新一代分子診斷技術(shù)在腫瘤精準醫(yī)學中的應(yīng)用

1、精準醫(yī)學（腫瘤領(lǐng)域）檢測方案設(shè)計和質(zhì)控精準醫(yī)學（腫瘤領(lǐng)域）檢測方案設(shè)計和質(zhì)控 基于新一代測序（基于新一代測序（NGS）的腫瘤精準醫(yī)學檢測產(chǎn)品設(shè)計）的腫瘤精準醫(yī)學檢測產(chǎn)品設(shè)計 全球多中心數(shù)據(jù)整合和數(shù)據(jù)質(zhì)控全球多中心數(shù)據(jù)整合和數(shù)據(jù)質(zhì)控基于基于iFISH-CTC技術(shù)的單細胞測序臨床應(yīng)用技術(shù)的單細胞測序臨床應(yīng)用 腫瘤早期檢測腫瘤早期檢測 腫瘤個體化診療方案腫瘤個體化診療方案整合國際腫瘤數(shù)據(jù)庫的基因整合國際腫瘤數(shù)據(jù)庫的基因Panel設(shè)計與應(yīng)用設(shè)計與應(yīng)用 超高深度基因測序超高深度基因測序Panel的設(shè)計的設(shè)計 腫瘤負荷、繼發(fā)耐藥監(jiān)控腫瘤負荷、繼發(fā)耐藥監(jiān)控精準醫(yī)學（腫瘤領(lǐng)域）檢測方案設(shè)計和質(zhì)控精準醫(yī)學（腫瘤

2、領(lǐng)域）檢測方案設(shè)計和質(zhì)控 基于新一代測序（基于新一代測序（NGS）的腫瘤精）的腫瘤精準醫(yī)學檢測產(chǎn)品設(shè)計準醫(yī)學檢測產(chǎn)品設(shè)計 全球多中心數(shù)據(jù)整合和數(shù)據(jù)質(zhì)控全球多中心數(shù)據(jù)整合和數(shù)據(jù)質(zhì)控新一代測序平臺互補融合性新一代測序平臺互補融合性 不同測序儀平臺可以使用不同基因富集方法 PCR 捕獲 PCR 捕獲IonIonIlluminaIllumina基因變異類型檢測基因變異類型檢測 I：突變：突變 基因全長（全外顯子）的所有點突變 影響因素：基因富集覆蓋度（90%） 影響因素：測序深度（200X） 報告中的突變 % 靈敏度：1%臨床研究發(fā)現(xiàn)癌組織中臨床研究發(fā)現(xiàn)癌組織中CNV與與Mutation共存共存腫瘤分

3、型新概念腫瘤分型新概念（C型腫瘤，型腫瘤，M型腫瘤）型腫瘤）2015年年CSCO年會廈門年會廈門 基因變異類型檢測基因變異類型檢測 II：拷貝數(shù)變異：拷貝數(shù)變異（Copy Number Variation, CNV）基因變異類型檢測基因變異類型檢測 III：融合基因：融合基因 NGS檢測融合基因 RNA轉(zhuǎn)錄混合法NCI-MATCH Sample Test -Oncomine Comprehensive Panel腫瘤發(fā)病機制學說腫瘤發(fā)病機制學說 腫瘤的器官發(fā)生學說 腫瘤的血液發(fā)生學說 腫瘤的泛感染發(fā)生學說泛感染發(fā)生學說及診療進展 50例接受纈更昔洛韋（Valcyte）治療的膠質(zhì)母細胞瘤患者2年

4、生存率為62%，對照組為18%（P0.001）。治療周期長的亞組預(yù)后更好，2年生存率可達70%和90% 上海xx醫(yī)院使用xxx抗病毒藥物治療xxx瘤獲得突破性進展N Engl J Med 2013 Sep 5;369(10):985-68基于基于iFISH-CTC技術(shù)的單細胞測序臨床應(yīng)用技術(shù)的單細胞測序臨床應(yīng)用腫瘤早期檢測腫瘤早期檢測 腫瘤個體化診療方案腫瘤個體化診療方案iFISH-CTC技術(shù)技術(shù)助力腫瘤精準醫(yī)學研究助力腫瘤精準醫(yī)學研究腫瘤早期腫瘤早期iFISH-CTC檢測數(shù)據(jù)檢測數(shù)據(jù)（2014.1-2015.8） 5363例統(tǒng)計癌種癌種靈敏度靈敏度乳腺癌 95.93%結(jié)直腸癌93.18%肝癌

