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Next generation sequencing-based analysis of mitochondrial DNA characteristics in plasma extracellular vesicles of patients with hepatocellular carcinoma.

ABSTRACT: Emerging evidence has revealed that mitochondrial DNA (mtDNA) is encapsulated in plasma extracellular vesicles (EVs). However, the characteristics of mtDNA in EVs from patients with cancer remain largely unexplored, which greatly limits its clinical application. Whole genome and capture-based sequencing found that EV mtDNA covered the whole mitochondrial genome. The medium fragment size in EV mtDNA was significantly larger compared with that in cell-free mtDNA [cfmtDNA; 159 vs. 109 base pairs (bp); P<0.001]. EV DNA appeared to have a higher mtDNA copy number compared with cfDNA. Of note, patients with hepatitis had >300-bp fragments in EV mtDNA compared with patients with hepatocellular carcinoma (HCC) and healthy controls. EV mtDNA fragments >300 bp in length exhibited a significantly higher proportion of EV mtDNA fragment ends than those that were ?300 bp in length in patients with hepatitis. The EV mtDNA copy number in patients with HCC and hepatitis were significantly lower compared with those in healthy controls. Furthermore, inconsistencies in the mtDNA heteroplasmic variant were observed among HCC tissues, plasma and EVs. In conclusion, EV mtDNA exhibited different characteristics among patients with HCC, hepatitis and healthy controls, indicating the potential value of EV mtDNA as a diagnostic biomarker that complements cfmtDNA.

SUBMITTER: Li Y

PROVIDER: S-EPMC7400774 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature

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Next generation sequencing-based analysis of mitochondrial DNA characteristics in plasma extracellular vesicles of patients with hepatocellular carcinoma.

Li Yijie Y Guo Xu X Guo Shanshan S Wang Yang Y Chen Lin L Liu Yang Y Jia Min M An Jiaze J Tao Kaishan K Xing Jinliang J

Oncology letters 20200708 3

Emerging evidence has revealed that mitochondrial DNA (mtDNA) is encapsulated in plasma extracellular vesicles (EVs). However, the characteristics of mtDNA in EVs from patients with cancer remain largely unexplored, which greatly limits its clinical application. Whole genome and capture-based sequencing found that EV mtDNA covered the whole mitochondrial genome. The medium fragment size in EV mtDNA was significantly larger compared with that in cell-free mtDNA [cfmtDNA; 159 vs. 109 base pairs (b ...[more]

PMID: 32782600

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Project description:BACKGROUND:Because imaging has a high sensitivity to diagnose hepatocellular carcinoma (HCC) and tissue biopsies carry risks such as bleeding, the latter are often not performed in HCC. Blood-derived circulating tumor DNA (ctDNA) analysis can identify somatic alterations, but its utility has not been characterized in HCC. MATERIALS AND METHODS:We evaluated 14 patients with advanced HCC (digital ctDNA sequencing [68 genes]). Mutant relative to wild-type allele fraction was calculated. RESULTS:All patients (100%) had somatic alterations (median?= 3 alterations/patient [range, 1-8]); median mutant allele fraction, 0.29% (range, 0.1%-37.77%). Mutations were identified in several genes: TP53 (57% of patients), CTNNB1 (29%), PTEN (7%), CDKN2A (7%), ARID1A (7%), and MET (7%); amplifications, in CDK6 (14%), EGFR (14%), MYC (14%), BRAF (7%), RAF1 (7%), FGFR1 (7%), CCNE1 (7%), PIK3CA (7%), and ERBB2/HER2 (7%). Eleven patients (79%) had ?1 theoretically actionable alteration. No two patients had identical genomic portfolios, suggesting the need for customized treatment. A patient with a CDKN2A-inactivating and a CTNNB1-activating mutation received matched treatment: palbociclib (CDK4/6 inhibitor) and celecoxib (COX-2/Wnt inhibitor); des-gamma-carboxy prothrombin level decreased by 84% at 2 months (1,410 to 242 ng/mL [normal: ?7.4 ng/mL]; alpha fetoprotein [AFP] low at baseline). A patient with a PTEN-inactivating and a MET-activating mutation (an effect suggested by in silico molecular dynamic simulations) received sirolimus (mechanistic target of rapamycin inhibitor) and cabozantinib (MET inhibitor); AFP declined by 63% (8,320 to 3,045 ng/mL [normal: 0-15 ng/mL]). CONCLUSION:ctDNA derived from noninvasive blood tests can provide exploitable genomic profiles in patients with HCC. IMPLICATIONS FOR PRACTICE:This study reports that blood-derived circulating tumor DNA can provide therapeutically exploitable genomic profiles in hepatocellular cancer, a malignancy that is known to be difficult to biopsy.

Project description:BackgroundAccumulating evidence implicates mitochondrial DNA (mtDNA) alleles, which are independent of the nuclear genome, in disease, especially in human metabolic diseases. However, this area of investigation has lagged behind in researching the nuclear alleles in complex traits, for example, in gout.MethodsNext-generation sequencing was utilized to investigate the relationship between mtDNA alleles and phenotypic variations in 52 male patients with gout and 104 age-matched male non-gout controls from the Taiwan Biobank whole-genome sequencing samples. Differences from a reference sequence (GRCh38) were identified. The sequence kernel association test (SKAT) was applied to identify gout-associated alleles in mitochondrial genes. The tools Polymorphism Phenotyping, Sorting Intolerant From Tolerant (SIFT), Predict the pathology of Mutations (PMUT), Human Mitochondrial Genome Database (mtDB), Multiple Alignment using Fast Fourier Transform (MAFFT), and Mammalian Mitochondrial tRNA Genes (Mamit-tRNA) were used to evaluate pathogenicity of alleles. Validation of selected alleles by quantitative polymerase chain reaction of single nucleotide polymorphisms (qPCR SNPs) was also performed.ResultsWe identified 456 alleles in patients with gout and 640 alleles in non-gout controls with 274 alleles shared by both. Mitochondrial genes were associated with gout, with MT-CO3, MT-TA, MT-TC, and MT-TT containing potentially pathogenic gout-associated alleles and displaying evidence of gene-gene interactions. All heteroplasmy levels of potentially pathogenic alleles exceeded metabolic thresholds for pathogenicity. Validation assays confirmed the next-generation sequencing results of selected alleles. Among them, potentially pathogenic MT-CO3 alleles correlated with high-density lipoprotein (HDL) levels (P?=?0.034).ConclusionThis study provided two scientific insights. First, this was the most extensive mitochondrial genomic profiling associated with gout. Second, our results supported the roles of mitochondria in gout and HDL, and this comprehensive analysis framework can be applied to other diseases in which mitochondrial dysfunction has been implicated.

Dataset Information

Next generation sequencing-based analysis of mitochondrial DNA characteristics in plasma extracellular vesicles of patients with hepatocellular carcinoma.

Publications

Next generation sequencing-based analysis of mitochondrial DNA characteristics in plasma extracellular vesicles of patients with hepatocellular carcinoma.

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