Hepatocellular carcinoma

Liver cancer is the second leading cause of cancer-related deaths globally and has an incidence of approximately 850,000 new cases per year. Hepatocellular carcinoma (HCC) represents approximately 90% of all cases of primary liver cancer. The main risk factors for developing HCC are well known and include hepatitis B and C virus infection, alcohol intake and ingestion of the fungal metabolite aflatoxin B1. Additional risk factors such as non-alcoholic steatohepatitis are also emerging. Advances in the understanding of the molecular pathogenesis of HCC have led to identification of critical driver mutations; however, the most prevalent of these are not yet druggable targets. The molecular classification of HCC is not established, and the Barcelona Clinic Liver Cancer staging classification is the main clinical algorithm for the stratification of patients according to prognosis and treatment allocation. Surveillance programmes enable the detection of early-stage tumours that are amenable to curative therapies - resection, liver transplantation or local ablation. At more developed stages, only chemoembolization (for intermediate HCC) and sorafenib (for advanced HCC) have shown survival benefits. There are major unmet needs in HCC management that might be addressed through the discovery of new therapies and their combinations for use in the adjuvant setting and for intermediate- and advanced-stage disease. Moreover, biomarkers for therapy stratification, patient-tailored strategies targeting driver mutations and/or activating signalling cascades, and validated measurements of quality of life are needed. Recent failures in the testing of systemic drugs for intermediate and advanced stages have indicated a need to refine trial designs and to define novel approaches

A novel biomarker TERTmRNA is applicable for early detection of hepatoma

<p>Abstract</p> <p>Backgrounds</p> <p>We previously reported a highly sensitive method for serum human telomerase reverse transcriptase (hTERT) mRNA for hepatocellular carcinoma (HCC). α-fetoprotein (AFP) and des-γ-carboxy prothrombin (DCP) are good markers for HCC. In this study, we verified the significance of hTERTmRNA in a large scale multi-centered trial, collating quantified values with clinical course.</p> <p>Methods</p> <p>In 638 subjects including 303 patients with HCC, 89 with chronic hepatitis (CH), 45 with liver cirrhosis (LC) and 201 healthy individuals, we quantified serum hTERTmRNA using the real-time RT-PCR. We examined its sensitivity and specificity in HCC diagnosis, clinical significance, ROC curve analysis in comparison with other tumor markers, and its correlations with the clinical parameters using Pearson relative test and multivariate analyses. Furthermore, we performed a prospective and comparative study to observe the change of biomarkers, including hTERTmRNA in HCC patients receiving anti-cancer therapies.</p> <p>Results</p> <p>hTERTmRNA was demonstrated to be independently correlated with clinical parameters; tumor size and tumor differentiation (P < 0.001, each). The sensitivity/specificity of hTERTmRNA in HCC diagnosis showed 90.2%/85.4% for hTERT. hTERTmRNA proved to be superior to AFP, AFP-L3, and DCP in the diagnosis and underwent an indisputable change in response to therapy. The detection rate of small HCC by hTERTmRNA was superior to the other markers.</p> <p>Conclusions</p> <p>hTERTmRNA is superior to conventional tumor markers in the diagnosis and recurrence of HCC at an early stage.</p

Aberrant reduction of telomere repetitive sequences in plasma cell-free DNA for early breast cancer detection.

