TWI516766B - A method for grouping glioblastoma multiforme - Google Patents
A method for grouping glioblastoma multiforme Download PDFInfo
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本發明係關於一種神經膠質瘤之診斷指標，特別是關於一種神經膠質瘤之分子指標，及其用於臨床分子診斷之用途，以及一種神經膠質瘤之分群判斷方法。 The present invention relates to a diagnostic index of glioma, in particular to a molecular index of glioma, and its use for clinical molecular diagnosis, and a method for judging the glioma.
神經膠質瘤(gliomas)係常見的腦部腫瘤，根據WHO分級制度，神經膠質瘤可分為毛狀星狀細胞瘤(pilocytic astrocytoma，簡稱PA，WHO分級為I)、星狀細胞瘤(astrocytoma，WHO分級為Ⅱ)、寡樹突膠質瘤(oligo-dendroglioma，WHO分級為Ⅱ)、退行性星狀細胞瘤(anaplastic astrocytoma，簡稱AA，WHO分級為Ⅲ)、寡樹突膠質瘤(anaplastic oligodendroglioma，WHO分級為Ⅲ)及多形性神經膠母細胞瘤(glioblastoma multiforme，簡稱GBM，WHO分級為Ⅳ)。毛狀星狀細胞瘤為良性腫瘤，而寡樹突膠質瘤、退行性寡樹突膠質瘤、星狀細胞瘤、退行性星狀細胞瘤及多形性神經膠母細胞瘤係屬於惡性腫瘤，惡性腫瘤患者的預後情形較差，其中50%的退行性星狀細胞瘤或多形性神經膠母細胞瘤患者，其存活時間僅有9~12個月。 Gliomas are common brain tumors. According to the WHO classification system, gliomas can be divided into pilocytic astrocytoma (PA, WHO classification I) and astrocytoma (astrocytoma). WHO classification is II), oligo-dendroglioma (WHO classification II), degenerative astrocytoma (AA, WHO classification III), anaplastic oligodendroglioma (anaplastic oligodendroglioma, The WHO classification is III) and glioblastoma multiforme (GBM, WHO classification is IV). Hairy astrocytoma is a benign tumor, while oligodendroglioma, degenerative oligodendroglioma, astrocytoma, degenerative astrocytoma and pleomorphic glioblastoma are malignant tumors. The prognosis of patients with malignant tumors is poor, and 50% of patients with degenerative astrocytoma or glioblastoma multiforme have a survival time of only 9 to 12 months.
一旦懷疑患者可能患有腦瘤，醫師會藉由顱骨X光檢查、電腦斷層掃描(computed tomography，簡稱CT)、磁振造影(magnetic resonance imaging，簡稱MRI)或正子斷層攝影(positron emission tomography，簡稱PET)等技 術診斷顱內腫瘤，再以神經外科手術摘取患者的腫瘤組織檢體，並對該腫瘤組織檢體進行病理學檢驗，判斷該腫瘤之病理型態與腫瘤惡性程度，以提供臨床醫師選擇適當的治療方式。 Once the patient is suspected of having a brain tumor, the physician will use a skull X-ray, computed tomography (CT), magnetic resonance imaging (MRI) or positron emission tomography (referred to as positron emission tomography). PET) Diagnosis of intracranial tumors, and then taking the tumor tissue of the patient by neurosurgery, and performing pathological examination on the tumor tissue to determine the pathological type and tumor malignancy of the tumor, so as to provide the clinician with appropriate choice. The treatment.
舉例而言，腫瘤組織檢體若判斷為良性腫瘤，通常可藉由手術完全切除，不需進行放射線或藥物化學治療，僅需定期以CT或MRI進行複檢，追蹤是否有復發情形即可；若判斷為惡性腫瘤，除了以手術切除該惡性腫瘤，尚需選擇適當的藥物進行治療，以提高患者的存活率及預後存活時間。 For example, if a tumor tissue is judged to be a benign tumor, it can usually be completely resected by surgery without radiotherapy or medicinal chemotherapy. It is only necessary to perform regular re-examination with CT or MRI to track whether there is a recurrence; If it is judged to be a malignant tumor, in addition to surgical removal of the malignant tumor, it is necessary to select an appropriate drug for treatment to improve the survival rate of the patient and the survival time of the prognosis.
