常用检测技术
在过去的十年里，各种新技术、新方法被广泛应用于病理学，使其得到了快速地发展，其中最具代表性的就是二代测序（NGS）。尽管这些技术优势显著，但是在病理科日常诊断中的应用普及程度并不高，更多是被分子病理学家应用到了与临床联系不太紧密的科学研究中。目前，病理科所采用的主要的分子检测手段还是IHC。
自18世纪晚期卵巢切除术被报道以来，乳腺癌是第一个运用了靶向治疗手段的恶性肿瘤{1}，Anti-HER2单克隆抗体治疗始于20世纪90年代，它代表了人类第一次在实体肿瘤中使用靶向药物{2} ，高侵袭性的基底样型乳腺癌是目前靶向治疗研究的重点{3}。乳腺癌也是第一个建立了分子分型的肿瘤{4}，并且已经被多种方法重复验证，肿瘤分型的主要意义在于每种亚型都会对应不同的临床指征及与之相对应的治疗方案{5-7}。在乳腺癌分子分型确立后的这15年中，虽然可用于乳腺癌分子分型的检测技术已经很多，但是IHC仍然是目前应用最广泛的技术手段{8}。IHC仅仅通过对ER、PR和HER2的染色，就可以为临床医生提供可靠的、可重复的乳腺癌临床预测性信息。IHC对Ki-67染色可以确定官腔型乳腺癌是否需要化疗{9}。基底样型乳腺癌在CK5/6和EGFR两个IHC指标上显示出了比单纯三阴型乳腺癌更高的特异性，这点对遗传性乳腺癌和卵巢癌筛查具有一定的帮助{10-11}。总之，IHC的6个指标就能够将一个复杂的看似没有头绪的乳腺癌做到精确的分子分型，基于这种检测方法的快速、准确及低成本，为广大病理工作者提供替代分子研究一种技术手段。
在乳腺癌分子分型确立之后，也陆续报道了其他的肿瘤的分子分型。通过对癌症基因组图谱分析发现：胃癌可以分成4个分子亚型（EB病毒阳性型、微卫星不稳定（MSI）型、基因组稳定型以及染色体不稳定型）{12}。虽然这些分子分型是采用全面分子属性得出，但是与临床密切相关的治疗效果和遗传性影响等指标可以应用以下蛋白标记物及方法得以重复获得（1）错配修复蛋白(MMR)的IHC检测。（2）HER2的IHC、FISH检测。（3）E-cadherin的 IHC检测。（4）EB病毒原位杂交检测（基于FFPE和形态学）{13-18}。所有这些检测在大多数的病理科都可开展。同样地，子宫内膜癌也被视为一种可以根据癌症基因图谱来进行临床相关分子分型的对象{19}。子宫内膜癌可以分为：高拷贝基因型（浆液样型）、低拷贝基因型（子宫内膜样型）、 高变异型（MMR缺失）、超变异型（聚合酶「DNA-directed」-ε；POLE）等四型，前三种可以通过应用p53和MMR的IHC染色来重复检测{20-23}。错配修复蛋白（MMR）的IHC染色不仅可以实现对胃癌和子宫内膜癌的分子分型，也是Lynch综合征（遗传性肿瘤）筛查中的重要指标{24-26}， 辅助性的IHC检测与以往的临床检测手段相比，更具优势性{27-28}。这个策略是改善Lynch相关肿瘤早期指标监测的关键（统计基于每年美国＞3000例Lynch相关直肠癌症患者）{29}。MMR的IHC检测不仅有助于于Lynch综合征的筛查，也可以大量节省直接基因测序的成本和时间{30}。此外，MMR在化疗和靶向治疗效果预测方面的应用前景也值得期待{31-32}。
通过MMR及其他IHC指标的检测可以辅助和指导肿瘤相关易感基因测序。胃肠间质瘤（GISTs）就是个很好的例子，这些肿瘤是由KIT突变驱动的{33-34}，对这些基因测序可以帮助指导靶向治疗和鉴别遗传性胃肠间质瘤{35}，相比直接基因测序，IHC也可以通过对KIT（CD117），DOG 1和 SDHB等蛋白的检测来进行筛选，并且能同时节省成本和时间{36}。通过以上例子说明，IHC可以同时应用于遗传性疾病筛查，指导基因测序研究以及直接指导治疗方案的确定。
IHC也被广泛地应用于预测基因改变方面。在乳腺癌和胃癌中，IHC检测的HER2过度
表达，高度预测潜在ERBB2扩增。在肺癌和其他恶性癌症肿瘤中，IHC可以检测异常ALK、的表达，因为基因ALK1的重排会导致重组蛋白的表达{37-38}。
最近，单克隆抗体制备工艺的提高也使得IHC检测特定基因突变成为可能。针对异柠檬酸脱氢酶1（IDH1）蛋白里R132H突变的抗体，已被广泛应用于神经胶质瘤的诊断中{39}。
参考文献：
1、Beatson GT. On the treatment of inoperable cases of carcinoma of the mamma: suggestions for a new method of treatment, with illustrative cases. Lancet 1896;148(3803):162–165.