Excessive telomere shortening is observed in breast cancer lesions when compared to adjacent non-cancerous tissues, suggesting that telomere length may represent a key biomarker for early cancer detection. Because tumor-derived, cell-free DNA (cfDNA) is often released from cancer cells and circulates in the bloodstream, we hypothesized that breast cancer development is associated with changes in the amount of telomeric cfDNA that can be detected in the plasma. To test this hypothesis, we devised a novel, highly sensitive and specific quantitative PCR (qPCR) assay, termed telomeric cfDNA qPCR, to quantify plasma telomeric cfDNA levels. Indeed, the internal reference primers of our design correctly reflected input cfDNA amount (R2 = 0.910, P = 7.82 × 10−52), implying accuracy of this assay. We found that plasma telomeric cfDNA levels decreased with age in healthy individuals (n = 42, R2 = 0.094, P = 0.048), suggesting that cfDNA is likely derived from somatic cells in which telomere length shortens with increasing age. Our results also showed a significant decrease in telomeric cfDNA level from breast cancer patients with no prior treatment (n = 47), compared to control individuals (n = 42) (P = 4.06 × 10−8). The sensitivity and specificity for the telomeric cfDNA qPCR assay was 91.49% and 76.19%, respectively. Furthermore, the telomeric cfDNA level distinguished even the Ductal Carcinoma In Situ (DCIS) group (n = 7) from the healthy group (n = 42) (P = 1.51 × 10−3). Taken together, decreasing plasma telomeric cfDNA levels could be an informative genetic biomarker for early breast cancer detection

Liquid biopsy - a review

Introduction and objective:Hepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths worldwide. Early detection is crucial for better outcomes. Traditional diagnostic methods, such as imaging and biopsies, often fail to detect HCC in early stages. Liquid biopsy, based on circulating free DNA (cfDNA) analysis, offers a promising, non-invasive approach, allowing frequent testing, addressing tumor heterogeneity, and reducing costs. Review methods:This article's databases were accessed through the WHO website, PubMed, and Google Scholar. A brief description of the state of knowledge:Early detection of HCC significantly improves survival. Biomarkers from cfDNA, including DNA fragment patterns, methylation markers (e.g., USP44), 5-hydroxymethylcytosine (5hmC), and digital PCR analysis, have shown potential in early-stage detection. Advanced cfDNA fragmentomics identifies tumor-specific DNA fragmentation patterns. Techniques like DELFI demonstrate high sensitivity (94%) and specificity (98%). Machine learning enhances cfDNA analysis by integrating multiple markers, improving accuracy in distinguishing cancerous from precancerous states. Combining methylation analysis with machine learning further addresses challenges of tumor heterogeneity. Summary:Studies highlight the high sensitivity and specificity of cfDNA biomarkers for HCC diagnosis, especially in high-risk groups like individuals with cirrhosis. Integrating technologies like 5hmC analysis and machine learning enables early diagnosis and treatment monitoring. These advancements represent a transformative step in cancer diagnostics, offering effective tools to improve patient outcomes

The landscape of mutation in plasma circulating tumor DNA sequencing as potential predictive biomarkers in hepatocellular carcinoma

Recently, the advancements in liquid biopsy have facilitated the techniques for clinical diagnosis and treatment monitoring of hepatocellular carcinoma. Genetic mutations are practical markers of distinguishing ctDNA, eliminating interference from cfDNA. Our study conducts a sequencing panel for ctDNA/cfDNA in patients with HCC and patients with benign liver disease, using NGS technology to identify mutant targets. After excluding germline and silent mutations, we obtain the mutation profiles of ctDNA in HCC, comprising solely functional mutations. Genetic mutations were found in free nucleic acid from 66.7% of HCC patients, while no mutant gene was in the control group. In our HCC cohort, ctDNA analysis was constituted of 49 genes and 91 exon mutations, with 15 genes (NCOR2, HGF, MECOM, ROBO1, MKI67, PEPN13, RANBP2, RELN, ALB, FAT4, KMT2B, MGAM, PAK5, PTPRB, ZFHX3) being identified for the first time in the ctDNA of HCC. NOCR2 and CTNNB1 were the highest frequent mutant genes in ctDNA, reaching 13.3%. The majority of these mutant genes were distributed in the classical molecular pathways of HCC, and the gene-enriched pathways showed a strong consistency between ctDNA and tDNA. A total of nineteen concordant mutations were detected in both ctDNA and matched tDNA, with 23 exons. We also found that the ratio of concordant mutation was highly correlated to tumor burden, especially vascular invasion. No mutations were found in the cfDNA of the control group, suggesting that mutant genes in ctDNA exhibit the potential to differentiate between benign and malignant liver diseases. Consequently, we further explored the diagnostic capabilities of ctDNA and discovered a great improvement in diagnostic accuracy of a combination of ctDNA mutation and AFP level over either one alone. Additionally, our research found the specific mutation-based gene set from ctDNA could contribute to predicting the prognosis of HCC patients. The mutation set screened according to TNM stages 2-4 consisted of twelve genes: NCOR2, ARID2, ERBB4, ERCC5, KMT2A, MSH6, PIK3CA, PIK3CG, POLQ, PEPRB, TERT, and TSC1. This analysis showed that the mutation of the NCOR2 gene was detected particularly frequently in HCC ctDNA and could therefore indicate a high potential for the prognosis of HCC patients. In total, these findings have demonstrated the potential of ctDNA mutation as a specific biomarker for liquid biopsy technique and deepened our understanding of the mutation profile in HCC. Our research supports the implications of mutations in ctDNA for precision medicine and illustrates the clinical prospect of ctDNA in the future

