New advances in precision medicine for hepatocellular carcinoma recurrence prediction and treatment
Authors: Silvia Vilarinho, Diego F. Calvisi
Journal: Hepatology (2014)

Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related death in men, affecting >700,000 individuals per year worldwide.1 Approximately 85% of HCCs arise in livers with chronic damage where the normal liver architecture has been replaced by nodular fibrotic tissue (cirrhosis). Around one-third of the patients with cirrhosis will develop HCC during their lifetime. Most of them will die from this complication since the 5-year survival remains a dismal 12% with currently available therapies.2 Specifically, there are five therapeutic options accepted in clinical guidelines: (1) surgical resection; (2) liver transplantation; (3) radiofrequency ablation (RFA); (4) transarterial chemoembolization (TACE); and (5) sorafenib. The first three treatments offer a high rate of potential cure, and resection is the first-line treatment option for patients with solitary tumors and well-preserved liver function, defined as a normal bilirubin level with either hepatic venous pressure gradient ≤10 mmHg or platelet count ≥100,000.3, 4 However, ∼70% of patients with HCC who undergo surgical resection develop tumor recurrence within 5 years postsurgery. Unlike most malignancies, HCC arises in a diseased organ; thus, all therapies must be considered with a view to the patient's liver function and performance status, and has two clear patterns of recurrence: (i) early dissemination (<2 years after curative treatment) of the primary tumor, and (ii) late recurrences or de novo tumors (>2 years after curative intervention) possibly due to new malignant clones.5 Based on our current understanding, cancer is a genetic disease that largely arises from somatic mutations. Several recent studies using massively parallel DNA sequencing technology have significantly contributed to uncover the mutational landscape of HCC.6-11 As of now, five major pathways have been identified as commonly altered by somatic mutations or homozygous gene deletions in HCC, namely, beta-catenin/Wnt signaling; p53/cell cycle control; chromatin remodeling; PI3K/AKT/Ras signaling; and oxidative and endoplasmic reticulum stress.7 Collectively, these studies have demonstrated that HCC is highly heterogeneous, not only at the clinical and pathological levels, but also at the molecular level. In this issue of Hepatology, Ahn et al.12 report the results of whole-exome sequencing (WES) and copy number variation analysis in 231 cases of resected frozen HCC specimens and matched nonneoplastic tissue, and provide its association with clinical outcomes: early tumor recurrence and survival. This is the largest study published to date analyzing the exomes of HCC normal/tumor pairs. In brief, WES consists of two major components. First, the exome capture and DNA sequencing pipeline, which includes the preparation of an exon enriched genomic DNA library, capture of library strands containing exons, and massively parallel DNA sequencing of the coding regions. Second, the computational pipeline, which encompasses nucleotide base calling, read mapping, alignment, and trimming, followed by single nucleotide variants (SNVs) and insertions and deletions (indels) calling, quality score variant filtering, and respective annotation. The exome corresponds to the coding part of the genome, and represents ∼1% of the entire genome. However, it is estimated that the exons harbor ∼85-90% of all mutations, with large effects on disease traits.13 Thus, WES represents a good balance between cost, amount of data, and sequencing duration. Nevertheless, a recent study reported telomerase reverse-transcriptase (TERT) promoter mutation (a variation in a noncoding region of the genome) as the most frequent genetic alteration in HCCs, arising from both cirrhotic and noncirrhotic livers.14 In the study by Ahn et al.,12 three-quarters of their cohort are hepatitis B virus (HBV)-associated HCCs, the median tumor size is 3.8 cm, 30% of patients had no vascular invasion, and 80% had advanced hepatic fibrosis (stage 3 or 4). At the genetic level, homozygous deletions of FAM123A, RB1, and CDKN2A genes and high-copy amplifications of MYC, RSPO2, CCND1, and FGF19 genes were found to occur often in this HCC collection. In addition, significantly mutated genes in the tumor as compared to matched nonneoplastic tissue (somatic mutations) included known HCC-related driver genes such as TP53, CTNNB1 (beta-catenin), and AXIN1. Interestingly, RB1 was newly identified as a significantly mutated gene in this study and the authors provide data to support that RB1 somatic protein-altering mutations, detected in <10% of patients, is an independent predictor of early recurrence of HCC after resection. This is the first demonstration that RB1 mutation in HCC showed negative prognostic significance for recurrence-free survival