由此可知，快速且正確判斷患者的腫瘤是否為惡性，對於患者的預後及術後的治療方式係為重要關鍵。 It can be seen that the rapid and correct determination of whether the patient's tumor is malignant is important for the prognosis of the patient and the post-operative treatment.
除了根據組織病理學判斷該腫瘤組織檢體為惡性或良性腫瘤族群外，以組織中特定的生物標記作為判斷腫瘤分群，係目前判斷病理組織檢體方法中最常被使用且準確度較高的，常用的生物標記包括MDM2及EGFR的異常增幅表現，CDKN2A(p16)及INK4A/ARF2的缺失，CDK4、TP53、RB或PTEN(MMAC1)之突變。此外，過度表現c-Met、上皮細胞生長因子受體(epidermal growth factor receptor，簡稱EGFR)及血小板衍生生長因子受體(platelet-derived growth factor receptor，簡稱PDGFR)也會促進神經膠質瘤之腫瘤存活及其血管新生。 In addition to judging that the tumor tissue is a malignant or benign tumor population according to histopathology, the specific biomarkers in the tissue are used to judge the tumor group, which is currently the most frequently used and accurate method for judging the pathological tissue method. Commonly used biomarkers include abnormal amplification of MDM2 and EGFR, deletion of CDKN2A (p16) and INK4A/ARF2, and mutation of CDK4, TP53, RB or PTEN (MMAC1). In addition, overexpression of c-Met, epidermal growth factor receptor (EGFR) and platelet-derived growth factor receptor (PDGFR) also promote tumor survival in gliomas. And its angiogenesis.
按，中華民國公開第200831536號專利案，揭示一種診斷神經膠質瘤之方法，必須先取得病理組織檢體與健康組織檢體，並且分別偵測各組織檢體的PIK3R3多肽表現 量，以該病理組織檢體之PIK3R3多肽表現量大於該健康組織檢體之PIK3R3多肽表現量，作為判斷該病理組織檢體為惡性腫瘤的依據。 According to the Patent No. 200831536 of the Republic of China, a method for diagnosing gliomas is disclosed. It is necessary to obtain a pathological tissue and a healthy tissue sample, and respectively detect the PIK3R3 polypeptide expression of each tissue sample. The amount of the PIK3R3 polypeptide expressed by the pathological tissue sample is greater than the expression amount of the PIK3R3 polypeptide of the healthy tissue sample, and is used as a basis for judging that the pathological tissue sample is a malignant tumor.
然而，該習用診斷神經膠質瘤之方法，係以統計分析之方法分別計算該病理組織檢體與該健康組織檢體之PIK3R3多肽表現量，再依據所得之計算值進行腫瘤分級判斷，然，統計分析之方法會因p值設定不同(如p<0.05或p<0.01)而影響判斷結果，又，該PIK3R3多肽表現量亦無法用於做為預測患者的預後存活分析工具。 However, the method for diagnosing glioma is to calculate the expression of the PIK3R3 polypeptide of the pathological tissue and the healthy tissue by statistical analysis, and then perform the tumor grading judgment according to the calculated value, and then, statistics The method of analysis will affect the judgment result due to different p- value settings (such as p <0.05 or p <0.01). Moreover, the PIK3R3 polypeptide expression cannot be used as a prognostic survival analysis tool for predicting patients.
由此可知，習用技術缺乏一具有高指標性及靈敏度並可應用於臨床神經膠質瘤分群及預後分析之工具，因此，有必要針對習用技術加以改進，以準確判斷神經膠質瘤為惡性族群及其預後狀況而改善其高致死率。 It can be seen that the conventional technology lacks a tool with high index and sensitivity and can be applied to clinical glioma grouping and prognosis analysis. Therefore, it is necessary to improve the conventional techniques to accurately judge gliomas as malignant groups and The prognosis status improves its high mortality rate.
本發明之主要目的係提供一種神經膠質瘤之分群診斷方法，係藉由一高指標性及靈敏度的神經膠質瘤之分子指標，準確判斷神經膠質瘤之分群，以提高惡性神經膠質瘤的診斷準確率。 The main object of the present invention is to provide a method for diagnosing glioma by accurately determining the glioma group by a molecular index of high index and sensitivity glioma to improve the diagnosis of malignant glioma. Confirmation rate.
