 제조될 수 있다.Antibodies specific for MCM3 can be produced by any method known in the art. For example, Seung-Kyu Yoon et al., "The human cervical cancer oncogene protein is a biomarker for human hepatocellular carcinoma"Cancer Research , 2004; 64: 5434-5441.
본 발명의 경우 토끼에서 생성한 모노클로날 항체를 사용할 수 있으며, 이에 나아가 이와 다른 종 유래의 모노클로날 항체 및 폴리클로날 항체도 사용될 수 있다In the case of the present invention, a monoclonal antibody produced in a rabbit can be used. Furthermore, a monoclonal antibody and a polyclonal antibody derived from another species can be used...
MCM3와 이에 특이적인 항체와의 복합체의 형성은 검출 표지를 사용하여 확인될 수 있다. 검출 표지로서 염색원, 형광물, 효소, 리간드, 발광물, 방사선동위원소를 비롯하여 당업자에게 알려져 있는 모든 검출표지가 사용될 수 있다. 본 발명의 구체예에서는 MCM3와의 복합체 형성을 확인하기 위해 비오틴이 표지된 항-마우스 면역글로불린(Ig) G를 2차 항체로 사용하고, 3,3-디아미노벤지딘-히드로겐 퍼옥 시드를 염색원(chromogen)으로 사용하였다.Formation of the complex of MCM3 with antibodies specific to it can be confirmed using a detection label. As the detection label, any detection label known to those skilled in the art may be used including a dye source, a fluorescent material, an enzyme, a ligand, a luminescent material, and a radioisotope. In an embodiment of the present invention, biotin-labeled anti-mouse immunoglobulin (Ig) G is used as a secondary antibody to confirm complex formation with MCM3, and 3,3-diaminobenzidine-hydrogen peroxide is used as a staining source. It was used as a (chromogen).
본 발명의 진단방법은 상기한 항체를 사용하여 당 분야에 공지된 통상의 방법에 따라 면역조직화학 분석법, 효소 면역측정법(ELISA), 방사선면역측정법(RIA), 샌드위치 측정법(sandwich assay), 폴리아크릴 겔 상의 웨스턴 블롯 또는 면역 블롯 등의 방법에 의해 개체의 시료 중 MCM3 단백질의 발현량을 확인함으로써 PTC를 진단할 수 있다.The diagnostic method of the present invention is an immunohistochemical assay, enzyme immunoassay (ELISA), radioimmunoassay (RIA), sandwich assay, polyacrylic acid according to a conventional method known in the art using the above-described antibody PTC can be diagnosed by confirming the expression level of MCM3 protein in a subject's sample by a method such as western blot or immunoblot on a gel.
본 발명의 진단키트는 MCM3에 특이적인 항체 및 검출 표지를 포함한다. 본 발명의 진단키트는 2차 항체를 더 포함할 수 있다. 상기 항체 및 검출 표지는 위에서 상술한 바와 같으며, 상기 2차 항체는 바람직하게는 면역글로불린 G(Ig G)이다.Diagnostic kits of the invention include antibodies and detection labels specific for MCM3. The diagnostic kit of the present invention may further comprise a secondary antibody. The antibody and detection label are as described above above and the secondary antibody is preferably immunoglobulin G (Ig G).
본 발명의 MCM3의 PTC에 대한 다양한 임상변리학적 파라미터와의 관련성을 통해 MCM3 단백질이 PTC 진단 마커로서의 갖는 유용성을 확인하기 위해서, PTC 조직의 MCM3에 대한 면역조직화학적(immunohistochemical) 분석, 및 PTC 환자의 조직과 정상조직간 MCM3 발현수준 비교를 위한 웨스턴 블롯 분석을 수행할 수 있다.In order to confirm the usefulness of the MCM3 protein as a PTC diagnostic marker through the relevance of the MCM3 to PTC for various clinical morphologic parameters, immunohistochemical analysis of MCM3 of PTC tissues, and of PTC patients Western blot analysis can be performed to compare MCM3 expression levels between tissue and normal tissue.
면역조직화학적 분석 결과, MCM3 발현수준은 PTC 환자들에게서 매우 높게 나타났으며, 특히 MCM3 단백질 수준은 종양크기 및 피막외 확장과 현저한 관련성을 보였다. 따라서, MCM3 발현정도가 세포군의 빠른 증식 구획을 검출하는데 있어서 보다 민감하고 신뢰할만한 마커가 될 수 있음을 알 수 있다.Immunohistochemical analysis revealed that MCM3 expression levels were very high in PTC patients, especially MCM3 protein levels were significantly associated with tumor size and extracapsular expansion. Therefore, it can be seen that the expression level of MCM3 may be a more sensitive and reliable marker for detecting the rapid proliferation section of the cell population.
항-MCM 항체의 PTC에서 유도된 악성종양 세포에 대한 증식 마커로서의 유용성을 파라핀에 잠긴 섹션상에 친화도-정제 폴리클로날 항체로 확인할 수 있는데, PTC 조직에 대해서 특징적인 섬유혈관 심지를 가진 유두엽(papillary fronds) 형상을 나타내는(도 1A, 왼쪽) 반면, 정상적인 갑상선 조직은 항-MCM3 항체와 양성 반응도 보이지 않는다(도 1A, 오른쪽). 항-MCM3 항체의 면역반응성에 의존한 염색 결과 주로 암 소(nest)의 둘레로 MCM3 단백질의 핵내 분포에 일치하는 염색 패턴을 보인다(도 1B). 또한, PTC 주변의 림프구 집합체에서도 증식중인 림프구내에서는 MCM3의 과발현이 확인된다(도 2).The usefulness of anti-MCM antibodies as a proliferation marker for PTC-induced malignant tumor cells can be confirmed with affinity-purified polyclonal antibodies on paraffin-clad sections, papillary lobes with fibrovascular wicks characteristic for PTC tissue. Normal thyroid tissue shows no positive reaction with anti-MCM3 antibody (FIG. 1A, right) while exhibiting papillary fronds shape (FIG. 1A, left). Staining dependent on the immunoreactivity of the anti-MCM3 antibody shows a staining pattern consistent with the intranuclear distribution of the MCM3 protein mainly around the cow (FIG. 1B). In addition, overexpression of MCM3 is observed in lymphocytes that are proliferating even in lymphocyte aggregates around PTC (FIG. 2).
MCM3 단백질 발현수준에 대한 웨스턴 블롯 분석 결과 모든 PTC 환자의 조직시료에서 MCM3 단백질이 과발현되었음을 확인하였다(도 3 참조). 대조적으로 모든 정상 조직에서는 MCM3의 발현수준이 매우 낮거나 거의 없었다.Western blot analysis of the MCM3 protein expression level confirmed that MCM3 protein was overexpressed in the tissue samples of all PTC patients (see FIG. 3). In contrast, all normal tissues had very low or very low levels of MCM3 expression.
결론적으로, PTC의 진단에 있어서 MCM3 단백질은 신뢰할 만한 표지가 될 수 있다. 특히, MCM3 단백질은 PTC 환자에게서 종양의 성장을 측정하고 종양 침투 및 예후 인자를 평가하는데 유용하다. 따라서, MCM3은 PTC의 종양성장성 및 종양침투성의 진단에 유용한 증식 마커라고 할 것이다.In conclusion, the MCM3 protein can be a reliable marker for the diagnosis of PTC. In particular, the MCM3 protein is useful for measuring tumor growth and assessing tumor penetration and prognostic factors in PTC patients. Therefore, MCM3 would be a proliferative marker useful for the diagnosis of tumor growth and tumor invasiveness of PTC.
이하 본 발명을 실시예에 의하여 더욱 상세히 설명하고자 한다. 단, 하기 실시예는 본 발명을 예시하기 위한 것일 뿐, 본 발명의 범위가 이들만으로 한정되는 것은 아니다.Hereinafter, the present invention will be described in more detail with reference to Examples. However, the following examples are only for illustrating the present invention, and the scope of the present invention is not limited thereto.
실시예 1: PTC 환자 조직시료 수집Example 1 PTC Patient Tissue Sample Collection
총 30명의 유두 갑상선 암(PTC) 환자들로부터 채취된 조직 시료와 임상기록자료를 분석하여 표 1에 나타내었다. 상기 임상기록은 연령, 성별 및 종양 크기를 포함한다. 이들 환자의 평균연령은 45세였다(20 내지 67세의 범위).Tissue samples and clinical records collected from 30 patients with papillary thyroid cancer (PTC) were analyzed and shown in Table 1. The clinical record includes age, sex and tumor size. The mean age of these patients was 45 years (range of 20-67 years).
림프절 전이, 림프구 침투, 피막외 확장, 근육침투, 종양 다양성 등을 비롯한 조직병리학적 인자들을 통상 인정되는 진단 표준을 사용해서 헤마톡실린(hematoxylin)과 에오신(eosin) 섹션(H-E)에서 평가했다. 상기 30명 중 16명은 초기환자(I기)이고, 14명은 후기환자였다.Histopathological factors, including lymph node metastasis, lymphocyte infiltration, extracapsular expansion, muscle infiltration, tumor diversity, etc., were evaluated in the hematoxylin and eosin sections (H-E) using commonly accepted diagnostic standards. Of the 30 patients, 16 were early patients (stage I) and 14 were late patients.