5、81%胰腺癌 97.73%肺癌91.67%胃癌92.59%甲狀腺癌87%食管癌81.8%2015年年CSCO年會廈門年會廈門 癌癥早期篩查受檢者信息檢測路徑病理檢查病理檢查結(jié)果：低分化腺癌結(jié)果：低分化腺癌手術(shù)切除腺癌手術(shù)切除腺癌12腫瘤早期綜合檢測及預(yù)防案例腫瘤早期綜合檢測及預(yù)防案例 -福建福建x州州xx醫(yī)院醫(yī)院循環(huán)腫瘤細胞檢測循環(huán)腫瘤細胞檢測體內(nèi)存在腫瘤細胞提示癌組織特征提示癌組織特征及用藥指導及用藥指導發(fā)現(xiàn) 22 個腫瘤細胞影像學顯示右肺彌影像學顯示右肺彌漫狀陰影漫狀陰影（纖維化（纖維化/癌變）癌變）激光顯微切割激光顯微切割單個單個CTC腫瘤細腫瘤細胞胞單細胞全基因組單細胞全基因組測序分析

6、測序分析12345定期隨訪和復(fù)查定期隨訪和復(fù)查6癌標癌標NSE ：72.6（ 99%每份樣本測序深度每份樣本測序深度10000XPhase 1 results 1,262 HCC related somatic mutations from original file (3,000,381 somatic mutations in total, for all phenotypes). 425 non-redundant HCC related somatic mutations, with unique genomic coordinates. These 425 mutations res

7、ide in 36 exons (covering 28 genes). Results would also be shown in figures later.Phase 1 results The covered 28 genes are (by official symbol): AACS,ADAM22,AKAP13,ARVCF,ATR,CDKN2A,CLCC1,CUL3,DMGDH,EGFR,ERBB2,GABBR1,GIF,GTPBP2,HNF1A,JAK2,LITAF,MET,MFSD8,MUC5B,MYCL,NOVA1,NTRK2,PCDH7,PLEKHG5,PSTK,RAF1

8、,USP25. OR (by ensembl ID): ENSG00000008277.10,ENSG00000036257.8,ENSG00000081760.12,ENSG00000096968.8,ENSG00000099889.9,ENSG00000105976.10,ENSG00000116990.9,ENSG00000117983.13,ENSG00000121940.11,ENSG00000132155.7,ENSG00000132837.10,ENSG00000134812.3,ENSG00000135100.13,ENSG00000139910.15,ENSG00000141

9、736.9,ENSG00000146648.11,ENSG00000147889.12,ENSG00000148053.11,ENSG00000155313.11,ENSG00000164073.5,ENSG00000169851.11,ENSG00000170776.15,ENSG00000171680.16,ENSG00000172432.14,ENSG00000175054.10,ENSG00000179988.9,ENSG00000189067.8,ENSG00000204681.6Phase 1 Results These 36 exons covered 12,415bp in t

10、otal, including: chr1 2061bp chr2 97bp chr3 201bp chr4 3218bp chr5 249bp chr6 322bp chr7 621bp chr8 0bp chr9 1986bp chr10 282bpchr11 350bpchr12 540bpchr13 0bpchr14 1400bpchr15 136bpchr16 157bpchr17 342bpchr18 0bpchr19 0bpchr20 0bpchr21 325bpchr22 128bpchrX 0bpchrY 0bp6-phase selectorMethod descripti

11、on:Phase 2: max coverage “For each exon with SNVs covering 5 patients in LUAD and SCC, we selected the exon with highest RI that identified at least 1 new patient when compared to the prior phase.” original paperData required: TCGA maf files (somatic mutation)6-phase selectorMethod description:Phase

12、 3: RI = 30 & =3 patients “For each remaining exon with an RI 30 and with SNVs covering 3 patients in LUAD and SCC, we identified the exon that would result in the largest reduction in patients with only 1 SNV.” original paperData required: TCGA maf files (somatic mutation)6-phase selectorMethod

13、 description:Phase 4: RI = 20 & = 3 patients “Same procedure as phase 3, but using RI 20.” original paperData required: TCGA maf files (somatic mutation)6-phase selectorMethod description:Phase 5: Predicted drivers “We included all exons from additional genes previously predicted to harbor drive

14、r mutations in NSCLC.” original paperData required: predicted driver gene in HCC (from publication, 15 genes in reference paper)6-phase selectorMethod description:Phase 6: Add fusions “For recurrent rearrangements in NSCLC involving the receptor tyrosine kinases ALK, ROS1, and RET, the introns most

15、frequently implicated in the fusion event and the flanking exons were included.” original paperNot included (not under our consideration by far).Results This panel includes: 604 genes (not necessarily have all exons of this gene included in panel) 782 exons 120.1 kb in total, 154 bp per exon 202 patients in total 199 patients covered by this panel 199 (99%), 181 (90%), 158 (78%) patients with at least 1, 2, 3 SNVs Mean RI is 22.67 (Recurrence Index, the higher this number th