长期生存的起点NSCLC诊断分子病理与组织病理并驾齐驱课件

1、长期生存的起点NSCLC诊断分子病理与组织病理并驾齐驱肺癌病理学诊断的演变: 分子病理诊断日渐重要WHO将定义了恶性肿瘤病理诊断的三大元素：(1) 解剖学部位；(2)组织学类型；(3)恶性程度分级WHO公布的第一版肺和胸膜肿瘤的组织学病理分型WHO公布的第二版肺和胸膜肿瘤的组织学病理分型1952年1967年1981年2011年2004年2001年2013年WHO首次联合IASLC公布的第三版肺和胸膜肿瘤的组织学病理分型，首次提出了肺癌分子病理的概念WHO更新了肺癌的病理学分型，并首次探讨了肺癌分子基因组学，但主要此时分子基因学还是从属于组织学诊断由于EGFR-TKIs，贝伐单抗等新靶向药物的问

2、世，IASLC肺腺癌新分类。分子诊断的作用已从辅助诊断肺癌跨越到帮助治疗决策。并首次在病理学诊断的流程中纳入了EGFR突变IASLCCAPAMP公布了EGFR&ALK检测指南，明确了腺癌患者需检测EGFRALK基因。首次给予分子病理与组织病理同等重要的地位病理诊断与临床治疗决策：小细胞癌&非小细胞癌预后：小细胞癌病情进展迅速，预后差；非小细胞肺癌相对生长较慢，预后相对好小细胞肺癌的标准化疗方案（EP）与非小细胞肺癌的化疗方案（三代新药+铂）不同小细胞肺癌与非小细胞肺癌的放疗原则不同病理诊断与临床治疗决策：鳞癌与非鳞癌鳞癌患者PFS（月）非鳞癌患者PFS（月）PFS率PFS率JMDB研究：培美曲

3、塞 /顺铂一线对照吉西他滨/顺铂治疗晚期NSCLC的前瞻性、随机、双盲、多中心III期研究Scagliotti GV,et al. J Clin Oncol.2008 Jul 20;26(21):3543-51.IASLC/ATS/ERS肺腺癌新分类Journal of Thoracic Oncology 2011 (February);6:24485IASLC腺癌新分类，首次提出肺腺癌需行EGFR检测Journal of Thoracic Oncology 2011 (February);6:24485Large Cell Carcinoma (LCC)New WHO Classificat

4、ion WHO 2004 Proposed WHO 2014/2015Variant LCCsLarge cell neuroendocrinecarcinoma (LCNEC)Baseloid carcinomaLymphoepithelloma-likecarcinoma Clear cell carcinomaLCC with rhabdoid phenotypeNeuroendocrine tumorsSquamous cell carcinoma Others and unclassified tumorsRemoved (cytology features)New WHO Lung

5、 Cancer ClassificationSquamous and Large Cell CarcinomasMajor changes were made to the classification of these lung tumors:Squamous cell carcinoma was divided in Keratinizating (“classic” histology) and non-keratiniziting (based on IHC phenotype). Basaloid carcinoma is regarded as a squamous cell tu

6、mor. Other variants of squamous cell carcinoma were removed as subtypes.Large cell carcinoma categories were reduced; this is a diagnosis of exclusion , using both histology and IHC.Tumor protein expression by IHC plays an important role on the new classification of these tumors.To elucidate the phe

7、notypic characteristics of lung cancer tumors is crucial for proper molecular testing decisions.Changing histopathology picture of lung cancer?P63-P63+P63+TTF-1+ADCADCADCTTF-1+ADCADC2SQCCTTF-1-ADC1INDETSQCCTTF-1p63(-)(+)(+)p63p63(-)(+)(+)ADC(10%)Indeterminate(1%ADC,1%SQCC)CK5/6+ SQCCSQCC(96%)(+/-)(+

8、)(any)ADC (6%)SQCC(3%)ADC(84%)New Terms The cribriform pattern - ? New pattern“Pseudosqumaoid” solid pattern TTF-1P63CK5/6分子病理诊断与临床治疗决策？UnknownKRASEGFRALKHER2BRAFPIK3CAAKT1MAP2K1NRASROS1RETOther11%Adenoca55%Squamous34%Histology-based SubtypingNSCLCas onediseaseAdenocarcinomaSquamous-cellFGFR1AmpUnkn