為達到前述發明目的，本發明所運用之技術內容包含有：一種神經膠質瘤之分群判斷方法，係包含：一HDGF測定步驟，係採檢一腦部的腫瘤組織檢體，測量該腫瘤組織檢體中，具有HDGF蛋白質表現之細胞個數百分率，獲得一HDGF數值；及一判斷步驟，該HDGF數值大於50%， 判斷該腫瘤組織檢體屬於惡性族群。 In order to achieve the foregoing object, the technical content of the present invention includes: a method for judging a glioma group, comprising: an HDGF measuring step, which is to take a tumor tissue of a brain and measure the tumor tissue test. In the body, the cells having the HDGF protein expression have a percentage of hundreds of HDGF values; and in a judging step, the HDGF value is greater than 50%. It is judged that the tumor tissue specimen belongs to a malignant group.
其中，該HDGF數值為一細胞核內HDGF數值，係指該神經膠質瘤的細胞核中，有表現HDGF蛋白質之細胞核數目，佔該腫瘤組織檢體之總細胞核數目之百分率。 Wherein, the HDGF value is a nuclear nucleus HDGF value, which refers to the number of nuclei representing the HDGF protein in the nucleus of the glioma, and the percentage of the total number of nuclei of the tumor tissue sample.
其中，該判斷步驟中，該HDGF數值小於20%，判斷該腫瘤組織檢體為良性族群。 Wherein, in the determining step, the HDGF value is less than 20%, and the tumor tissue is determined to be a benign group.
其中，該HDGF測定步驟之前另包含一鑑定步驟，係根據組織型態或病理學鑑定該腫瘤組織檢體為多形性神經膠母細胞瘤，其中該判斷步驟中，該HDGF數值大於50%，則係判斷該腫瘤組織檢體之所屬患者的存活時間小於7個月。 Wherein, the HDGF measuring step further comprises an identifying step of identifying the tumor tissue sample as a pleomorphic glioblastoma according to a tissue type or pathology, wherein the HDGF value is greater than 50% in the determining step. Then, it is determined that the survival time of the patient to which the tumor tissue sample belongs is less than 7 months.
為讓本發明之上述及其他目的、特徵及優點能更明顯易懂，下文特舉本發明之較佳實施例，並配合所附圖式，作詳細說明如下： The above and other objects, features and advantages of the present invention will become more <RTIgt;
本發明所述之「腫瘤分群」，係依照病理組織型態及腫瘤分子標記鑑定為良性族群及惡性族群，其中，該良性族群係包含正常(健康)細胞或WHO分級為I之腫瘤細胞；該惡性族群係包含WHO分級為Ⅱ、Ⅲ或Ⅳ之腫瘤細胞。 The "tumor grouping" according to the present invention is identified as a benign group and a malignant group according to a pathological tissue type and a tumor molecular marker, wherein the benign group includes normal (healthy) cells or tumor cells of WHO class I; The malignant group contains tumor cells with WHO classification of II, III or IV.
本發明係分析神經膠質瘤患者之腫瘤組織中的肝癌衍生生長因子(hepatoma-derived growth factor，簡稱HDGF)表現程度，特別係相對於該腫瘤組織檢體之總細胞核個數中有表現HDGF蛋白質的細胞程度，作為神經膠 質瘤患者之惡性腫瘤及其預後存活時間的判斷依據，本發明以HDGF作為神經膠質瘤分群之判斷指標，係針對臨床神經膠質瘤患者之腫瘤分群及預後狀況具有高度指標性及靈敏度，因此，可進一步應用於臨床神經膠質瘤之分子診斷，並提供一種神經膠質瘤之分群判斷方法，以提高神經膠質瘤之診斷率，並改善神經膠質瘤之高致死率。 The present invention relates to the analysis of the degree of expression of hepato-derived growth factor (HDGF) in the tumor tissue of a glioma patient, in particular, the expression of HDGF protein in the total number of nuclei of the tumor tissue sample. Cell degree, as a nerve glue The malignant tumor of the tumor of the tumor and the judgment of the survival time of the prognosis, the HDGF is used as the judgment index of the glioma group, and has high index and sensitivity for the tumor group and the prognosis of the clinical glioma patients. It can be further applied to the molecular diagnosis of clinical glioma, and provides a method for judging the glioma to improve the diagnosis rate of glioma and improve the high mortality rate of glioma.