after hepatectomy. Furthermore, the authors replicated this finding in a previously published HCC cohort,11 in which RB1 somatic nonsynonymous mutations were only observed in 3.4% of cases (three subjects in a total of 88 patients with HCC). The present evidence further supports a crucial role of deregulation of RB1 in HCC. Besides somatic mutations, various molecular mechanisms have been shown to be responsible for inactivation of RB1 in human HCC, including loss of heterozygosity, promoter hypermethylation, loss of imprinting, deregulated phosphorylation, and accelerated degradation.15, 16 These multiple modes of RB1 inactivation imply the existence of a selective pressure toward the disruption of cell cycle control by RB1 in human HCC.15, 16 In accordance with the findings in human liver cancer, it has been recently demonstrated that hepatic-specific ablation of RB1 predisposes mice to chemically induced hepatocarcinogenesis.17 Of note, liver tumors developing in RB1-depleted mice exhibited a gene expression pattern closely resembling that of a human HCC subset characterized by elevated biological aggressiveness and patients' short survival,17 further substantiating the link between RB1 deregulation and poor HCC outcome. In addition, transgenic overexpression in the mouse liver of E2F1, the transcription factor whose pro-oncogenic activity is specifically inhibited by RB1, triggers HCC development.18 Thus, inactivation of RB1 seems to be an essential step for malignant hepatocytes to express their full oncogenic potential. Another important finding of the study by Ahn et al.12 is the identification of a subgroup of HBV-related HCCs characterized by FGF19 amplification. Intriguingly, this HCC subset is distinct from HCCs harboring CTNNB1 or AXIN1 mutations, and does not show nuclear accumulation of CTNNB1. Additional studies will be necessary to identify the crucial downstream effectors of FGF19 in human HCC. Nonetheless, as FGF19 and its receptor, FGF4R, have been demonstrated to be druggable targets,19 patients with HCCs harboring FGF19 amplification might significantly benefit from molecular-based therapies. Even when the first HCC nodule(s) are successfully treated, most of these patients remain at an increased risk for HCC “recurrence” due to their underlying liver disease. Together with other predictors of prognosis, such as the recently established 5-gene score,20 the finding by Ahn et al. may significantly contribute to stratify the risk of relapse following “curative” liver resection. In particular, based on their observation, more aggressive adjuvant approaches should be implemented in HCC cases harboring RB1 mutations (Fig. 1). However, currently there is no evidence to support the use of adjuvant therapy postresection in liver cancer. The STORM trial (clinicalTrials.gov identifier: NCT00692770) was designed to evaluate the efficacy and safety of sorafenib versus placebo in the adjuvant treatment of HCC after potentially curative treatment, which includes surgical resection. The trial has been completed and the official results will likely be publicly shared in the near future. Due to the molecular heterogeneity of human HCC, confirmed in the study by Ahn et al., it is envisioned that sorafenib administration would be of benefit only in some patients as adjuvant treatment. To significantly improve the effectiveness of targeted therapies in HCC, we are in high need of reliable biomarkers able to predict the response to a given drug/treatment. Encouraging results in this regard come from recent retrospective studies. For instance, it has been recently shown in an HCC collection that amplification of FGF3/FGF4 genes is frequently observed in responders to sorafenib.21 A similar, favorable response to sorafenib has been identified in human HCC displaying VEGFA amplification.22 Furthermore, treatment with tivantinib, a selective oral inhibitor of MET, has been found to be beneficial primarily in HCC cases with high c-MET expression.23 Although the identified molecular alterations associated with a favorable response to a given treatment occur in a relatively low number of HCC patients, they are the proof of principle that a personalized medicine can be achieved in HCC. Large-scale genomic and proteomic approaches will significantly contribute to identify the relevant biomarkers for effective molecularly targeted therapies in distinct HCC subsets. Proposed algorithm for patient stratification posthepatectomy of resectable HCC, based on Ahn et al.'s most recent findings. We believe that efforts that focus on the implementation of Precision Medicine approaches in HCC will probably dominate research in this field in the next several years. Silvia Vilarinho, M.D., Ph.D.1 Diego F. Calvisi, M.D.2 1Department of Internal Medicine Section of Digestive Diseases Yale University School of Medicine New Haven, CT 2Institut für Pathologie Universitätsmedizin Greifswald Greifswald, Germany