1) 총 100개 악성종양 세포들에대해 산출된 양성 종양 세포의 비율.1) Proportion of benign tumor cells calculated against a total of 100 malignant tumor cells.
실시예 2: MCM3 단백질에 대한 항체 생산Example 2: Antibody Production Against MCM3 Proteins
MCM3 유전자 생성물에 대한 래빗(rabbit) 폴리클로날 항체를 형성하였다. MCM3의 PCR 생성물을 pBAD/TOPO 발현벡터(pBAD/TOPO 티오퓨전 발현키트, 인비트로젠 사,미국)에 붙여 6x 히스티딘 태그와 융합된 전체 MCM3를 발현하는 플라스미드를 제작하였다. 이렇게 제작된 플라스미드를 E.Coli에 도입하고 아라비노스로 상기 E.Coli를 유도하여 융합단백질을생성하고, Ni-니트릴로테트라아세트산-아가로스 칼럼(ProBond 정제시스템, 인비트로젠 사, 미국)을 사용하여 정제하였다. 뉴질랜드 화이트 래빗(New Zealand White rabbits)에서 상기 정제된 단백질에 대한 항체를 생성했다. 결과 얻어진 토끼의 혈청을 CNBr-활성화 세파로스 비드(Amersham Biosciences사, 미국)에 교차공유결합된 MCM3 단백질과의 친화도를 이용하여 정제했다.Rabbit polyclonal antibodies to the MCM3 gene product were formed. The PCR product of MCM3 was attached to a pBAD / TOPO expression vector (pBAD / TOPO thiofusion expression kit, Invitrogen,USA ) to prepare a plasmid expressing the entire MCM3 fused with a 6x histidine tag. The plasmid thus prepared was introduced into E.Coli and the arabinose was used to induce the E.Coli to generate a fusion protein, and a Ni-nitrilotetraacetic acid-agarose column (ProBond Purification System, Invitrogen, USA) Purified using. Antibodies to the purified proteins were generated in New Zealand White rabbits. The resulting rabbit serum was purified using affinity with MCM3 protein cross-linked to CNBr-activated Sepharose beads (Amersham Biosciences, USA).
실시예 3: 면역조직화학 염색Example 3: Immunohistochemical Staining
스트렙타비딘-비오틴 복합체 면역과산화효소 방법(Dako사, 덴마크)을 이용하여 면역조직화학 염색을 수행하였다. 항원 복구를 위해 마이크로파 전처리를 10mM 농도의 시트레이트(citrate) 완충액 내에서 9분간 수행하였다. 내생(endogenous) 과산화효소의 활성은 메탄올 내 0.3% H2O2를 사용하여 30분간 차단하였다. MCM3에 대한 항체(Dako 사, 덴마크)를 1:50의 비율로 희석하여PTC 환자로부터 얻은 조직시료와 정상인의 갑상선 조직 시료에 각각 적용한 후, 4℃에서 밤새 인큐베이트하였다. 비오틴이 표지된 항-마우스 면역글로불린(Ig) G를 2차 항체로 사용하고, 3,3-디아미노벤지딘-히드로겐 퍼옥시드를 염색원(chromogen)으로 사용하였다. 구획들이 헤마톡실린으로 대비 염색된 후, 탈수시키고, 정화시킨 다음 고정하였다. 트리스 완충 식염수로 1차 항체를 대체한 음성대조군을 준비했다.Immunohistochemical staining was performed using the streptavidin-biotin complex immunoperoxidase method (Dako, Denmark). Microwave pretreatment was performed for 9 minutes in citrate buffer at 10 mM concentration for antigen recovery. Endogenous peroxidase activity was blocked for 30 minutes using 0.3% H2O2 in methanol. Antibodies against MCM3 (Dako, Denmark) were diluted 1:50and appliedto tissue samples obtained from PTC patients and thyroid tissue samples from normal subjects, and then incubated overnight at 4 ° C. Biotin-labeled anti-mouse immunoglobulin (Ig) G was used as secondary antibody and 3,3-diaminobenzidine-hydrogen peroxide was used as chromogen. Compartments were counterstained with hematoxylin, then dehydrated, clarified and fixed. A negative control was prepared by replacing the primary antibody with Tris buffered saline.
비교예: Ki-67 항체를 사용한 면역조직화학 염색Comparative Example: Immunohistochemical Staining with Ki-67 Antibody
스트렙타비딘-비오틴 복합체 면역과산화효소 방법(Dako사, 덴마크)을 이용하여 면역조직화학 염색을 수행하였다. 항원 복구를 위해 마이크로파 전처리를 10mM 농도의 시트레이트(citrate) 완충액 내에서 9분간 수행하였다. 내생(endogenous) 과산화효소의 활성은 메탄올 내 0.3% H2O2를 사용하여 30분간 차단하였다. Ki-67에 대한 항체(Dako 사, 덴마크)를 1:100의 비율로 희석하여 적용한 후, 4℃에서 밤새 인큐베이트하였다. 비오틴이 표지된 항-마우스 면역글로불린(Ig) G를 2차 항체로 사용하고, 3,3-디아미노벤지딘-히드로겐 퍼옥시드를 염색원(chromogen)으로 사용하였다. 구획들이 헤마톡실린으로 대비 염색된 후, 탈수시키고, 정화시킨 다음 고정하였다. 트리스 완충 식염수로 1차 항체를 대체한 음성대조군을 준비했다.Immunohistochemical staining was performed using the streptavidin-biotin complex immunoperoxidase method (Dako, Denmark). Microwave pretreatment was performed for 9 minutes in citrate buffer at 10 mM concentration for antigen recovery. Endogenous peroxidase activity was blocked for 30 minutes using 0.3% H2O2 in methanol. Antibodies against Ki-67 (Dako, Denmark) were applied at a dilution of 1: 100 and then incubated at 4 ° C. overnight. Biotin-labeled anti-mouse immunoglobulin (Ig) G was used as secondary antibody and 3,3-diaminobenzidine-hydrogen peroxide was used as chromogen. Compartments were counterstained with hematoxylin, then dehydrated, clarified and fixed. A negative control was prepared by replacing the primary antibody with Tris buffered saline.
결과분석Result Analysis
실시예 3 및 비교예에서 얻은 면역조직화학 염색 결과에 대해 분석을 수행하였다. 구획들은 고해상도(400배)로 관찰되었다. 뚜렷한 핵 염색 상태를 보여야 양성으로 인정되었다. MCM3 및 Ki-67 라벨 인덱스를 대부분의 활성영역 내에 있는 1,000개의 종양세포에 대한 퍼센티지로 평가되었다. 종양의 라벨인덱스(MCM3 및 Ki-67에 대한 양성 종양 세포들의 퍼센티지)를 1군(0-25%), 2군(26-50%), 3군(51-75%) 및 4군(76-100%)의 네 개의 등급으로 분류하였다(표 1; MCM3 등급).Analysis was performed on the immunohistochemical staining results obtained in Example 3 and Comparative Examples. Compartments were observed at high resolution (400 times). Positive nuclear staining was required to show positive status. MCM3 and Ki-67 label indices were assessed as a percentage of 1,000 tumor cells within most active regions. Tumor label indexes (percentage of positive tumor cells for MCM3 and Ki-67) were divided into Group 1 (0-25%), Group 2 (26-50%), Group 3 (51-75%), and Group 4 (76). -100%) and four grades (Table 1; MCM3 grade).
PTC의 항-MCM3 항체 반응 및 면역염색 결과 특징적인 섬유혈관 심지를 가진 유두엽(papillary fronds) 형상을 관찰하였다(도 1A, 왼쪽). 항-MCM3 항체의 면역반응성에 대해서는, 이러한 유두상(papillary) 구조의 종양세포는 주로 암의 소(nest)의 둘레로 핵 염색 패턴을 보였는데, 이는 MCM3 단백질의 핵내 분포에 일치하는 것이다(도 1B). 그러나, PTC에 대조적으로, 정상적인 갑상선 조직에서는 염색결과 항-MCM3 항체와의 어떠한 양성 반응도 보이지 않았다(도 1A, 오른쪽). 이는 MCM3 발현이 PTC와 강한 견련성이 있음을 보여주는 것이다. 도 2는 또한 PTC 주변의 림프구 집합체에서 증식중인 림프구내MCM3를 면역염색한 결과를 보여주고 있다.Anti-MCM3 antibody response and immunostaining of PTC showed papillary fronds shape with characteristic fibrovascular wicks (FIG. 1A, left). For the immunoreactivity of anti-MCM3 antibodies, these papillary tumor cells showed a nuclear staining pattern mainly around the cancer's nest, which is consistent with the intranuclear distribution of the MCM3 protein (Fig. 1B). However, in contrast to PTC, normal thyroid tissue did not show any positive reaction with anti-MCM3 antibody in staining (FIG. 1A, right). This shows that MCM3 expression is strongly related to PTC. Figure 2 also shows the results of immunostaining of lymphocyte MMC3 proliferating in lymphocyte aggregates around PTC.