請參照第1圖所示，本發明之神經膠質瘤之分群判斷方法，包含一HDGF測定步驟S11及一判斷步驟S12。 Referring to Fig. 1, the method for judging the glioma of the present invention comprises an HDGF measuring step S11 and a determining step S12.
該HDGF測定步驟S11，係採檢一腦部的腫瘤組織檢體，測量該腫瘤組織檢體中，具有HDGF蛋白質表現之細胞個數百分率，獲得一HDGF數值。更詳言之，本實施例係取一腦部腫瘤組織檢體，利用一免疫組織化學(immunohistochemistry)染色法偵測該腫瘤組織中HDGF蛋白質的表現程度。 In the HDGF measurement step S11, a tumor tissue sample of a brain is collected, and the number of cells having HDGF protein expression in the tumor tissue sample is measured to obtain a HDGF value. More specifically, in this embodiment, a brain tumor tissue sample is taken, and the degree of expression of HDGF protein in the tumor tissue is detected by an immunohistochemistry staining method.
舉例而言，本實施例以一親和性染料(Hematoxylin)進行複染法(counterstain)，標記該腫瘤組織檢體中的細胞核，以計算於該腫瘤組織檢體中一單位面積下的細胞個數，再利用一HDGF抗體標定該腫瘤組織檢體中的HDGF蛋白質，計算該腫瘤組織檢體中，有表現該HDGF蛋白質之細胞個數，獲得一細胞核內的HDGF數值(labeling intensity of nuclear HDGF，簡稱LIN值)，再依據該LIN值之高低，對該腫瘤組織檢體進行腫瘤分群；本實施例另提供一健康組織檢體作為對照組，同樣以複染法及該HDGF抗體分別標定該健康組織檢體之細胞核HDGF蛋白質，該健康組織檢體之細胞核及細胞質中幾乎沒有HDGF 蛋白質之表現(或者HDGF蛋白質表現量低於5%)。 For example, in this embodiment, counterstaining is performed by an affinity dye (Hematoxylin), and the nuclei in the tumor tissue are labeled to calculate the number of cells per unit area in the tumor tissue sample. The HDGF protein in the tumor tissue is calibrated by using an HDGF antibody, and the number of cells expressing the HDGF protein in the tumor tissue is calculated, and the HDGF value in a nucleus is obtained (labeling intensity of nuclear HDGF, referred to as According to the LIN value, the tumor tissue is subjected to tumor grouping according to the height of the LIN value; in this embodiment, a healthy tissue sample is further provided as a control group, and the healthy tissue is also separately labeled by the counterstaining method and the HDGF antibody. The nuclear HDGF protein of the sample, and there is almost no HDGF in the nucleus and cytoplasm of the healthy tissue sample. The performance of the protein (or HDGF protein performance is less than 5%).
舉例而言，該LIN值係以百分率標記之：於一固定放大的顯微視野下，計算該腫瘤組織檢體中細胞核之數目(nuclear-staining counting，簡稱NC)，及標記有HDGF之細胞核的數目(HDGF-staining in nuclear counting，簡稱HNC)，並根據下列公式I計算，獲得該腫瘤組織檢體之LIN值。 For example, the LIN value is labeled as a percentage: the number of nuclei in the tumor tissue (nuclear-staining counting, NC) and the nuclei labeled with HDGF are calculated under a fixed magnification microscopic field of view. HDGF-staining in nuclear counting (HNC), and calculated according to the following formula I, the LIN value of the tumor tissue sample is obtained.