2、Pegram MD, Lipton A, Hayes DF, et al. Phase II study of receptor-enhanced chemosensitivity using recombinant humanized anti-p185HER2/neu monoclonal antibody plus cisplatin in patients with HER2/new-overxpressing metastatic breast cancer refractory to chemotherapy treatment. J Clin Oncol. 1998;16(8): 2659–2671.
3、Park Y, Moryama A, Kithara T, Yoshida Y, Urita T, Kato R. Triple-negative breast cancer and poly(ADP-ribose) polymerase inhibitors. Anticancer Agents Med Chem. 2012;12(6):672–677.
4、Perou CM, Srlie T, Eisen MB, et al. Molecular portraits of human breast tumours. Nature. 2000;406(6797):747–752.
5、Parker JS, Mullins M, Cheang MC, et al. Supervised risk predictor of breast cancer based on intrinsic subtypes. J Clin Oncol. 2009;27(8):1160–1167.
6、Srlie T, Perou CM, Tibshirani R, et al. Gene expression patterns of breast carcinomas distinguish tumor subclasses with clinical implications. Proc Natl Acad Sci U S A. 2001;98(19):10869–10874.
7、Chia SK, Bramwell VH, Tu D, et al. A 50-gene intrinsic subtype classifier for prognosis and prediction of benefit from adjuvant tamoxifen. Clin Cancer Res. 2012;18(16):4465–4472
8、Kittaneh M, Montero AJ, Gluck¨ S. Molecular profiling for breast cancer: a comprehensive review. Biomark Cancer. 2013;5(1):61–70.
9、Polley MY, Leung SC, Gao D, et al. An international study to increase concordance in Ki67 scoring. Mod Pathol. 2015;18(6):778–786.
10、Lakhani SR, Reis-Filho JS, Fulford L, et al; Breast Cancer Linkage Consortium. Prediction of BRCA1 status in patients with breast cancer using estrogen receptor and basal phenotype. Clin Cancer Res. 2005;11(14):5175– 5180.
11、Won JR, Gao D, Chow C, et al. A survey of immunohistochemical biomarkers for basal-like breast cancer against a gene expression profile gold standard. Mod Pathol. 2013;26(11):1438–1450.
12、Bass AJ, Thorsson V, Shmulevich I, et al; for Cancer Genome Atlas Research Network. Comprehensive molecular characterization of gastric adenocarcinoma [published correction online September 10, 2014]. Nature. 2014;513(7517):202–209.
13、Takeichi M. Cadherin cell adhesion receptors as a morphogenetic regulator. Science. 1992;251(5000):1451–1455.
14、Hansford S, Kaurah P, Li-Chang H, et al. Hereditary diffuse gastric cancer syndrome: CDH1 mutations and beyond [published correction appears in JAMA Oncol. 2015;1(1):110]. JAMA Oncol. 2015;1(1):23–32.
15、Sheffield BS, Garratt J, Kalloger SE, et al. HER2/neu testing in gastric cancer by immunohistochemistry: assessment of interlaboratory variation. Arch Pathol Lab Med. 2014;138(11):1495–1502.
16、Bang YJ, Van Cutsem E, Feyereislova A, et al; ToGA Trial Investigators. Trastuzumab in combination with chemotherapy versus chemotherapy alone for treatment of HER2-positive advanced gastric or gastro-oesophageal junction cancer (ToGA): a phase 3, open-label, randomized controlled trial. Lancet. 2010; 376(9742):687–697.
17、Halling KC, Harper J, Moskaluk CA, et al. Origin of microasatellite instability in gastric cancer. Am J Pathol. 1999;155(1):205–211.
18、Shibata D, Weiss LM. Epstein-Barr virus-associated gastric adenocarcino-ma. Am J Pathol. 1992;140(4):769–774.
19、Kandoth C, Schultz N, Cherniack AD, et al; Cancer Genome Atlas Research Network. Integrated genomic characterization of endometrial carcino-ma. Nature. 2013;497(7447):67–73.
20、Garg K, Leitao MM Jr, Wynveen CA, et al. p53 overexpression in morphologically ambiguous endometrial carcinomas correlates with adverse clinical outcomes. Mod Pathol. 2010;23(1):80–92.