한편, MCM3와 Ki-67 발현과, 환자의 변수(clinical variables) (나이, 성별, 종양크기) 및 임상적인 지표(histopathologic parameters)(림프절 전이, 림프절 침투, 피막외 침습, 근육 침투, 종양의 다중성)와의 상관관계를 분산분석방법으로 결정하였다. 단순회귀분석방법(simple regression analysis)을 사용하여 MCM3 및 Ki-67 라벨 지수간의 상관관계를 분석하였다.Meanwhile, MCM3 and Ki-67 expression, patient variables (age, gender, tumor size) and clinical parameters (lymph node metastasis, lymph node invasion, extracapsular invasion, muscle infiltration, tumor multiplicity) ) Was determined by analysis of variance. Simple regression analysis was used to analyze the correlation between MCM3 and Ki-67 label indices.
표 2에 30명의 PTC 환자들에서 얻은 조직들의 임상병리학적 파라미터들과 MCM3 및 Ki-67 라벨 지수들간의 관계를 정리하였다.Table 2 summarizes the relationship between the clinicopathological parameters of the tissues obtained from 30 PTC patients and the MCM3 and Ki-67 label indices.
상기 표 2는 면역조직화학적 분석에 있어서, 대조군으로서 당해기술분야에서 보편적으로 종양의 마커로 사용되고 있는 Ki-67에 대한 면역조직화학적 분석을 함께 수행하여 MCM3에 대한 분석결과와 비교한결과이다. 이러한 분석 결과를 보면, MCM3 라벨인덱스(LI)는 종양크기(P=0.031) 및 피막외 확장(P=0.037)에 대해 현저한 통계적 관련성을 보인반면, Ki-67 LI는 실험대상 PCT 환자들 30명에서 8가지의 임상병리학적 파라미터들과 통계학적으로 중요한 관련성을 보이지 않았다. 그러므로, Ki-67은 PTC 증식을 검사하는데 좋은 지표는 아닌 것으로 여겨진다.Table 2 shows the results of immunohistochemical analysis of Ki-67, which is commonly used as a marker for tumors in the art as a control, and compared with the analysis results for MCM3. These analyzes show that MCM3 label index (LI) has a significant statistical relationship between tumor size (P = 0.031) and extracapsular expansion (P = 0.037), whereas Ki-67 LI shows 30 PCT patients. There was no statistically significant relationship with the eight clinicopathological parameters in. Therefore, Ki-67 is not considered a good indicator for testing PTC proliferation.
일차종양에서, MCM3와Ki-67라벨지수 사이에현저한 상호관련성이 관찰되지는 않았으나, MCM3 LI는 상당히 더 높았다.In primary tumors, no significant correlation was observed between MCM3 and Ki-67 label index, but MCM3 LI was significantly higher.
MCM3과 Ki-67 단백질의 PTC에서의 발현수준을 조사한 결과, Ki-67 LI가 PTC에서 낮게 나타남을 확인했다. 이러한 조사 결과와 일치하게, 대부분의 연구들에서 잘 분화된 갑상선 암종 내 낮은 Ki-67 LI 값을 확인하고 있다[티믈러 디(Timler D) 등, "Expression of proteins: D1 cyclin and Ki-67 in papillary thyroid carcinomas"Folia Histochem Cytobiol2001, 39:201-202 크젤만 피(Kjellman P) 등, "MIB-1 index in thyroid tumors: a predictor of the clinical course in papillary thyroid carcinoma"Thyroid2003, 13:371-380 시이로넨 피(Siironen P) 등, "Prognostic factors in papillary thyroid cancer: an evaluation of 601 consecutive patients"Tumour Biol 2005, 26:57-64.]. 본 발명자들에 의해서 Ki-67 LI 값이 종양의 임상병리학적 파라미터들과 상당한 관련성을 갖지 아니한다는 사실이 명백해졌다.As a result of examining the expression level of MCM3 and Ki-67 protein in PTC, it was confirmed that Ki-67 LI was low in PTC. Consistent with these findings, most studies have identified low Ki-67 LI values in well-differentiated thyroid carcinoma [Timler D et al., "Expression of proteins: D1 cyclin and Ki-67 in papillary thyroid carcinomasFolia Histochem Cytobiol 2001, 39: 201-202 Kjellman P et al., "MIB-1 index in thyroid tumors: a predictor of the clinical course in papillary thyroid carcinoma"Thyroid 2003, 13: 371- 380 Siironen P et al., "Prognostic factors in papillary thyroid cancer: an evaluation of 601 consecutive patients"Tumour Biol 2005, 26: 57-64. It was clear by the inventors that Ki-67 LI values do not have a significant correlation with the clinicopathological parameters of the tumor.
위와 같이, MCM3 LI가 Ki-67 LI보다 상당히 높다는 사실은 Ki-67이 MCM3 보다 짧은 세포주기 간격 동안 발현될 수 있음을 보여주는 것이다. 구체적으로, MCM3은 세포주기 전체에 걸쳐 존재하다가, MCM3의 mRNA 수준은 G1-S의 경계면에서 극적으로 증가한다. Ki-67이 S, G2 그리고 M기에서 주로 발현됨에 반해, MCM3는 정상 세포와 신생중인 세포에서도 G1 초기에 발현이 이루어지는데, Ki-67은 이 시기에 검출되지 않는다.As above, the fact that MCM3 LI is significantly higher than Ki-67 LI shows that Ki-67 can be expressed during cell cycle intervals shorter than MCM3. Specifically, MCM3 is present throughout the cell cycle, while the mRNA level of MCM3 increases dramatically at the interface of G1-S. While Ki-67 is mainly expressed in S, G2 and M phases, MCM3 is expressed early in G1 in normal and neoplastic cells, but Ki-67 is not detected at this time.
또한, 임상병리학적 특징들간의 상호관계를 분석해보면, 표 2의 8가지 임상병리학적 특징들 중, 연령과 종양의 다중도는 다른 파라미터들과 유의할만한 관계를 보이지 않았다. 반면, 종양의 크기는 피막외 확장과 상당한 관련이 있었다(P=0.016). 본 발명자들은 또한 피막외 확장과 근육 침투 사이에 상당한 관련성을 확인했다(P=0.019). 부가하여, 림프절 침투와 국소 림프절 전이 사이의 상관관계 역시 상당했다(P=0.000). 최종적으로, 성별(남자)은 종양 크기(P=0.015), 림프절 전이(P=0.007), 그리고 림프절 침투(P=0.015)와 상당히 상관관계를 가지고 있었다.In addition, the analysis of the correlation between the clinicopathologic features showed that, among the eight clinicopathological features of Table 2, age and multiplicity of the tumor did not show a significant relationship with other parameters. On the other hand, tumor size was significantly associated with extracapsular expansion (P = 0.016). We also found a significant relationship between extracapsular expansion and muscle penetration (P = 0.019). In addition, the correlation between lymph node infiltration and regional lymph node metastasis was also significant (P = 0.000). Finally, gender (male) correlated significantly with tumor size (P = 0.015), lymph node metastasis (P = 0.007), and lymph node infiltration (P = 0.015).
실시예 4: 웨스턴 블롯 분석Example 4: Western Blot Analysis
웨스턴 블롯 분석을 위해, 조직시료를 프로테아제저해제 혼합물(Sigma사, 미국)을 함유한 라이시스 완충액(lysis buffer)[20mM 트리스(pH7.4), 1% NP40, 5mM EDTA, 10% 글리콜, 0.1% SDS, 및 150mM NaCl] 내에서 호모게나이즈 시켰다. 상기 호모게나이즈된 시료를5,000G에서 10분간 원심분리하여 맑게 하였다. 총 단백질 중 20㎍을 함유하는 세포 라이세이트를 동일한 부피로 10% SDS-폴리아크릴아미드 겔상에 로딩하고 전기영동으로 분리하였다. 그 후 단백질 샘플을 니트로셀룰로스 멤브레인으로 옮겼다. 상기 블롯들을 폴리클로날 항-MCM3 혈청과 함께 인큐베이션하고 개량된 화학발광 검출 키트(Pierce 사, 미국)로 현상하였다.For Western blot analysis, tissue samples were subjected to Lysis buffer [20 mM Tris (pH7.4), 1% NP40, 5 mM EDTA, 10% glycol, 0.1% containing a protease inhibitor mixture (Sigma, USA). SDS, and 150 mM NaCl]. The homogenized sample was cleared by centrifugation at 5,000 G for 10 minutes. Cell lysates containing 20 μg of total protein were loaded on a 10% SDS-polyacrylamide gel in equal volume and separated by electrophoresis. The protein sample was then transferred to the nitrocellulose membrane. The blots were incubated with polyclonal anti-MCM3 serum and developed with an improved chemiluminescence detection kit (Pierce, USA).
6개 PTC와 정상 갑상선 조직의 MCM3 발현수준을 비교한 결과를 도 3에 나타냈다. 도 1 및 도 2에서와 같이, MCM3 단백질은 정상조직에서보다 PTC에서 높은 수준으로 검출되었고, 정상 갑상선 조직에서 MCM3 단백질 수준은 매우 낮거나 거의 없었다.The results of comparing MCM3 expression levels of six PTCs and normal thyroid tissues are shown in FIG. 3. As shown in Figures 1 and 2, MCM3 protein was detected at higher levels in PTC than in normal tissues, with very low or almost no MCM3 protein levels in normal thyroid tissue.