該判斷步驟S12，該HDGF數值大於50%，判斷該腫瘤組織檢體屬於惡性族群。更詳言之，由於正常(健康)的腦部組織中，不會表現HDGF；相對於惡性族群而言，該細胞核內HDGF數值小於20%，則判斷該腫瘤組織檢體為良性族群。因此，藉由測定組織檢體中HDGF的存在，作為準確判斷且高靈敏度的腫瘤組織檢體分群的判斷標準。 In the determining step S12, the HDGF value is greater than 50%, and it is determined that the tumor tissue sample belongs to a malignant group. More specifically, HDGF is not expressed in normal (healthy) brain tissue; compared to the malignant population, the HDGF value in the nucleus is less than 20%, and the tumor tissue is judged to be a benign group. Therefore, by measuring the presence of HDGF in a tissue sample, it is a criterion for judging the tumor tissue sample group which is accurately judged and highly sensitive.
請參照第2圖，為本發明神經膠質瘤之分群判斷方法的第二實施例，係於該HDGF測定步驟S11前，進行一鑑定步驟S10，本實施例之其餘步驟與第一實施例相同，在此容不贅述。 Referring to FIG. 2, a second embodiment of the method for judging the glioma of the present invention is performed before the HDGF measurement step S11, and an identification step S10 is performed. The remaining steps of the embodiment are the same as the first embodiment. I will not repeat them here.
該鑑定步驟S10，係根據組織型態或病理學鑑定該腫瘤組織檢體之種類為多形性神經膠母細胞瘤，並經前述該判斷步驟S12確認該HDGF數值大於50%，則判斷該腫瘤組織檢體之所屬患者的存活時間小於7個月。 In the identification step S10, the type of the tumor tissue sample is identified as a pleomorphic glioblastoma according to the tissue type or pathology, and the HDGF value is greater than 50% after the determining step S12, and the tumor is judged. The survival time of the patient to which the tissue sample belongs is less than 7 months.
為證實本發明神經膠質瘤之分群判斷方法，確實能夠 藉由一腫瘤組織中HDGF蛋白質表現百分率大於50%，作為惡性腫瘤及預後存活時間的判斷依據，本實施例係取一篩檢族群作為本發明神經膠質瘤之分群判斷方法的驗證，該篩檢族群係來自高雄地區榮民總醫院的105名神經膠質瘤患者的腫瘤組織檢體。 In order to confirm the method for judging the glioma of the present invention, it is indeed possible to Since the percentage of HDGF protein expression in a tumor tissue is greater than 50%, as a basis for judging the survival time of the malignant tumor and the prognosis, this embodiment adopts a screening population as a verification method for the method of judging the glioma of the present invention, and the screening is performed. The ethnic group is a tumor tissue from 105 glioma patients from the Veterans General Hospital in Kaohsiung.
本實施例取得該篩檢族群之腫瘤組織檢體及5件正常(健康)組織檢體，該等檢體係以石蠟包埋後保存以備用，再進行本發明之HDGF測定步驟S11前，先將該等檢體進行一去蠟程序(deparaffinization)，再以一HDGF抗體(1：200 dilution)標定該檢體之HDGF，用以生產該HDGF抗體之融合瘤細胞株，係寄存於中華民國新竹食品科學工業發展研究所，其寄存編號為BCRC 960444，本實施例係選擇但不限定以一商用偵測套組(NovolinkTM polymer detection system,Leica,USA)偵測該HDGF抗體之標記。 In this embodiment, the tumor tissue sample of the screening group and five normal (health) tissue samples are obtained, and the detection system is stored in paraffin and stored for use, and before the HDGF measurement step S11 of the present invention is performed, The specimens were subjected to a deparaffinization, and the HDGF of the specimen was calibrated with an HDGF antibody (1:200 dilution) to produce a fusion tumor cell line of the HDGF antibody, which was deposited in the Republic of China Hsinchu Food. industrial development Research Institute, which is a register number BCRC 960444, selection of the present embodiment is not limited to detecting a commercial kit (Novolink TM polymer detection system, Leica , USA) detecting the labeled antibody HDGF.
本實施例將組織檢體以該HDGF抗體標記後，以包含有HRP(Horseradish Peroxidase)之二抗對該HDGF抗體進行顯色，再以Hematoxylin對細胞核進行複染，並於放大倍率為200倍之顯微視野下，分別計算該腫瘤組織檢體之細胞個數及友表現HDGF蛋白質之細胞個數，即該細胞核內的HDGF數值(LIN值)。本發明篩檢族群之所有腫瘤組織檢體中，不論是何種種類之腫瘤皆偵測到HDGF之表現。 In this example, after the tissue sample is labeled with the HDGF antibody, the HDGF antibody is colored with a secondary antibody containing HRP (Horseradish Peroxidase), and the nuclei are counterstained with Hematoxylin at a magnification of 200 times. Under the microscopic field of view, the number of cells in the tumor tissue and the number of cells expressing the HDGF protein, that is, the HDGF value (LIN value) in the nucleus, were calculated. In all tumor tissue samples of the screening population of the present invention, the expression of HDGF was detected regardless of the type of tumor.