The clinical role of circulating free tumor DNA in gastrointestinal malignancy

Circulating cell-free DNA (cfDNA) is DNA released from necrotic or apoptotic cells into the bloodstream. While both healthy cells and cancer cells release cfDNA, tumors are associated with higher levels of tumor-derived circulating cell-free DNA (ctDNA) detectable in blood. Absolute levels of ctDNA and its genetic mutations and epigenetic changes show promise as potentially useful biomarkers of tumor biology, progression, and response to therapy. Moreover, studies have demonstrated the discriminative accuracy of ctDNA levels for diagnosis of gastrointestinal cancer compared with benign inflammatory diseases. Therefore, ctDNA detected in blood offers a minimally invasive and easily repeated “liquid biopsy” of cancer, facilitating real-time dynamic analysis of tumor behavior that could revolutionize both clinical and research practices in oncology. In this review, we provide a critical summary of the evidence for the utility of ctDNA as a diagnostic and prognostic biomarker in gastrointestinal malignancies

Sensitive droplet digital PCR method for detection of TERT promoter mutations in cell free DNA from patients with metastatic melanoma

Background: Currently mainly BRAF mutant circulating tumor DNA (ctDNA) is utilized to monitor patients with melanoma. TERT promoter mutations are common in various cancers and found in up to 70% of melanomas, including half of BRAF wildtype cases. Therefore, a sensitive method for detection of TERT promoter mutations would increase the number of patients that could be monitored through ctDNA analysis. Methods: A droplet digital PCR (ddPCR) assay was designed for the concurrent detection of chr5:1,295,228 C \u3e T and chr5:1,295,250 C \u3e T TERT promoter mutations. The assay was validated using 39 melanoma cell lines and 22 matched plasma and tumor samples. In addition, plasma samples from 56 metastatic melanoma patients and 56 healthy controls were tested for TERT promoter mutations. Results: The established ddPCR assay detected TERT promoter mutations with a lower limit of detection (LOD) of 0.17%. Total concordance was demonstrated between ddPCR and Sanger sequencing in all cell lines except one, which carried a second mutation within the probe binding-site. Concordance between matched plasma and tumor tissue was 68% (15/22), with a sensitivity of 53% (95% CI, 27%- 79%) and a specificity of 100% (95% CI, 59%-100%). A significantly longer PFS (p=0.028) was evident in ctDNA negative patients. Importantly, our TERT promoter mutations ddPCR assay allowed detection of ctDNA in 11 BRAF wild-type cases. Conclusions: The TERT promoter mutation ddPCR assay offers a sensitive test for molecular analysis of melanoma tumors and ctDNA, with the potential to be applied to other cancers