9、ownEGFRvIIIP13KCAEGFRDDR2Li T. J Clin Oncol.2013 (adapted from Pao W. Lancet Oncology.2011;12:175-80.)NSCLC亚型正在从组织学分型向分子分型转换SCLCRB1RLF-MYCL1MYCL1MYCNMYCFGFR1SOX2SquamousCarcinomaNFE2L2TP63NOTCH1KEAP1TP53CDKN2APIK3CAPTENAdenocarcinomaEGFRKRASERBB2BRAFALK fusionsROS1 fusionsRET fusionsSTK11UnknownKRAS

10、EGFRALKfusionsHER2BRAFPIK3CAAKT1NRASMAP2K1ROS1 fusionsKIF58-RETLung AdenocarcinomaPao W. Nat Med.2012;18:349-51.Sos M. Oncogene.2012;331:4811-4.肺癌的分子基因谱EGFR突变与ALK融合基因，靶向药物疗效显著IPASS: EGFR mutation positiveGefitinib (n=132)Carboplatin / paclitaxel (n=129)048121620240.00.20.40.60.81.0Probability of pro

11、gression-free survivalMonthsPROFILE 1007: ALK fusion positiveHR (95% CI) = 0.48 (0.36, 0.64) p0.0001No. events gefitinib, 97 (73.5%)No. events C / P, 111 (86.0%)ALK阳性NSCLC具有鲜明的临床及病理特征特征EGFR突变EML4/ALK组织学腺癌 TTF1+腺癌TTF1+亚型非粘液型粘液型吸烟状态不吸烟不吸烟人种东亚所有人种发病年龄66岁52岁性别女性无差异Shaw AT, et al. J Clin Oncol 2009; 27:4

12、247-4253.85.4%的ALK阳性NSCLC年龄65岁(PROFILE1007)Frequency (%)East AsianNon-Asian肺鳞癌患者中，EGFR突变比例低Adapted from Lindeman et al, J Thorac Oncol 2013 Jul;8(7):823-59 不同NSCLC组织学类型中ALK的发生率Number of Tumors TestedEstimated Prevalence Rate (% tested)NSCLC: non-small cell lung cancer; ADC: adenocarcinoma; SCC: squa

13、mous cell carcinoma; ADSC: adenosquamous carcinomaAdapted from IASLC ATLAS of ALK Testing in Lung Cancer , Chapter 2, 2013EGFR突变检测平台及其敏感性Adapted from Pao W, Ladanyi M. Clin Cancer Res 2007;13:495455MethodSensitivityMutations IdentifiedDirect sequencing25%Known and newPCR-SSCP10%Known and newTaqMan P

14、CR10%Known onlyLoop-hybrid mobility shift assay7.5%Known onlyCycleave PCR5%Known onlyPCR-RLFP (fragment length analysis)5%Known onlyMassARRAY genotyping5%Known onlyLNA-PCR clamp1%Known onlyScorpion ARMS (DxS)1%Known onlydHPLC1%Known onlyCOLD-TaqMan PCR0.05%Known onlyParallel (Next Generation) Sequen

15、cing0.01%Known and UnknownSSCP, single-strand conformation polymorphism; RLFP, restriction fragment length polymorphism; LNA, locked nucleic acid; ARMS, Amplification Refractory Mutation System; dHPLC, denaturing high performance liquid chromatographyTesting Methods Available for ALK Rearranged Lung

16、 CancersFluorescent in situ hybridization (FISH)Chromogenic in situ hybridization (CISH)Immunohistochemistry (IHC, Ventana IHC)RT-PCRNanoStringRT-qPCRNext generation sequencingEGFR、ALK检测国内外指南对照适宜人群诊断方法中国EGFR与ALK阳性NSCLC诊断及治疗指南（2014）所有含腺癌成分的NSCLCFISH, 经权威机构批准的IHC与RT-PCR，其余IHC可做为筛选CSCO中国专家共识所有含腺癌成分的NSC