請參照第3至8圖所示，係本發明篩檢族群中，不同種類的腫瘤組織檢體免疫染色顯微照片，其中，第3圖為腦部正常(健康)組織，第4圖為毛狀星狀細胞瘤，第5 圖為寡樹突膠質瘤，第6圖為星狀細胞瘤，第7圖為術後存活時間大於10個月的多形性神經膠母細胞瘤，第8圖為存活時間小於10個月的多形性神經膠母細胞瘤組織切片結果；各照片之放大倍率皆為100倍；比較第3至8圖可知，該HDGF蛋白質表現量確實與神經膠質瘤的生成具有正相關性。 Please refer to Figures 3 to 8 for the immunostaining micrographs of different types of tumor tissue in the screening population of the present invention, wherein Figure 3 is the normal (healthy) tissue of the brain, and Figure 4 is the hair of the brain. Astrocytoma, 5th The picture shows oligodendroglioma, Figure 6 is astrocytoma, Figure 7 is a polymorphic glioblastoma with a survival time of more than 10 months, and Figure 8 is a survival time of less than 10 months. The results of tissue section of pleomorphic glioblastoma; the magnification of each photograph was 100 times; comparing the figures 3 to 8, the HDGF protein expression was indeed positively correlated with the generation of glioma.
本發明篩檢族群中，各腫瘤組織檢體之臨床病理資料及其LIN值係如第1表所示，本實施例係利用統計分析的方式，如cox迴歸分析(cox regression analysis)進行統計。其中，第A1至A6組分別為不同腫瘤種類之組織檢體，其中，第A1組的正常組織檢體，其LIN值為1%；屬於良性族群的第A2及A3組，其LIN值為20%以下；相較於惡性族群的第A4至A6組，其LIN值約為50%以上。 In the screening population of the present invention, the clinicopathological data and the LIN value of each tumor tissue specimen are shown in Table 1, and the present example is statistically analyzed, such as cox regression analysis. Among them, groups A1 to A6 are tissue samples of different tumor types, wherein the normal tissue of group A1 has a LIN value of 1%; in groups A2 and A3 of a benign group, the LIN value is 20 % or less; the LIN value is about 50% or more compared to the group A4 to A6 of the malignant group.
由此可知，該腫瘤組織檢體之LIN值大於50%，則代表該腫瘤組織檢體屬於惡性族群，即該組織檢體於WHO分級中屬於Ⅱ、Ⅲ或Ⅳ者；反之，LIN值小於20%者，則代表該腫瘤組織檢體屬於良性族群。而該腫瘤組織檢體之LIN值介於20~50%之間，則表示該腫瘤組織檢體為惡性族群的可能性高。 It can be seen that the LIN value of the tumor tissue sample is greater than 50%, which means that the tumor tissue sample belongs to a malignant group, that is, the tissue sample belongs to II, III or IV in the WHO classification; otherwise, the LIN value is less than 20 %, it means that the tumor tissue is a benign group. The LIN value of the tumor tissue sample is between 20% and 50%, indicating that the tumor tissue is highly likely to be a malignant group.
請參照第D1及D2組，係追蹤39名多形性神經膠母細胞瘤患者之存活率，其中該LIN值大於50%者，代表該惡性族群檢體之預後存活率係小於10個月。 Please refer to groups D1 and D2 to track the survival rate of 39 patients with polymorphic glioblastoma, wherein the LIN value is greater than 50%, indicating that the prognosis survival rate of the malignant group is less than 10 months.