Tissue- and Serum-Associated Biomarkers of Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC), one of the leading causes of cancer deaths in the world, is offering a challenge to human beings, with the current modes of treatment being a palliative approach. Lack of proper curative or preventive treatment methods encouraged extensive research around the world with an aim to detect a vaccine or therapeutic target biomolecule that could lead to development of a drug or vaccine against HCC. Biomarkers or biological disease markers have emerged as a potential tool as drug/vaccine targets, as they can accurately diagnose, predict, and even prevent the diseases. Biomarker expression in tissue, serum, plasma, or urine can detect tumor in very early stages of its development and monitor the cancer progression and also the effect of therapeutic interventions. Biomarker discoveries are driven by advanced techniques, such as proteomics, transcriptomics, whole genome sequencing, micro- and micro-RNA arrays, and translational clinics. In this review, an overview of the potential of tissue- and serum-associated HCC biomarkers as diagnostic, prognostic, and therapeutic targets for drug development is presented. In addition, we highlight recently developed micro-RNA, long noncoding RNA biomarkers, and single-nucleotide changes, which may be used independently or as complementary biomarkers. These active investigations going on around the world aimed at conquering HCC might show a bright light in the near future

Circulating nucleic acids in plasma and serum (CNAPS): Applications in oncology

The presence of small amounts of circulating nucleic acids in plasma and serum (CNAPS) is not a new finding. The verification that such amounts are significantly increased in cancer patients, and that CNAPS might carry a variety of genetic and epigenetic alterations related to cancer development and progression, has aroused great interest in the scientific community in the last decades. Such alterations potentially reflect changes that occur during carcinogenesis, and include DNA mutations, loss of heterozygosity, viral genomic integration, disruption of microRNA, hypermethylation of tumor suppressor genes, and changes in the mitochondrial DNA. These findings have led to many efforts toward the implementation of new clinical biomarkers based on CNAPS analysis. In the present article, we review the main findings related to the utility of CNAPS analysis for early diagnosis, prognosis, and monitoring of cancer, most of which appear promising. However, due to the lack of harmonization of laboratory techniques, the heterogeneity of disease progression, and the small number of recruited patients in most of those studies, there has been a poor translation of basic research into clinical practice. In addition, many aspects remain unknown, such as the release mechanisms of cell-free nucleic acids, their biological function, and the way by which they circulate in the bloodstream. It is therefore expected that in the coming years, an improved understanding of the relationship between CNAPS and the molecular biology of cancer will lead to better diagnosis, management, and treatmen

Potential of modern circulating cell-free DNA diagnostic tools for detection of specific tumour cells in clinical practice

Personalized medicine is a developing field of medicine that has gained in importance in recent decades. New diagnostic tests based on the analysis of circulating cell-free DNA (cfDNA) were developed as a tool of diagnosing different cancer types. By detecting the subpopulation of mutated DNA from cancer cells, it is possible to detect the presence of a specific tumour in early stages of the disease. Mutation analysis is performed by quantitative polymerase chain reaction (qPCR) or the next generation sequencing (NGS), however, cfDNA protocols need to be modified carefully in preanalytical, analytical, and postanalytical stages. To further improve treatment of cancer the Food and Drug Administration approved more than 20 companion diagnostic tests that combine cancer drugs with highly efficient genetic diagnostic tools. Tools detect mutations in the DNA originating from cancer cells directly through the subpopulation of cfDNA, the circular tumour DNA (ctDNA) analysis or with visualization of cells through intracellular DNA probes. A large number of ctDNA tests in clinical studies demonstrate the importance of new findings in the field of cancer diagnosis. We describe the innovations in personalized medicine: techniques for detecting ctDNA and genomic DNA (gDNA) mutations approved Food and Drug Administration companion genetic diagnostics, candidate genes for assembling the cancer NGS panels, and a brief mention of the multitude of cfDNA currently in clinical trials. Additionally, an overview of the development steps of the diagnostic tools will refresh and expand the knowledge of clinics and geneticists for research opportunities beyond the development phases