본 발명은 유두갑상선암에 대한 새롭고 효과적인 진단마커로서 MCM3 단백질을 제공한다. 본 발명에 따르면 유두갑상선암에 대한 정확한 조기진단이 가능할 것이다.The present invention provides MCM3 protein as a new and effective diagnostic marker for papillary thyroid cancer. According to the present invention, accurate early diagnosis of papillary thyroid cancer will be possible.
<110> KIM, Hyun Kee<120> NOVEL PROLIFERATION MARKER IN PAPILLARY THYROID CARCINOMA AND NOVEL METHOD OF DIAGNOSIS DETECTING THE MARKER<140> 10-2007-0003463<141> 2007-01-11<160> 2<170> KopatentIn 1.71<210> 1<211> 3091<212> DNA<213> Homo sapiens<220><221> CDS<222> (54)..(2081)<400> 1attaccaatc gcgaaaccgc gactttggtg gaggtagttc tttggcagcg ggc 53atg gcg ggt acc gtg gtg ctg gac gat gtg gag ctg cgg gag gct cag 101Met Ala Gly Thr Val Val Leu Asp Asp Val Glu Leu Arg Glu Ala Gln 1 5 10 15 aga gat tac ctg gac ttc ctg gac gac gag gaa gac cag gga att tat 149Arg Asp Tyr Leu Asp Phe Leu Asp Asp Glu Glu Asp Gln Gly Ile Tyr 20 25 30 cag agc aaa gtt cgg gag ctg atc agt gac aac caa tac cgg ctg att 197Gln Ser Lys Val Arg Glu Leu Ile Ser Asp Asn Gln Tyr Arg Leu Ile 35 40 45 gtc aat gtg aat gac ctg cgc agg aaa aac gag aag agg gct aac cgg 245Val Asn Val Asn Asp Leu Arg Arg Lys Asn Glu Lys Arg Ala Asn Arg 50 55 60 ctt ctg aac aat gcc ttt gag gag ctg gtt gcc ttc cag cgg gcc tta 293Leu Leu Asn Asn Ala Phe Glu Glu Leu Val Ala Phe Gln Arg Ala Leu 65 70 75 80 aag gat ttt gtg gcc tcc att gat gct acc tat gcc aag cag tat gag 341Lys Asp Phe Val Ala Ser Ile Asp Ala Thr Tyr Ala Lys Gln Tyr Glu 85 90 95 gag ttc tac gta gga ctg gaa ggc agc ttt ggc tcc aag cac gtc tcc 389Glu Phe Tyr Val Gly Leu Glu Gly Ser Phe Gly Ser Lys His Val Ser 100 105 110 ccg cgg act ctt acc tcc tgc ttc ctc agc tgt gtg gtc tgt gtg gag 437Pro Arg Thr Leu Thr Ser Cys Phe Leu Ser Cys Val Val Cys Val Glu 115 120 125 ggc att gtc act aaa tgt tct cta gtt cgt ccc aaa gtc gtc cgc agt 485Gly Ile Val Thr Lys Cys Ser Leu Val Arg Pro Lys Val Val Arg Ser 130 135 140 gtc cac tac tgt cct gct act aag aag acc ata gag cga cgt tat tct 533Val His Tyr Cys Pro Ala Thr Lys Lys Thr Ile Glu Arg Arg Tyr Ser 145 150 155 160 gat ctc acc acc ctg gtg gcc ttt ccc tcc agc tct gtc tat cct acc 581Asp Leu Thr Thr Leu Val Ala Phe Pro Ser Ser Ser Val Tyr Pro Thr 165 170 175 aag gat gag gag aac aat ccc ctt gag aca gaa tat ggc ctt tct gtc 629Lys Asp Glu Glu Asn Asn Pro Leu Glu Thr Glu Tyr Gly Leu Ser Val 180 185 190 tac aag gat cac cag acc atc acc atc cag gag atg ccg gag aag gcc 677Tyr Lys Asp His Gln Thr Ile Thr Ile Gln Glu Met Pro Glu Lys Ala 195 200 205 cca gcc ggc cag ctc ccc cgc tct gtg gac gtc att ctg gat gat gac 725Pro Ala Gly Gln Leu Pro Arg Ser Val Asp Val Ile Leu Asp Asp Asp 210 215 220 ttg gtg gat aaa gcg aag cct ggt gac cgg gtt cag gtg gtg gga acc 773Leu Val Asp Lys Ala Lys Pro Gly Asp Arg Val Gln Val Val Gly Thr 225 230 235 240 tac cgt tgc ctt cct gga aag aag gga ggc tac acc tct ggg acc ttc 821Tyr Arg Cys Leu Pro Gly Lys Lys Gly Gly Tyr Thr Ser Gly Thr Phe 245 250 255 agg act gtc ctg att gcc tgt aat gtt aag cag atg agc aag gat gct 869Arg Thr Val Leu Ile Ala Cys Asn Val Lys Gln Met Ser Lys Asp Ala 260 265 270 cag ccc tct ttc tct gct gag gat ata gcc aag atc aag aag ttc agt 917Gln Pro Ser Phe Ser Ala Glu Asp Ile Ala Lys Ile Lys Lys Phe Ser 275 280 285 aaa acc cga tcc aag gat atc ttt gac cag ctg gcc aag tca ttg gcc 965Lys Thr Arg Ser Lys Asp Ile Phe Asp Gln Leu Ala Lys Ser Leu Ala 290 295 300 cca agt atc cat ggg cat gac tat gtc aag aaa gca atc ctc tgc ttg 1013Pro Ser Ile His Gly His Asp Tyr Val Lys Lys Ala Ile Leu Cys Leu 305 310 315 320 ctc ttg gga ggg gtg gaa cga gac cta gaa aat ggc agc cac atc cgt 1061Leu Leu Gly Gly Val Glu Arg Asp Leu Glu Asn Gly Ser His Ile Arg 325 330 335 ggg gac atc aat att ctt cta ata gga gac cca tcc gtt gcc aag tct 1109Gly Asp Ile Asn Ile Leu Leu Ile Gly Asp Pro Ser Val Ala Lys Ser 340 345 350 cag ctt ctg cgg tat gtg ctt tgc act gca ccc cga gct atc ccc acc 1157Gln Leu Leu Arg Tyr Val Leu Cys Thr Ala Pro Arg Ala Ile Pro Thr 355 360 365 act ggc cgg ggc tcc tct gga gtg ggt ctg acg gct gct gtc acc aca 1205Thr Gly Arg Gly Ser Ser Gly Val Gly Leu Thr Ala Ala Val Thr Thr 370 375 380 gac cag gaa aca gga gag cgc cgt ctg gaa gca ggg gcc atg gtc ctg 1253Asp Gln Glu Thr Gly Glu Arg Arg Leu Glu Ala Gly Ala Met Val Leu 385 390 395 400 gct gac cga ggc gtg gtt tgc att gat gaa ttt gac aaa atg tct gac 1301Ala Asp Arg Gly Val Val Cys Ile Asp Glu Phe Asp Lys Met Ser Asp 405 410 415 atg gat cgc aca gcc atc cat gaa gtg atg gag cag ggt cga gtg acc 1349Met Asp Arg Thr Ala Ile His Glu Val Met Glu Gln Gly Arg Val Thr 420 425 430 att gcc aag gct ggc atc cat gct cgg ctg aat gcc cgc tgc agt gtt 1397Ile Ala Lys Ala Gly Ile His Ala Arg Leu Asn Ala Arg Cys Ser Val 435 440 445 ttg gca gct gcc aac cct gtc tac ggc agg tat gac cag tat aag act 1445Leu Ala Ala Ala Asn Pro Val Tyr Gly Arg Tyr Asp Gln Tyr Lys Thr 450 455 460 cca atg gag aac att ggg cta cag gac tca ctg ctg tca cga ttt gac 1493Pro Met Glu Asn Ile Gly Leu Gln Asp Ser Leu Leu Ser Arg Phe Asp 465 470 475 480 ttg ctc ttc atc atg ctg gat cag atg gat cct gag cag gat cgg gag 1541Leu Leu Phe Ile Met Leu Asp Gln Met Asp Pro Glu Gln Asp Arg Glu 485 490 495 atc tca gac cat gtc ctt cgg atg cac cgt tac aga gca cct ggg gag 1589Ile Ser Asp His Val Leu Arg Met His Arg Tyr Arg Ala Pro Gly Glu 500 505 510 cag gat ggc gat gct atg ccc ttg ggt agt gct gtg gat atc ctg gcc 1637Gln Asp Gly Asp Ala Met Pro Leu Gly Ser Ala Val Asp Ile Leu Ala 515 520 525 aca gat gat ccc aac ttt agc cag gaa gat cag cag gac acc cag att 1685Thr Asp Asp Pro Asn Phe Ser Gln Glu Asp Gln Gln Asp Thr Gln Ile 530 535 540 tat gag aag cat gac aac ctt cta cat ggg acc aag aag aaa aag gag 1733Tyr Glu Lys His Asp Asn Leu Leu His Gly Thr Lys Lys Lys Lys Glu 545 550 555 560 aag atg gtg agt gca gca ttc atg aag aag tac atc cat gtg gcc aaa 1781Lys Met Val Ser Ala Ala Phe Met Lys Lys Tyr Ile His Val Ala Lys 565 570 575 atc atc aag cct gtc ctg aca cag gag tcg gcc acc tac att gca gaa 1829Ile Ile Lys Pro Val Leu Thr Gln Glu Ser Ala Thr Tyr Ile Ala Glu 580 585 590 gag tat tca cgc ctg cgc agc cag gat agc atg agc tca gac acc gcc 1877Glu Tyr Ser Arg Leu Arg Ser Gln Asp Ser Met Ser Ser Asp Thr Ala 595 600 605 agg aca tct cca gtt aca gcc cga aca ctg gaa act ctg att cga ctg 1925Arg Thr Ser Pro Val Thr Ala Arg Thr