Circumventing intratumoral heterogeneity to identify potential therapeutic targets in hepatocellular carcinoma.
Identifying target genetic mutations in hepatocellular carcinoma (HCC) for therapy is made challenging by intratumoral heterogeneity. Circulating cell-free DNAs (cfDNA) may contain a more complete mutational spectrum compared to a single tumor sample. This study aimed to identify the most efficient strategy to identify all the mutations within heterogeneous HCCs.



Whole exome sequencing (WES) and targeted deep sequencing (TDS) were carried out in 32 multi-regional tumor samples from five patients. Matched preoperative cfDNAs were sequenced accordingly. Intratumoral heterogeneity was measured using the average percentage of non-ubiquitous mutations (present in parts of tumor regions). Profiling efficiencies of single tumor specimen and cfDNA were compared. The strategy with the highest performance was used to screen for actionable mutations.



Variable levels of heterogeneity with branched and parallel evolution patterns were observed. The heterogeneity decreased at higher sequencing depth of TDS compared to measurements by WES (28.1% vs. 34.9%, p<0.01) but remained unchanged when additional samples were analyzed. TDS of single tumor specimen identified an average of 70% of the total mutations from multi-regional tissues. Although genome profiling efficiency of cfDNA increased with sequencing depth, an average of 47.2% total mutations were identified using TDS, suggesting that tissue samples outperformed it. TDS of single tumor specimen in 66 patients and cfDNAs in four unresectable HCCs showed that 38.6% (26/66 and 1/4) of patients carried mutations that were potential therapeutic targets.



TDS of single tumor specimen could identify actionable mutations targets for therapy in HCC. cfDNA may serve as secondary alternative in profiling HCC genome.



Targeted deep sequencing of single tumor specimen is a more efficient method to identify mutations in hepatocellular carcinoma made from mixed subtypes compared to circulating cell-free DNA in blood. cfDNA may serve as secondary alternative in profiling HCC genome. Identifying mutations may help clinicians choose targeted therapy for better individual treatments.
Liver Surgery Department, Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion (Fudan University), Ministry of Education, Shanghai 200032, China.
Ao
A
Huang
BGI-Shenzhen, Shenzhen 518083, China; China National Genebank-Shenzhen, BGI-Shenzhen, Shenzhen 518083, China.
Xin
X
Zhao
Liver Surgery Department, Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion (Fudan University), Ministry of Education, Shanghai 200032, China.
Xin-Rong
XR
Yang
BGI-Shenzhen, Shenzhen 518083, China; China National Genebank-Shenzhen, BGI-Shenzhen, Shenzhen 518083, China.
Fu-Qiang
FQ
Li
BGI-Shenzhen, Shenzhen 518083, China; China National Genebank-Shenzhen, BGI-Shenzhen, Shenzhen 518083, China.
Xin-Lan
XL
Zhou
BGI-Shenzhen, Shenzhen 518083, China; China National Genebank-Shenzhen, BGI-Shenzhen, Shenzhen 518083, China.
Kui
K
Wu
Liver Surgery Department, Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion (Fudan University), Ministry of Education, Shanghai 200032, China.
Xin
X
Zhang
Liver Surgery Department, Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion (Fudan University), Ministry of Education, Shanghai 200032, China.
Qi-Man
QM
Sun
Cancer Research Institute, Central South University, Key Laboratory of Carcinogenesis and Cancer Invasion, Ministry of Education, Changsha 410078, China.
Ya
Y
Cao
BGI-Shenzhen, Shenzhen 518083, China; BGI-Tianjin, Tianjin 300308, China.
Hong-Mei
HM
Zhu
Zhongshan Hospital Institute of Clinical Science, Shanghai Institute of Clinical Bioinformatics, Fudan University Center for Clinical Bioinformatics, Shanghai 200032, China.
Xiang-Dong
XD
Wang
BGI-Shenzhen, Shenzhen 518083, China; James D. Watson Institute of Genome Sciences, Hangzhou 310058, China.
Huan-Ming
HM
Yang
BGI-Shenzhen, Shenzhen 518083, China; James D. Watson Institute of Genome Sciences, Hangzhou 310058, China.
Jian
J
Wang
Liver Surgery Department, Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion (Fudan University), Ministry of Education, Shanghai 200032, China.
Zhao-You
ZY
Tang
BGI-Shenzhen, Shenzhen 518083, China; China National Genebank-Shenzhen, BGI-Shenzhen, Shenzhen 518083, China. Electronic address: houyong@genomics.cn.
Yong
Y
Hou
Liver Surgery Department, Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion (Fudan University), Ministry of Education, Shanghai 200032, China; Institute of Biomedical Sciences, Fudan University, Shanghai 200032, China; State Key Laboratory of Genetic Engineering, Fudan University, Shanghai 200433, China. Electronic address: fan.jia@zs-hospital.sh.cn.
Jia
J
Fan
Liver Surgery Department, Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion (Fudan University), Ministry of Education, Shanghai 200032, China; Institute of Biomedical Sciences, Fudan University, Shanghai 200032, China; State Key Laboratory of Genetic Engineering, Fudan University, Shanghai 200433, China. Electronic address: zhou.jian@zs-hospital.sh.cn.
Jian
J
Zhou
D006528:Carcinoma, Hepatocellular / Q000235:genetics* / Q000628:therapy*; D056915:DNA Copy Number Variations; D004252:DNA Mutational Analysis; D004273:DNA, Neoplasm / Q000097:blood / Q000235:genetics; D005260:Female; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics* / Q000628:therapy*; D008297:Male; D058990:Molecular Targeted Therapy; D009154:Mutation; D010802:Phylogeny; D017422:Sequence Analysis, DNA; D000073359:Exome Sequencing
D004273:DNA, Neoplasm
Circulating cell-free DNA (cfDNA); Hepatocellular carcinoma (HCC); Intratumoral heterogeneity (ITH); Target therapy; Targeted deep sequencing (TDS)