請參照第2表所示，本實施例係取41名多形性神經膠母細胞瘤患者之腫瘤組織檢體，以寄存於中華民國新竹食品科學工業發展研究所之HDGF單株抗體(寄存編號為BCRC 960444)進行組織免疫染色分析，獲得該LIN值，接著再以接受特徵曲線(receiver operating characteristic curve，又稱ROC曲線)分析，可將病患分為高LIN值表現群組(即LIN>50%)與低LIN值表現群組(LIN≦50%)，此兩大群組再與其臨床病理資料(存活率)進行比對，推得該41名患者存活率分析。 Please refer to the second table. In this example, 41 tumor tissues of patients with pleomorphic glioblastoma were collected from HDGF monoclonal antibody deposited in the Hsinchu Food Science and Industrial Development Research Institute of the Republic of China. Tissue immunostaining analysis for BCRC 960444), obtaining the LIN value, and then analyzing the receiver operating characteristic curve (ROC curve), the patient can be divided into high LIN value group (ie LIN> 50%) and low LIN value performance group (LIN ≦ 50%), the two groups were compared with their clinical pathology data (survival rate), and the 41 patients were analyzed for survival.
請參照第9圖所示，係根據第2表之數值，以Kaplan-Meier法(p<0.001)分析各患者存活率，顯示LIN值高於50%者，確實能夠代表該患者的預後存活率(特別係存活率低於7個月)，相對而言，LIN值低於50%者，可以預估其存活時間能夠達17個月以上；此外，由第2表亦可知，年齡高於65歲之多形性神經膠母細胞瘤患者之存活時間亦較短。 Please refer to Figure 9 for the survival rate of each patient by Kaplan-Meier method ( p < 0.001) according to the value of Table 2. It shows that the LIN value is higher than 50%, which can represent the prognosis survival rate of the patient. (Specially, the survival rate is less than 7 months). Relatively speaking, if the LIN value is less than 50%, the survival time can be estimated to be more than 17 months. In addition, it can be known from the second table that the age is higher than 65. The survival time of patients with pleomorphic glioblastoma is also shorter.
由此可得知，LIN值對於多形性神經膠母細胞瘤的預後存活率預測具有良好的專一性及靈敏度，具有應用於臨床多形性神經膠母細胞瘤檢測之潛力。 It can be seen that the LIN value has good specificity and sensitivity for the prediction of prognosis survival rate of polymorphic glioblastoma, and has the potential to be applied to the detection of clinical polymorphic glioblastoma.
綜上所述，本發明係利用腫瘤組織檢體中於細胞核內之HDGF蛋白質表現量(LIN值)，判斷該腫瘤組織檢體為惡性族群或良性族群之分群，特別係該腫瘤組織檢體於細胞核內之HDGF表現量，對於臨床神經膠質瘤患者的惡性腫瘤分群及預後狀況具有高度指標性及靈敏度，因此，可應用於臨床神經膠質瘤分子診斷之開發，以提升神經膠質瘤之診斷率，並且改善神經膠質瘤之高致死率。 In summary, the present invention utilizes the HDGF protein expression (LIN value) in the nucleus of the tumor tissue sample to determine whether the tumor tissue sample is a malignant group or a benign group, especially the tumor tissue sample. The amount of HDGF in the nucleus is highly indicative and sensitive to the malignant tumor population and prognosis of clinical glioma patients. Therefore, it can be applied to the development of clinical molecular diagnosis of glioma to improve the diagnosis rate of glioma. And improve the high mortality rate of glioma.
本發明神經膠質瘤之分子標記，乃係利用免疫組織化學染色分析法，綜合本發明之臨床病理資料，經由嚴謹的統計方式計算而獲得，該HDGF於細胞核內的蛋白質表現 量與神經膠質瘤的惡性腫瘤分群及預後存活時間具有高度相關性，HDGF對於臨床神經膠質瘤患者的惡性腫瘤分群及預後判斷具有高度指標性、專一性以及良好的靈敏度之功效。 The molecular marker of the glioma of the present invention is obtained by immunohistochemical staining analysis method, synthesizing the clinicopathological data of the present invention, and obtaining the protein expression of the HDGF in the nucleus by rigorous statistical calculation. The amount is highly correlated with the malignant tumor group and prognosis survival time of glioma. HDGF has high index, specificity and good sensitivity for the malignant tumor grouping and prognosis of clinical glioma patients.