Leu Glu Thr Leu Ile Arg Leu 610 615 620 gcc aca gcc cat gcg aag gcc cgc atg agc aag act gtg gac ctg cag 1973Ala Thr Ala His Ala Lys Ala Arg Met Ser Lys Thr Val Asp Leu Gln 625 630 635 640 gat gca gag gaa gct gtg gag ttg gtc cag tat gct tac ttt aag aag 2021Asp Ala Glu Glu Ala Val Glu Leu Val Gln Tyr Ala Tyr Phe Lys Lys 645 650 655 gtt ctg gag aag gag aag aaa cgt aag aag cga agt gag gat gaa tca 2069Val Leu Glu Lys Glu Lys Lys Arg Lys Lys Arg Ser Glu Asp Glu Ser 660 665 670 gag aca gaa gat tgaagagga gaaaagccaa gaggaccagg agcagaagag 2120Glu Thr Glu Asp 675 gaagagaagg aagactcgcc agccagatgc caaagatggg gattcatacg acccctatga 2180cttcagtgac acagaggagg aaatgcctca agtacacact ccaaagacgg cagactcaca 2240ggagaccaag gaatcccaga aagtggagtt gagtgaatcc aggttgaagg cattcaaggt 2300ggccctcttg gatgtgttcc gggaagctca tgcgcagtca atcggcatga atcgcctcac 2360agaatccatc aaccgggaca gcaaagagcc cttctcttca gttgagatcc aggctgctct 2420gagcaagatg caggatgaca atcaggtcat ggtgtctgag ggcatcatct tcctcatctg 2480aggaggcctc gtctctgaac ttgggttgtg ccgagagagt ttgttctgtg tttcccaccc 2540tctccctgac ccaagtcttt gcctctactc ccttaacagt gttgaattca actgaaggcg 2600aggaatgttg gtgatgaagc tgagttcagg actcggtgga ccctttggga atgggtcatg 2660aaagctgcca tggggtgagg aaagaggaga cagtgggaga ggacaatgac tattgcatct 2720tcattgcaaa agcactggct catccgccct acttcccatc ccacacaaac ccaattgtaa 2780ataacatatg acttctgagt acttttgggg gcacaactgt tttctgtttg ctgttttttt 2840gttttgtttt ttttctccag agcactttgg tctagactag gctttgggtg gttccaattg 2900gtggagagaa gctctgaggc acgtcatgca ggtcaagaaa gctttctttg cagtagcacc 2960agttaaggtg aatatgtatt gtatcacaaa acaaacccaa tatccagatg aatatccgag 3020atgttgaata aacttagcca tttcgtacac atggaaaaga aaaaaaaaaa aaaaaaaaaa 3080aaaaaaaaaa a 3091<210> 2<211> 676<212> PRT<213> Homo sapiens<400> 2Met Ala Gly Thr Val Val Leu Asp Asp Val Glu Leu Arg Glu Ala Gln 1 5 10 15 Arg Asp Tyr Leu Asp Phe Leu Asp Asp Glu Glu Asp Gln Gly Ile Tyr 20 25 30 Gln Ser Lys Val Arg Glu Leu Ile Ser Asp Asn Gln Tyr Arg Leu Ile 35 40 45 Val Asn Val Asn Asp Leu Arg Arg Lys Asn Glu Lys Arg Ala Asn Arg 50 55 60 Leu Leu Asn Asn Ala Phe Glu Glu Leu Val Ala Phe Gln Arg Ala Leu 65 70 75 80 Lys Asp Phe Val Ala Ser Ile Asp Ala Thr Tyr Ala Lys Gln Tyr Glu 85 90 95 Glu Phe Tyr Val Gly Leu Glu Gly Ser Phe Gly Ser Lys His Val Ser 100 105 110 Pro Arg Thr Leu Thr Ser Cys Phe Leu Ser Cys Val Val Cys Val Glu 115 120 125 Gly Ile Val Thr Lys Cys Ser Leu Val Arg Pro Lys Val Val Arg Ser 130 135 140 Val His Tyr Cys Pro Ala Thr Lys Lys Thr Ile Glu Arg Arg Tyr Ser145 150 155 160 Asp Leu Thr Thr Leu Val Ala Phe Pro Ser Ser Ser Val Tyr Pro Thr 165 170 175 Lys Asp Glu Glu Asn Asn Pro Leu Glu Thr Glu Tyr Gly Leu Ser Val 180 185 190 Tyr Lys Asp His Gln Thr Ile Thr Ile Gln Glu Met Pro Glu Lys Ala 195 200 205 Pro Ala Gly Gln Leu Pro Arg Ser Val Asp Val Ile Leu Asp Asp Asp 210 215 220 Leu Val Asp Lys Ala Lys Pro Gly Asp Arg Val Gln Val Val Gly Thr225 230 235 240 Tyr Arg Cys Leu Pro Gly Lys Lys Gly Gly Tyr Thr Ser Gly Thr Phe 245 250 255 Arg Thr Val Leu Ile Ala Cys Asn Val Lys Gln Met Ser Lys Asp Ala 260 265 270 Gln Pro Ser Phe Ser Ala Glu Asp Ile Ala Lys Ile Lys Lys Phe Ser 275 280 285 Lys Thr Arg Ser Lys Asp Ile Phe Asp Gln Leu Ala Lys Ser Leu Ala 290 295 300 Pro Ser Ile His Gly His Asp Tyr Val Lys Lys Ala Ile Leu Cys Leu305 310 315 320 Leu Leu Gly Gly Val Glu Arg Asp Leu Glu Asn Gly Ser His Ile Arg 325 330 335 Gly Asp Ile Asn Ile Leu Leu Ile Gly Asp Pro Ser Val Ala Lys Ser 340 345 350 Gln Leu Leu Arg Tyr Val Leu Cys Thr Ala Pro Arg Ala Ile Pro Thr 355 360 365 Thr Gly Arg Gly Ser Ser Gly Val Gly Leu Thr Ala Ala Val Thr Thr 370 375 380 Asp Gln Glu Thr Gly Glu Arg Arg Leu Glu Ala Gly Ala Met Val Leu385 390 395 400 Ala Asp Arg Gly Val Val Cys Ile Asp Glu Phe Asp Lys Met Ser Asp 405 410 415 Met Asp Arg Thr Ala Ile His Glu Val Met Glu Gln Gly Arg Val Thr 420 425 430 Ile Ala Lys Ala Gly Ile His Ala Arg Leu Asn Ala Arg Cys Ser Val 435 440 445 Leu Ala Ala Ala Asn Pro Val Tyr Gly Arg Tyr Asp Gln Tyr Lys Thr 450 455 460 Pro Met Glu Asn Ile Gly Leu Gln Asp Ser Leu Leu Ser Arg Phe Asp465 470 475 480 Leu Leu Phe Ile Met Leu Asp Gln Met Asp Pro Glu Gln Asp Arg Glu 485 490 495 Ile Ser Asp His Val Leu Arg Met His Arg Tyr Arg Ala Pro Gly Glu 500 505 510 Gln Asp Gly Asp Ala Met Pro Leu Gly Ser Ala Val Asp Ile Leu Ala 515 520 525 Thr Asp Asp Pro Asn Phe Ser Gln Glu Asp Gln Gln Asp Thr Gln Ile 530 535 540 Tyr Glu Lys His Asp Asn Leu Leu His Gly Thr Lys Lys Lys Lys Glu545 550 555 560 Lys Met Val Ser Ala Ala Phe Met Lys Lys Tyr Ile His Val Ala Lys 565 570 575 Ile Ile Lys Pro Val Leu Thr Gln Glu Ser Ala Thr Tyr Ile Ala Glu 580 585 590 Glu Tyr Ser Arg Leu Arg Ser Gln Asp Ser Met Ser Ser Asp Thr Ala 595 600 605 Arg Thr Ser Pro Val Thr Ala Arg Thr Leu Glu Thr Leu Ile Arg Leu 610 615 620 Ala Thr Ala His Ala Lys Ala Arg Met Ser Lys Thr Val Asp Leu Gln625 630 635 640 Asp Ala Glu Glu Ala Val Glu Leu Val Gln Tyr Ala Tyr Phe Lys Lys 645 650 655 Val Leu Glu Lys Glu Lys Lys Arg Lys Lys Arg Ser Glu Asp Glu Ser 660 665 670 Glu Thr Glu Asp 675<110> KIM, Hyun Kee<120> NOVEL PROLIFERATION MARKER IN PAPILLARY THYROID CARCINOMA AND NOVEL METHOD OF DIAGNOSIS DETECTING THE MARKER<140> 10-2007-0003463<141> 2007-01-11<160> 2<170> KopatentIn 1.71<210> 1<211> 3091<212> DNA<213> Homo sapiens<220><221> CDS(222) (54) .. (2081)<400> 1attaccaatc gcgaaaccgc gactttggtg gaggtagttc tttggcagcg ggc 53atg gcg ggt acc gtg gtg ctg gac gat gtg gag ctg cgg gag gct cag 101Met Ala Gly Thr Val Val Leu Asp Asp Val Glu Leu Arg Glu Ala Gln 1 5 10 15aga gat tac ctg gac ttc ctg gac gac gag gaa gac cag gga att tat 149Arg Asp Tyr Leu Asp Phe Leu Asp Asp Glu Glu Asp Gln Gly Ile Tyr 20 25 30cag agc aaa gtt cgg gag ctg atc agt gac aac caa tac cgg ctg att 197Gln Ser Lys Val Arg Glu Leu Ile Ser Asp Asn Gln Tyr Arg Leu Ile 35 40 45gtc aat gtg aat gac ctg cgc agg aaa aac gag aag agg gct aac cgg 245Val Asn Val Asn Asp Leu Arg Arg Lys Asn Glu Lys Arg Ala Asn Arg 50 55 60ctt ctg aac aat gcc ttt gag gag ctg gtt gcc ttc cag cgg gcc tta 293Leu Leu Asn Asn Ala Phe Glu Glu Leu Val Ala Phe Gln Arg Ala Leu 65 70 75 80aag gat ttt gtg gcc tcc att gat gct acc tat gcc aag cag tat gag 341Lys Asp Phe Val Ala Ser Ile Asp Ala Thr Tyr Ala Lys Gln Tyr Glu 85 90 95gag ttc tac gta gga ctg gaa ggc agc ttt ggc tcc aag cac gtc tcc 389Glu