21、Chiaravalli AM, Furlan D, Facco C, et al. Immunohistochemical pattern of hMSH2/hMLH1 in familial and sporadJournal style is to include both the percentage and the numbers that make up that percentage in all proportions. In the sentence that begins, astrocytosis, please provide the percentage, where indicated. ic colorectal, gastric, endometrial and ovarian carcinomas with instability in microsatellite sequences. Virchows Arch. 2001;438(1):39–48.
22、Bakhsh S, Kinloch M, Hoang LN, et al. Histopathological features of endometrial carcinomas associated with POLE mutations: implications for decisions about adjuvant therapy. Histopathology. 2016;68(6):916–924.
23、Hoang LN, McConechy MK, Kobel M, et al. Polymerase epsilon exonuclease domain mutations in ovarian endometrioid carcinoma. Int J Gynecol Cancer. 2015;25(7):1187–1193.
24、Lynch HT, Lynch PM. Clinical selection of candidates for mutational testing for cancer susceptibility. Oncology. 2006;20(1):29–34.
25、Jin M, Hampel H, Zhou X, Schunemann L, Yearsley M, Frankel WL. BRAV V600E mutation analysis simplifies the testing algorithm for Lynch syndrome. Am J Clin Pathol. 2013;140(2):177–183.
26、Cui D, Cao D, Yang Y, Qui M, Huang Y, Yi C. Effect of BRAF V600E mutation on tumor response of anti-EGFR monoclonal antibodies for first-line metastatic colorectal cancer treatment: a meta-analysis of randomized studies. Mol Biol Rep. 2014;41(3):1291–1298.
27、Vasen HFA, Mecklin JP, Meera Khan P, Lynch HT. The International Collaborative Group on Hereditary Non-Polyposis Colorectal Cancer (ICG-HNPCC). Dis Colon Rectum. 1991;34(1):424–425.
28、Umar A, Boland CR, Terdiman JP, et al. Revised Bethesda Guidelines for hereditary nonpolyposis colorectal cancer (Lynch syndrome) and microsatellite instability. J Natl Cancer Inst. 2004;96(4):261–268.
29、American Cancer Society. Cancer Facts & Figures 2013. Atlanta, GA: American Cancer Society; 2013.
30、Caldes´ T, Godino J, Sanchez A, et al. Immunohistochemistry and microsatellite instability testing for selecting MLH1, MSH2 and MSH6 mutation carriers in hereditary non-polyposis colorectal cancer. Oncol Rep. 2004;12(3): 621–629.
31、Riazy M, Kalloger SE, Sheffield BS, et al. Mismatch repair status may predict response to adjuvant chemotherapy in resectable pancreatic ductal adenocarcinoma. Mod Pathol. 2015;28(10):1383–1389.
32、Le DT, Uram JN, Wang H, et al. PD-1 blockade in tumors with mismatch-repair deficiency. N Eng J Med. 2015;372(26):2509–2520.
33、Hirota S, Isozaki K, Moriyama Y, et al. Gain-of-function mutations of c-kit in human gastrointestinal stromal tumors. Science. 1998;279(5350):577–580.
34、Heinrich MC, Corless CL, Duensing A, et al. PDGFRA activating mutations in gastrointestinal stromal tumors. Science. 2003;299(5607):708–710.
35、Valsangkar N, Sehdev A, Misra S, Zimmers TA, O’Neil BH, Koniaris LG. Current management of gastrointestinal stromal tumors: surgery, current biomarkers, mutations, and therapy. Surgery. 2015;158(5):1149–1164.
36、Gill AJ, Chou A, Vilain R, et al. Immunohistochemistry for SDHB divides gastrointestinal tumors (GISTs) into 2 distinct types. Am J Surg Pathol. 2010;24(5): 636–644.
37、Cutz JC, Craddock KJ, Torlakovic E, et al. Canadian anaplastic lymphoma kinase study: a model for multicenter standardization and optimization of ALK testing in lung cancer. J Thorac Oncol. 2014;9(9):1255–1263.
38、Sholl LM, Sun H, Butaney M, et al. ROS1 immunohistochemistry for detection of ROS1-rearranged lung adenocarcinomas. Am J Surg Pathol. 2013; 37(9):1441–1449.
39、Camelo-Piragua S, Jansen M, Ganguly A, et al. A sensitive and specific diagnostic panel to distinguish diffuse astrocytoma from astrocytosis: chromo-some 7 gain with mutant isocitrate dehydrogenase 1 and p53. J Neuropathol Exp Neurol. 2011;70(2):110–115.
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