17、LC，富集人群可优先检测FISH, Ventana IHC, RT-PCR, 常规IHC可做初筛ATLAS ALK Testing Guideline所有含腺癌成分的NSCLCFISH, Ventana IHC与FISH吻合度高CAP/AMP/IASLC EGFR&ALK detection guideline所有含腺癌成分的NSCLCFISH, 常规IHC可作为初筛CFDA（中国国家药监局）Ventana IHCAmoy RT-PCRNSCLC分子病理诊断与治疗决策密切相关HER2EGFR mutantsALKROS/RETb-rafK-rasChemotherapy and PCIAden

18、oLCC/NOSSCCSCLCK-ras大部分(70%)亚裔肺腺癌患者可检测到驱动基因1.Seo JS, et al. Genome Res 2012; 22:2109-2119.基因突变靶向药物亚裔患者的发生率1EGFR突变erlotinib,gefitinib,afatinib60.5%ALK重排crizotinib4.0%HER2突变trastuzumab,afatinib2%BRAF突变vemurafenib,dabrafenib1.0%MET扩增crizotinib1%ROS1融合crizotinib1.5%RET融合cabozantinib1%组织 or 细胞学标本的诊断流程鳞癌组

19、织or细胞学标本（活检，胸腔积液，细胞学检查）腺癌非小细胞小细胞NSCLC, NOSNSCLC ?LCNEC神经内分泌标记：CD56，嗜铬粒蛋白，突触素典型组织 or 细胞学标本的诊断方法鳞癌阳性肿瘤（活检，胸腔积液，细胞学检查）腺癌非小细胞小细胞NSCLCNSCLC ?LCNEC神经内分泌标记：CD56，嗜铬粒蛋白，突触素典型NSCLC 倾向鳞癌NSCLC倾向腺癌P63+Mucin+, TTF1+NSCLCNOS? 腺癌鳞癌标记 -veTTF1+ p63+ 粘蛋白&免疫组化标记EGFR和ALK检测鳞癌阳性肿瘤（活检，胸腔积液，细胞学检查）腺癌非小细胞小细胞NSCLCNSCLC ?LCNEC神

20、经内分泌标记：CD56，嗜铬粒蛋白，突触素典型NSCLC 倾向鳞癌NSCLC 倾向腺癌P63+Mucin+, TTF1+NSCLCNOS? 腺癌鳞癌标记 -veTTF1+ p63+ 粘蛋白&免疫组化标记对于NSCLC患者来说：组织病理区分腺鳞癌可有助于分子病理诊断的精准亚裔肺腺癌患者60%-70%的驱动基因已知，分子病理对于临床治疗的指导意义更大总结组织病理诊断=分子病理诊断NSCLCWhats the problem? I gave you at least 10 cells!Fine Needle Aspiration (FNA)Core Needle Biopsy (CNB)Advanc

21、ed Tumor“THE TISSUE IS STILL THE ISSUE”谢谢Frequency (%)East AsianNon-Asian东亚、女性、非吸烟、腺癌患者，EGFR突变比例高Adapted from Lindeman et al, J Thorac Oncol 2013 Jul;8(7):823-59 V1E13;A20E20;A20E20ins18;A20E6;A20E6ins33;A20E14;ins11del49A20T3;A20KI24;A20KI15;A20E6;A19E2;A20E2;ins117A20E3;ins69A20E14;del12A20E14;del

22、36A20E17;ins30A20E17ins61;ins34A20E15del19;del20A20E18;A20V2V3aV3bV4V5bV5aV7V6V8aV8b“V5”“V4”E17ins68;A20EML4-ALKKIF5B-ALKTFG-ALKKI17;A20ALK-PTPN3KLC1-ALKKL9;A20ALK Kinase domainOu et al., Oncologist 2012; 2012;17:1351-75非小细胞肺癌中ALK融合基因变异免疫组织化学检测ALK融合蛋白 优势: 可用于常规检测、快速、信价比高IASLC ATLAS of ALK Testing in Lung Cancer , Chapter 4, 2013生物学基础：ALK mRNA正常状态下，仅微量存在与小肠大脑等，肺部未检出；未发生ALK融合的肺癌，ALK蛋白几乎不表达抗体患者数IHC阴性IHC阳性IHC结果可疑（结果不明确)FISH-FISH+FISH-FI