Phe Tyr Val Gly Leu Glu Gly Ser Phe Gly Ser Lys His Val Ser 100 105 110ccg cgg act ctt acc tcc tgc ttc ctc agc tgt gtg gtc tgt gtg gag 437Pro Arg Thr Leu Thr Ser Cys Phe Leu Ser Cys Val Val Cys Val Glu 115 120 125ggc att gtc act aaa tgt tct cta gtt cgt ccc aaa gtc gtc cgc agt 485Gly Ile Val Thr Lys Cys Ser Leu Val Arg Pro Lys Val Val Arg Ser 130 135 140gtc cac tac tgt cct gct act aag aag acc ata gag cga cgt tat tct 533Val His Tyr Cys Pro Ala Thr Lys Lys Thr Ile Glu Arg Arg Tyr Ser145 150 155 160gat ctc acc acc ctg gtg gcc ttt ccc tcc agc tct gtc tat cct acc 581Asp Leu Thr Thr Leu Val Ala Phe Pro Ser Ser Ser Val Tyr Pro Thr 165 170 175aag gat gag gag aac aat ccc ctt gag aca gaa tat ggc ctt tct gtc 629Lys Asp Glu Glu Asn Asn Pro Leu Glu Thr Glu Tyr Gly Leu Ser Val 180 185 190tac aag gat cac cag acc atc acc atc cag gag atg ccg gag aag gcc 677Tyr Lys Asp His Gln Thr Ile Thr Ile Gln Glu Met Pro Glu Lys Ala 195 200 205cca gcc ggc cag ctc ccc cgc tct gtg gac gtc att ctg gat gat gac 725Pro Ala Gly Gln Leu Pro Arg Ser Val Asp Val Ile Leu Asp Asp Asp 210 215 220ttg gtg gat aaa gcg aag cct ggt gac cgg gtt cag gtg gtg gga acc 773Leu Val Asp Lys Ala Lys Pro Gly Asp Arg Val Gln Val Val Gly Thr225 230 235 240tac cgt tgc ctt cct gga aag aag gga ggc tac acc tct ggg acc ttc 821Tyr Arg Cys Leu Pro Gly Lys Lys Gly Gly Tyr Thr Ser Gly Thr Phe 245 250 255agg act gtc ctg att gcc tgt aat gtt aag cag atg agc aag gat gct 869Arg Thr Val Leu Ile Ala Cys Asn Val Lys Gln Met Ser Lys Asp Ala 260 265 270cag ccc tct ttc tct gct gag gat ata gcc aag atc aag aag ttc agt 917Gln Pro Ser Phe Ser Ala Glu Asp Ile Ala Lys Ile Lys Lys Phe Ser 275 280 285aaa acc cga tcc aag gat atc ttt gac cag ctg gcc aag tca ttg gcc 965Lys Thr Arg Ser Lys Asp Ile Phe Asp Gln Leu Ala Lys Ser Leu Ala 290 295 300cca agt atc cat ggg cat gac tat gtc aag aaa gca atc ctc tgc ttg 1013Pro Ser Ile His Gly His Asp Tyr Val Lys Lys Ala Ile Leu Cys Leu305 310 315 320ctc ttg gga ggg gtg gaa cga gac cta gaa aat ggc agc cac atc cgt 1061Leu Leu Gly Gly Val Glu Arg Asp Leu Glu Asn Gly Ser His Ile Arg 325 330 335ggg gac atc aat att ctt cta ata gga gac cca tcc gtt gcc aag tct 1109Gly Asp Ile Asn Ile Leu Leu Ile Gly Asp Pro Ser Val Ala Lys Ser 340 345 350cag ctt ctg cgg tat gtg ctt tgc act gca ccc cga gct atc ccc acc 1157Gln Leu Leu Arg Tyr Val Leu Cys Thr Ala Pro Arg Ala Ile Pro Thr 355 360 365act ggc cgg ggc tcc tct gga gtg ggt ctg acg gct gct gtc acc aca 1205Thr Gly Arg Gly Ser Ser Gly Val Gly Leu Thr Ala Ala Val Thr Thr 370 375 380gac cag gaa aca gga gag cgc cgt ctg gaa gca ggg gcc atg gtc ctg 1253Asp Gln Glu Thr Gly Glu Arg Arg Leu Glu Ala Gly Ala Met Val Leu385 390 395 400gct gac cga ggc gtg gtt tgc att gat gaa ttt gac aaa atg tct gac 1301Ala Asp Arg Gly Val Val Cys Ile Asp Glu Phe Asp Lys Met Ser Asp 405 410 415atg gat cgc aca gcc atc cat gaa gtg atg gag cag ggt cga gtg acc 1349Met Asp Arg Thr Ala Ile His Glu Val Met Glu Gln Gly Arg Val Thr 420 425 430att gcc aag gct ggc atc cat gct cgg ctg aat gcc cgc tgc agt gtt 1397Ile Ala Lys Ala Gly Ile His Ala Arg Leu Asn Ala Arg Cys Ser Val 435 440 445ttg gca gct gcc aac cct gtc tac ggc agg tat gac cag tat aag act 1445Leu Ala Ala Ala Asn Pro Val Tyr Gly Arg Tyr Asp Gln Tyr Lys Thr 450 455 460cca atg gag aac att ggg cta cag gac tca ctg ctg tca cga ttt gac 1493Pro Met Glu Asn Ile Gly Leu Gln Asp Ser Leu Leu Ser Arg Phe Asp465 470 475 480ttg ctc ttc atc atg ctg gat cag atg gat cct gag cag gat cgg gag 1541Leu Leu Phe Ile Met Leu Asp Gln Met Asp Pro Glu Gln Asp Arg Glu 485 490 495atc tca gac cat gtc ctt cgg atg cac cgt tac aga gca cct ggg gag 1589Ile Ser Asp His Val Leu Arg Met His Arg Tyr Arg Ala Pro Gly Glu 500 505 510cag gat ggc gat gct atg ccc ttg ggt agt gct gtg gat atc ctg gcc 1637Gln Asp Gly Asp Ala Met Pro Leu Gly Ser Ala Val Asp Ile Leu Ala 515 520 525aca gat gat ccc aac ttt agc cag gaa gat cag cag gac acc cag att 1685Thr Asp Asp Pro Asn Phe Ser Gln Glu Asp Gln Gln Asp Thr Gln Ile 530 535 540tat gag aag cat gac aac ctt cta cat ggg acc aag aag aaa aag gag 1733Tyr Glu Lys His Asp Asn Leu Leu His Gly Thr Lys Lys Lys Lys Glu545 550 555 560aag atg gtg agt gca gca ttc atg aag aag tac atc cat gtg gcc aaa 1781Lys Met Val Ser Ala Ala Phe Met Lys Lys Tyr Ile His Val Ala Lys 565 570 575atc atc aag cct gtc ctg aca cag gag tcg gcc acc tac att gca gaa 1829Ile Ile Lys Pro Val Leu Thr Gln Glu Ser Ala Thr Tyr Ile Ala Glu 580 585 590gag tat tca cgc ctg cgc agc cag gat agc atg agc tca gac acc gcc 1877Glu Tyr Ser Arg Leu Arg Ser Gln Asp Ser Met Ser Ser Asp Thr Ala 595 600 605agg aca tct cca gtt aca gcc cga aca ctg gaa act ctg att cga ctg 1925Arg Thr Ser Pro Val Thr Ala Arg Thr Leu Glu Thr Leu Ile Arg Leu 610 615 620gcc aca gcc cat gcg aag gcc cgc atg agc aag act gtg gac ctg cag 1973Ala Thr Ala His Ala Lys Ala Arg Met Ser Lys Thr Val Asp Leu Gln625 630 635 640gat gca gag gaa gct gtg gag ttg gtc cag tat gct tac ttt aag aag 2021Asp Ala Glu Glu Ala Val Glu Leu Val Gln Tyr Ala Tyr Phe Lys Lys 645 650 655gtt ctg gag aag gag aag aaa cgt aag aag cga agt gag gat gaa tca 2069Val Leu Glu Lys Glu Lys Lys Arg Lys Lys Arg Ser Glu Asp Glu Ser 660 665 670gag aca gaa gat tgaagagga gaaaagccaa gaggaccagg agcagaagag 2120Glu Thr Glu Asp 675gaagagaagg aagactcgcc agccagatgc caaagatggg gattcatacg acccctatga 2180cttcagtgac acagaggagg aaatgcctca agtacacact ccaaagacgg cagactcaca 2240ggagaccaag gaatcccaga aagtggagtt gagtgaatcc aggttgaagg cattcaaggt 2300ggccctcttg gatgtgttcc gggaagctca tgcgcagtca atcggcatga atcgcctcac 2360agaatccatc aaccgggaca gcaaagagcc cttctcttca gttgagatcc aggctgctct 2420gagcaagatg caggatgaca atcaggtcat ggtgtctgag ggcatcatct tcctcatctg 2480aggaggcctc gtctctgaac ttgggttgtg ccgagagagt ttgttctgtg tttcccaccc 2540tctccctgac ccaagtcttt gcctctactc ccttaacagt gttgaattca actgaaggcg 2600aggaatgttg gtgatgaagc tgagttcagg actcggtgga ccctttggga atgggtcatg 2660aaagctgcca tggggtgagg aaagaggaga cagtgggaga ggacaatgac tattgcatct 2720tcattgcaaa agcactggct catccgccct acttcccatc ccacacaaac ccaattgtaa 2780ataacatatg acttctgagt acttttgggg gcacaactgt tttctgtttg ctgttttttt 2840gttttgtttt ttttctccag agcactttgg tctagactag gctttgggtg gttccaattg 2900gtggagagaa gctctgaggc acgtcatgca ggtcaagaaa gctttctttg cagtagcacc 2960agttaaggtg aatatgtatt gtatcacaaa acaaacccaa tatccagatg aatatccgag 