本發明之神經膠質瘤之分群判斷方法，係藉由腫瘤組織檢體中細胞內的HDGF蛋白質表現量作為惡性腫瘤分群之判斷依據，特別係以一腫瘤組織檢體之細胞內HDGF蛋白質表現百分率大於50%，作為明確界定其為惡性腫瘤之分群指標，具有提高該惡性神經膠質瘤之診斷準確率之功效。 The method for judging the glioma of the present invention is based on the expression of HDGF protein in the tumor tissue as a basis for judging the malignant tumor group, in particular, the percentage of intracellular HDGF protein expression in a tumor tissue sample is greater than 50%, as a clustering index that clearly defines it as a malignant tumor, has the effect of improving the diagnostic accuracy of the malignant glioma.
本發明神經膠質瘤之分子標記可應用於臨床神經膠質瘤新興檢測方法的開發，作為神經膠質瘤惡性腫瘤分群及預後存活的評估指標，可藉由偵測患者檢體中HDGF之表現程度，作為神經膠質瘤預後狀況之判斷依據，並且建立一個具有高專一性、靈敏度，並且可廣泛應用於臨床神經膠質瘤早期診斷及預後分析之工具，改善神經膠質瘤之高致死率，為本發明之功效。 The molecular marker of the glioma of the invention can be applied to the development of an emerging detection method for clinical glioma, and as an evaluation index of glioma malignant tumor group and prognosis survival, the degree of HDGF expression in the patient sample can be detected as The basis for judging the prognosis of glioma, and establishing a tool with high specificity and sensitivity, which can be widely used in the early diagnosis and prognosis analysis of clinical glioma, improve the high lethal rate of glioma, and is the efficacy of the present invention. .
雖然本發明已利用上述較佳實施例揭示，然其並非用以限定本發明，任何熟習此技藝者在不脫離本發明之精神和範圍之內，相對上述實施例進行各種更動與修改仍屬本發明所保護之技術範疇，因此本發明之保護範圍當視後附之申請專利範圍所界定者為準。 While the invention has been described in connection with the preferred embodiments described above, it is not intended to limit the scope of the invention. The technical scope of the invention is protected, and therefore the scope of the invention is defined by the scope of the appended claims.
S10‧‧‧鑑定步驟 S10‧‧‧ Identification steps
S11‧‧‧HDGF測定步驟 S11‧‧‧HDGF determination steps
S12‧‧‧判斷步驟 S12‧‧‧ judgment steps
第1圖：本發明神經膠質瘤之分群判斷方法的步驟方塊圖(一)。 Fig. 1 is a block diagram (1) of the method for judging the glioma of the present invention.
第2圖：本發明神經膠質瘤之分群判斷方法的步驟方塊圖(二)。 Fig. 2 is a block diagram (2) of the method for judging the glioma of the present invention.
第3圖本實施例之腦部正常組織切片顯微照片。 Fig. 3 is a photomicrograph of a normal tissue section of the brain of the present embodiment.
第4圖：本實施例之毛狀星狀細胞瘤組織切片顯微照片。 Figure 4: Micrograph of tissue sections of hairy astrocytoma in this example.
第5圖：本實施例之寡樹突膠質瘤組織切片顯微照片。 Figure 5: Micrograph of oligodendroglioma tissue sections of this example.
第6圖：本實施例之星狀細胞瘤組織切片顯微照片。 Fig. 6: Micrograph of histomorphology of astrocytoma tumor of this example.
第7圖：本實施例之多形性神經膠母細胞瘤組織切片顯微照片(存活時間>10個月者)。 Figure 7: Micrograph of tissue sections of polymorphic glioblastoma of the present example (survival time > 10 months).
第8圖：本實施例之多形性神經膠母細胞瘤組織切片顯微照片(存活時間<10個月者)。 Figure 8: Micrograph of tissue sections of the polymorphic glioblastoma of this example (survival time < 10 months).
第9圖：本實施例LIN值為50%與GBM患者存活率之折線關係圖。 Figure 9: The relationship between the LIN value of 50% of this example and the survival rate of GBM patients.
S11‧‧‧HDGF測定步驟 S11‧‧‧HDGF determination steps
S12‧‧‧判斷步驟 S12‧‧‧ judgment steps
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