3020atgttgaata aacttagcca tttcgtacac atggaaaaga aaaaaaaaaa aaaaaaaaaa 3080aaaaaaaaaa a 3091<210> 2<211> 676<212> PRT<213> Homo sapiens<400> 2Met Ala Gly Thr Val Val Leu Asp Asp Val Glu Leu Arg Glu Ala Gln 1 5 10 15Arg Asp Tyr Leu Asp Phe Leu Asp Asp Glu Glu Asp Gln Gly Ile Tyr 20 25 30Gln Ser Lys Val Arg Glu Leu Ile Ser Asp Asn Gln Tyr Arg Leu Ile 35 40 45Val Asn Val Asn Asp Leu Arg Arg Lys Asn Glu Lys Arg Ala Asn Arg 50 55 60Leu Leu Asn Asn Ala Phe Glu Glu Leu Val Ala Phe Gln Arg Ala Leu 65 70 75 80Lys Asp Phe Val Ala Ser Ile Asp Ala Thr Tyr Ala Lys Gln Tyr Glu 85 90 95Glu Phe Tyr Val Gly Leu Glu Gly Ser Phe Gly Ser Lys His Val Ser 100 105 110Pro Arg Thr Leu Thr Ser Cys Phe Leu Ser Cys Val Val Cys Val Glu 115 120 125Gly Ile Val Thr Lys Cys Ser Leu Val Arg Pro Lys Val Val Arg Ser 130 135 140Val His Tyr Cys Pro Ala Thr Lys Lys Thr Ile Glu Arg Arg Tyr Ser145 150 155 160Asp Leu Thr Thr Leu Val Ala Phe Pro Ser Ser Ser Val Tyr Pro Thr 165 170 175Lys Asp Glu Glu Asn Asn Pro Leu Glu Thr Glu Tyr Gly Leu Ser Val 180 185 190Tyr Lys Asp His Gln Thr Ile Thr Ile Gln Glu Met Pro Glu Lys Ala 195 200 205Pro Ala Gly Gln Leu Pro Arg Ser Val Asp Val Ile Leu Asp Asp Asp 210 215 220Leu Val Asp Lys Ala Lys Pro Gly Asp Arg Val Gln Val Val Gly Thr225 230 235 240Tyr Arg Cys Leu Pro Gly Lys Lys Gly Gly Tyr Thr Ser Gly Thr Phe 245 250 255Arg Thr Val Leu Ile Ala Cys Asn Val Lys Gln Met Ser Lys Asp Ala 260 265 270Gln Pro Ser Phe Ser Ala Glu Asp Ile Ala Lys Ile Lys Lys Phe Ser 275 280 285Lys Thr Arg Ser Lys Asp Ile Phe Asp Gln Leu Ala Lys Ser Leu Ala 290 295 300Pro Ser Ile His Gly His Asp Tyr Val Lys Lys Ala Ile Leu Cys Leu305 310 315 320Leu Leu Gly Gly Val Glu Arg Asp Leu Glu Asn Gly Ser His Ile Arg 325 330 335Gly Asp Ile Asn Ile Leu Leu Ile Gly Asp Pro Ser Val Ala Lys Ser 340 345 350Gln Leu Leu Arg Tyr Val Leu Cys Thr Ala Pro Arg Ala Ile Pro Thr 355 360 365Thr Gly Arg Gly Ser Ser Gly Val Gly Leu Thr Ala Ala Val Thr Thr 370 375 380Asp Gln Glu Thr Gly Glu Arg Arg Leu Glu Ala Gly Ala Met Val Leu385 390 395 400Ala Asp Arg Gly Val Val Cys Ile Asp Glu Phe Asp Lys Met Ser Asp 405 410 415Met Asp Arg Thr Ala Ile His Glu Val Met Glu Gln Gly Arg Val Thr 420 425 430Ile Ala Lys Ala Gly Ile His Ala Arg Leu Asn Ala Arg Cys Ser Val 435 440 445Leu Ala Ala Ala Asn Pro Val Tyr Gly Arg Tyr Asp Gln Tyr Lys Thr 450 455 460Pro Met Glu Asn Ile Gly Leu Gln Asp Ser Leu Leu Ser Arg Phe Asp465 470 475 480Leu Leu Phe Ile Met Leu Asp Gln Met Asp Pro Glu Gln Asp Arg Glu 485 490 495Ile Ser Asp His Val Leu Arg Met His Arg Tyr Arg Ala Pro Gly Glu 500 505 510Gln Asp Gly Asp Ala Met Pro Leu Gly Ser Ala Val Asp Ile Leu Ala 515 520 525Thr Asp Asp Pro Asn Phe Ser Gln Glu Asp Gln Gln Asp Thr Gln Ile 530 535 540Tyr Glu Lys His Asp Asn Leu Leu His Gly Thr Lys Lys Lys Lys Glu545 550 555 560Lys Met Val Ser Ala Ala Phe Met Lys Lys Tyr Ile His Val Ala Lys 565 570 575Ile Ile Lys Pro Val Leu Thr Gln Glu Ser Ala Thr Tyr Ile Ala Glu 580 585 590Glu Tyr Ser Arg Leu Arg Ser Gln Asp Ser Met Ser Ser Asp Thr Ala 595 600 605Arg Thr Ser Pro Val Thr Ala Arg Thr Leu Glu Thr Leu Ile Arg Leu 610 615 620Ala Thr Ala His Ala Lys Ala Arg Met Ser Lys Thr Val Asp Leu Gln625 630 635 640Asp Ala Glu Glu Ala Val Glu Leu Val Gln Tyr Ala Tyr Phe Lys Lys 645 650 655Val Leu Glu Lys Glu Lys Lys Arg Lys Lys Arg Ser Glu Asp Glu Ser 660 665 670Glu Thr Glu Asp 675
Claims (13)
Translated fromKoreanPriority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1020070003463AKR100886670B1 (en) | 2007-01-11 | 2007-01-11 | New diagnostic markers and methods of papillary thyroid cancer |
| PCT/KR2008/000190WO2008085007A1 (en) | 2007-01-11 | 2008-01-11 | Novel proliferation marker in papillary thyroid carcinoma and novel method of diagnosis detecting the marker |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1020070003463AKR100886670B1 (en) | 2007-01-11 | 2007-01-11 | New diagnostic markers and methods of papillary thyroid cancer |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| KR20080066252A KR20080066252A (en) | 2008-07-16 |
| KR100886670B1true KR100886670B1 (en) | 2009-03-04 |
Family
ID=39608847
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| KR1020070003463AExpired - Fee RelatedKR100886670B1 (en) | 2007-01-11 | 2007-01-11 | New diagnostic markers and methods of papillary thyroid cancer |
Country Status (2)
| Country | Link |
|---|---|
| KR (1) | KR100886670B1 (en) |
| WO (1) | WO2008085007A1 (en) |
Families Citing this family (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101049583B1 (en)* | 2009-06-30 | 2011-07-14 | 한국과학기술연구원 | A marker for diagnosing papillary thyroid cancer containing 3-indole acetonitrile as an active ingredient |
| RU2485517C2 (en)* | 2011-12-13 | 2013-06-20 | Федеральное Государственное Бюджетное Учреждение "Московский Научно-Исследовательский Онкологический Институт Им. П.А. Герцена Министерства Здравоохранения И Социального Развития России" | Diagnostic technique for degree of thyroid carcinoma |
| WO2016198835A1 (en) | 2015-06-08 | 2016-12-15 | Arquer Diagnostics Limited | Methods and kits |
| JP6854246B2 (en) | 2015-06-08 | 2021-04-07 | アーケア ダイアグノスティクス リミテッド | How to analyze urine sample |
| IT201600069927A1 (en)* | 2016-07-05 | 2018-01-05 | Fondazione Univ Niccolò Cusano Per La Ricerca Medico Scientifica | Method for in vitro diagnosis of thyroid carcinoma. |
| CN118501477B (en)* | 2024-07-16 | 2024-10-11 | 浙江省肿瘤医院 | Radioactive iodine resistance thyroid cancer biomarker and screening method and application thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030225528A1 (en) | 2002-03-13 | 2003-12-04 | Baker Joffre B. | Gene expression profiling in biopsied tumor tissues |
| KR20070088979A (en)* | 2006-02-27 | 2007-08-30 | 사회복지법인 삼성생명공익재단 | Marker protein for cancer diagnosis, cancer diagnosis method and cancer diagnostic kit using the same |
- 2007
- 2007-01-11KRKR1020070003463Apatent/KR100886670B1/ennot_activeExpired - Fee Related
- 2008
- 2008-01-11WOPCT/KR2008/000190patent/WO2008085007A1/enactiveApplication Filing
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030225528A1 (en) | 2002-03-13 | 2003-12-04 | Baker Joffre B. | Gene expression profiling in biopsied tumor tissues |
| KR20070088979A (en)* | 2006-02-27 | 2007-08-30 | 사회복지법인 삼성생명공익재단 | Marker protein for cancer diagnosis, cancer diagnosis method and cancer diagnostic kit using the same |
Non-Patent Citations (1)
| Title |
|---|
| Genebank accession number NP_002379. |
Also Published As
| Publication number | Publication date |
|---|---|
| KR20080066252A (en) | 2008-07-16 |
| WO2008085007A1 (en) | 2008-07-17 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Lynch et al. | Human DNA topoisomerase II-alpha: a new marker of cell proliferation in invasive breast cancer | |
| US20220283168A1 (en) | Method for the detection of cancer cells by localization of peptides in nucleus as compared to cytoplasm | |
| Heidebrecht et al. | p100: a novel proliferation-associated nuclear protein specifically restricted to cell cycle phases S, G2, and M | |
| US20070053896A1 (en) | Diagnostic marker for ovarian cancer | |
| US8759004B2 (en) | Compounds and methods for detection of carcinomas and their precursor lesions | |
| US20070161064A1 (en) | Antibodies as a cancer diagnostic | |
| EP2021794B1 (en) | Use of protein s100a 12 as a marker for colorectal cancer | |
| KR100886670B1 (en) | New diagnostic markers and methods of papillary thyroid cancer | |
| EP2473621B1 (en) | Methods for detection of lethal cell and uses thereof | |
| WO2012141285A1 (en) | Biomarker for breast cancer | |
| JP4948728B2 (en) | Antibodies for cancer diagnosis | |
| US10126311B2 (en) | Methods and compositions for detecting endometrial or ovarian cancer | |
| US20210018504A1 (en) | Shon as a prognostic biomarker for cancer and as a predictor of response to endocrine therapy | |
| AU2017201499A1 (en) | Her3 protein SRM/MRM assay | |
| JP2020522501A (en) | Prediction method of cancer treatment results by T-DM1 | |
| EP1470239B1 (en) | Psoriasin expression by breast epithelial cells | |
| Frolova et al. | A shift from nuclear to cytoplasmic breast cancer metastasis suppressor 1 expression is associated with highly proliferative estrogen receptor-negative breast cancers | |
| KR101270761B1 (en) | Transaldolase, a marker for diagnosing radiation exposure, a composition for diagnosing radiation exposure to measure the level of expression of the marker, a kit for diagnosing radiation exposure comprising the composition, and a method for diagnosing radiation exposure using the marker | |
| EP1627230B1 (en) | Predicting cancer progression | |
| Göhring et al. | Prognostic value of cathepsin D in breast cancer: comparison of immunohistochemical and immunoradiometric detection methods. | |
| CA2567987A1 (en) | Use of protein pdx1 as a marker for breast cancer | |
| Migaldi et al. | p27Kip1 Expression and Survival in N0 Gastric Carcinoma | |
| JP2009535033A (en) | Compositions and methods for detection of Cripto-3 | |
| KR101594287B1 (en) | A kit for pancreatic cancer diagnosis comprising complememt factor b-specific binding antibody | |
| KR101403019B1 (en) | Novel biomarker for the diagnosis of lung cancer |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| A201 | Request for examination | ||
| PA0109 | Patent application | St.27 status event code:A-0-1-A10-A12-nap-PA0109 | |
| PA0201 | Request for examination | St.27 status event code:A-1-2-D10-D11-exm-PA0201 | |
| D13-X000 | Search requested | St.27 status event code:A-1-2-D10-D13-srh-X000 | |
| D14-X000 | Search report completed | St.27 status event code:A-1-2-D10-D14-srh-X000 | |
| E902 | Notification of reason for refusal | ||
| PE0902 | Notice of grounds for rejection | St.27 status event code:A-1-2-D10-D21-exm-PE0902 | |
| E13-X000 | Pre-grant limitation requested | St.27 status event code:A-2-3-E10-E13-lim-X000 | |
| P11-X000 | Amendment of application requested | St.27 status event code:A-2-2-P10-P11-nap-X000 | |
| P13-X000 | Application amended | St.27 status event code:A-2-2-P10-P13-nap-X000 | |
| E902 | Notification of reason for refusal | ||
| PE0902 | Notice of grounds for rejection | St.27 status event code:A-1-2-D10-D21-exm-PE0902 | |
| PG1501 | Laying open of application | St.27 status event code:A-1-1-Q10-Q12-nap-PG1501 | |
| T11-X000 | Administrative time limit extension requested | St.27 status event code:U-3-3-T10-T11-oth-X000 | |
| T11-X000 | Administrative time limit extension requested | St.27 status event code:U-3-3-T10-T11-oth-X000 | |
| T11-X000 | Administrative time limit extension requested | St.27 status event code:U-3-3-T10-T11-oth-X000 | |
| R17-X000 | Change to representative recorded | St.27 status event code:A-3-3-R10-R17-oth-X000 | |
| E13-X000 | Pre-grant limitation requested | St.27 status event code:A-2-3-E10-E13-lim-X000 | |
| P11-X000 | Amendment of application requested | St.27 status event code:A-2-2-P10-P11-nap-X000 | |
| P13-X000 | Application amended | St.27 status event code:A-2-2-P10-P13-nap-X000 | |
| E701 | Decision to grant or registration of patent right | ||
| PE0701 | Decision of registration | St.27 status event code:A-1-2-D10-D22-exm-PE0701 | |
| GRNT | Written decision to grant | ||
| PR0701 | Registration of establishment | St.27 status event code:A-2-4-F10-F11-exm-PR0701 | |
| PR1002 | Payment of registration fee | St.27 status event code:A-2-2-U10-U11-oth-PR1002 Fee payment year number:1 | |
| PG1601 | Publication of registration | St.27 status event code:A-4-4-Q10-Q13-nap-PG1601 | |
| FPAY | Annual fee payment | Payment date:20120323 Year of fee payment:4 | |
| PR1001 | Payment of annual fee | St.27 status event code:A-4-4-U10-U11-oth-PR1001 Fee payment year number:4 | |
| FPAY | Annual fee payment | Payment date:20130813 Year of fee payment:5 | |
| PR1001 | Payment of annual fee | St.27 status event code:A-4-4-U10-U11-oth-PR1001 Fee payment year number:5 | |
| LAPS | Lapse due to unpaid annual fee | ||
| PC1903 | Unpaid annual fee | St.27 status event code:A-4-4-U10-U13-oth-PC1903 Not in force date:20140226 Payment event data comment text:Termination Category : DEFAULT_OF_REGISTRATION_FEE | |
| PC1903 | Unpaid annual fee | St.27 status event code:N-4-6-H10-H13-oth-PC1903 Ip right cessation event data comment text:Termination Category : DEFAULT_OF_REGISTRATION_FEE Not in force date:20140226 | |
| PN2301 | Change of applicant | St.27 status event code:A-5-5-R10-R13-asn-PN2301 St.27 status event code:A-5-5-R10-R11-asn-PN2301 |